gluten free diet research

• Open access
• Published: 08 September 2023

Mapping the knowledge structure of a gluten-free diet: a global perspective

• Sa ’ed H. Zyoud 1 , 2 , 3 ,
• Muna Shakhshir 4 ,
• Amani S. Abushanab 1 ,
• Amer Koni 1 , 5 ,
• Moath Hamdallah 6 , 7 &
• Samah W. Al-Jabi 1  

Translational Medicine Communications volume  8 , Article number:  18 ( 2023 ) Cite this article

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Metrics details

A gluten-free diet (GFD) has become one of the most popular eating plans and is essential for managing gluten-related medical conditions, signs, and symptoms. Therefore, we performed a bibliometric analysis of the scientific literature on the GFD to describe the research landscape.

The Scopus database was searched for publications on the GFD from 1952 to 2021. A bibliometric analysis of the data was performed. VOSviewer software was used to perform visualization analysis, co-occurrence analysis, and publication trends in GFD.

A total of 3,258 publications were retrieved. In terms of publications, Italy ( n  = 468, 14.36%) led in the number of publications, followed by the USA ( n  = 398, 12.22%) and Spain ( n  = 274, 8.41%). The retrieved documents earned an average of 22.89 citations per document, for a total of 74,560 citations. Since 2001, there has been a gradual growth in the number of articles published, going from 23 to more than 370 in 2021. Using the mapping terms in the title/abstract a minimum of 50 times, 291 terms were divided into two main clusters: ‘ adherence to a gluten-free diet in celiac disease’ and ‘ improvement of the nutritional and sensory quality of gluten-free products.’

Conclusions

Over the past six decades, there has been a growing need for gluten-free bakery products and a noticeable increase in related publications. This study indicates that the “ improvement of the nutritional and sensory quality of gluten-free products” will remain a hotspot in this research field for upcoming years.

Introduction

According to the World Health Organization (WHO), a healthy diet is the best way to protect against malnutrition, cardiovascular diseases, diabetes and some types of cancer [ 1 ]. In addition, various worldwide dietary guidelines have reported that a healthy diet should be balanced and varied. However, some medical conditions, food allergies, or intolerance require a special diet to be considered healthy, such as the diet of Dietary Approaches to Stop Hypertension (DASH) for hypertensive patients, a renal diet for chronic kidney diseases, and a gluten-free diet (GFD) for intolerant patients or other medical reasons. These examples exclude any food components that could harm some people [ 2 ].

A GFD requires complete gluten exclusion, a protein complex soluble in ethanol in food products such as wheat, rye, barley, and triticale. There are many naturally available gluten-free (GF) food products, such as vegetables and fruits, dairy products, eggs, fish, and meat. In addition, GF alternatives manufactured specifically for wheat-based foods can be used as a GFD [ 3 , 4 ].

Many conditions require treatment with a GFD, including allergies and intolerances such as gluten sensitivity, wheat allergy, celiac disease (CD), and others. Allergies occur as an immunologic reaction in individuals upon ingestion of wheat proteins; CD is a chronic autoimmune disorder triggered by gluten ingestion, resulting in histological changes in the small intestine due to the autoimmune reaction. Individuals with CD experience malabsorption, other gastrointestinal and extraintestinal symptoms [ 5 , 6 ]. Strict adherence to the GFD is the only effective first-line treatment for CD that leads to duodenal mucosa healing along with the resolution of CD symptoms and signs of malabsorption of CD [ 6 ]. GFD is also an interesting therapeutic option for preventing and treating type 1 diabetes, depending on many promising animal studies. Gluten has multiple effects on the gastrointestinal tract, affecting the composition of the microbiota, inducing enteropathy in type 1 diabetes, and increasing intestinal permeability, all of which can be improved using a GFD [ 7 ].

Other studies shed light on the possible effect of the use of a GFD with probiotics in patients with major depressive disorders depending on the fact that a diet free of gluten has great potential to reduce the severity of depression symptoms in gluten-related disordered subjects [ 8 ]. Furthermore, supplementing a combination containing probiotics and a GFD might be crucial to inhibiting the immune-inflammatory cascade, which can regulate the central nervous system and digestive tract functions in patients with major depressive disorder [ 9 ].

While a GFD is recognized as productive for numerous conditions, patients must receive guidance and education about diet from qualified professionals. This is especially important due to the tendency of some variations of this diet to be high in carbohydrates and lipids while lacking in essential vitamins and fiber [ 10 ]. Furthermore, individuals adopting a GFD might encounter challenges related to excessive weight gain and obesity, as they often consume energy-rich gluten-free products [ 11 ]. Some gluten-free food items include quinoa, brown rice, almond flour, chickpea pasta, and gluten-free bread. Consequently, the food industry continuously expands its offerings by introducing innovative cereal-based gluten-free options. Unfortunately, a notable portion of gluten-free products falls short when compared to their gluten-containing counterparts, particularly with regard to nutritional composition and sensory attributes. Nutritionally, gluten-free breads tend to lack essential macronutrients and micronutrients such as protein, iron, calcium, and vitamins. This deficiency can lead to nutritional inadequacies for individuals with celiac disease [ 12 , 13 , 14 , 15 , 16 , 17 , 18 ].

However, it is important to acknowledge that not all gluten-free products are equal, and some may contain high levels of sugar or unhealthy fats [ 19 ]. Therefore, it is advisable to carefully review product labels and opt for items that are rich in nutrients and crafted from whole, unprocessed ingredients [ 20 ].

In recent years, the GFD has received substantial interest in a range of clinical research fields, including those indicated above, and an increasing number of studies are being published on the topic [ 21 , 22 , 23 , 24 , 25 ]. Therefore, we predicted that there might be numerous hotspots and research focuses in the area of the GFD. However, only a few attempts have been made to comprehensively evaluate the CD area's scientific output and current condition from a global perspective [ 26 , 27 , 28 , 29 ]. As a result, it is extremely important to shed light on the current state of GFD research and its application on a global scale, as well as prospective research trends and hotspots.

The bibliometric technique is the best method to analyze specific research trends that affect a given subject over time and to compare the contributions made between countries, institutions, and journals [ 30 , 31 ]. Therefore, our bibliometric study of the literature on this topic will help to solve research gaps and increase understanding of the most recent viewpoints of the GFD. Thus, bibliometrics were performed to investigate potential focuses within this area of research for a thorough analysis of the present state of global GFD research using data from Scopus. Overall, a bibliometric analysis can offer insightful information about global research patterns and the organization of the knowledge base surrounding gluten-free diets. This can be aided by making wise choices about the direction of research and the distribution of resources for researchers, decision-makers, and other stakeholders.

Materials and methods

Search strategies and data collection.

A bibliometric approach was applied. SciVerse Scopus was used to carry out the current study. Scopus is the most popular and authoritative database of research publications and citations, containing publications from journals with the highest global impact. The bibliometric indicators used in the current study were the same as those used in previously published studies [ 32 , 33 , 34 ]. To improve the accuracy of the retrieved data, the search was restricted to the title and abstract of publications in the Scopus database because if extended to all fields of search, such as keywords or the full text of publications, many irrelevant publications would be obtained (i.e., false positive data). Scopus does not consider keywords as authors. Instead, Scopus uses various algorithms to match documents to relevant keywords, which can sometimes lead to the generation of false-positive results. In addition, Scopus also uses indexed keywords such as "EMTRE drug terms," "EMTREE medical terms," and "Medline keywords." These keywords are pre-defined by Scopus and can limit the search results to a specific field, but they can also lead to false positives if the search terms are too broad or not relevant to the research question. Using this approach will result in a considerable improvement in the level of specificity achieved, while the level of sensitivity may suffer slightly as a result. "Gluten-free" was used as a search term to search Scopus titles for all prior years up to 31 December 2021. We chose the keyword "gluten-free" because we are more interested in gluten-free as a concept than related terms. The productivity of scientific research beyond 2021 was omitted from the analysis because this time was still available for new journal issues. All data extraction was performed on a single day (4 August 2022) to avoid daily Scopus updates. The search strategy was validated for the absence of false positive documents by scanning the abstract of the top 500 cited documents in the retrieved literature.

Bibliometric analysis

We conducted bibliometric analysis from the following aspects: countries' contribution to publications, citations, and H-Index, growth trends of publications, types of publications, and contributions of institutions, funding agencies, and journals.

Visualization analysis

VOSviewer software version 1.6.8 was used to map the retrieved literature [ 35 , 36 , 37 ]. VOSviewer was used to display and develop a network of terms derived from titles and abstracts. The terms were simultaneously separated into clusters based on co-occurrence analysis and color-coded by time course. Furthermore, an average appearance year was established to evaluate emerging topics and detect a developing trend.

Description of publications

Based on an analysis of the Scopus database, 3,258 GFD-related documents published between 1952 and 2021 were retrieved. Research articles ( n  = 2514, 77.16%) constituted the majority of the retrieved documents, followed by reviews ( n  = 237, 7.27%) and letters ( n  = 121, 3.71%). Other types of documents included 11.84% ( n  = 386).

Growth trends of publications

The first article on a GFD was published in 1952, entitled ‘Gluten-free diet in idiopathic steatorrhoea: report of a case’ [ 38 ]. Before 2001, this research area received little attention from researchers. Since 2001, there has been a gradual growth in the number of articles published, going from 23 to more than 370 in 2021 (Fig.  1 ).

The global number of publications related to the gluten-free diet from 1952 to 2021

Active countries and research collaboration

Authors from 91 different countries contributed to the retrieved documents. The top ten active countries are shown in Table 1 . The top ten countries contributed approximately 60.93% ( n  = 1984) of the documents retrieved. Italy ( n  = 468, 14.36%) was the leader in the number of publications, followed by the USA ( n  = 398, 12.22%), Spain ( n  = 274, 8.41%), and Brazil ( n  = 204, 6.26%). The mapping of the research collaboration of the leading active countries showed that the USA, Italy and Spain had the strongest research collaboration with other countries (Fig.  2 ).

International research collaboration among the main active countries (20 documents per country was established as a threshold ( n  = 36). The thickness of the connecting line represents the strength of research collaboration, whereas the node size is a relative representation of the research output

Top ten active institutions

Table 2 shows the top ten active institutions in research on the GFD. The top ten countries contributed to approximately 9.82% ( n  = 320) of the retrieved documents. Again, institutions from the European Union dominated the list. However, the Universita degli Studi di Milano , an Italian research institute, was the main active institution (n = 79, 2.42%), followed by the University College Cork- Ireland ( n  = 75, 2.30%), the Universidad de Valladolid- Spain ( n  = 61, 1.87%) and CSIC—Instituto de Agroquimica y Tecnologia de los Alimentos IATA- Spain ( n  = 60, 1.84%). The top ten list included two institutions from Italy, Spain, Finland, and Poland.

Analysis of research funding agencies

Table 3 lists the top ten funding agencies in terms of GFD publications. The European Regional Development Fund (EU) funded a large number of publications ( n  = 67; 2.06%). The European Commission (EU) came second ( n  = 56; 1.72%), followed by the Ministerio de EconomÃa y Competitividad (Spain) ( n  = 46; 1.41%).

Journal analysis

We identified the ten most productive journals in this field (Table 4 ). Nutrients ranked first in the number of publications ( n  = 104, 3.19%), followed by Lebensmittel-Wissenschaft & Technologie ( n  = 80, 2.46%) and the International Journal of Food Science and Technology ( n  = 69, 2.12%).

Citation analysis

The retrieved documents earned an average of 22.89 citations per document, for a total of 74,560 citations. 105 was the H-index of the retrieved documents. Five hundred forty-two of the retrieved documents did not have any citations, but 143 of the documents received 100 or more citations. In terms of the number of times they were cited, the top ten articles received a total of 4,214 citations [ 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 ]. There was a wide range in the total number of citations for these GFD publications, from 257 to 936 (Table 5 ).

Co-occurrence term analysis

The terms in the title/abstract were used a minimum of 50 times, and of the 41,738 terms used, 291 terms were divided into two main clusters: the green cluster focused on ‘adherence to a gluten-free diet in celiac disease’, while the red cluster focused on the results of ‘ improvement of the nutritional and sensory quality of gluten-free products’ (Fig.  3 ).

Cluster map based on analysis of terms appearing in titles or abstracts. The size of the circle indicates the occurrences of the terms, and the different colors indicate the variety of clusters. The map was created using VOSviewer software version 1.6.18

Future research direction analysis

Each term in Fig.  4 was colored differently by VOSviewer based on the average frequency with which it appeared in all the retrieved publications. Overlay visualization revealed that the yellow group represented recent research in this field, while the blue cluster represented relatively older research. Before 2014, the primary focus of this field was "adherence to a gluten-free diet in patients with celiac disease .” The “improvement of the nutritional and sensory quality of gluten-free products” was focused on later (after 2014), reflecting the most recent research advances.

A network visualization map of the analysis of terms in titles based on their frequency of appearance. Blue represents earlier occurrences of the terms, while yellow represents later occurrences. The map was created using VOSviewer software version 1.6.18

In this work, we used bibliometric analysis to illustrate the global research landscape of the GFD for all previous years. There were 3,258 items in total. Research on GFDs has blossomed and attracted the world's attention, particularly in Italy, the United States, and Spain. We identified a group of notable contributors, including institutions, journals, and funding agencies. Trends and hotspots in the field of research were outlined, and future developments were forecast.

The list of the ten most prolific countries published in the GFD includes countries unfamiliar with the ranking of scientific productivity in other disciplines [ 49 , 50 , 51 , 52 ]. Specifically, existing statistics revealed that Italy had been the leading contributor to GFD research, possibly because Italy has a rapidly expanding economy, which generates more finances to conduct research [ 53 ], contributing to the rise in GFD-related publications. In addition, Italy ranks first in the consumption of pasta and for pasta quality worldwide. Whole-grain pasta, along with bread and other starch from cereals, is an important food in Italy because it is the foundation of the Mediterranean diet inspired by the eating habits of people who live near the Mediterranean Sea. Pasta is also a part of Italian culture and its gastronomic history. Eating pasta meets both the nutritional and hedonistic and social needs linked to food. Therefore, manufacturers put more effort into researching how well GF products in supermarkets meet the needs of celiac people in terms of variety, prices, and safety to ensure adequate intake of nutrients and fiber necessary for well-being [ 54 ], which may explain why more research has emphasized the GFD over that time in Italy.

Increasing the allocation of financial resources toward research on GFD requirements can yield many substantial advantages for a particular geographic area. Enhanced financial support allocated toward study endeavours can facilitate a more comprehensive comprehension of gluten-related diseases, encompassing celiac disease and non-celiac gluten sensitivity. Consequently, this might offer prospective benefits and advantages. Understanding this concept is of utmost importance to achieve an exact diagnosis, effective therapy, and sustainable illness management [ 55 , 56 ]. Furthermore, the provision of increased research funding has the potential to facilitate the advancement of sophisticated diagnostic tools, biomarkers, and tests that can effectively detect and intervene in individuals who are at risk of developing gluten-related diseases [ 57 , 58 ]. Additionally, allocating resources toward research endeavors would contribute to a more comprehensive comprehension of gluten-related diseases, including celiac disease and nonceliac gluten sensitivity [ 59 ]. Additionally, it is worth noting that regions with a greater incidence of gluten-related diseases may encounter a substantial burden on their healthcare systems [ 60 ]. The allocation of resources toward research endeavors has the potential to deliver better preventative techniques, therefore justifying the total burden of these illnesses on the well-being of the general population. Additionally, it has the potential to facilitate the implementation of focused public health initiatives, endorsing the implementation of suitable eating practices and the cultivation of better lifestyle preferences [ 61 ]. The proficiency possessed by individuals can catalyze fostering innovation, facilitating cooperation, and promoting the flow of information, therefore enhancing the region's standing within the respective domain. Additionally, it has the potential to foster collaborations among academia, healthcare institutions, and industry, thereby facilitating the advancement of state-of-the-art therapies, diagnostic tools, and dietary products. Furthermore, research funding can contribute to educational initiatives to enhance public knowledge regarding gluten-related diseases, their associated symptoms, and the significance of proper dietary management [ 62 ]. This enables individuals to make well-informed decisions regarding their health. Additionally, this bibliometric study has the potential to facilitate the development of evidence-based dietary recommendations for individuals affected by gluten-related conditions [ 27 , 63 ]. Furthermore, the research findings derived from this bibliometric study can contribute to the existing pool of scientific knowledge on a global scale, potentially resulting in significant advancements in the comprehension of autoimmune disorders, gastrointestinal health, and nutritional science that extend beyond gluten-related diseases [ 3 , 64 ].

The terms that are used in the title, as well as the abstracts, represent the primary focus themes. The co-occurrence of GFD terms is an essential indicator that shows the trending subjects and advancements in a research field. The research on GFD can be broken up into the following aspects based on the terms: (1) ‘adherence to a gluten-free diet in celiac disease patients’ ; and (2) ‘ improvement of the nutritional and sensory quality of gluten-free products’.

One of the main hot topics in our study was ‘ adherence to a gluten-free diet in patients with celiac disease’. Strong research evidence indicates that all celiac patients should follow a strict GFD for life. Patients with CD should avoid permanently ingesting food or other substances containing wheat, barley, or rye, as a small amount of these substances will trigger the immune system reaction and damage the small intestine. Therefore, monitoring dietary changes should become part of routine celiac follow-up [ 65 ]. Despite diet being the only treatment for CD, a diet regimen can be difficult to maintain for economic, palatability, and social reasons. Specifically, diet can act as a source of bullying, isolating patients from social life and reducing their quality of life. Therefore, many researchers highlight the importance of joining patients with multiple support groups and encouraging the provision of ‘alternate diets’ in social settings and supermarkets as a key to adherence to a GFD [ 66 ]. On the other hand, a GFD cannot be considered a healthy diet for those who do not have CD, as it is low in fiber, protein, iron, folate, and other B vitamins [ 67 ]. Hence, all those confirmed to have CD should be referred to dietitians for education and to limit exposure to gluten cross-contact in home and restaurant settings [ 68 , 69 ].

Our research concluded that adherence to a GFD in patients with CD is among the hot topics globally, even though little is known about CD patients and adherence to a GFD in low- to middle-income countries. Therefore, different types of research are needed on this underestimated important issue [ 70 , 71 ]. A GFD is required as part of the treatment for CD; however, much research is being done on alternative pharmacological treatments due to the high psychological load associated with such a diet [ 72 ].

Another hot topic is the ‘ improvement of the nutritional and sensory quality of gluten-free products ’. This issue occurred as a major hotspot in our investigation since gluten-free food is essential for consumption by people with celiac disease, gluten intolerance, or wheat allergy, while the related products are those that do not contain gluten, a protein found in wheat, barley, and rye. Regular bread and bakery are the major parts of meals worldwide, while regular bread flour has been reported to have the highest amount of gluten [ 73 ]. Gluten is essential to provide structure and elasticity to the product; therefore, GF bakery products are considered a great challenge, as they are often unattractive, undesirable, unavailable, and approximately 160% more expensive than regular products [ 74 , 75 ].

Patients on a gluten-free diet may face sensory challenges, including issues with palatability, texture, and appearance of gluten-free foods. Clinicians should encourage patients to discuss these issues and provide guidance to overcome these barriers by suggesting recipes or alternatives that may improve the sensory experience of the diet [ 76 ]. In addition, the study suggests that there is a need for the development of high-quality, nutritious, and palatable gluten-free products. Clinicians and dietitians could provide feedback to manufacturers to develop products that meet the specific nutritional needs of patients while also addressing the sensory challenges they face [ 77 , 78 ].

In the last seven years, there has been an increase in the demand for GF products, which has required the production of high-quality and nutritious GF baked goods using a variety of available substitutes, such as almond and coconut flour, which are rich in protein, healthy fats, and fiber and are considered a friendly choice for diabetic patients due to their low glycemic index [ 16 , 77 , 79 ]. This has improved the quality of life for patients with gluten sensitivity [ 16 , 67 ]. Brown rice flour is also a good alternative, rich in micronutrients, fiber and complex carbohydrates that can provide sustained energy [ 14 ]. Pseudo grains include amaranth, quinoa, which are high in protein, especially with essential amino acids, minerals such as iron and magnesium, and fiber [ 80 ], and buckwheat, which is rich in fiber, protein, micronutrients and antioxidants [ 17 ]. Corn, montina, millet and teff flour have also been used as possible base ingredients. In addition, alternative hydrocolloids, enzymes, and fiber sources have been used to give superior properties [ 81 ].

Overall, gluten-free foods such as rice, corn, fruits, vegetables, legumes, beans and peas and GF products can be a healthy addition to one's diet, especially if they are consumed as part of a balanced diet that includes all food groups with a variety of nutrient-rich meals. However, GFD products remain challenging across the board and contain fewer sensory and nutritional ingredients than regular products. Therefore, producing affordable and high-quality GF products and labeling gluten are urgent issues that need to be considered in low- and middle-income countries to manage this public health problem related to gluten disorders [ 77 , 82 ].

It would be very helpful to have a thorough bibliometric analysis of the most cited papers, as this would shed light on the future development direction in this field. Due to the clinical importance of GFD and the significance of highly cited publications, we conducted a qualitative and quantitative study of the ten GFD articles that garnered the most citations. This was done in light of the importance of GFD and highly cited articles. Our objective was to improve researchers' understanding of research quality and trends, facilitate more effective use of classic publications on the GFD, and serve as a reference for future research in this area. The most-cited publication out of 936 total citations was "Malignancy in celiac disease—Effect of a gluten-free diet, " written by Holmes et al. and published in Gut journal in 1989 [ 45 ]. The findings of this study have shown that celiac patients who have been on a GFD for five years or more have no increased risk of developing cancer at all sites compared to the general population. However, the risk of mouth, pharynx, and esophagus (relative risk = 22.7, p  = 0.001) and lymphoma (relative risk = 77.8, p  = 0.001) increases in those who follow a GF or normal diet. In addition, a significant inverse correlation existed between increased GFD use and the morbidity rate. The findings suggest that a GFD may protect against celiac disease malignancy and further support the recommendation that all patients follow a strict GFD for the rest of their lives [ 45 ].

The article that was second in the list of citations, which was entitled ‘Effects of hydrocolloids on dough rheology and bread quality parameters in gluten-free formulations’, had a total of 664 citations; it was published in the Journal of Food Engineering in 2007 and was written by Lazaridou et al. [ 45 ]. This study thoroughly investigated various technological parameters and formulations to produce high-quality GF bread. In recent years, interest in GF bread has increased [ 18 , 83 , 84 , 85 ]. As a result, many different types of flour and starches, as well as a number of additives such as gums, enzymes, and soybean proteins, have been used to mimic the viscoelastic characteristics of gluten and improve the structure, texture, acceptability, and shelf life of GF bread.

The article with the third-most citations, titled "Recent advances in the formulation of gluten-free cereal-based products", was published in 2004 in Trends in Food Science and Technology by Gallagher et al. [ 48 ]. It received a total of 518 citations. This article provides an overview of the prevalence of celiac disease and recent developments in creating GF products through the utilization of hydrocolloids, starches, gums, and other innovative ingredient processes [ 48 ].

Strengths and limitations

This is the first study to identify and evaluate the properties of documents related to GFD. The bibliometric analysis conducted by VOSviewer is more comprehensive and objective than the traditional literature review. However, when interpreting our findings, certain limitations must be considered. First, world regions with journals that are not indexed in the Scopus database will be underrepresented. As a result, the presence of false negative results remains a possibility. A second limitation is the list of active countries and institutions, which must be carefully interpreted due to overlap in publications, research networking, and self-citations. Third, there is an inherent flaw in the fact that we only included publications on gluten-free in the article title. Our previous experience has shown that including search items in the abstract has a much lower sensitivity [ 49 , 50 , 52 , 86 ]. It would have only found a small number of additional papers, if any at all. This is something that we should have avoided. If we do not place any constraints on including phrases from the abstract in our search query, we will receive many articles that do not pertain to the topic we are interested in.

This study provided a comprehensive bibliographic analysis by reviewing research published over 60 years on the GFD from a global perspective using bibliometric analysis. The study has revealed that the majority of studies are related to research articles, and our findings demonstrated significant advances in GFD research and several hot topics during the previous decades. Italy supplied the most works, followed by the United States and Spain. Institutions from the European Union dominated the list with the most funded agencies. Diet is the only remedy for CD and is difficult to maintain; therefore, ‘adherence to a gluten-free diet in celiac disease’ has been found to be the most frequent occurrence issue, followed by ‘ improvement of the nutritional and sensory quality of gluten-free products ,’ which has gradually become the focus of GFD research. These findings may provide valuable indications for future research paths and scientific decision-making in this domain. The study highlights the importance of continuing research in this field. Clinicians may need to stay up-to-date with the latest research to provide patients with the most accurate and current information regarding gluten-free diets.

Availability of data and materials

All data generated or analyzed during this study are included in this published article. In addition, other data sets used during the current study are available from the corresponding author upon reasonable request.

Abbreviations

• Gluten-free diet

Gluten-free

World Health Organization

Dietary Approaches to Stop Hypertension

Celiac disease

Healthy diet [ https://www.who.int/news-room/fact-sheets/detail/healthy-diet ]

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The author thanks An-Najah National University for all administrative assistance during the implementation of the project.

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Sa ’ed H. Zyoud, Amani S. Abushanab, Amer Koni & Samah W. Al-Jabi

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Sa ’ed H. Zyoud

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Muna Shakhshir

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Zyoud SH designed the study, collected the data, analyzed the data, made major contributions to the manuscript’s literature search and interpretation, and drafted the manuscript; Shakhshir M contributed to the conceptualization and methodology of the study, was involved in the interpretation of the data, contributed to the manuscript writing, and made revisions to the initial draft; Abushanab AS, Koni A, Hamdallah M, and Al-Jabi SW were involved in the interpretation of the data, contributed to the manuscript writing, and made revisions to the initial draft; all authors provided a critical review and approved the final manuscript before submission.

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Zyoud, S.’.H., Shakhshir, M., Abushanab, A.S. et al. Mapping the knowledge structure of a gluten-free diet: a global perspective. transl med commun 8 , 18 (2023). https://doi.org/10.1186/s41231-023-00152-w

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Recent practical researches in the development of gluten-free breads

• Hiroyuki Yano   ORCID: orcid.org/0000-0002-0910-854X 1  

npj Science of Food volume  3 , Article number:  7 ( 2019 ) Cite this article

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Wheat bread is consumed globally and has played a critical role in the story of civilization since the development of agriculture. While the aroma and flavor of this staple food continue to delight and satisfy most people, some individuals have a specific allergy to wheat or a genetic disposition to celiac disease. To improve the quality of life of these patients from a dietary standpoint, food-processing researchers have been seeking to develop high-quality gluten-free bread. As the quality of wheat breads depends largely on the viscoelastic properties of gluten, various ingredients have been employed to simulate its effects, such as hydrocolloids, transglutaminase, and proteases. Recent attempts have included the use of redox regulation as well as particle-stabilized foam. In this short review, we introduce the ongoing advancements in the development of gluten-free bread, by our laboratory as well as others, focusing mainly on rice-based breads. The social and scientific contexts of these efforts are also mentioned.

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Introduction.

The aroma emanating from a bread bakery is unmistakably alluring. The flavor and crunchy texture of wheat breads sharpen our appetite and satisfy our basic human cravings for comfort as well as nutrition. Indeed, human beings are so enchanted by bread that it is much more than a “staple food”; it has been called “the staff of life”. Breadmaking has a long and fascinating story. 1 , 2 , 3 , 4 It is generally accepted that breadmaking dates back to the New Stone Age, from 8000 to 10,000 BC, and originated around the Fertile Crescent and consisted of emmer and einkorn wheat grains. 1 At first the grains were consumed as porridge. Then, grains that had been hand-crushed using knocking stones were mixed with water and baked on a heated stone with a cover of hot ash, resulting in an unfermented, flat bread. Later, around 6000 BC, people in southern Mesopotamia started using sourdough, 5 speculated to have been developed accidently in an abandoned mixture of flour and water. This first leavened bread dough, which contained fermentation gas, swelled up in the baking process. In ~3000 BC, the Egyptians improved bread by adding yeast, developing what would become the prototype of modern bread. They dehulled and milled wheat grains using saddle querns, the most ancient type of quern stones, 6 which were later replaced by rotary querns and are used even today. Breadmaking and beer production in Egypt are closely related and are considered evidence of a high degree of civilization. 7 Bread was made not only with flour prepared from raw grains, but sometimes also with malt (germinated grains). Moreover, water with a blend of cooked and uncooked malt was used in brewing. The mixture was strained free of husk before inoculation with yeast.

The precise origin of bread has still not been determined. Recent reports show archaeobotanical evidence that the origins of bread date back to 14,400 years ago. 8 Progress in archaeology will eventually clarify the origin of bread, along with some sense of how bread fits into the larger culture of ancient civilizations. Wheat bread is now one of the most representative food in the world. A unique property of wheat gluten realizes bread with high quality. However, some genetically predisposed people cannot eat wheat bread, because gluten causes harmful reactions to them. In this short review, we will summarize the gluten-dependent swelling mechanism of wheat bread and the recent scientific effort to make bread without gluten.

Modern wheat breadmaking

Simply stated, breadmaking is composed of three steps: mixing/sheeting, fermenting, and baking processes. 9 In the mixing process, wheat flour, water, yeast, sugar, salt, oil, and other components are mixed and kneaded. Here, the ingredients are blended homogeneously and hydrated, resulting in the development of the all-important gluten network. 10 Gluten is made from two major wheat proteins together comprising 85% of wheat endosperm protein: gliadin and glutenin. Kneading of wheat dough promotes the hydrogen bonding and disulfide cross-linking interactions of these proteins, eventually producing a viscoelastic and highly conformational protein network termed “gluten”. 11 Yeast grows fast in the dough, feeding on supplemental sugar, until it consumes all available oxygen. Then, it shifts metabolism from aerobic respiration to anaerobic fermentation. In the subsequent fermentation process, yeast generates fermentation gas, mainly composed of carbon dioxide and other components, such as ethanol:

In wheat dough, the gas is confined in the continuous “gluten matrix”, 12 which is composed of the viscoelastic gluten network and other components, such as starch granules and water (Fig. 1a ). Thus, in the beginning of the fermentation process, many small gas cells are produced throughout the dough, like so many small balloons. As the fermentation proceeds, each small gas cell grows bigger, and the dough rises. In the following baking process, the gas cell inflates further by heat, resulting in the expansion, namely, “oven spring” of the dough. 13 The starch molecules are gelatinized by heat, so that the gluten matrix forming the envelopes of the “balloons” become hardened, thus constructing the stable crumb framework. 14 Concurrently, the crust, or surface of the bread dough, is hardened as well as browned by the Maillard reaction between the sugars and amino acids. 15 Finally, the breadmaking is completed, emitting a fresh aroma. 16

Comparison of the swelling mechanism ( a ) and appearance ( b ) of fermenting wheat dough and additive-free, gluten-free (GF) rice batter

The preparation of ingredients, especially flour, is also a critical step. Wheat grain is composed mainly of three parts: the endosperm, germ, and bran. 17 In the endosperm, which is the major constituent of the polished grain, starch granules are embedded in a protein matrix. 18 Wheat flour is produced by grinding whole-wheat grains or polished ones mechanically. 19 Impact mills, such as hammer mills and pin mills, accomplish particle size reduction by exposing seeds to a set of rotating hammer or pins that fracture the seeds, while roller and stone mills compress the seeds between two hardened surfaces. 20 During the milling of wheat grains, a portion of the starch granules are mechanically damaged. 21 The extent of the damage depends on wheat variety (hard or soft type) as well as milling conditions. In the mixing and fermentation steps of breadmaking, damaged starch accelerates the absorption of water to the starch granules, resulting in the activation of local amylases, leading to the degradation of starch molecules into dextrin and maltose. 22 Consequently, yeast activity and the final bread volume is increased. However, excessive starch damage produces wet or sticky dough and bread with poor quality. Thus, control of flour quality in terms of the starch damage is critical in the milling industry. 23

In other words, intact and damaged starch granules each have their respective role in the making of wheat bread—and, as we will show, in rice-flour breads as well. In the case of wheat dough, intact starch granules constitute the gluten matrix, while damaged ones activate fermentation. Generally, the extent of starch damage in commercially available wheat flours is 10–15%. 19

Social demand for gluten-free food

Gluten intolerance.

While the unique viscoelastic property of gluten realizes wheat bread with high quality, some people choose to or must follow a gluten-free diet. Recent reviews well summarize the background and status quo of gluten-free diets, 24 , 25 so only the outline will be mentioned here. Gluten intolerance includes autoimmune celiac disease (CD), wheat allergy, and non-celiac gluten sensitivity (NCGS). Celiac disease is an autoimmune disorder caused by genetic as well as environmental factors. 26 In CD patients, ingestion of gluten leads to small intestinal damage, typically leading to malabsorption. Its prevalence in the United States and Europe is estimated to reach about 1%. Gluten protein has protease-resistant regions in its structure. 27 Digestion of gluten in the human gastrointestinal tract generates “pathogenic” peptides that occasionally reach the lamina propria, where the peptides are deamidated by local transglutaminase. 28 The modified gluten peptides have a higher affinity to human leukocyte antigen (HLA)–DQ2 as well as HLA–DQ8 molecules, 29 which are present only in the small percentage of people carrying the HLA–DQ2 or the DQ8 haplotype. 30 This bonding results in the presentation of the gluten peptides to T cells, thereby triggering further malignant immune response in those with CD. In addition, tissue transglutaminase cross-links covalently to gliadin molecules. The protein complexes with new epitopes are considered to trigger the primary immune response as well. Antibodies against tissue transglutaminase are characteristic of CD. 31

In contrast, food allergy to wheat is characterized by T helper type 2 (Th2) activation, which can result in immunoglobulin E (IgE) and non-IgE-mediated reactions. 32 The IgE-mediated wheat allergy reactions usually occur immediately after contact of wheat, and are characterized by the occurrence of wheat-specific IgE antibodies in serum. Ingestion of wheat causes food allergy, while inhalation of wheat causes respiratory allergy to genetically predisposed individuals. A food allergy to wheat may cause a life-threatening reaction, such as anaphylaxis and wheat-dependent, exercise-induced anaphylaxis. 33 In contrast, repetitive exposure to wheat flour may cause baker’s asthma or rhinitis, mostly characterized as occupational allergic diseases. 34 Non-IgE- mediated food allergy reactions to wheat usually occur hours or even days after ingestion of wheat products and are characterized by chronic eosinophilic inflammation of the gastrointestinal tract. 35 There is a variability among reports of wheat allergy prevalence due to the differences in the diagnostic criteria, methodology, age, and geography. 36 The prevalence of wheat allergy is estimated to be 0.9% in the United Kingdom (based on questionnaire response), 37 3.6% in the United States (based on measurement of anti-wheat-specific IgE antibodies), 38 and 0.2% in Japan (based on a combination of questionnaire-based examination, skin prick test, and serum omega-5 gliadin-specific IgE test). 39

Non-celiac gluten sensitivity (NCGS) is a recently proposed, increasingly recognized clinical condition in patients in whom celiac disease and wheat allergy have been ruled out. It is characterized by intestinal and extra-intestinal symptoms triggered by the ingestion of gluten-containing foods. 40 Due to the lack of a reliable biomarker, confirmation of an NCGS diagnosis relies only on a double-blind placebo-controlled (DBPC) gluten challenge. 41

So far, a gluten-free diet is the only safe and effective treatment for the above conditions of gluten intolerance. 32

Gluten-free “lifestylers”

Demand for gluten-free foods is not limited to the gluten-intolerant population. Although it is not clear whether a gluten-free diet is beneficial for one’s health, some gluten-tolerant consumers believe that gluten-free food products are simply healthier. 42 , 43 This can be partly explained by a kind of “health halo” effect, making consumers believe that products with “free-from” label are healthier overall. 44 Besides, some popular books by bestseller authors, athletes, and celebrities have encouraged a gluten-free diet. An online questionnaire survey demonstrated that 41% of non-celiac athletes, including Olympic medalists, follow a gluten-free diet 50–100% of the time, and that adoption of the diet in most cases was not based on a medical rationale and may have been driven by the perception that gluten removal provides health benefits and an ergogenic edge. 45 Approximately 13% of young adults are reported to value gluten-free food; this population is more likely to engage in other healthy dietary behaviors, such as eating breakfast daily and eating more fruits/vegetables while simultaneously pursuing questionable behaviors, such as using diet pills to control weight. 42

A double-blind randomized study found that the supposed health benefit of a gluten-free diet has no evidence base in individuals who do not have celiac disease or irritable bowel syndrome, demonstrating that gluten is unlikely to be the culprit for gastrointestinal symptoms or fatigue in otherwise healthy individuals. 43 Moreover, commercially available gluten-free food products tend to contain ingredients with less diversity and less nutritional quality compared with their gluten-containing counterparts. 46 , 47 Other studies claim that despite recent improvements in the formulation and availability of gluten-free foods, they still are less available and more expensive than gluten-containing versions. 48 They generally have adequate levels of fiber and sugar, but lower levels of protein and higher levels of fat compared with their gluten-containing counterparts. 48 Also, very few gluten-free foods are fortified with micronutrients. 48

The gluten-free products market was valued at USD 4.18 billion in 2017 and this is projected to reach USD 6.47 billion by 2023, at a compound average growth rate of 7.6% during the forecast period. 49 The gluten-free diet has become the mainstream rather than just supporting a niche market.

Developments of gluten-free breads

As mentioned in the previous sections, demand for the development of gluten-free foods is growing. 50 Much of the focus is on bread products, as bread is an important staple food. Rice is considered a suitable substitute for wheat, as it is available worldwide and is less allergenic. So, development of rice-based gluten-free breads is the main topic of this review. It is not easy to make bread without using wheat flour or gluten, as bread’s quality depends on the properties and functionality of gluten. 25 In a wheat flour dough, the gluten matrix, composed mainly of the protein network of gluten, starch granules, and water (Fig. 1a ), encloses the fermentation gas, making small “balloons”. Thus, the dough rises as the fermentation proceeds. On the other hand, hydration of flour from gluten-free cereals, such as rice, results in a runny “batter” rather than viscoelastic “dough” as their proteins do not possess the network-forming properties typically found in gluten. 51 Therefore, the fermentation gases rise to the surface while starch granules and yeast settle. 52 Generally, a gluten-free batter without a thickening agent, such as hydrocolloids, becomes foamy. 53 , 54

Several efforts have been made in the development of gluten-free breads. Typical gluten-free breads contain hydrocolloids (e.g., xanthan gum, guar gum, etc.) which increase the viscosity of the liquid phase, keeping the starch granules, yeast, and gas bubbles suspended in the fermentation process. 52 , 55 The subsequent baking process gelatinizes the starch and hardens around the hydrocolloid membrane surrounding the air bubbles to set the crumb structure. As a surface-active hydrocolloid, hydroxypropyl methylcellulose (HPMC) behaves somewhat differently. It has hydrophobic methyl ester/hydroxypropyl groups in addition to hydrophilic cellulose regions. Thus, HPMC stays at the gas/liquid interface, uniquely stabilizing the bubbles and preventing coalescence. 52 , 56 Moreover, as HPMC is thermoreversible, 57 it also helps harden the bubble membrane in the baking process. 58

Another recent approach includes enzymatic treatment of gluten-free batter. 51 Transglutaminase (EC 2.3.2.13) catalyzes the acyl-transfer reaction between primary amino groups on protein-bound lysine residues and γ-carboxyamide groups on protein-bound glutamine residues. 59 Thus, transglutaminase is capable of introducing covalent cross-links between proteins. 60 The protein cross-linking ability has been shown to transform weak gluten into a strong gluten, with measurable effects on rheological behavior. 61 The addition of transglutaminase, along with HPMC, to a gluten-free rice batter resulted in its improved elastic and viscous behavior, as well as a higher specific volume and softer crumbs in the resulting bread. 62 The improvement in the viscoelastic properties of the rice batter appeared to be associated with the enhanced capability of the rice flour to retain the carbon dioxide produced during proofing. The quantitative decrease of free amino groups of proteins suggested that this improvement was due to the cross-linking of protein, that is, the generation of a gluten substitute, supplementing the role of HPMC in the baking of rice bread. 62 Microstructure analyses of a rice-based bread fortified with skim milk or egg powder using confocal laser-scanning microscopy (CLSM) verified that addition of transglutaminase promoted the formation of a protein network in the gluten-free bread that mimicked the gluten network in wheat breads. 63 The networking efficiency of transglutaminase depends on both the correct protein substrates and the level of enzyme addition. Thus, formation of the appropriate protein network under the right conditions should improve the overall quality of gluten-free bread by enhancing loaf volume and crumb characteristics, as well as appearance.

Improvement of the gas-retaining capability of gluten-free batter using protease, a seemingly paradoxical strategy for cross-linking, is also in progress. Protease has actually been used to weaken wheat dough by cleaving a portion of the gluten network. 64 However, treatment of a brown rice batter with bacterial protease improved bread quality by significantly increasing the specific volume while decreasing crumb hardness and chewiness. 65 CLSM images of the bread crumbs suggested that the gelatinized starch phase was the main structure component in the protease-treated bread. Thus, protease may partially degrade the large macromolecular protein complex embedding starch granules, 66 , 67 resulting in improved continuity of the starch phase as well as better rheological properties of the batter. Treatment of rice batter with a protease from Aspergillus oryzae increased its viscosity and resulted in bread with a high specific volume. Optical microscopic observation of the batter suggested that partially degraded protein, possibly glutelin, and starch granules formed aggregations containing voids. 54 This fine network of interlinked protein‒starch aggregates resulted in gas cell stabilization. 54 Proteases are mainly categorized into four classes based on the catalytic mechanism: metallo, serine, cysteine, and aspartyl proteases. 68 Comparative analyses of the proteases 69 , 70 demonstrated that metallo, serine, and cysteine proteases, but not aspartyl protease, are effective additives for improving the quality of gluten-free rice breads.

Application of the redox regulation

Addition of glutathione, a ubiquitous natural peptide, facilitated the deformation of rice batter, thus increasing its elasticity in the early stages of bread baking and increasing the volume of the resulting bread. 53 , 71 Below, we would like to introduce briefly how glutathione can be used in making gluten-free rice bread. The disulfide bond is a cross-link between two cysteine residues and plays an important role in the structure/function of proteins. 72 Redox regulation, control of reduction/oxidation of the disulfide bonds, as well as phosphorylation are the two major post-translational modifications of proteins. 73 Thioredoxin (Trx), 74 a small 12 -kDa protein, and glutathione, 75 a natural tripeptide, play central roles in the redox-dependent regulatory mechanisms.

Trx reduces the disulfide bond of its target protein specifically. In the reactions below, oxidative status is abbreviated as “OX” and reduced status is abbreviated as “RED”:

Glutathione (GSH) is a tripeptide with a free SH group. Two molecules of glutathione occasionally cross-link with an intermolecular disulfide bond to make “oxidized” glutathione (GSSG). Glutathione’s reaction occasionally entails glutathionylation (GL): 76

Redox regulation has been a key target of Dr. Bob Buchanan’s laboratory, University of California, Berkeley, after he clarified the Trx-dependent regulatory mechanism in photosynthesis. 77 , 78 In the proteomic analyses of plant biochemistry mostly performed by the Berkeley group, 79 , 80 , 81 , 82 we have found that redox regulation occurs in many aspects of plant life and plays critical roles in plant biology: seed germination/maturation, photosynthesis, defense against oxidative stress/pathogens, and others. 83 Then, thinking in the opposite direction, modification of the disulfide bonds in biology, that is, artificial activation of the redox regulatory mechanism, might lead to the production of a new, useful plant. Following this hypothesis, overexpression of Trx in plants was first tried in the starchy endosperm of barley. 84 The transformant germinated earlier than the wild type. Also, enzymes in charge of starch mobilization appeared earlier. As fast germination of barley seeds reduces the production cost and improves the quality of beer, 85 the results suggest the practical utility of Trx transformants. Conversely, underexpression of Trx in white wheat seed has been tried. White wheat has received increasing attention, as it is naturally white and needs no bleaching for uses, such as breadmaking. However, white wheat grains tend to germinate on the spike before harvest. 86 The preharvest sprouting (PHS) reduces the crop yield as well as the quality of the seeds and the flour. Rainfall or high humidity in the grain-filling season leads to PHS, and causes farmers significant financial losses. 87 Suppression of Trx in the starchy endosperm led to improved PHS resistance in the transformants 88 without affecting the crop yield or flour quality. 89

These two findings reported by the Berkeley group are the first discovery that control of Trx expression, that is, artificial redox regulation, affects the physiological processes of plants. Although risk assessment of genetically modified organisms (GMOs) is a critical issue, 90 the characteristics of these and other trial model plants provide the possibility of the industrial application of redox regulation. 91

More recently, we have sought to use this strategy to enable rice batter to confine fermentation gas. Glutathione was added to rice batter in an attempt to transform the intramolecular disulfide bonds of rice proteins into intermolecular disulfide bonds and eventually form a gluten-like network. Both reduced glutathione (GSH) and oxidized glutathione (GSSG) were found to be successful in swelling gluten-free rice batter and bread. 53 , 71 However, contrary to our expectations, analysis of the proteins revealed that no gluten-like protein network was formed. In contrast, microstructure and biochemical analyses suggested that glutathione cleaved the disulfide-linked glutelin polymers embedding the starch granules. The glutelin polymer has been suggested to work as a hindrance to the absorption of water by starch molecules when water is added to a rice flour; 66 glutathione may fray this barrier to make the batter more consistent and viscous, thereby improving its gas-holding capability in the fermentation process, 53 as is the case with protease-treated rice batter. 65 Although the number of its applications in food processing has been limited so far, 91 glutathione appears to be a promising tool for developing food with new properties. Glutathione is usable as a food ingredient in the United States 92 and some east Asian countries. For example, glutathione-based oral dietary supplements have been accorded the status of a Generally Recognized as Safe (GRAS) constituent with Section 201(s) of the Federal Food, Drug, and Cosmetic Act of the US Food and Drug Administration (US-FDA). 93

On the other hand, usage of glutathione for food has some limitations. First, glutathione is not usable as a food in all countries. In Japan, for instance, it is recognized as medicine, and cannot be incorporated as a food additive. 94 Second, GSH-added rice batter has been shown to yield a slight amount of hydrogen sulfide (0.43 ppm) and methyl mercaptan (0.106 ppm) in the headspace gas of the bread. 71 Generation of hydrogen sulfide in heated meat or purified GSH is well known; 95 indeed, a slight amount of hydrogen sulfide contributes to the pleasant aroma of cooked meat 96 and rice. 97 Usage of GSSG in breadmaking instead of GSH significantly reduced the generation of these sulfur compounds, 71 and sensory evaluation demonstrated that the aroma of GSSG-added rice bread was almost equivalent to that of non-added bread. 98 However, we sought to develop rice bread without glutathione or any other additives.

In the process of developing glutathione-added rice bread, we found that the control sample, that is, “non-added bread”, occasionally swelled in fermentation. Although it collapsed mostly in the following baking process, we expected that if optimal conditions could be found, we could make an additive-free, gluten-free rice bread from solely the basic ingredients: rice flour, water, yeast, sugar, salt, and oil.

Additive-free, gluten-free rice bread

The development of additive-free, gluten-free rice bread has taken a trial-and-error rather than a strategic approach. 99 , 100 First, we tried several commercially available rice flours and found that flours with low-starch damage (<5%) were the most suitable. The physical property of the gluten-free rice batter appeared quite different from the familiar viscoelastic wheat dough. It had an appearance and texture of a slurry with low viscosity. So, lots of “cooking tips” have been discerned for the breadmaking process. For example, as rice batter tends to make lumps, we paid attention in the mixing procedure to avoid lumps. Also, the dried yeast needs to be dissolved completely. Generation of bubbles of different sizes due to heterogeneous distribution of dried yeast may result in their coalescence 101 and a sudden shrinkage of the batter in the fermentation process. The breadmaking processes, i.e., mixing of the batter, fermentation and baking, as well as tips for successful making in the respective processes, are mentioned in a later section.

To clarify how the gluten-free batter swells without additives, we sought to investigate the microstructure of the fermenting batter. The fermenting batter appeared like a meringue and was quite different from wheat dough, which is so viscoelastic that its full mass can be lifted with a scoop (Fig. 1b ). As it was not easy to freeze the fragile batter without destroying the delicate structure, a sectioned specimen for microscope observation could not be made. Instead, freshly made batter was sandwiched between a microscope slide and a coverslip and the batter was left at room temperature to ferment there. Optical microscopic observation revealed the microstructure: bubbles covered by starch granules (Fig. 2 ). The structure was entirely different from that of the typical wheat dough, in which gas cells are surrounded by the gluten matrix made by a network of gluten protein and starch granules. 102 In contrast, it had a similar structure to a “particle emulsion” 101 in which rice granules stabilize the interface between oil and water (Fig. 2 ). 103 Thus, it was suggested that the bubble observed in an additive-free, gluten-free rice batter had the structure of a “particle foam” (Figs. 1a , 2 ). 101

Explanatory figure of particle emulsion/foam. Adapted from refs. 99 , 100 . Scale bar: 30 µm. Copyright (2017), with permission from Elsevier

The hypothetical mechanism is illustrated in Fig. 2 . Generally, oil and water do not mix. However, when they are mixed well in the presence of a detergent, microscopic oil droplets covered by detergent molecules disperse throughout water. This is a classic emulsion. Likewise, aeration of water in the presence of detergent results in a foam. A small amount of air is surrounded by a thin film of water, in which detergent molecules stabilize the boundary.

At the beginning of the 20th century, solid particles were found able to adsorb onto the interface between oil and water, and play a similar role to that of detergent molecules. 104 , 105 This is called a “particle-stabilized emulsion” or “particle emulsion”. Starch granules of native rice, maize, wheat, 103 quinoa, 106 high-pressure treated corn starch granules, 107 chemically modified waxy maize and tapioca, 108 as well as rice starch granules 109 have been reported to form particle emulsions. A particle-stabilized foam occurs in the same manner. Particle emulsions/foams have received renewed attention during the past decade, as recent advancement in nanoparticle technology accelerates research trends. 110 , 111 Moreover, such foams have potential applications in a wide variety of industries, including foods, pharmaceuticals, and cosmetics. One of the key advantages of the mechanism for foodstuff applications is that microparticles of biological origin, such as starch granules, cellulose, or protein particles, work as stabilizers. 101 Our report showed for the first time that rice starch granules stabilize particle “foam” in an additive-free, gluten-free rice batter. 99

The breadmaking processes and tips for the successful gluten-free breadmaking from rice flour are summarized in Fig. 3 . In the early stage of fermentation, yeast produces fermentation gas, composed mainly of carbon dioxide and alcohol. Ordinarily, the batter cannot hold the gas and becomes foamy. 53 , 54 However, if rice flour with low-starch damage is used and breadmaking is performed with the right conditions, the fermentation gas is trapped in the batter. 99 Thus, small bubbles appear throughout the batter. The small bubbles are particle foams in which fermentation gas is surrounded by starch granules. As the fermentation proceeds, the fragile bubbles gradually grow bigger, making the whole batter rise. Here, it is critical to keep the temperature stable, as fragile bubbles tend to burst in fluctuating temperatures. In the late stage of fermentation, the swollen bubbles should be heated rapidly to make the starch granules gelatinize, that is, to solidify the bubble walls. The most swollen bubbles are the most fragile, so rapid heating is the key.

Summary of the procedures for making additive-free rice bread and “cooking tips” for each step. Adapted from ref., 100 with permission

The overall process resembles the synthesis of a polyacrylamide hydrogel, in which modified nanoparticles stabilize an air/water (acrylamide solution) emulsion, and the macroporous structure is fixed by thermal-induced polymerization. 112

We have investigated several commercially available rice flours and found that rice flours with less starch damage (<5%) make bread with a higher specific volume. 99 Higher starch damage tends to facilitate greater absorption of water by starch granules. 113 The hydrophobicity/hydrophilicity ratio determines the aptitude of starch granules to form particle foam. 114 Thus, to prevent destabilization of the fragile bubbles in the fermentation process, it is important to maintain the proper hydrophobicity/ hydrophilicity ratio. Our success in making bread using flour with less starch damage, that is, less water absorption, seems consistent with the hypothetical mechanism. In this context, reduction of surface tension by hydrophobic treatment of rice starch granules was successful in making a stable particle emulsion. 108 , 109

From another point of view, if rice starch granules are capable of constituting a particle foam, they should have the ability to mimic the function of detergents, that is, to reduce the surface tension of water. Starch granules with less starch damage (4.7 w/w%) effectively reduced the surface tension of water from 73 to 35 mN/m. In contrast, starch granules with higher starch damage (9.8 w/w%) were not as effective, reducing the surface tension to only 47 mN/m. 99

Starch granules show emulsion-forming ability by stabilizing the water/tetradecane interface. 108 So, similar experiments were conducted using starch granules with low- and high-starch damage (Fig. 4 ). Both starch granules made stable water/tetradecane emulsions (Fig. 4a ). However, the microstructures of the emulsions were somewhat different (Fig. 4b ). Optical microscopic analyses of the emulsions showed that starch granules with less starch damage (LD) covered the oil droplets densely. In contrast, in the case of rice granules with higher starch damage (HD), swollen granules were occasionally seen, and the oil droplets were not covered completely. Thus, rice granules with low-starch damage demonstrated better particle-emulsion-forming ability compared with the high-starch-damage counterparts. This was consistent with the observation that rice starch granules with low-starch damage were suitable for constructing particle foam, that is, to make additive-free rice bread.

a Water/tetradecane emulsions formed by starch granules at different rice flour concentrations. From left to right: control (no flour), addition of rice flour with low-starch damage (20% w/w, 50% w/w), as well as high-starch damage (20% w/w, 50% w/w). b Optical microscopic analyses of the emulsion. Rice flour with low- (LD) and high- (HD) starch damage was compared. Adapted from ref. 99 Scale bar: 100 µm for ×100, and 30 µm for ×400, respectively. Copyright (2017), with permission from Elsevier

All these three observations support the hypothetical particle foam theory. Verification studies are in progress in our lab.

Several approaches in the development of gluten-free bread by our own laboratory and others have been introduced in this review, together with the social and scientific context of these efforts. The research is aimed to improve the quality of life of celiac disease or wheat allergy patients. Better bread quality (flavor, texture, and volume), reduced production cost, and wider availability are all important issues. 115 For example, so far, rice bread lacks the mouth-watering aroma of freshly baked wheat bread. It is not clear whether this is inevitable or whether a better selection of ingredients or an improved breadmaking procedure could lead to improvement of the aroma and flavor of rice bread, such that it becomes comparable with that of wheat bread. Besides, rice breads tend to be sticky compared with wheat bread. Also, gelatinized rice starch tends to retrograde faster, 116 so the bread is prone to become stale and hardened faster, 117 resulting in a shorter shelf life. 118 Using rice varieties with intermediate amylose content and a low water absorption index may give superior crumb properties. 119

Recent wide availability of household breadmaking countertop appliances has prompted our laboratory and others to develop gluten-free bread recipes suitable for these machines. Providing specific ingredients, such as fitted rice flour sold along with the breadmaker, may help consumers experience success in making custom gluten-free bread at home. Improving the machines by incorporating an induction-heating (IH) system may be suitable for making “particle-foam” type rice bread, as an IH system guarantees stable temperature control in fermentation as well as rapid heating in the baking process. 120 Addition of micronutrients and functional food ingredients is also an important theme. Further studies may thus improve the bread quality to be comparable to that of wheat bread and to improve the quality of wheat-sensitive patients’ life through providing a satisfactory diet.

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Acknowledgements

We appreciate Dr. Bob Buchanan and Dr. Peggy Lemaux, University of California, and Dr. Wallace Yokoyama and Dr. James Pan, USDA, for useful discussions. Dr. Shigeru Kuroda is also appreciated for his encouragement throughout this work.

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Yano, H. Recent practical researches in the development of gluten-free breads. npj Sci Food 3 , 7 (2019). https://doi.org/10.1038/s41538-019-0040-1

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Gluten: A Benefit or Harm to the Body?

The quick answer is that it can be either, but it all depends on the individual.

What is Gluten?

Gluten is a protein naturally found in some  grains  including wheat, barley, and rye. It acts like a binder, holding food together and adding a “stretchy” quality—think of a pizza maker tossing and stretching out a ball of dough. Without gluten, the dough would rip easily.

Other grains that contain gluten are wheat berries, spelt, durum, emmer, semolina, farina, farro, graham, khorasan wheat, einkorn, and triticale (a blend of wheat and rye). Oats—though naturally gluten free—often contain gluten from cross-contamination when they are grown near, or  processed in the same facilities as the grains listed above. Gluten is also sold as wheat gluten, or seitan, a popular vegan high-protein food. Less obvious sources of gluten include soy sauce and modified food starch, however gluten-free options of these products are available and labeled as such to comply with the U.S. Food and Drug Administration’s gluten-free labeling rule.

Gluten and Health Benefits

Gluten is most often associated with wheat and wheat-containing foods that are abundant in our food supply. Negative media attention on wheat and gluten has caused some people to doubt its place in a healthful diet. There is little published research to support these claims; in fact published research suggests the opposite.

In a 2017 study of over 100,000 participants without celiac disease, researchers found no association between long-term dietary gluten consumption and heart disease risk. [1] In fact, the findings also suggested that non-celiac individuals who avoid gluten may increase their risk of heart disease, due to the potential for reduced consumption of whole grains.

• Many studies have linked whole grain consumption with improved health outcomes. For example, groups with the highest intakes of whole grains including wheat (2-3 servings daily) compared with groups eating the lowest amounts (less than 2 servings daily) were found to have significantly lower rates of heart disease and stroke, development of type 2 diabetes, and deaths from all causes. [2-5]

Gluten may also act as a prebiotic, feeding the “good” bacteria in our bodies. Arabinoxylan oligosaccharide is a prebiotic carbohydrate derived from wheat bran that has been shown to stimulate the activity of bifidobacteria in the colon. These bacteria are normally found in a healthy human gut. Changes in their amount or activity have been associated with gastrointestinal diseases including inflammatory bowel disease, colorectal cancer, and irritable bowel syndrome. [6,7]

When Gluten Is a Problem

What’s not great about gluten is that it can cause serious side effects in certain individuals. Some people react differently to gluten, where the body senses it as a toxin, causing one’s immune cells to overreact and attack it. If an unknowingly sensitive person continues to eat gluten, this creates a kind of battle ground resulting in inflammation. The side effects can range from mild (fatigue, bloating, alternating constipation and diarrhea) to severe (unintentional weight loss, malnutrition, intestinal damage) as seen in the autoimmune disorder celiac disease . Estimates suggest that 1 in 133 Americans has celiac disease, or about 1% of the population, but about 83% of them are undiagnosed or misdiagnosed with other conditions. [8,9] Research shows that people with celiac disease also have a slightly higher risk of osteoporosis and anemia (due to malabsorption of calcium and iron, respectively); infertility; nerve disorders; and in rare cases cancer. [10] The good news is that removing gluten from the diet may reverse the damage. A gluten-free diet is the primary medical treatment for celiac disease. However, understanding and following a strict gluten-free diet can be challenging, possibly requiring the guidance of a registered dietitian to learn which foods contain gluten and to ensure that adequate nutrients are obtained from gluten-free alternatives. Other conditions that may require the reduction or elimination of gluten in the diet include:

• Non-celiac gluten sensitivity, also referred to as gluten sensitive enteropathy (GSE) or gluten intolerance —An intolerance to gluten with similar symptoms as seen with celiac disease, but without the accompanying elevated levels of antibodies and intestinal damage. There is not a diagnostic test for GSE but is determined by persistent symptoms and a negative diagnostic celiac test.
• Wheat allergy —An allergy to one or more of the proteins (albumin, gluten, gliadin, globulin) found in wheat, diagnosed with positive immunoglobulin E blood tests and a food challenge. Compare this with celiac disease, which is a single intolerance to gluten. Symptoms range from mild to severe and may include swelling or itching of the mouth or throat, hives, itchy eyes, shortness of breath, nausea, diarrhea, cramps, and anaphylaxis. People who test negative for this condition may still have gluten sensitivity. This condition is most often seen in children, which most outgrow by adulthood.
• Dermatitis herpetiformis (DH) —A skin rash that results from eating gluten. It is an autoimmune response that exhibits itself as a persistent red itchy skin rash that may produce blisters and bumps. Although people with celiac disease may have DH, the reverse is not always true.

It is important to note that gluten is a problem only for those who react negatively to it, or test positive for celiac disease. Most people can and have eaten gluten most of their lives, without any adverse side effects.

Does gluten cause brain fog?

But does this side effect occur in people without a true gluten intolerance, and can the reverse be suggested in that the avoidance of gluten might sharpen the mind? A large cohort study disagrees. Almost 13,500 middle-aged women from the Nurses’ Health Study II without celiac disease were followed for 28 years to observe any potential links between gluten intake and mental ability. [15]   No significant differences were found in cognitive scores (measuring reaction time, attention, memory, etc.) comparing women with the highest and lowest gluten intakes. The lack of association remained even after excluding women with a dementia or cancer diagnosis.

Unless a person has diagnosed celiac disease, a wheat allergy, or a gluten sensitivity, current evidence does not support that eating gluten increases inflammation in the brain or negatively affects brain health.

What Is a “Gluten-Free Diet”?

This is essentially a diet that removes all foods containing or contaminated with gluten . However, since gluten-containing whole grains contain fiber and nutrients including B vitamins , magnesium , and iron , it’s important to make up for these missing nutrients. Along with consuming naturally gluten-free foods in their whole form like fruits , vegetables , legumes, nuts , seeds, fish, eggs , and poultry, the following whole grains are also inherently gluten-free:

• Brown, black, or red rice
• Gluten-free oats

It’s also key not to rely on processed gluten-free foods that may be high in calories, sugar, saturated fat, and sodium and low in nutrients, such as gluten-free cookies, chips, and other snack foods. Often, these foods are made with processed unfortified rice, tapioca, corn, or potato flours.

The gluten-free food industry has grown 136% from 2013 to 2015 with almost $12 billion in sales in 2015. Interestingly, studies show that people who do not have celiac disease are the biggest purchasers of gluten-free products. [11] Consumer surveys show that the top three reasons people select gluten-free foods are for “no reason,” because they are a “healthier option,” and for “digestive health.” [12] For those who are not gluten-intolerant, there is no data to show a specific benefit in following a gluten-free diet, particularly if processed gluten-free products become the mainstay of the diet. In fact, research following patients with celiac disease who change to a gluten-free diet shows an increased risk of obesity and metabolic syndrome. This could be partly due to improved intestinal absorption, but speculation has also focused on the low nutritional quality of processed gluten-free foods that may contain refined sugars and saturated fats and have a higher glycemic index. [13,14]

• Diet Review: Gluten-Free for Weight Loss
• Whole Grains
• Lebwohl B, Cao Y, Zong G, Hu FB, Green PHR, Neugut AI, Rimm EB, Sampson L, Dougherty L, Giovannucci E, Willett WC, Sun Q, Chan AT. Long term gluten consumption in adults without celiac disease and risk of coronary heart disease: prospective cohort study.  BMJ . 2017 May 2;357:j1892.
• Liu S, Stampfer MJ, Hu FB, et al. Whole-grain consumption and risk of coronary heart disease: results from the Nurses’ Health Study. Am J Clin Nutr . 1999;70:412-9.
• Mellen PB, Walsh TF, Herrington DM. Whole grain intake and cardiovascular disease: a meta-analysis. Nutr Metab Cardiovasc Dis . 2008;18:283-90.
• de Munter JS, Hu FB, Spiegelman D, Franz M, van Dam RM. Whole grain, bran, and germ intake and risk of type 2 diabetes: a prospective cohort study and systematic review. PLoS Med . 2007;4:e261.
• Johnsen, N.F., et al. Whole-grain products and whole-grain types are associated with lower all-cause and cause-specific mortality in the Scandinavian HELGA cohort. British Journal of Nutrition , 114(4), 608-23.
• Neyrinck, A.M., et al. Wheat-derived arabinoxylan oligosaccharides with prebiotic effect increase satietogenic gut peptides and reduce metabolic endotoxemia in diet-induced obese mice. Nutr Diabetes . 2012 Jan; 2(1): e28.
• Tojo, R., et al. Intestinal microbiota in health and disease: role of bifidobacteria in gut homeostasis. World J Gastroenterol . 2014 Nov 7;20(41):15163-76.
• Beyond Celiac. Celiac Disease: Fast Facts https://www.beyondceliac.org/celiac-disease/facts-and-figures/ Accessed 4/1/2017.
• Riddle, M.S., Murray, J.A., Porter, C.K. The Incidence and Risk of Celiac Disease in a Healthy US Adult Population. Am J Gastroenterol . 2012;107(8):1248-1255.
• N., Freeman, H.J., Thomson, A.B.R. Celiac disease: Prevalence, diagnosis, pathogenesis and treatment. World J Gastroenterol . 2012 Nov 14; 18(42): 6036–6059.
• Topper A. Non-celiacs Drive Gluten-Free Market Growth. Mintel Group Ltd. Web. http://www.mintel.com/blog/food-market-news/gluten-free-consumption-trends . Accessed Mar 27, 2017.
• Reilly, N.R. The Gluten-Free Diet: Recognizing Fact, Fiction, and Fad. The Journal of Pediatrics. Volume 175, August 2016, pages 206–210.
• Tortora, R., et al. Metabolic syndrome in patients with celiac disease on a gluten-free diet. Aliment Pharmacol Ther . 2015 Feb;41(4):352-9.
• Kabbani, T.A., et al. Body mass index and the risk of obesity in coeliac disease treated with the gluten-free diet. Aliment Pharmacol Ther . 2012 Mar;35(6):723-9.
• Wang Y, Lebwohl B, Mehta R, Cao Y, Green PHR, Grodstein F, Jovani M, Lochhead P, Okereke OI, Sampson L, Willett WC, Sun Q, Chan AT. Long-term Intake of Gluten and Cognitive Function Among US Women. JAMA Netw Open. 2021 May 3;4(5):e2113020. Disclosures: B Lebwohl reported receiving personal fees from Takeda and Kanyos outside the submitted work. OI Okereke reported receiving royalties from Springer Publishing outside the submitted work and receiving honoraria from the AARP for participation at the Global Council on Brain Health meetings. AT Chan reported receiving personal fees from Pfizer, Boehringer Ingelheim, Bayer Pharma, and Zoe Global outside the submitted work.

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Gluten-Free Diet: Is It Right for Me?

Featured Expert:

Selvi Rajagopal, M.D., M.P.H.

There’s a lot of buzz around going gluten-free, with everyone from celebrities to pro athletes touting the benefits of a gluten-free diet. But these diets aren’t for everyone. Selvi Rajagopal, M.D ., a specialist in internal medicine and obesity, explains the risks and benefits of cutting gluten and shares how you can make a healthy eating plan.

What is a gluten-free diet?

A gluten-free diet excludes any foods that contain gluten , which is a protein found in wheat and several other grains. It means eating only whole foods that don’t contain gluten, such as fruits, vegetables, meat and eggs, as well as processed gluten-free foods like gluten-free bread or pasta.

“Gluten is a protein naturally occurring in certain foods, but it can also be added to foods during processing for texture,” explains Rajagopal. Gluten can be used as a binding agent and flavoring, so you can sometimes find it in foods you wouldn’t expect. In addition to foods like pizza, pasta, cereal and baked goods, gluten can be in everything from soy sauce and ice cream to certain medications, beauty products and dietary supplements.

Some people think going gluten-free means not eating any carbohydrates, but this isn’t the case. Lots of foods that contain carbs, such as rice, potatoes and beans, don’t contain gluten.

Who should eat a gluten-free diet?

People with celiac disease.

A gluten-free diet is necessary for people with celiac disease , an autoimmune response to gluten that causes the body to attack the small intestine, causing belly pain, nausea, bloating or diarrhea. People with celiac disease can’t tolerate gluten in any form, and need to follow a gluten-free diet for the rest of their lives. If you have celiac and accidentally eat gluten, you’ll probably experience the same symptoms you did before you went gluten-free.

People with gluten sensitivity

Another condition that may prompt someone to cut gluten from their diets is a non-celiac gluten sensitivity, sometimes called gluten intolerance. “We don’t have a clear definition for gluten intolerance or a clear way to explain it,” says Rajagopal. “We know that some people eat something that contains gluten and then they don’t feel well.”

It’s important not to assume that gastrointestinal irritation is the result of gluten. If you think you may have a gluten intolerance, Rajagopal recommends working with a physician and a registered dietitian to get to the bottom of your symptoms.

“There isn’t a test for gluten intolerance, so we might try a process of elimination such as the low FODMAP diet ,” says Rajagopal. This is a temporary eating plan that eliminates lots of foods that can irritate the gut, including wheat-based products. If gluten is the source of the irritation, you may notice an improvement in symptoms such as:

• Constipation or diarrhea
• Stomach pain

People who are allergic to wheat

People with a wheat allergy should avoid certain foods containing gluten, but not because of the gluten. Wheat triggers an immune response in their bodies, which can cause symptoms such as a skin rash, headache or sneezing. They can still eat gluten in other grains, including barley and rye.

Can you go gluten-free to lose weight?

People who adopt a gluten-free diet often lose weight, but it’s usually because they also cut out a lot of processed foods and refined carbohydrates that contain gluten. If you stop eating gluten to lose weight, it’s important to watch your portion sizes, get regular exercise and eat plenty of whole foods such as fruits, vegetables and lean proteins.

Are there risks to trying a gluten-free diet if you don’t have celiac disease?

If you cut all gluten out of your diet, there’s a risk that you could miss out on nutritious whole grains, fiber and micronutrients. Getting enough whole grains in your diet is especially important if you’re at risk for heart disease or diabetes. Whole grains can lower cholesterol levels and even help regulate your blood sugar. In addition, some gluten-containing foods are sources of important vitamins and minerals, such as B vitamins, iron and magnesium.

Keep in mind that some processed gluten-free foods contain high amounts of unhealthy ingredients such as sodium, sugar and fat. Consuming these foods can lead to weight gain, blood sugar swings, high blood pressure and other problems. So, a gluten-free label doesn’t necessarily make a food healthy.

If you don’t have celiac disease or gastrointestinal irritation, Rajagopal recommends removing highly processed foods from your diet before removing gluten. Add in more fruits, vegetables, whole-grain bread or pasta, and lean proteins. Many people find they feel better just by eating better, not by removing gluten.

Will I go through gluten withdrawal if I start eating gluten-free?

There’s no scientific evidence to suggest that people actually go through “withdrawal” when they stop eating gluten. Some people report feeling dizziness, nausea, extreme hunger and even anxiety and depression when they suddenly go from eating a lot of gluten to being gluten-free. These symptoms usually go away after a few weeks on a gluten-free diet, but talk to your health care provider if they persist.

How do I get started with a gluten-free diet?

If you’re interested in trying a gluten-free diet, talk to a physician or a registered dietitian. They can guide you toward a balanced eating plan that meets your unique nutritional needs.

Tips for making dietary changes if you have celiac disease include:

• Check for warnings on packages. Many products that don’t contain gluten may have been processed in a facility where there are gluten products.
• Keep kitchen utensils, dishes and other food prep items that are used for gluten-containing foods separate from your utensils.
• Read ingredient labels carefully to check for any traces of wheat. Some artificial colors and seasonings also contain gluten.
• Substitute oat, buckwheat, quinoa or other gluten-free or alternative grain flours for wheat flour in cooking and baking.

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The gluten-free diet for celiac disease and beyond.

1. Introduction

2. gluten and celiac disease, 3. gluten-free diet for celiac disease, 3.1. efficacy of gluten-free diet in celiac disease, 3.2. skepticism of the gluten-free diet, 3.3. challenges of a gluten-free diet, 3.4. how to monitor a gluten-free diet for celiac disease, 3.4.1. symptom assessment, 3.4.2. dietetic interview, 3.4.3. serology, 3.4.4. stool and urine markers, 3.4.5. small bowel biopsy and pathology, 4. gluten-free diet for other health problems, 4.1. gluten ataxia, 4.2. cognitive impairment and neurological and mental illnesses, 4.3. inflammatory bowel disease and irritable bowel syndrome, 4.4. dermatitis herpetiformis, 4.5. non-celiac gluten sensitivity (ncgs) and people who avoid gluten, 5. adverse events of gfd, 5.1. gluten and the gut microbiome, 5.2. nutritional deficiencies, 5.4. social and psychological impact, 6. conclusions, author contributions, institutional review board statement, informed consent statement, data availability statement, conflicts of interest.

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Share and Cite

Aljada, B.; Zohni, A.; El-Matary, W. The Gluten-Free Diet for Celiac Disease and Beyond. Nutrients 2021 , 13 , 3993. https://doi.org/10.3390/nu13113993

Aljada B, Zohni A, El-Matary W. The Gluten-Free Diet for Celiac Disease and Beyond. Nutrients . 2021; 13(11):3993. https://doi.org/10.3390/nu13113993

Aljada, Bara, Ahmed Zohni, and Wael El-Matary. 2021. "The Gluten-Free Diet for Celiac Disease and Beyond" Nutrients 13, no. 11: 3993. https://doi.org/10.3390/nu13113993

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Researchers Find Link between Gluten-Free Diet and Metabolic Syndrome

There are different levels of gluten sensitivity, the most severe form being the autoimmune disorder, celiac disease. Normally, as food travels through the digestive system, small hair-like structures called microvilli help absorb nutrients from food. Unfortunately, when gluten enters the digestive system of a celiac disease patient, the microvilli are attacked by the immune system, causing them to flatten and lose their ability to absorb nutrients. 1 This can result in symptoms, including: an inability to gain weight, bloating, headaches, and fatigue, which can lead to long-term health conditions, like dementia or infertility. 2 The only way to reduce such symptoms is by strict adherence to a gluten-free diet (GFD). However, recent studies have found that living on a GFD may come with a cost.

Today, gluten is used as a thickening agent or filler in almost anything from soy sauce to salad dressing. To compensate for the lack of gluten, most gluten-free foods contain excessive amounts of sugar, carbohydrates, sodium, and calories. For some, a GFD at first may seem like a healthy alternative to the normal diet. However, for those with celiac disease (CD) it is an obligation that must be followed to improve one’s health. Unfortunately, this consistent diet of sugary foods can cause a person to gain weight and, more significantly, can “lead to deficiencies in iron, calcium, thiamine, riboflavin, niacin, and folate.” 3

A study conducted from July 2012 to July 2013 at the Department of Clinical Medicine and Surgery of the University  of  Naples Federico II,  followed 98 newly diagnosed celiac disease patients as they progressed with their gluten-free diet for one year. The objective was to identify a link between a GFD and Metabolic Syndrome (MS), as well as Hepatic Steatosis (HS). MS is a combination of diseases that includes diabetes, high blood pressure, and obesity. Conditions such as physical inactivity or a diet filled with saturated fats and cholesterol increase a person’s risk for getting MS. 4

During the same time as diagnosis, the patient’s waist circumference, BMI, blood pressure, lipid profile and glucose levels were recorded. These measurements were then recorded again after one year on a GFD. The results showed a significant increase in all fields except lipid profile, which stayed fairly constant. Of the 98 patients, 29 were diagnosed with MS. Other medical conditions like anemia, hypoalbuminemia, and hypocalcaemia were also noted in some of the patients. 5

Risk of HS is also possible for a CD patient on a GFD for at least one year. Hepatic Steatosis describes a buildup of fats and triglycerides on the liver. Certain risk factors, such as type 2 diabetes, insulin resistance, and high cholesterol can increase a person’s chance of developing HS. 6 Diagnosis of HS was done with an ultrasound, and experimental results confirmed that the chance of getting HS after starting a GFD is highly possible. HS is most commonly found in people with MS who have celiac disease and are on a GFD.

Gluten-free diets are complicated to follow and can sometimes become frustrating for patients. Newly diagnosed CD patients are encouraged to meet with a Registered Nutritionist or Dietitian to go over meal plans and strategies when tackling the GFD. Many experiments have confirmed that CD patients who start a GFD are at a greater risk for developing MS than before they started the diet. However, there are experiments that present conflicting results where a GFD is not directly linked to weight gain. For this reason, more research is needed to understand the connection between the gluten-free diet and MS. In any case, newly diagnosed CD patients, as well as formerly diagnosed patients, must be wary of what they eat and the long-term effects of these foods on their body.

1 Fasano, A., & Catassi, C. (2012). Celiac Disease.  Clinical Practice,  (367), 2419-2426.

2 Silvester, J., & Rashid, M. (2007). Evaluation Of Current Practice Guidelines For Long-Term Follow-Up Of Individuals With Celiac Disease.  Journal Of Pediatric Gastroenterology And Nutrition,   21 (9), 557–564.

3 The Reality Behind Gluten-Free Diets, Nutrition and Health Library. (2015, January 12). Retrieved April 2, 2015, from http://www.uwhealth.org/nutrition-diet/the-reality-behind-gluten-free-diets/31084

4 Grundy, S., Cleeman, J., Daniels, S., Donato, K., Eckel, R., Franklin, B., . . . Costa, F. (2005). Interactions among Hepatic Steatosis, Inflammation, and Insulin Resistance: Beyond Common Sense Circulation, (112), 2735-2752.

5 Tortora, R., Capone, P., De Stefano, G., Imperatore, N., Gerbino, N., Donetto, S., . . . Rispo, A. (2015). Metabolic Syndrome in Patients With Coeliac Disease on a Gluten-free Diet. Alimentary Pharmacology & Therapeutics,   41 (4), 352-359.

6 Wu C (2012) Interactions among Hepatic Steatosis, Inflammation, and Insulin Resistance: Beyond Common Sense. J Nutr Food Sci 2:e106. doi: 10.4172/2155-9600.1000e106

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Your small intestine is lined with tiny hairlike projections called villi, which absorb sugars, fats, proteins, vitamins, minerals and other nutrients from the food you eat. Gluten exposure in people with celiac disease damages the villi, making it hard for the body to absorb nutrients necessary for health and growth.

Celiac disease is an illness caused by an immune reaction to eating gluten. Gluten is a protein found in foods containing wheat, barley or rye.

If you have celiac disease, eating gluten triggers an immune response to the gluten protein in your small intestine. Over time, this reaction damages your small intestine's lining and prevents it from absorbing nutrients, a condition called malabsorption.

The intestinal damage often causes symptoms such as diarrhea, fatigue, weight loss, bloating or anemia. It also can lead to serious complications if it is not managed or treated. In children, malabsorption can affect growth and development in addition to gastrointestinal symptoms.

There's no definite cure for celiac disease. But for most people, following a strict gluten-free diet can help manage symptoms and help the intestines heal.

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The symptoms of celiac disease can vary greatly. They also may be different in children and adults. Digestive symptoms for adults include:

• Weight loss.
• Bloating and gas.
• Abdominal pain.
• Nausea and vomiting.
• Constipation.

However, more than half the adults with celiac disease have symptoms that are not related to the digestive system, including:

• Anemia, usually from iron deficiency due to decreased iron absorption.
• Loss of bone density, called osteoporosis, or softening of bones, called osteomalacia.
• Itchy, blistery skin rash, called dermatitis herpetiformis.
• Mouth ulcers.
• Headaches and fatigue.
• Nervous system injury, including numbness and tingling in the feet and hands, possible problems with balance, and cognitive impairment.
• Joint pain.
• Reduced functioning of the spleen, known as hyposplenism.
• Elevated liver enzymes.

Children with celiac disease are more likely than adults to have digestive problems, including:

• Chronic diarrhea.
• Swollen belly.
• Pale, foul-smelling stools.

The inability to absorb nutrients might result in:

• Failure to thrive for infants.
• Damage to tooth enamel.
• Irritability.
• Short stature.
• Delayed puberty.
• Neurological symptoms, including attention-deficit/hyperactivity disorder (ADHD), learning disabilities, headaches, lack of muscle coordination and seizures.

Dermatitis herpetiformis

Gluten intolerance can cause this blistery skin disease. The rash usually occurs on the elbows, knees, torso, scalp or buttocks. This condition is often associated with changes to the lining of the small intestine identical to those of celiac disease, but the skin condition might not cause digestive symptoms.

Health care professionals treat dermatitis herpetiformis with a gluten-free diet or medicine, or both, to control the rash.

When to see a doctor

Consult your health care team if you have diarrhea or digestive discomfort that lasts for more than two weeks. Consult your child's health care team if your child:

• Is irritable.
• Is failing to grow.
• Has a potbelly.
• Has foul-smelling, bulky stools.

Be sure to consult your health care team before trying a gluten-free diet. If you stop or even reduce the amount of gluten you eat before you're tested for celiac disease, you can change the test results.

Celiac disease tends to run in families. If someone in your family has the condition, ask a member of your health care team if you should be tested. Also ask about testing if you or someone in your family has a risk factor for celiac disease, such as type 1 diabetes.

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Your genes, combined with eating foods with gluten and other factors, can contribute to celiac disease. However, the precise cause isn't known. Infant-feeding practices, gastrointestinal infections and gut bacteria may contribute, but these causes have not been proved. Sometimes celiac disease becomes active after surgery, pregnancy, childbirth, viral infection or severe emotional stress.

When the body's immune system overreacts to gluten in food, the reaction damages the tiny, hairlike projections, called villi, that line the small intestine. Villi absorb vitamins, minerals and other nutrients from the food you eat. If your villi are damaged, you can't get enough nutrients, no matter how much you eat.

Risk factors

Celiac disease tends to be more common in people who have:

• A family member with celiac disease or dermatitis herpetiformis.
• Type 1 diabetes.
• Down syndrome, William syndrome or Turner syndrome.
• Autoimmune thyroid disease.
• Microscopic colitis.
• Addison's disease.

Complications

Celiac disease that is not treated can lead to:

• Malnutrition. This occurs if your small intestine can't absorb enough nutrients. Malnutrition can lead to anemia and weight loss. In children, malnutrition can cause slow growth and short stature.
• Bone weakening. In children, malabsorption of calcium and vitamin D can lead to a softening of the bone, called osteomalacia or rickets. In adults, it can lead to a loss of bone density, called osteopenia or osteoporosis.
• Infertility and miscarriage. Malabsorption of calcium and vitamin D can contribute to reproductive issues.
• Lactose intolerance. Damage to your small intestine might cause you abdominal pain and diarrhea after eating or drinking dairy products that contain lactose. Once your intestine has healed, you might be able to tolerate dairy products again.
• Cancer. People with celiac disease who don't maintain a gluten-free diet have a greater risk of developing several forms of cancer, including intestinal lymphoma and small bowel cancer.
• Nervous system conditions. Some people with celiac disease can develop conditions such as seizures or a disease of the nerves to the hands and feet, called peripheral neuropathy.

Nonresponsive celiac disease

Some people with celiac disease don't respond to what they consider to be a gluten-free diet. Nonresponsive celiac disease is often due to contamination of the diet with gluten. Working with a dietitian can help you learn how to avoid all gluten.

People with nonresponsive celiac disease might have:

• Bacterial overgrowth in the small intestine.
• Poor pancreas function, known as pancreatic insufficiency.
• Irritable bowel syndrome.
• Difficulty digesting sugar found in dairy products (lactose), table sugar (sucrose), or a type of sugar found in honey and fruits (fructose).
• Truly refractory celiac disease that is not responding to a gluten-free diet.

Refractory celiac disease

In rare instances, the intestinal injury of celiac disease doesn't respond to a strict gluten-free diet. This is known as refractory celiac disease. If you still have symptoms after following a gluten-free diet for 6 months to 1 year, you should talk to your health care team to see if you need further testing to look for explanations for your symptoms.

Celiac disease care at Mayo Clinic

• Rubio-Tapia A, et al. American College of Gastroenterology guidelines update: Diagnosis and management of celiac disease. American Journal of Gastroenterology. 2023; doi:10.14309/ajg.0000000000002075.
• Catassi C, et al. Coeliac disease. The Lancet. 2022; doi:10.1016/S0140-6736(22)00794-2.
• Singh P, et al. Who to screen and how to screen for celiac disease. World Journal of Gastroenterology. 2022; doi:10.3748/wjg.v28.i32.4493.
• What is celiac disease? Celiac Disease Foundation. https://celiac.org/about-celiac-disease/what-is-celiac-disease/. Accessed April 26, 2023.
• Feldman M, et al., eds. Celiac disease. In: Sleisenger and Fordtran's Gastrointestinal and Liver Disease: Pathophysiology, Diagnosis, Management. 11th ed. Elsevier; 2021. https://www.clinicalkey.com. Accessed April 26, 2023.
• Celiac disease. National Institute of Diabetes, Digestive and Kidney Diseases. https://www.niddk.nih.gov/health-information/digestive-diseases/celiac-disease. Accessed April 26, 2023.
• Ami TR. AllScripts EPSi. Mayo Clinic. March 24, 2023.
• Khanna S (expert opinion). Mayo Clinic. May 12, 2023.

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Gluten and Autism Spectrum Disorder

Iain d. croall.

1 Department of Infection, Immunity & Cardiovascular Disease, University of Sheffield/INSIGENO, Sheffield S10 2JF, UK; [email protected]

Nigel Hoggard

Marios hadjivassiliou.

2 Academic Departments of Neurosciences and Neuroradiology, Sheffield Teaching Hospitals NHS Trust, Sheffield S10 2JF, UK; [email protected]

An expanding body of literature is examining connections between Autism Spectrum Disorder (ASD) and dietary interventions. While a number of specialist diets have been suggested as beneficial in ASD, gluten has received particularly close attention as a potentially exacerbating factor. Reports exist suggesting a beneficial effect of the gluten-free diet (GFD) in ameliorating behavioural and intellectual problems associated with ASD, while epidemiological research has also shown a comorbidity between ASD and coeliac disease. However, both caregivers and clinicians have expressed an uncertainty of the value of people with ASD going gluten-free, and as the GFD otherwise receives considerable public attention a discussion which focuses specifically on the interaction between ASD and gluten is warranted. In this review we discuss the historical context of ASD and gluten-related studies, and expand this to include an overview of epidemiological links, hypotheses of shared pathological mechanisms, and ultimately the evidence around the use and adoption of the GFD in people with ASD.

1. Motivation and Literature Search Methods

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder characterised primarily by deficits in social communication and restricted/repetitive patterns of behaviour (DSM-5) [ 1 ]. As the name implies its phenotype exists on a spectrum, and overall it is estimated to affect as many as one in 69 children [ 2 ]. Interest in the use of specialist diets in ASD is increasing, as a way to alleviate its behavioural and intellectual outcomes. Though many dietary interventions have been suggested, the gluten-free diet (GFD) is among the most notable. Clinically, the GFD is well recognised as the primary treatment for patients with a gluten-related disorder. The most prominent of these is coeliac disease (CD) which is predominantly expressed as a gastrointestinal (GI) condition. However, physiological sensitivity to gluten is known to exist in other forms. These include other immune-mediated disorders (e.g., dermatitis herpetiformis and gluten ataxia), allergic reactions (wheat allergy), and non-coeliac gluten sensitivity (a condition characterized by self-reported gastrointestinal and extra-intestinal symptoms subjectively improving upon a GFD in subjects in whom other major organic gluten related disorders have been excluded) [ 3 ]. The clinical utility of the GFD outside of these contexts is debatable, and elsewhere its adoption within the general public as a sometimes “fad” diet heightens such scrutiny [ 4 ]. However, generally increased rates of GI problems have been reported in people with ASD, as has evidence of an apparent comorbidity between ASD and CD specifically. As adoption rates of specialty diets which include a gluten-free component are very high in ASD we were motivated to conduct a literature review which focused specifically on the interaction between ASD and gluten.

Searches were made on Pubmed on the 12th of November 2020. Terms included:

• Autism coeliac
• Autism celiac
• Autism gluten
• Autism wheat
• Autistic coeliac
• Autistic celiac
• Autistic gluten
• Autistic wheat

These terms were designed to capture a range of relevant terminology, for example “autism spectrum disorder” or “gluten free diet” would each be picked up by these. Regional variation in spelling of coeliac/celiac would also be accounted for. This returned 237 unique articles. The abstracts of all of these were read to determine eligibility for inclusion in the main review. Criteria for this were that the paper must be original research (i.e., not another review or systematic review) which in some way directly explored links between ASD and GRDs, the use of the GFD in ASD, or any other relevant interaction between ASD and gluten. Case reports were excluded, as were papers where the main article was not available in English. Seventy-nine articles were deemed eligible for inclusion. These were read, and any additional relevant citations found through this were added for discussion. Figure 1 shows these included papers according to their year of publication.

A histogram of eligible papers found in the primary pubmed search according to their year of publication.

Throughout the study of the literature, common research methods/sub-topics were noted. The remainder of this review synthesises these papers according to those themes. Additional literature was searched for and referenced where necessary to elucidate on a relevant key concept which was not adequately covered by the initial searches.

2. Historical Context

The first observation of a possible link between gluten and ASD was reported in 1969 by Goodwin & Goodwin [ 5 ], who noted in a cohort of 65 children with ASD that one 6 year old boy also had CD. This child’s subsequent treatment with a GFD appeared to improve outcomes relating to his ASD. It is relevant to note that at that time the prevalence of CD was considered to be far less than the 1-in-100 that it is sometimes reported as today [ 6 ], largely due to the lack of effective diagnostic methods such as serological testing. In a commentary piece on the then-emerging topic, Dohan references in their 1970 paper [ 7 ] that CD is thought to affect approximately 1 in 3000 people, making suspicion after finding it in one of a cohort of 65 children with ASD understandable. Another point of interest emerges from this paper, which was purely focused on links between CD and schizophrenia but in which Dohan also found it relevant to include an anecdotal report of increased CD rates in ASD groups (“at least 2 coeliac patients among 140 severely autistic children”). While limited phenotypic similarities between schizophrenia and ASD are still discussed now, initial characterisation of the conditions involved such overlap that full clinical separation did not occur until 1980 with the publication of the DSM-III [ 8 ]. To a modern reader, this explains why there are a number of early articles which combine ASD with schizophrenia and otherwise draw potentially confusing links between the two.

Goodwin et al. published another study in 1971 [ 9 ] which is arguably the first trial investigating how gluten modified behaviour in a cohort of children with ASD. Also included were controls and a group of participants with schizophrenia for further comparison. Here, the participants followed the “sprue diet” (GFD) for a single day, in which they were also given a cherry drink which had added to it either gliadin or a placebo (sugar). They were then subsequently monitored and tested via investigation of blood counts and electrophysiological recordings by trans cephalic direct current. Although the authors note some findings, these predominantly focus on differences which appear to separate ASD and schizophrenia participants, providing only one comment on the effect of gliadin where it appeared to reduce plasma cortisol levels. However, this was observed across both control and ASD participants, and when coupled with the small sample sizes (the ASD group had 9 participants for that portion of the results) and extremely short period of dieting, it is difficult to extrapolate any meaningful conclusions. Further studies in the 1970’s included a comparison of serum alpha-1-antitrypsin levels between children with ASD vs. children with CD [ 10 ] (finding them comparably abnormal and suggesting a shared pathology), an experiment published in a book [ 11 ] describing 72 patients with ASD in whom CD was diagnosed (without biopsy) in 8, and another trial of gluten in eight children with ASD who already followed a GFD and were purportedly better for it [ 12 ]. This latter study saw the participants stop their diet to undergo a gluten challenge for 1 month, hypothesising this would worsen their phenotype, but finding no change in bodily measurements (weight, bowel habit etc.) or behaviour (measured by parental reports and observation from a specialist paediatrician).

To summarise, early interest in the topic was driven by essentially anecdotal reports of apparently comorbid cases of CD with ASD. Direct experimentation of this resulted in largely negative findings. These studies had small samples sizes and, when viewed with a contemporary lens, suffered from experimental designs and measurement techniques which would now be considered extremely insensitive in targeting relevant outcomes. Following these papers, little relevant research activity appeared until the mid 1990’s when the topic appeared to become more popular once again.

3. Gastrointestinal Symptoms in ASD

A heightened rate of gastrointestinal (GI) symptoms in people with ASD is well documented. While this particular topic in its entirety falls outside of the scope of the present systematic review, the observation of these symptoms is a major motivator for gluten-specific research. Relevant studies from the review are therefore included here, as well as other key literature.

In 2014 Chaidez et al. [ 13 ] conducted a large study in which 499 children with ASD were compared to typically-developing (TD) children ( N = 324) and children with developmental delay ( N = 137) in terms of GI symptoms measured by 10 Likert scales (abdominal pain, constipation etc.). After controlling for age, sex, maternal education and medications which may lead to GI side effects, children with ASD had significantly heightened odds ratios (OR) compared to controls for 8 outcomes, the lowest being 3.14 (abdominal pain) and the highest being 8.61 (sensitivity to foods). The children with ASD and developmental delay were not significantly different from one another.

This is one of a number of similar studies who’s findings are supported by meta-analyses; a pubmed search of “gastrointestinal autism” found the most recent meta-analysis was performed in 2014 [ 14 ]. This included 15 studies which gave a combined sample of 2215 children with ASD in which four variables were included; general GI concerns, diarrhoea, constipation and abdominal pain. Each of these was found to be significantly more prevalent in the ASD group compared to TD children, with overall OR’s of 4.42, 3.63, 3.86 and 2.45 respectively.

Relevant studies from the current review include a report [ 15 ] of a higher frequency of constipation in children with ASD, a study [ 16 ] which found children with ASD and regression more often had abnormal stool than those without regression, and another experiment [ 17 ] which found GI symptoms to be more common in children and adolescents with ASD than in TD controls, and for these symptoms to be weakly correlated to behavioural measures. Correlations such as these have been documented elsewhere [ 18 , 19 , 20 ]. These studies reference the often non-specific nature of GI symptoms with one explaining that “a GI pathology specific to ASD had not been established” (Babinska et al., 2020 [ 17 ]). Regardless, a notable body of literature has investigated for a comorbidity between ASD and specific GI conditions, often finding significant results.

4. The Co-Morbidity between ASD and CD

Following early literature from the 1970’s, the first study to investigate for an increased rate of CD in ASD was Pavone et al. in 1997 [ 21 ]. Pavone examined a cohort of children with ASD ( N = 11) to detect the rate of CD (by antibody and ultimately biopsy testing), and similarly a cohort of children with CD ( N = 120) to detect the rate of those with features of ASD (as reported by parents and according to the DSM III-R). None of the children with ASD had biopsy-proven CD, while none of the children with CD met criteria for a full ASD diagnosis (though a limited few did show isolated features). This was therefore overall a negative study, but the limitations of examining in such small cohorts as 11 are evident.

Since then, a limited number of large epidemiological studies have been conducted which generally do show an effect indicating CD and ASD to be comorbid to one another. One of 2009 [ 22 ] which focused specifically on comorbidities to ASD within parental medical history, used the Danish Civil Registration System to identify all children born between 1993 and 2004 with ASD ( N = 3325). Here, maternal history of CD led to a significant, overall incidence rate ratio (IRR) of 2.97 in terms of the child having ASD. Other studies examining comorbidities within the same participant have also found significant results while using medical databases. A 2017 study [ 23 ] examined for the risk of psychiatric sequalae in children with CD ( N = 10,903), finding a hazard ratio (HR) of 1.5 (univariate analysis) of being diagnosed with ASD after their CD diagnosis but before the age of 18 (adult data was not included). The same research group has replicated this more recently [ 24 ] with a larger cohort of people diagnosed with CD while a child ( N = 19,189), but which this time did also include psychiatric diagnoses obtained after 18. Here, the HR of developing ASD was 1.47, the highest of all disorders included in analyses.

These papers do however contrast an earlier study, again by the same research group [ 25 ], which examined specifically for the likelihood of an ASD diagnosis preceding a CD diagnosis in children and adults with CD ( N = 26,995). This OR was non-significant, though potentially of interest was a finding wherein previous ASD was still associated with an increased risk of having normal mucosa on biopsy, but positive CD serological test results (tissue transglutaminase; TTG, endomysial; EMA or gliadin; AGA antibodies, reported as a single grouping). While the immediate clinical implication of positivity to these antibodies varies, i.e., TTG/EMA positivity indicates CD with high sensitivity/specificity while many generally-healthy individuals may exhibit AGA positivity, it should be highlighted that within the study of wider “gluten sensitivity” heightened rates of any of these may be considered pathologically-relevant when compared to an appropriate “control” such as in this discussed study. This is therefore an important study as it highlights the link between serological markers of gluten sensitivity and ASD in the absence of enteropathy.

Other prevalence research has been conducted with less stringent diagnostic criteria and/or smaller sample sizes, finding mixed results. Studies with significant findings include Calderoni et al. [ 26 ] who examined a cohort of children with ASD ( N = 382) and found the rate of CD within the sample was 2.62%, although it should be noted this sometimes relied only on a positive serological (TTG/EMA) result and formal CD diagnosis was not always confirmed. Valicenti-McDermott et al. [ 16 ] found an increased family history of CD and/or inflammatory bowel disease in children with ASD who also exhibited regression ( N = 24), compared to children with ASD without regression ( N = 71). In a letter to the editor, Barcia et al. [ 27 ] report an experiment where of 91 “randomly selected” children with ASD, 4 had “biopsy-proven” CD (the authors reference diagnostic guidelines for diagnosis where Marsh grade 3 denotes CD). This is a rate of 4.4% which is considerably higher than might be expected. Mazzone et al. [ 28 ] found in a cohort of 100 children with CD that 2 had ASD (while none of a control group did); whether this is a positive result or not is arguable.

Studies with negative findings include Alabaf et al. [ 29 ] who via parental reporting of 91 children with ASD did not find an association with CD (not reported but this was measured, implying a negative finding). Juneja et al. [ 30 ] screened children with ASD ( N = 150) for CD defined by IgA TTG testing, finding no positive tests. In 2012, Batista et al. [ 31 ] examined children and adolescents with either ASD ( N = 147) or biopsy-proven CD (Marsh grade 3, N = 211) for the rate of the other. The ASD group was found to be entirely negative for CD (although one subject did have a weakly-positive TTG result with negative EMA), while two cases of ASD were found in the CD group; this was concluded to not be above chance. Zelnik et al. [ 32 ] examined a cohort of CD patients ( N = 111) for a range of neurological outcomes including ASD, although as this was reported mixed in with other learning disabilities and ADHD (which overall affected 20.7% of the group) how common ASD specifically was is not known. Finally, Black et al. [ 33 ] examined medical records to identify 96 children with ASD and reported on all diagnosed gastrointestinal comorbidities, failing to identify any CD cases.

A recent meta-analysis [ 34 ] combined some of the above studies (where eligible) to find a significant odds ratio of 1.53 in terms of CD patients having ASD, but a non-significant likelihood of ASD participants having CD. Overall therefore, the majority of studies relevant to the question of comorbidity have used variable sample sizes and diagnostic methods, making definitive conclusions difficult in most individual instances. However, the strongest powered are undoubtedly those from Sweden which studied large cohorts and established an increased risk of a subsequent ASD diagnosis in people with CD, therefore showing a convincing comorbidity. This is further supported by the meta-analysis finding, which showed the same. Studies which examine the “reverse” of this, where initial ASD features may increase risk of subsequent CD, have led to more negative findings however do demonstrate an association with the development of gluten antibodies in the absence of clinical CD. It should also be highlighted that a very large epidemiological study which principally studies an ASD cohort for the rate of CD is absent, which may introduce a sampling bias when interpreting the findings of this overall field. Overall therefore, ASD does appear comorbid to CD, and while an increased risk of CD in ASD is not currently supported a suspicion of ASD being linked to subsequent, immune-mediated “gluten sensitivity” may be warranted.

5. Hypothetical Mechanisms of Action

With a comorbidity between ASD and CD established, a natural question is of what shared pathophysiology may drive these associations. Further, as non-specific GI symptoms are also seen to be generally more prevalent in ASD and that gluten sensitivity is increasingly understood to be a spectrum that extends beyond the clinical criteria for CD specifically, it is important to consider any mechanism of action between gluten and ASD.

From an early point in the literature, hypotheses have frequently related in some way to heightened autoimmunity in ASD. While a predisposition towards autoimmunity was noted as early as 1971 [ 35 ], enquiries of this nature gathered pace after a key publication in 2001 [ 36 ] which showed children with ASD (with regression) to have increased markers of innate and adaptive immune response (TNF-A, cytokines etc.). This was investigated at the time partially in response to parental reports of children with ASD suffering apparently high rates of reactions to dietary irritants, and this autoimmune phenotype was subsequently hypothesised to be part of the aetiology of ASD. The authors presented this idea in terms of environmental stimuli triggering an immune response which exacerbates ASD features, and in the specific case of their study hypothesised it may stimulate regression.

In the early 2000’s a series of publications by Vojdani et al. [ 37 , 38 , 39 ] built on this evidence by focusing on more specific dietary triggers and hypothetical knock-on effects they would lead to in terms of molecular pathways. Here, focusing mainly on gliadin (a gluten-specific protein) and casein (a protein in dairy products), it was demonstrated that children with ASD have high rates of antibodies against these (i.e., anti-gliadin and anti-casein) as well as antibodies against DPP4, a digestive enzyme. DPP-4 is important in the processing of gliadin. Initially, gliadin is degraded into various peptides which include gliadinomorphin-7 [ 40 ], an immune reactive substance with “opioid activity”, i.e., which stimulates opioid receptors in the body [ 41 ]. Further degradation of gliadinomorphin-7 is therefore required, which is where DPP4 functions by cleaving such peptides [ 42 ]. As Vojdani et al. reported, the existence of anti-DPP4 would hypothetically reduce the amount of circulating DPP4, increasing the abundance of gliadinomorphin-7 and the likelihood of downstream, opioid-like effects. It should be highlighted that casein and other dietary peptides are similarly degraded to intermediary substances with opioid properties (e.g., casomorphin [ 43 ]), and together these potentially harmful peptides have been termed “exorphins” [ 44 ].

Stimulation of the opioid system has been studied in the context of ASD features. An early proponent of this link, Panksepp outlined a theory in 1979 [ 45 ] (based largely on his earlier animal model experiments) wherein excess opioid activity may lead to the decreased social behaviour seen in ASD. This theory has persisted until today, with numerous publications concerned with evidence of opioid overactivity in people with ASD. Animal studies have continued to show the importance of a balanced opioid system in maintaining social behaviours which are similar to those impacted in ASD, while experiments investigating for levels of relevant, opioid-like peptides in the sera, CSF or urine of people with ASD have generally shown raised titres albeit with some notable exceptions where decreases have been reported [ 46 ]. Indeed, measurement of urine peptides has become a common tool in this field, with high levels being seen as an indication of insufficient digestion of food which may lead to excess exorphins [ 47 ].

An alternative theory has focused on the role that oxidative stress may play in ASD, which may lead to a state of inflammation in the brain. It has for example been reported that people with ASD have an impaired antioxidant defence in the cerebellum [ 48 ], while problems metabolising nitrous oxide (which may lead to increased oxidative stress [ 49 ]) has also been proposed as driver of ASD pathophysiology [ 50 ]. This holds a relevance to gluten sensitivity, where increased oxidative stress has also been demonstrated, for example as triggered by gliadin [ 51 ] or as demonstrated generally by raised markers of oxidative stress across untreated children with CD [ 52 ].

Studies have also noted the potential for shared genetic predisposition. One recent paper by Bennabi et al. [ 53 ] compared genotyping data between ASD and control cohorts, finding that the haplotype HLA-DRB1*11-DQB1*07 was more common in the ASD group, with this being more prevalent still in those ASD patients with the most pronounced behavioural symptoms. A different haplotype (HLA-DRB1*17-DQB1*02) was conversely more common in the control group. The *07 haplotype was therefore concluded to be potentially causative and the *02 one protective. Of relevance is that the *07 haplotype is additionally recognised as associated with CD, leading to a suggestion that there may be a sub-group of people with ASD holding a genetic risk for both [ 54 ]. However, other genetic research has produced negative findings, such as a meta-analysis of genome-wide association studies [ 55 ] which did identify regions associated with ASD but noted only overlap between these and schizophrenia.

The potential of reactivity of antibodies to gluten products should be discussed. Antibodies against tissue-transglutaminase (TTG) have very high sensitivity and specificity in diagnosing CD, meaning that as a comorbidity has been demonstrated they may have a relevance in ASD pathology. These antibodies have been reported to lead to apoptosis of neuroblast cells in vitro [ 56 ]. Other antibodies which may indicate gluten sensitivity but not CD specifically include transglutaminase 6 (TG6) antibodies, which have been indicated in the diagnosis of gluten ataxia [ 57 ] (where the cerebellum is the primary site of damage), with this supported by animal research showing TG6 to be distributed throughout the central nervous system including brain regions such as the cerebellum and thalamus [ 58 ]. TG6 antibodies have been reported at a rate of 4.4% in a group of 77 children with ASD [ 59 ]; this experiment lacked a control group and as this is a relatively novel marker it is difficult to evaluate if this is abnormal. Finally, gliadin antibodies have been shown to react with brain blood vessel structures [ 60 ], show cross-reactivity with neuronal synapsin 1 [ 61 ], and to be associated with rates of depression in people with CD and healthy controls [ 62 ]. Gliadin antibodies have been measured across a number of studies in ASD, frequently finding them to be raised. Those found in the current review are summarised in Table 1 .

A summary of studies which have investigated if gliadin antibodies (AGA) are affected by ASD. * : full text not available.

The potential for antibodies and other irritants to travel from the gut to the brain is raised by a number of studies which have demonstrated generally inflamed/abnormal intestinal findings in ASD [ 68 , 69 , 70 , 71 ], and others showing compromised intestinal permeability specifically [ 66 , 72 ]. Indeed, one other publication [ 73 ] examined gene and protein expression of brain and intestinal tissue of human ASD subjects, finding evidence of impaired intestinal permeability in combination with altered blood-brain barrier integrity. It should be noted that not all studies support an impacted intestinal permeability [ 59 , 74 ], but nonetheless these phenomena raise the possibility of a gut-brain axis interaction being relevant in ASD. Here, a negative feedback loop between the brain and the gut would lead to exacerbation of both neurological and GI outcomes. Arguably most studies which have investigated how gluten can impact people with ASD may fall within this broader concept, where irritants enter the bloodstream from the gut to cause downstream negative consequences for the brain. A loop may be completed if the effect on the brain leads to alterations of behaviour and appetite which may maintain or exacerbate the cycle [ 75 ].

In summary, pathological interactions between ASD and gluten have focused on opioid activity from improperly digested gluten products, inflammation caused by oxidative stress and/or reactivity with anti-gluten antibodies, and some indications of shared genetic factors. These hypotheses provide some explanation for the previously discussed comorbidity between ASD and CD and also make it appear reasonable that gluten may exacerbate bodily stress in other groups of people with ASD who do not have CD. However, it remains unclear to what extent ASD populations and sub-populations are affected, and to what degree these findings represent a unique interaction with gluten specifically or are a consequence of a generally-raised autoimmune profile in ASD.

6. Trials of the GFD in ASD

Establishing that gluten is potentially harmful for people with ASD leads to the question of if a GFD would then bring any benefits. After the early studies of the 1970’s, the first trial which involved gluten in any capacity was conducted in 1990 and is detailed in two publications [ 76 , 77 ] by Knivsberg et al. Here, fifteen children with ASD in combination with abnormal urine peptide results engaged with a gluten and casein-free diet (GCFD) for four years. Measurements were generally taken at baseline, one year and four year time points, and included scales which characterised psychotic behaviour in children, psycholinguistic ability and fluid intelligence. However, this data was not all collected consistently (e.g., the psychotic behaviour measurements were not made at 4 years), and authors note variable dietary success between the children. Regardless, significant findings suggested improvement across multiple outcomes, including normalisation of urine peptides. The 1990’s saw one other trial [ 78 ], the primary analyses of which concerned a group of 22 children with mixed spectrum disorders (the most common being ASD) who undertook a GFD for 5 months. Other groups were also examined, e.g., children with ASD already on a GFD took a gluten challenge, although these sample sizes were very small. Similar to the Knivsberg study, improvement in behavioural outcomes was noted in response to the GFD although no change in urinary peptide levels were seen.

The first randomised trial was conducted in 2002 [ 79 ]. Here, 20 children with ASD and abnormal urinary peptides were randomised into parallel groups to receive either the GCFD or a regular diet for 12 months. Following this, improvements were noted across behavioural and intellectual outcomes. A number of randomised trials have been conducted since and those that utilise an intervention that in any way involves gluten are summarised in Table 2 .

A summary of randomised trials which have in some way included a gluten-free diet as an intervention in treating ASD.

Findings described in the final column relate to any analysis which indicates with statistical significance that the intervention affected an outcome. 6-GSI; 6-Item Gastrointestinal Symptom Index, ABC; Abberant Behaviour Checklist, ADHD-IV; Attention-Deficit Hyperactivity Disorder—IV rating scale, ADOS; Autism Diagnostic Observation Schedule, AQ; Autism Spectrum Quotient, ASD; Autistic Spectrum Disorder, ASRS; Autism Spectrum Rating Scale, ATEC; Autism Treatment Evaluation Checklist, AWPC; Approach Withdrawal Problems Composite (a subset of the PDD-BI), CARS; Childhood Autism Rating Scale, CARSA; Conners Abbreviated Rating Scale and Actigraphy, CBC; Child Behavior Checklist, CPRS-R; Connor’s Parent Rating Scale-Revised, DIPAB; Diagnose af Psykotisk Atfærd hos Børn, ECO; Ecological Communication Orientation, ERC-III; The Behavioral Summarized Evaluation, EQ-SQ; Empathy and Systemising Quotient, GARS; Gilliam Autism Rating Scale, GI; Gastro-intestinal, ITPA; Illinois Test of Psycholinguistic Abilities, LIPS; Leiter International Performance Scale, MABC; Movement Assessment Battery for Children, MSEL; Mullen Scales of Early Learning AGS edition, PDD-BI; Pervasive Developmental Disorders Behaviour Inventory, RIAS; Reynolds Intellectual Assessment Scales, RRLRS; Ritvo-Freeman Real Life Rating Scales, SAS Pro; Severity of Autism scale-Professional Evaulation, SCAS-P; Spence’s Children Anxiety Scale-Parent version, SCQ; Social Communication Questionnaire, SRS; Social Responsiveness Scale, SSP; Short Sensory Profile, VABS-2; Vineland Adaptive Behavior Scale, Second Edition.

Examining this literature reveals a very mixed picture of findings. Of the 13 RCT’s found in the current review, improvements of some kind were noted in 6 [ 79 , 82 , 83 , 84 , 85 , 90 ], no findings were observed in another 6 [ 80 , 81 , 86 , 88 , 89 , 91 ], while a worsening of GI symptoms (in response to the GCFD) was observed in one study [ 87 ]. All 6 studies which noted a positive effect from the interventional diet included improvements in intellectual/behavioural outcomes, and sometimes also in physiological measurements (e.g., GI symptoms).

Consolidation of these studies is difficult even beyond the mixed findings. One immediate observation is that the exact dietary intervention employed is variable. This increases heterogeneity between studies and makes commenting on the effect of gluten specifically impossible in most cases. Only 3 of the 13 trials had a group design which in some way tested the GFD in isolation; one of these reported improvements in outcomes [ 85 ]. Others typically focus on the GCFD (the most investigated of all interventions), while some use unique interventions such as Grimaldi et al. [ 82 ] who primarily tested a probiotic mixture (in combination with the GCFD). The study by Adams et al. [ 83 ] is also notable for the approach of sequentially accumulating interventions over a year which included the likes of dietary supplementation and epsom salt baths, with the GCFD being added at day 210.

The majority of studies are also unblinded (8 of 13) which raises a risk of placebo/nocebo effects. Some trials which are blinded use as a placebo gluten-free versions of food (bread etc.) given to participants on the assumption that they will not be able to tell the difference, meaning that a degree of skepticism is warranted even for those with such an experimental design. Authors of non-blinded trials that achieve significant results acknowledge this limitation but highlight the practical difficulty of effective blinding for a GFD over a long period of time, or blinding of the other mixed interventions employed. This often leads to varying levels of “blindedness” within a trial, depending on the specific intervention/outcome examined. For example Adams et al. [ 83 ] write “A strength of the study is that it was a randomized, controlled study, but a major limitation of this study is that implementation of a healthy, HGCSF (healthy/gluten/casein/soy-free diet) does not allow blinding of participants. The RIAS evaluation was single-blinded, and the CARS and SAS-Pro were semi-blinded (the evaluators were blinded, the participants were not), so those results are fairly robust. The parent evaluations certainly are subject to some placebo-effect but provide an upper-bound on possible benefits. The laboratory measurements were conducted in a blinded manner, so those results should be reliable.”

Studies variably do or do not use dietary “run-in” periods, which would be generally advisable to account for delays in physiological adjustment between different regimens when taking experimental measurements. Some trials are conducted over very short timeframes, such as Navarro et al. [ 88 ] which ran for 4 weeks or Pusponegoro et al. [ 87 ] which ran for 1 week. The implication of this will vary depending on the outcomes measured, but regarding gluten it is for example known that resolution of symptoms due to gluten exposure can take a number of weeks in patients with CD [ 92 ], while achieving gliadin antibody negativity can take 6 months or longer [ 93 ]. This emphasises the need for long term trials if adequate time is to be given for changes to be captured. In terms of assessing change, the measurement scales used are also scarcely replicated between studies. The majority of RCTs employ a set of tools which are unique to that particular trial, further complicating comparisons or synthesis of findings. An effort to arrive at an agreed-upon set of outcomes would benefit these trials greatly, as would purposeful replication of already-reported significant findings using the same measurement techniques. Otherwise, it also remains an open question as to how much current findings are driven by e.g., different tool sensitivities.

Taken together, it is very difficult to identify a single trial which arguably addresses all of these concerns. Ghalichi et al. [ 85 ] is the largest of those identified (80 subjects randomised), but this ran for 6 weeks and was not blinded. The longest running trials were Whiteley et al. [ 90 ] and Adams et al. [ 83 ], which took principle measurements over 12 months. Each of these did have modest sample sizes ( N = 73 and N = 67 randomised, respectively), but neither were blinded and as discussed Adams et al. included a wide range of accumulative interventions. This highlights a real gap, wherein a well-powered, long-duration and placebo-controlled trial of either the GFD or GCFD has not yet been conducted. Such an experiment would ideally be run after a community consensus is reached regarding what outcomes should be focused on. Until such a trial is conducted a confident, overall conclusion cannot be made. Currently therefore, the overall pattern of the available literature does not support a proved benefit of the GFD in people with ASD (who do not have a clinical diagnosis of CD).

7. Adoption of the GFD and GCFD in ASD

Regardless of there being inconclusive evidence of a benefit to the GFD or GCFD in ASD, adoption of speciality diets is high. Studies assessing this also frequently attempt assessment of possible benefits of the diet primarily via cross-sectional analyses utilising symptom scales/survey responses, or anecdotal reporting from caregivers.

Bowers [ 94 ] reported that a majority of ASD referrals to their diet service regarded a suggestion to go on a GCFD (54.1%). Two of these 14 referrals later saw families of the patient report a “transformation” following adoption of the GFD diet (“One family described a 90% improvement and another family described an ‘awakening’ from a different level of consciousness”). A small comparison study [ 95 ] of children with ASD who were and were not following the GCFD reported that 7 of 13 children with ASD were already on a GCFD when recruited (outcome measures did not differ significantly from the 6 of 13 who were not on the diet, however parents of all children on the GCFD reported that it had improved symptoms and behaviour). Babinska et al. [ 20 ] found 20.7% of children and adolescents with ASD to follow a diet which in some way restricted gluten (either GFD or GCFD); it was not found that the following of speciality diet correlated with GI symptom severity. Another study [ 96 ] found that 12% of their cohort of children with ASD consumed a GCFD, with these children also more likely to take supplements and overall showing better intake of nutrients including vitamin E, D and magnesium.

Hopf et al. [ 97 ] surveyed parents of children with ASD to identify reasons why they engaged with “complimentary and alternative medicine” (CAM). The GCFD had been used at some point by 54.8% of responders, although this was not rated among the interventions which were perceived as having had the greatest effectiveness (which included sensory integration therapy, melatonin and prescription antifungal medication). A similar study [ 98 ] also focused on the use of CAM in children with ASD as measured by caregiver report. Here, the GFD was followed at a lower rate (10% of the whole group), but was still the most common speciality diet followed. Rubenstein et al. [ 99 ] found 20.4% of children with ASD had ever used a GFD; those currently engaging with it had started on the suggestion of a medical professional in 50.7% of cases. Self-reported (from caregivers) data which predicted use of the GFD included GI conditions and developmental regression. Another experiment [ 100 ] examined all inpatients at a university medical centre, who did not have CD but who followed a GFD, to find predictors as to why in terms of comorbidities. In this, it was observed that having ASD led to an odds ratio of being on the diet of 23.42; by far the highest of all significant conditions reported (the next being irritable bowel syndrome with an OR of 6.16).

Studies which focus more directly on matching parental reporting of dietary practice to behavioural outcomes include Pennesi et al. [ 101 ]. Here, reports from 387 parents/caregivers of children with ASD were examined which focused on GI symptoms, suspected food sensitivities and adoption of speciality diets (primarily GCFD). Within these reports statistical effects were noted wherein greater suspicion of GI problems predicted greater improvement in ASD outcomes following adoption of speciality diets. Strict diet engagement was also observed to be significantly related to better outcomes. Another study [ 102 ] found no associations between dietary intake (which included measurement of gluten) and GI symptoms. However, these authors compared intake of gluten in grams against study outcome and a critical observation may be that gluten often needs to be eliminated entirely to usually see any benefit.

Some research has also focused more on the motivation in parents of children with ASD to adopt a GFD or GCFD. Marsden et al. [ 103 ] noted that parents who adopted these diets for their children with ASD were most influenced by “anticipated regret, positive outcomes and attitude”. Perceived control was also relevant as a factor (with more predicting use of the diet). Tarnowska et al. [ 104 ] also investigated a similar question of what influenced parents of children with ASD to purchase GCFD foods. Packing features such as clear labelling that the food was e.g., gluten-free made them more likely to buy, while social issues around following exclusion diets (e.g., going out for a meal) and the expense/limited range of GCFD foods were seen as negative points. A survey study [ 105 ] found that approximately three quarters of clinical professionals who care for people with ASD had been asked at some point about the GCFD, while 29.5% of parents reported use of the GCFD specifically. Inadequacies with the knowledgebase regarding the use of speciality diets were noted by respondents.

Adoption of the GFD or GCFD in children with ASD is therefore quite pronounced, with lower estimates starting at 10%, and multiple studies reporting >50%. Motivations to engage with the diet appear to revolve around anticipated regret of negative outcomes should it not be tried, as well as a parent having a higher degree of perceived control. A recurring theme in a number of studies is the anecdotal reporting (e.g., by parents) of improvement in ASD outcomes, which are often isolated in incidence but apparently dramatic in effect. Caregivers and clinicians each highlight that greater understanding of how these diets interact with ASD is required.

8. Nutritional Considerations

Limited research has also studied the impact of the GFD or GCFD on the nutritional health of children with ASD. Studies which indicate a positive consequence of following the GFD/GCFD on health include one by Herndon et al. [ 106 ]. While this focused on comparisons between (all) children with ASD compared to TD children, a subgroup analysis revealed those with ASD who followed a GCFD had higher vitamin E intake than those who did not. As already discussed, Stewart et al. [ 96 ] found following a GCFD led to higher levels of vitamin D, E and magnesium, possibly relating to a higher likelihood of simultaneously using supplements compared to those following a regular diet. Supporting this, another study [ 107 ] found that those on a GCFD were far more likely to take vitamin D and calcium supplements; no child who followed the GCFD had a deficiency of 25(OH)D (a marker of bone health), compared to 24% of those on a regular diet who did.

Studies of no or mixed outcomes include one [ 108 ] where no difference was found in nutritional intake between children with ASD who did and did not follow a GCFD (although an overall effect of suboptimal intake was noted across the whole cohort). Analysing food diaries, Mari-Bauset et al. [ 109 ] also reported mixed outcomes wherein children who followed a GCFD had lower BMI and energy, and lower intake of some nutrients (sodium, calcium, phosphorus and pantothenic acid). Conversely however, they had better intake of fiber, legumes, vegetables and fat.

Studies reporting negative impacts of the diet include Arnold et al. [ 110 ], who found a trend for children with ASD who followed the GCFD to have more deficiencies relating to essential amino acids, including tryptophan. This was replicated by another investigation [ 111 ] which also detected lower tryptophan in children with ASD compared to TD controls (it was lowest in those with ASD who followed a restricted diet). These authors hypothesised this may lead to a worsening of ASD symptoms.

9. Synthesis of Literature and Future Directions for Research

The interest of speciality diets in ASD, and particularly the GCFD, has increased markedly in the last 2 decades both in terms of adoption in the ASD community as well as scientific study. Research does convincingly demonstrate certain effects and associations which justify this. Most notably is a modest comorbidity between CD and ASD. Also shown is physiological evidence of inadequate digestion of gluten in people with ASD, leading to elevated “exorphins”/gluten antibodies around which reasonable hypotheses exist regarding downstream negative consequences on the central nervous system. Whether these associations are because of a unique relationship between CD and ASD or because of a predisposition in people with ASD to have a generally higher rate of autoimmune-like features, is yet to be resolved. Further research which directly examines the effects of these gluten products is needed in people with ASD, while additional studies examining shared genetic predispositions would also be warranted and beneficial. There is also a scarcity of epidemiological research characterising the comorbidity of ASD and CD; while one very well powered study does exist and supports this to be the case replication elsewhere is desirable. The availability of newer gluten-related antibodies (e.g., TG6) and the use of native antigliadin antibodies that are known to be sensitive to the whole spectrum of gluten-related disorders, may provide a good opportunity for further large scale epidemiological studies.

Arguably the greatest gap in current literature relates to the lack of tightly designed trials of the GFD, or GCFD in people with ASD. Those that are currently available suffer from very pronounced heterogeneity regarding the intervention followed, sample size, trial duration, blinding and outcomes measured. There does not yet exist an RCT which combines an at-least modest sample size with a placebo-controlled design over a long duration. This would be highly valuable to address, accepting the limitations of the difficulties of such an intervention (gluten free diet) in the context of what is a behaviourally-complex cohort of patients.

Adoption of the GFD and GCFD appears to be very high amongst people with ASD. An impression is gained of strong anecdotal evidence of a benefit in relevant studies, although statistical associations do not as often bear this out. It is also unclear if any reported behavioural benefits would be because of a direct interaction between physiological gluten/casein-related impacts on the brain, or if engaging with speciality diets simply reduce non-specific GI symptoms and thus improve quality of life in a more general sense.

The nutritional impact of a GCFD on the child with ASD generally seems to be slight, or even associated with improved intake. However, some studies showing deficiencies of certain nutrients highlight the need to still maintain a balanced diet once on a restricted one. Finally, a general observation is the abundance of research which focuses on children. Of the initial papers found in the literature review, 68 (of 79) studied groups which were exclusively children, or mixed children and adolescents. While this is likely an outcome of opportunity and convenience sampling effects, it is nonetheless a strong bias within the available literature and means that generalisation of anything discussed in this review to adults with ASD is difficult.

10. Conclusions

This review highlights a modest comorbidity between ASD and CD, and a base of evidence on which reasonable hypotheses may be built to explore if gluten has a generally adverse effect in exacerbating the symptoms and quality of life in children with ASD. However, a negative effect of gluten ingestion in ASD has not been proved. Trials which have sought to demonstrate this are variable in their findings, and suffer from issues with experimental design and execution which means that an overall interpretation cannot yet be made. Efforts should focus on future studies which address limitations detailed here to create an RCT from which confident conclusions can be drawn. As diets which restrict gluten see a very high adoption rate among people with ASD, such further research is certainly warranted.

Author Contributions

I.D.C.: methodology, data curation, writing-original draft preparation, writing-review and editing; N.H.: writing-review and editing, supervision; M.H.: conceptualization, writing-review and editing, supervision. All authors have read and agreed to the published version of the manuscript.

This research received no external funding.

Conflicts of Interest

The authors declare no conflict of interest.

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