Review Article

Exploring the villus

Arzu Ensari, Michael N Marsh

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 181-190

The small intestinal villus and its associated epithelium includes enterocytes as the main cell type and differentiated goblet and argentaffin cells, while the invaginated crypt epithelium is the site of cell division and hence the origin of all epithelial components. Enterocytes form a cohesive monolayer which acts both as a permeability barrier between lumen and the interior, and an important gateway for nutrient digestion, absorption and transport. Differentiation and polarisation of enterocytes depends on cytoskeletal proteins that control cell shape and maintain functionally specialised membrane domains; extracellular matrix (ECM) receptors; channels and transporters regulating ion/solute transfer across the cell. The mesenchymally-derived basement membrane dynamically controls morphogenesis, cell differentiation and polarity, while also providing the structural basis for villi, crypts and the microvasculature of the lamina propria so that tissue morphology, crucially, is preserved in the absence of epithelium. Mucosal re-organisation requires immense cooperation between all elements within the lamina, including marked revisions of the microvasculature and extensive alterations to all basement membranes providing support for endodermal and mesenchymal components. In this context, subepithelial myofibroblasts fulfil important regulatory activities in terms of tissue morphogenesis; remodelling; control of epithelial cell development, polarity and functional attributes; and an intimate involvement in repair, inflammation and fibrosis.

This paper reviews the main structural and functional aspects of the villus, including the epithelium and its outer glycocalyx and microvillous border; and subjacent to the epithelium, the basement membrane with its attached web of myo-fibroblasts together with the lamina propria core of the villi, and its microvasculature and lacteals. Finally, some comments on the rapidity with which the overall structure of the villi changes in their response to both external, and internal, influences.

Keywords: Small intestinal villus, Permeability, Epithelium.

(Please cite as: Ensari A, Marsh MN. Exploring the villus. Gastroenterol Hepatol Bed Bench 2018;11(3):181-190).

Role of biopsy in diagnosis and treatment of adult celiac disease

Hugh James Freeman

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 191-196

Celiac disease is an immune-mediated enteropathy that characteristically responds to treatment with a gluten-free diet.  In most, clinical features improve with resolution of diarrhea and weight loss.  Serological studies also tend to normalize.  Despite this, a mucosal inflammatory process may persist, especially in the proximal small intestine.  In celiac disease, resolution of histopathological changes can occur within 6 months, but often, more than a year is required, sometimes, 2 years or more.  Changes are not only time-dependent, but appear to be gender-dependent with resolution more readily achieved in females compared to males, and age-dependent with more persistence of the inflammatory process in the elderly compared to younger patients.  Future studies need to take into account the individual nature of the normal mucosal healing process in celiac disease treated with a gluten-free diet.

Non Celiac Gluten Sensitivity and diagnostic challenges

G Casella, V Villanacci, C Bella Di, G Bassotti, J Bold, Kamran Rostami

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 197-202

Non-celiac gluten sensitivity (NCGS), also referred to as non-celiac wheat sensitivity (NCWS), is a clinical syndrome characterized by both intestinal and extra-intestinal symptoms responsive to the withdrawal of gluten-containing food from the diet. The aim of this review is to summarize recent advances in research and provide a brief overview of the history of the condition for the benefit of professionals working in gastroenterology. Academic databases such as PubMed and Google Scholar were searched using key words such as ”non-celiac gluten sensitivity”, “gluten related disorders”, and the studies outlined in reference page were selected and analysed.

 Most of the analysed studiers agree that NCGS would need to be diagnosed only after exclusion of celiac disease and wheat allergy, and that a reliable serological marker is not available presently. The mechanisms causing symptoms in NCGS after gluten ingestion are largely unknown, but recent advances have begun to offer novel insights. The estimated prevalence of NCGS, at present, varies between 0.6 and 6%. There is an overlap between irritable bowel syndrome and NCGS with regard to the similarity of gastrointestinal symptoms. The histologic characteristics of NCGS are still under investigation, ranging from normal histology to slight increase in the number of T lymphocytes in the superficial epithelium of villi. Positive response to gluten free diet for a limited period (e.g., 6 weeks), followed by the reappearance of symptoms after gluten challenge appears, at this moment, to be the best approach for confirming diagnosis. The Salerno expert criteria may help to diagnose NCGS accurately in particular for research purposes but it has limited applicability in clinical practice.

Keywords: Celiac disease, Non celiac gluten sensitivity, Wheat allergy.

(Please cite as: Casella G, Villanacci V, Di Bella C, Bassotti G, Bold J, Rostami K. Non celiac gluten sensitivity and diagnostic challenges. Gastroenterol Hepatol Bed Bench 2018;11(3):197-202).

Biological markers for non celiac gluten sensitivity: a question awaiting for a convincing answer

Enzo Ierardi, Giuseppe Losurdo, Domenico Piscitelli, Floriana Giorgio, Annacinzia Amoruso, Andrea Iannone, Mariabeatrice Principi, Alfredo Di Leo

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 203-208

Non Celiac Gluten Sensitivity (NCGS) is characterized by immunological, morphological or symptomatic manifestations precipitated by gluten ingestion in individuals without celiac disease (CD). The most important challenge in NCGS is the diagnosis, currently based only on clinical observation. The “Salerno criteria” have been pointed out to achieve a reliable diagnosis even if they lack immediacy and practicality, thus making questionable patient’s adherence. Therefore, biological indicators supporting the clinical diagnosis of NCGS are advisable. For these reasons, many attempts have been performed in order to identify possible serological, immunological, histopathological, immunohistochemical and pathophysiological aspects characterizing this condition with the aim of using them for diagnostic purposes. In the present narrative review, we carried out an update of the current scenario of potential markers of NCGS. The main fault of available studies is that, in most cases investigations have been pointed out towards molecules, which cannot be searched in the current laboratories of clinical analysis. Therefore, the matter has been confined within basic research. Additionally, in these studies, sensitivity and specificity of biological markers were not computable. This is a relevant limit, since an ideal test for NCGS should have a good discriminative power against both CD and other causes of microscopic enteritis. Until now, serological tests have failed, while the search for a soluble marker indicative of activation of innate immune system as well as immunohistochemistry could be the promising bases for the development of appropriate investigations in the future.

Keywords: Gluten sensitivity, Marker, Diagnosis, Cytokines, Immunohistochemistry.

(Please cite as: Ierardi E, Losurdo G, Piscitelli D, Giorgio F, Amoruso A, Iannone A, et al. Biological markers for non-celiac gluten sensitivity: a question awaiting for a convincing answer. Gastroenterol Hepatol Bed Bench 2018;11(3):203-208).

Short Review

The serological diagnosis of coeliac disease - a step forward

geoffrey Holmes, Carolina Ciacci

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 209-215

The development of highly performing serological tests to identify patients with coeliac disease (CD), allowed large scale screening studies to be carried out and the results transformed our understanding of the prevalence of the condition in the general population. The next logical step was to ask whether CD could be reliably diagnosed by these tests without the need for small intestinal biopsies. This was shown to be the case. Studies from Derby, UK, indicated that about half of adult patients can be diagnosed in this way and similar figures have been provided for children. When considering this approach, it is essential that laboratories only use highly performing test kits that they have validated to measure tissue transglutaminase antibodies because all kits do not function to the same high standard. There remains a place for biopsy when criteria for serological diagnosis are not met, if the diagnosis of CD is strongly suspected but serological tests are negative or in patients not showing the expected responses to gluten free diet or otherwise causing concern, when not only small bowel biopsy will be indicated but also other investigations. Those with refractory CD should not be compromised by this diagnostic strategy. As serological tests become more refined and information accumulates, it is likely that this mode of diagnosis will gather momentum for the benefit of patients and carers. This brief review looks at the evidence for making the diagnosis of CD in some cases by serological tests alone.

Keywords: Celiac disease, Tissue transglutaminase, Serology, Diagnosis.

(Please cite as: Holmes G, Ciacci C. The serological diagnosis of coeliac disease - a step forward. Gastroenterol Hepatol Bed Bench 2018;11(3):209-215).

Original Article

Celiac disease microarray analysis based on system biology approach

Mostafa Rezaei –Tavirani, Davood Bashash, Fatemeh Tajik Rostami, Sina Rezaei Tavirani, Abdolrahim Nikzamir, Majid Rezaei Tavirani, Mohammad Hossain Haidari

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 216-224

Aim: Aim of this study is screen of the large numbers of related genes of CD to find the key ones.

Background: Celiac disease (CD) is known as a gluten sensitive and immune system dependent disease. There are several high throughput investigations about CD but it is necessary to clarify new molecular aspects mechanism of celiac.

Methods: Whole-genome profile (RNA) of the human peripheral blood mononuclear cells (PBMCs) as Gene expression profile GSE113469 was retrieved Gene Expression Omnibus (GEO) database.  The significant genes were selected and analyzed via protein-protein interaction (PPI) network by Cytoscape software. The key genes were introduced and enriched via ClueGO to find the related biochemical pathways.

Results: Among 250 significant genes 47 genes with expressed change above 2 fold change (FC) were interacted and the constructed network were analyzed. The network characterized by poor connections so it was promoted by addition 50 related nodes and 18 crucial nodes were introduced. Two clusters of biochemical pathways were identified and discussed.

Conclusion: There is an obvious conflict between microarray finding and the well-known related genes of CD. This problem can be solve by more attention to the interpretation of PPI ntwork analysis results.

Keywords: Celiac disease, System biology, Crucial genes, Cytoscape, ClueGO.

(Please cite as: Rezaei Tavirani M, Bashash D, Tajik Rostami F, Rezaei Tavirani S, Nikzamir A, Rezaei Tavirani M, et al. Celiac Disease Microarray Analysis based on System Biology Approach. Gastroenterol Hepatol Bed Bench 2018;11(3):216-224).

Pentraxin 3 and biopsy status in celiac patients

Roberto Assandri, Alessandro Montanelli

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 225-232

Aim: In our study we explored a possible relationship between PTX3 and CD.

Background: Gluten sensitivity is known as a hallmark of celiac disease (CD). The diagnosis of CD requires demonstration of a typical enteropathy, and positive serology supports the diagnosis. The CD immune response involves the adaptive, as well as the innate immunity and is characterized by the presence of anti-gliadin (AGA) and anti-transglutaminase 2 antibodies (tTGA), lymphocytic infiltration in the intestinal epithelial membrane and expression of multiple cytokines. The long pentraxin 3 (PTX3), an acute-phase inflammatory molecule, plays an important role in innate immunity.

Methods: 108 CD patients were divided according to Marsh Histological grade following Marsh criteria classification in three groups: Group 1: Marsh 0, patients with a known history of CD under gluten free diet, complete remission; Group 2: Marsh1 and Marsh 2; Group 3: Marsh 3.

Healthy age-matched controls without a known history of CD or gastrointestinal symptoms (n=30) served as controls. PTX3 serum levels were measured by sandwich ELISA on an automated platform.

Results: PTX3 serum levels were significantly elevated in group 3 and group 2 compared with HC (mean 3.31± 1.27 ng/mL and 3.97 ± 0.54 ng/mL versus 1.06 ± 0.59 ng/mL; P < 0.005), with group 1 (0.76±0.31 ng/mL). No statistically significant differences were found between group 1 and HC group. We found a strong linear correlation between PTX3 serum levels and AGA levels in group 2 (r=0.78, P <0.0001), and group 3 (r =0.63, P < 0.005) but no correlations were detected between PTX3 serum levels and tTGA levels (group 2, r= 0.04; group 3, r=0.24). Serological data revealed that PTX3 correlated with major gastrointestinal damage patients.

Conclusion: PTX3 is a component of the humoral arm of the innate immune system. Our data showed that PTX3 serum levels were high in active disease patients with pathological levels of AGA. We also demonstrated that patients with normal AGA IgA levels had PTX3 serum levels compared to healthy control. We hypothesized that PTX3 is able to modulate the innate response to gliadin in CD and it could regulate the adaptive immune response.

Keywords: Gluten-sensitivity, Innate immune response, Adaptive immune response, Pentraxin 3, Antigliadin antibodies, Celiac disease.

(Please cite as: Assandri R, Montanelli A. Pentraxin 3 and biopsy status in celiac patients. Gastroenterol Hepatol Bed Bench 2018;11(3):225-232).

The influences of dried Chicory root and White lupine added to food on jejunal morphology: experimental study

Peter Makovicky, Zdenek Volek, Linda Uhlirova, Pavol Makovicky

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 233-238

Aim: The objective of this work was to test the effects of adding dried Chicory root and White lupine food on small bowel morphology and compare it to a standard commercial diet.

Background: Various commercial gluten-free products, gluten-free raw materials and gluten-free plants are this time available on the food market, but there are still not enough information about their effect on the small bowel morphology.

Methods: Altogether thirty rabbits were used in this study. The control diet (C) contained common feed components. The first experimental diet (E1) contained (per kg) 60 g of dried chicory roots instead of barley, whereas the second experimental diet (E2) was based on white lupine seeds (cv. Amiga; 120 g per kg diet) instead of the soybean meal used in the control diet. The experiment started when the rabbits were 34-days old and lasted until they were 55-days old. At the end, one jejunal small bowel tissue was sampled, and both the heights and depths of the villi and crypts were measured.

Results: The highest villi were measured in the E1 (598.99 µm) group, mean in the C (590.30 µm) group and the lowest were in the E2 (563.74 µm) group. The most intense mucin villous positivity was observed in the E2 group, followed by the E1 group, and the weakest positivity was found in the visible C group.

Conclusion: Chicory root has practical uses in gluten-free industries.

Keywords: Celiac disease, Gluten-free diet, Nutrition, Small bowel, Villi intestinalis.

(Please cite as: Makovicky P, Volek Z, Uhlirova L, Makovicky P. The influences of dried Chicory root and White lupine added to food on jejunal morphology: experimental study. Gastroenterol Hepatol Bed Bench 2018;11(3):233-238).

Prevalence of celiac disease serological markers in a cohort of Italian rheumatological patients

Giacomo Caio, Roberto De Giorgio, Francesco Ursini, Silvia Fanaro, Umberto Volta

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 239-243

Aim: To assess the prevalence of celiac disease (CD) serological markers in a cohort of patients referred to an Italian rheumatological outpatient clinic.

Background: Current guidelines do not suggest CD screening in patients with rheumatological diseases and these subjects are not considered to be at high risk for CD.

Methods: A total of 230 sera of rheumatological patients referred to the Division of Internal Medicine at the Department of Medical and Surgical Sciences between January 2005 and December 2013 were screened for CD by testing IgA antitransglutaminase (TTG IgA), IgG deamidated gliadin peptides (DGP IgG) and IgA antiendomysium (EMA) antibodies. Of the 230 patients tested, 67 had a diagnosis of rheumatoid arthritis (RA), 52 Sjögren’s syndrome (SjS), 42 systemic sclerosis (SCL), 35 systemic lupus erythematosus (SLE), 15 mixed connective tissue disease, 11 polymyositis and 10 dermatomyositis.

Results: TTG IgA antibodies were identified in 7/230 cases (3%), 3 in SjS (3/42 – 5.8%), 2 in SCL (2/42 – 4.8%), 1 in RA (1/67 – 1.5%) and 1 in SLE sera (1/35 – 2.8%). All the seven sera were also positive for DGP IgG and EMA IgA. DGP IgG were the most frequent antibody detected, being found in 16 (7%) sera.

Conclusion: This study identified a high prevalence of CD antibodies in adult patients referred to a rheumatology outpatient clinic. These results highlight the importance of CD screening in subjects presenting with rheumatological features.

Keywords: Celiac disease, Rheumatological disorders, Screening, Anti-transglutaminase antibodies, Anti-deamidated gliadin peptide antibodies, Anti-endomysium antibodies.

(Please cite as: Caio G, De Giorgio R, Ursini F, Fanaro S, Volta U. Prevalence of celiac disease serological markers in a cohort of Italian rheumatological patients. Gastroenterol Hepatol Bed Bench 2018;11(3):244-249).

A rapid and sensitive assay to identify HLA-DQ2/8 risk alleles for celiac disease using real-time PCR method

Kazem Mashayekhi, Mohammad Rostami Nejad, Davar Amani, Mostafa Rezaei-Tavirani, Hamid Mohaghegh Shalmani, Mohammad Reza Zali

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 250-258

Aim: To perform a simple, rapid and sensitive Real-time PCR based SYBR Green method to determine the human leukocyte antigen (HLA)-DQ 2/8 alleles in celiac disease (CD) patients.

Background: Many molecular techniques are available to determine the HLA-DQ2 and DQ8 alleles, but they are too expensive and have many steps that make them difficult to use.

Methods: To determine the HLA-DQ 2/8 alleles we have developed a new real-time PCR assay, using SYBR Green technique with melting curve analysis on genomic DNA isolated from 75 CD patients and 94 healthy controls. The specific primers to examine HLA-DQA1*05, HLA-DQB1*02 and HLA-DQB1*0302 alleles were used and results were compared with commercially available kits.

Results: Using this method, the presence of HLA-DQ2 and HLA-DQ8 alleles were determined with sensitivity and specificity 80% and 100% respectively and compared to low resolution commercially available kits, the results of this method were more efficient. The frequency of DQ2 and DQ8 in patients was 76% and 29%, respectively and overall 96% of patients were carries DQ2 and/or DQ8 alleles.

Conclusion: The result of this study showed that Real-time PCR using SYBR Green method with melting curve analysis has good efficiency to identify the HLA-DQ2/8 risk alleles.

Keywords: Celiac disease, Real-time PCR, Melting curve analysis, HLA-DQ2/8 alleles, HLA typing.

(Please cite as: Mashayekhi K, Rostami-Nejad M, Amani D, Rezaei-Tavirani M, Mohaghegh-Shalmani H, Zali MR. A rapid and sensitive assay to identify HLA-DQ2/8 risk alleles for celiac disease using real-time PCR method. Gastroenterol Hepatol Bed Bench 2018;11(3):250-258).

An exploration into the motivation for gluten avoidance in the absence of coeliac disease

Lucy Harper, Justine Sara Bold

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 259-268

Aim: To explore the motivation for gluten avoidance in the absence of coeliac disease (CD) and ascertain what symptoms are triggered by gluten and what beliefs/reasons influence this decision.

Background: Links between physical/psychological symptoms and gluten in CD are well known but less is known about those who self-select a gluten-free diet (GFD) in the absence of CD.

Methods: An empirical study using responses to an anonymous on-line questionnaire. Closed questions were used as a screening tool to exclude participants who had CD, wheat allergy or were following a low FODMAP diet. Data from participants using a GFD in the absence of a medical diagnosis was then analysed using thematic analysis.

Results: 120 initial responses, 87 were completed in full. 23 respondents fulfilled the inclusion criteria for thematic analysis. 7 different themes emerged, including one for signs/symptoms. Other themes identified included difficulties of a GFD, health beliefs, feelings and influence on decision to follow a GFD. Responses indicate that the reasons for gluten avoidance are in the most part reasoned and logical and were based around participants’ self-management of symptoms.

Conclusion: Symptoms included those typical of irritable bowel syndrome (IBS), but also infertility, low mood/energy, immune function and weight management and visual and auditory hallucinations. It appears the majority of responses analysed thematically could fit into the spectrum of non-coeliac gluten sensitivity (NCGS).   Findings also suggest more support at all levels of medical care may help patients establish if it is gluten, rather than wheat or FODMAPs particularly fructans that are contributing to signs/symptoms.

Keywords: Non-coeliac gluten sensitivity, Non-celiac wheat sensitivity, Gluten intolerance, Gluten-free, Coeliac disease, Self-management of symptoms.

(Please cite as: Harper L, Bold J. An exploration into the motivation for gluten avoidance in the absence of coeliac disease. Gastroenterol Hepatol Bed Bench 2018;11(3):259-268).

Case Report

Detection of asymptomatic celiac disease in two siblings from a mother with non-celiac gluten sensitivity

Umberto Volta, Giacomo Caio, Roberto Manfredini, Roberto De Giorgio

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 269-272

Non-celiac gluten sensitivity and celiac disease are known to be two distinct clinical entities, however, non-celiac gluten sensitivity has been detected in a proportion of first-degree relatives of celiac patients. Herein for the first time we describe the occurrence of asymptomatic celiac disease in two siblings, a girl and a boy, whose mother suffered from a proven non-celiac gluten sensitivity. Both the 12-year old girl and 9-year old boy were positive for anti-endomysial and anti-tissue transglutaminase antibodies of IgA class at a very high and low titer, respectively.  Duodenal biopsy confirmed the diagnosis of active celiac disease (severe villous flattening) in the girl, whereas her brother had Marsh 1 lesion consistent with a potential celiac disease. This case report indicates that antibody screening for celiac disease can be recommended in any symptomatic or asymptomatic first-degree relatives of patients with non-celiac gluten sensitivity.

Keywords: Celiac disease, Non-celiac gluten sensitivity, Screening, Serology, Relatives, Familiarity.

(Please cite as: VoltaU, CaioG, ManfrediniR, De Giorgio R. Detection of asymptomatic celiac disease in two siblings from a mother with non-celiac gluten sensitivity. Gastroenterol Hepatol Bed Bench 2018;11(3):269-272).

Post Pancreatitis/Cholecystectomy Gluten Intolerance

Ivan Tang, George MacFaul, Ravi Madhotra, Kamran Rostami

Gastroenterology and Hepatology from Bed to Bench, Vol. 11 No. 3 (2018), 2 June 2018, Page 273-275

This case report describes the journey of a patient who suffered from life-limiting gastrointestinal symptoms after an acute bout of pancreatitis following ERCP for cholelithiasis bile following a ductal stone, and subsequent cholecystectomy. She was diagnosed and treated for IBS with medication without significant improvement.  On implementation of a simple gluten and lactose exclusion diet she recovered to her premorbid state, and trials of gluten challenge triggered flares of symptoms. This case report will go on to discuss current evidence for use of gluten and lactose exclusion diets in some gluten sensitive patients misdiagnosed with IBS.

Keywords: Gluten sensitivity, Non-coeliac, Post-pancreatitis, Post-cholecystectomy gluten intolerance.

(Please cite as: Tang I, MacFaul G, Madhotra R, Rostami K. Post pancreatitis/cholecystectomy gluten intolerance. Gastroenterol Hepatol Bed Bench 2018;11(3):273-275).