Hereditary α-tryptasemia; a review of mechanisms linking α-tryptase gene dosage to intestinal homeostasis and immunopathology.

Covering the Cover

A Common Barrier Defect for Celiac Disease and Ulcerative Colitis

Celiac Disease.

Bone mass and bone metabolism in pediatric gastrointestinal disorders.

Objective To explore the function and significance of interleukin(IL)-25 in the pathogenesis of inflammatory bowel disease (IBD) through testing its expression in the intestinal mucosal and serum of patients with IBD. Methods Intestinal or colonic mucosal biopsies of 12 patients with ulcerative colitis (UC), 16 patients with Crohn's disease (CD) and 10 healthy controls were collected. The expression of IL-25 at mRNA level was detected by real-time PCR and the situ expression of IL-25 in intestinal mucosa was analyzed with immunohistochemistry. At same time,serum of 20 UC patients, 24 CD patients and 20 controls was collected, and IL-25 concentration in the serum was determined by enzyme-linked immunosorbent assay (ELISA). Results Compared with healthy controls, the expression of IL-25 at mRNA level in inflamed mucosa of CD and UC patients was significantly decreased (P＜0.05), no statistic difference between UC and CD groups (P＞0.05).The immunohistochemistry results indicated that more IL-25 positive cells in normal lamina propria,the expression of IL-25 in mucosal epithelia cells was low, the expression of IL-25 protein in the intestinal or colonic mucosa of UC and CD patients significantly decreased ( P ＜ 0.05), no statistic difference between UC and CD groups (P＞0. 05). ELISA results showed that the level of IL-25 in the serum of UC and CD groups was significantly lower than healthy controls (P ＜ 0.05). Conclusion The expression of IL-25 in the intestinal mucosa and serum of IBD patients was significantly decreased,which suggested that IL-25 expression defects may closely related to the pathogenesis of IBD. IL-25 may be a new target for the IBD treatment. Key words: Crohn's disease; Inflammatory bowel disease; Ulcerative colitis; Interleukin-25

Objective To investigate the expression of microRNA (miRNA)-10a in the intestinal mucosa,serum and peripheral blood mononuclear cell (PBMC) of patients with inflammatory bowel disease (IBD) and explore its role and relevance in the pathogenesis of the disease.Methods The intestinal or colonic mucosal biopsy specimens of nine active ulcerative colitis (UC) patients,11 active Crohn's disease (CD) patients and eight patients with negative colonoscopy result as control were collected.The sera of 12 active UC patients,13 active CD patients and nine healthy controls were collected.The PBMC of nine active UC patients,11 active CD patients and eight healthy controls were collected.The expression of miRNA-10a in the intestinal mucosa,sera and PBMC and the expression of IL-12/IL-23 p40 in the intestinal mucosa were detected by real-time polymerase chain reaction (PCR).Each 8 cases of active UC and CD patients were collected.The intestinal mucosa before infliximab (IFX) treatment and six weeks after three times of IFX treatment were collected.And at same time,the intestinal mucosa of 11 active UC patients and 10 active CD patients were collected and cultured for 18 hours stimulated with IFX in vitro and then the expression of miRNA-10a in the intestinal mucosa was tested.One-way analysis of variance was used for comparison in three samples.Paired t-test was used for two samples comparison.Spearman test was used for correlation analysis.Results Compared with healthy controls,the expression of miRNA-10a in the intestinal mucosa,serum and PBMC of UC and CD patients significantly decreased (F=38.45,30.46 and 14.74,all P＜0.05).There was no statistic significance between UC and CD groups.The expression of IL-12/IL-23 p40 in the intestinal mucosa of UC and CD patients significantly increased (F=32.90,P＜0.05).The expression of IL-12/IL-23 p40 was negatively correlated with the expression of miRNA-10a in the intestinal mucosa of CD patients.After three times of IFX treatment,the expression of miR-10a in the intestinal mucosa of IBD patients significantly increased (t=3.341,3.382,both P＜0.05).After stimulated with IFX in vitro,the expression of miRNA-10a in the intestinal mucosa significantly increased (t=3.095,7.193,both P＜0.05).Conclusions miRNA-10a was closely correlated with the inflammation of IBD patients and with the role of targeting IL-12/IL-23 p40.miRNA-10a might be a new target for the IBD treatment. Key words: Crohn disease; Colitis, ulcerative; MicroRNAs; Interleukin-12; Interleukin-12 subunit p40

BACKGROUND A small fraction of Inflammatory bowel disease (IBD) subjects have celiac disease (CeD). We conducted a retrospective case-control study to compare disease characteristics of subjects with IBD vs IBD+ CeD. METHODS A retrospective study was conducted from January 2017 to June 2022 using appropriate ICD-10 codes for IBD and CeD. Data was collected including demographics, disease phenotype, and medication use patterns. Disease groups included IBD, IBD+ CeD with subgroup analyses being further performed as follows: Crohn’s disease (CD), CD+ CeD, ulcerative colitis (UC), UC+ CeD. RESULTS Females consisted of 59% of IBD (N=72) and 64% of IBD+ CeD (N=36) (p=0.62). Median age at diagnosis of IBD was 24.5 years in the IBD+CeD group compared to IBD group (43.5 years), p=0.003. Median duration of IBD since IBD diagnosis was longer (IBD:16.4 yrs vs IBD +CeD: 8.4 yrs, p< 0.001) with the age at the end of study being older in IBD subjects (64 yrs) vs. IBD+ CeD (34 yrs), p< 0.001. Subgroup analysis (Table 1) based on Montreal classification revealed a higher prevalence of A2 in CD+ CeD (56%) vs CD (24%) and A3 in CD (68%) vs CD+ CeD (22%), p=0.007. There was a higher prevalence of autoimmune conditions in the IBD+ CeD group (27%) in comparison to IBD group (6.9%), p< 0.003 with rheumatoid arthritis (RA) and thyroiditis occurring more commonly in the IBD+ CeD group (Fig 1). Immuno-modulator and biologic use (anti-TNF, anti-integrin and anti-IL12/23) use was significantly higher in the UC+ CeD group at 33%, 39%, 33%, 11% respectively when compared to 10%, 6.5%, 3.2% and 3.2% in UC subjects (p < 0.05 to < 0.001). Immuno-modulator and biologic use (anti-integrin and anti-IL12/23) use were significantly higher in the CD+ CeD group at 50%, 33%, and 33% respectively when compared to 24%, 2.4%, and 10% in CD subjects (p < 0.05 to < 0.001). CONCLUSIONS The prevalence of autoimmune diseases, especially autoimmune thyroiditis and RA is higher in IBD+ CeD subjects with a greater preponderance in UC+ CeD. Further, IBD+ CeD subjects tend to be younger at disease onset than subjects with IBD alone. Use of steroids and escalation to biologics is more common in IBD+ CeD subjects than IBD subjects. These findings should encourage clinicians to assess younger IBD patients for CeD and autoimmune conditions including thyroiditis and RA. Further, early use of biologics may become necessary in this subset of IBD patients.

Despite improved therapies targeting inflammation in inflammatory bowel disease, persistent gastrointestinal and extraintestinal symptoms in those believed to be in remission are highly prevalent with significant impact on patient function and quality of life. This review aims to summarize the scale of the problem, to identify the limitations of previous studies, and to propose how these may be addressed. The published literature was extensively reviewed. Persistent gastrointestinal symptoms are reported in up to 40% of patients with inflammatory bowel disease in apparent remission and have mostly been characterized using the Rome criteria, which are unvalidated in this population. They are also closely linked to anxiety/depression and fatigue. Importantly, studies have frequently failed to use tests with high negative predictive values to exclude intestinal inflammation and have assumed severe symptoms are inflammatory in nature. The presence of ongoing inflammation and different mechanisms underlying the development of symptom-generating abnormalities, including visceral hypersensitivity and central sensitization, that are shared by disorders of gut-brain interaction are evident in the small number of studies performed. Thus, the concept that inflammatory and noninflammatory symptoms are mutually exclusive may be fallacious. Persistent gastrointestinal symptoms are common in patients with inflammatory bowel disease in apparent remission, but variable criteria to define inflammatory remission may introduce a high risk of bias within existing literature. Further research using objective and robust measures of inflammatory remission is key to better defining this population and to clarifying pathophysiological mechanisms so that effective management strategies can be developed.

Objective To investigate the expression of CD27-CD70 co-stimulatory pathway in peripheral circulation and intestinal mucosa of patients with inflammatory bowel disease, and to find the difference between the expression of CD27-CD70 in patients with inflammatory bowel disease and in healthy controls. Methods A total of 62 patients with Crohn's disease, 64 patients with ulcerative colitis and 56 healthy controls were enrolled. Enzyme-linked immunosorbent assay was applied to evaluate plasma CD27-CD70 protein expression in patients with inflammatory bowel disease and healthy controls. SYBR-green real time PCR was applied to access CD27-CD70 mRNA expression in peripheral blood mononuclear cells in patients with inflammatory bowel disease and healthy controls.And CD27-CD70 protein expression in intestinal mucosa was determined by immunohitochemistry.Results Plasma levels of CD27 (P=0. 025) and CD70 (P=0. 000) were significantly higher in patients with Crohn's disease than in healthy controls. However, CD27 (r= 0. 055, P= 0. 673) and CD70 (r= 0. 024, P = 0. 852) were not significantly associated with endoscopic disease activity in patients with Crohn's disease. Similarly, CD27 (P=0. 001) and CD70 (P=0. 000) were significantly higher in patients with ulcerative colitis than in healthy controls. And CD27 (r=0. 077, P=0. 547)and CDT0 (r=0.021, P=0. 869) were not significantly associated with endoscopic disease activity in patients with ulcerative colitis. Moreover, CD27 and CD70 mRNA expression in peripheral blood mononuclear cells were significantly higher in patients with Crohn's disease and ulcerative colitis than in healthy controls (all P=0. 000), and immunostaining indicated that CD27 and CD70 expression in intestinal mucosa were significantly higher in patients with Crohn's disease and ulcerative colitis than in healthy controls (all P=0. 000). Conclusions CD27-CD70 pathway activated in plasma, peripheral blood mononuclear cells and intestinal mucosa of patients with inflammatory bowel disease. However,plasma levels of CD27 and CD70 can not reflect endoscopic disease activity. Key words: Crohn's disease; Ulcerative colitis; Antigens; CD27; Antigens; CD70

Introduction: Inflammatory bowel disease (IBD) has been associated with large-vessel vasculitis (LVV), with the diagnosis of IBD preceding that of LVV by years. We present for the first time in known literature a triad of concurrent ulcerative colitis (UC), aortitis and celiac disease. Case Description/Methods: A 58 year old Hispanic man with a history of hypertension and gout presented with two weeks of intractable temporomandibular headaches, and two months of non-bloody diarrhea and weight loss. Physical exam was unremarkable. Labs showed hemoglobin 6.9 g/dL, erythrocyte sedimentation rate 120 mm/hr, C-reactive protein 281 mg/dL and IgA tissue transglutaminase antibody 23 U/mL. ANA, C3, C4, proteinase-3 and myeloperoxidase antibodies were within normal limits. Colonoscopy showed pancolitis from rectum to ascending colon. The terminal ileum was normal. Abdominal MRI found aortic wall hyperintensity from the renal arteries to common iliac bifurcation. CT angiogram showed wall thickening of the left carotid artery, aortic arch, descending thoracic and abdominal aorta, consistent with vasculitis. Patient was given stress dose steroids with improvement in headache and normalization of ESR and CRP. Temporal artery biopsy was unremarkable. Four months after hospitalization, repeat colonoscopy with duodenal biopsies for celiac disease revealed mild increase in intraepithelial lymphocytes with preserved villous architecture. He was started on a gluten free diet and adalimumab in combination with methotrexate for UC and LVV. Discussion: About 10 case reports of patients with both UC and either Takayasu (TAK) or giant cell arteritis (GCA) have been described, with UC typically diagnosed 15-45 years before the vasculitis. Vasculitis in the GI tract can mimic IBD, making colonoscopy and biopsy crucial for diagnosis. HLA haplotypes A24, B52, and DR2 are associated with both UC and aortitis and Interleukin-9, observed in temporal arteritis lesions, may be implicated in the pathogenesis of UC. Shared chromosomal variants between patients with UC and celiac disease may explain why IBD risk is up to 9-fold higher in patients with celiac disease. Our patient may have presented with isolated aortitis or an early form of GCA. Methotrexate is used to treat LVV and is combined with an anti-TNF agent to treat UC, as in our case. This is the first known report of co-occurring UC, celiac disease and aortitis; however, whether the three inflammatory conditions are mechanistically related warrants further research.

Rationale Accumulation and activation of mast cells and eosinophils have been implicated in the pathogenesis of several chronic inflammatory gastrointestinal (GI) diseases, including eosinophilic gastrointestinal diseases (EGIDs) and inflammatory bowel disease (IBD). Despite the strong association of mast cell and eosinophil numbers and activation with the pathogenesis of IBD, no further characterization of these cells has been performed. Current treatment options for IBD include aminosalicylates, antibiotics, immunomodulators, biologic agents and small molecules. These therapies are only moderately effective. A significant proportion of patients fail to respond, do not fully respond, or lose response over time. Therefore, there is significant need for more selective and effective therapy options. Siglec-8 is an inhibitory receptor selectively expressed on human eosinophils and mast cells and represents a novel target for the treatment of IBD with the anti-Siglec-8 mAb, antolimab (AK002). We aimed to quantify and evaluate the activation state of mast cells and eosinophils in colonic tissue from IBD or non-diseased patients. In addition, we quantified the production of TNFa from human colon tissue mast cells and evaluated the inhibitory activity of antolimab (AK002) on these cells. Methods Single-cell suspensions were prepared by enzymatic digestion of fresh colonic biopsies from patients clinically diagnosed with IBD (n=29) or non-diseased control tissues (n=16). Multi-color flow cytometry was performed to identify major immune cell populations and evaluate the activation state of mast cells and eosinophils. Mast cells were FACS-sorted from human colon tissue to evaluate cytokine production and inhibitory activity of antolimab. Results The percentage of mast cells and the expression of the mast cell degranulation marker CD107a were significantly increased in ulcerative colitis (UC) patient biopsy tissue compared to Crohn’s disease (CD) and non-diseased colonic tissue (Figure 1A and B). Furthermore, FACS-sorted mast cells from human colon tissue produced significant quantities of TNFa that was reduced after ex vivo antolimab treatment. Colonic tissue eosinophils were also elevated in a subset of UC and CD patient biopsies, and all UC and CD tissue eosinophils displayed increased expression of the activation marker CD11b compared to control colonic tissue. Conclusions Mast cells and eosinophils may play a significant role in driving the pathogenesis of ulcerative colitis through the production of inflammatory mediators. The high expression of Siglec-8 and the inhibitory activity against mast cells suggests that antibodies that target this receptor, such as antolimab (AK002) represent a potential novel approach for the treatment of IBD.

BACKGROUND AND OBJECTIVES:Celiac disease (CD) is an immune-mediated enteropathy, induced by gluten in genetically susceptible individuals. The objective of this study was to describe the clinical pattern of CD in children from the western region of Saudi Arabia.DESIGN AND SETTING:Retrospective, hospital-based.PATIENTS AND METHODS:This study included children with a biopsy-proven diagnosis of CD made between September 2002 and July 2007. Children were admitted to the endoscopy unit for a small-bowel biopsy if they had gastrointestinal symptoms suggestive of CD or if they were positive for a CD-antibody screen performed for the high-risk groups.RESULTS:Eighty children were identified with a diagnosis of CD. Their mean (SD) age was 9.6 (4.9) years (range, 0.5-18 years). There were 44 (55%) female patients. Forty-one (51%) patients were detected during screening of high-risk groups, while 39 (49%) patients had classical symptoms of malabsorption. The screening also detected asymptomatic patients. Of 65 patients tested, 11 (17%) had elevated liver function tests, which reverted to normal after introduction of a gluten-free diet (GFD) except in one case. Seventy-three (91%) patients were positive for anti-tissue transglutaminase antibodies, 18 (23%), for IgG anti-gliadin antibodies; and 46 (58%), for IgA anti-gliadin antibodies. Forty-one (56%) patients showed good adherence to GFD as assessed by dietary history and the decline in anti-tTG level.CONCLUSION:CD may present with classical symptoms or be identified through screening programs. Growth and laboratory abnormalities usually improve after introduction of a GFD. Adherence to a GFD remains a problem; therefore, thorough assessment and counseling at the time of diagnosis and ongoing care are crucial.

Celiac Disease: From Pathogenesis to Novel Therapies

Celiac disease (CD) is a chronic inflammatory disease characterized by small intestinal mucosal injury and nutrient malabsorption in genetically susceptible individuals following the dietary ingestion of gluten1. The disease is characterized by villous atrophy, intraepithelial lymphocyte infiltration, chronic inflammation and activation of lamina propria T cells. The common genetic background in CD is the presence of heterodimeric HLA class II molecules DQ2 or DQ8 that account for ~40% of the genetic predisposition in CD2; several susceptibility loci not related HLA have been identified, but their contribution is only 3–4%3. Further, in addition to DNA variations, also gene transcription regulation, such as microRNAs (miRNAs) mechanism, could be involved in CD pathogenesis. MiRNAs are small non coding RNAs regulating basic cellular functions including proliferation, differentiation and death4. Indeed, it is easy to conceive that a defective miRNA-based mRNA regulation may compromise normal cell function and cause genetic diseases. The aim of this project is to perform an extensive study of the different miRNAs expression pattern in intestinal mucosa from CD patients at different stages of the disease [17 with active CD and 9 at gluten free diet (GFD)] and from 11 controls, to investigate the role of these molecules in transcriptional regulation in CD. We detect and quantify the expression profiling of 365 mature miRNAs using TaqMan Low Density Array methodology and Comparative CT method. Expression profiling revealed that the 25% of miRNAs tested are not expressed in jejunal intestine. Among the large set of expressed microRNAs over 50% of miRNAs are expressed at similar levels in CD and in control patients whereas respectively, the 23% and 19% of miRNAs were differently expressed in active CD and in GFD patients vs controls. Among these miRNAs upregulated, miR-449a shows very high levels: 55.18±16.45 and 15.43±7.69 (RQ±SEM) in active CD and in GFD children respectively. The quantitative RT-PCR confirmed the high level of miR-449a in active CD patients vs controls (mean RQ±SEM: 2.8±0.9). The bioinformatic analysis identified several target genes of miR-449a, belonging to Notch pathway such as NOTCH1, KLF4, DLL, LEF1 and NUMBL. Notch signaling is a gatekeeper of the progenitor/stem cell compartment of the intestine5. The luciferase reporter assay confirmed that the effective, specific and functional interaction between miR-449a and NOTCH1. The quantitative RT-PCR confirmed the expression of NOTCH1 and HES1 (a target gene of NOTCH1) mRNAs in active CD patients (RQ±SEM: 3.4±1.3 and 2.2±0.6, respectively) and in GFD patients (RQ±SEM: 6.5±4.7 and 4.2±2.9, respectively). In addition, immunohistochemistry showed that NOTCH1- and HES1- positive cells were significantly fewer in CD patients vs controls. Because NOTCH1 signals interact with the WNT pathway to influence the intestinal stem cell fate, we investigated both the WNT pathway and secretory goblet cells. We showed a similar β-catenin expression in CD and in control children, suggesting that the WNT pathway is not altered in CD, and a reduced number of goblet cells indicating an altered differentiation in celiac small intestine. In conclusion, we investigated miRNA expression in celiac small intestine and detected very high miR-449a expression levels in both active CD and GFD patients. These data demonstrate an altered miRNA gene regulation in CD patients than in controls, in association with a reduced NOTCH1 pathway and with a decrease differentiation of intestinal cells towards the secretory goblet cell lineage. Our findings suggest a novel pathogenic event in CD.

Introduction. Violated integrity of the local epithelial barrier is considered to contribute significantly to the occurrence of inflammatory bowel diseases. Its subepithelial part includes the components of the lamina propria consisting of granulocytes, macrophages, and lymphocytes, which are directly involved in damaging, inflaming, and regenerating the intestinal mucosa. The aim of the study was to determine the cellular composition of the infiltrate of the lamina propria of various intestinal parts. Materials and methods. We performed 300 histological and 15 immunohistochemical examinations of biopsies of the ileum, rectum, and ascending and sigmoid colons of patients with Crohn’s disease, ulcerative colitis, and irritable bowel syndrome. The biopsies of patients with inflammatory bowel syndrome were included in the comparison group. We determined CD163+ macrophages, CD3+ lymphocytes, CD20+ lymphocytes, CD138+ plasmocytes, and CD117+ mast cells. Results. When comparing the number of infiltrate cells in the rectal lamina propria in inflammatory bowel diseases, we detected that an acute stage of irritable bowel syndrome is characterized by a greater number of neutrophils, eosinophils, macrophages, T-lymphocytes, plasmocytes, mast cells in ulcerative colitis; macrophages in Crohn’s disease were noted. A comparison between nosologies in the acute stage revealed a greater number of macrophages in Crohn’s disease; mast cells in ulcerative colitis. A comparison of the stages of the disease showed a greater number of macrophages in ulcerative colitis in remission and a Crohn’s disease in the acute stage; neutrophils, T-lymphocytes, mast cells in ulcerative colitis in the acute stage. Conclusion. The cellular composition of the infiltrate of the intestinal mucosa varies and depends on the location, nosology, and the phase of the disease. The following diagnostic criteria for inflammatory bowel diseases can be considered as additional ones: a 2-fold increase in the number of macrophages in the ileum in acute Crohn’s disease and a 4-fold and 16-fold increase in the number of mast cells in the ascending and sigmoid colons, respectively, in ulcerative colitis. Keywords: inflammatory bowel diseases, Crohn’s disease, ulcerative colitis, irritable bowel syndrome, infiltrate