Human Leukocyte Antigen Signatures as Pathophysiological Discriminants of Microscopic Colitis Subtypes.

BACKGROUND AND AIMS: Microscopic colitis [MC] is currently regarded as an inflammatory bowel disease that manifests as two subtypes: collagenous colitis [CC] and lymphocytic colitis [LC]. Whether these represent a clinical continuum or distinct entities is, however, an open question. Genetic investigations may contribute important insight into their respective pathophysiologies. METHODS: We conducted a genome-wide association study [GWAS] meta-analysis in 1498 CC, 373 LC patients, and 13 487 controls from Europe and the USA, combined with publicly available MC GWAS data from UK Biobank and FinnGen [2599 MC cases and 552 343 controls in total]. Human leukocyte antigen [HLA] alleles and polymorphic residues were imputed and tested for association, including conditional analyses for the identification of key causative variants and residues. Genetic correlations with other traits and diagnoses were also studied. RESULTS: We detected strong HLA association with CC, and conditional analyses highlighted the DRB1*03:01 allele and its residues Y26, N77, and R74 as key to this association (best p = 1.4 × 10-23, odds ratio [OR] = 1.96). Nominally significant genetic correlations were detected between CC and pneumonia [rg = 0.77; p = 0.048] and oesophageal diseases [rg = 0.45, p = 0.023]. An additional locus was identified in MC GWAS analyses near the CLEC16A and RMI2 genes on chromosome 16 [rs35099084, p = 2.0 × 10-8, OR = 1.31]. No significant association was detected for LC. CONCLUSION: Our results suggest CC and LC have distinct pathophysiological underpinnings, characterised by an HLA predisposing role only in CC. This challenges existing classifications, eventually calling for a re-evaluation of the utility of MC umbrella definitions.

Lymphocytic Colitis With Increased Apoptosis: A Marker of Mutation in T-Cell-Mediated Immunity?

Lymphocytic colitis is a subtype of microscopic colitis that is mostly seen in adults. It presents mainly as chronic nonbloody diarrhea, with the hallmark of normal or near-normal endoscopy. In this case series, we are presenting 4 pediatric patients with lymphocytic colitis with prominent apoptosis of the colonic gland epithelium. Remarkably, all the patients have genetic mutations known to be associated with autoimmune enteropathy. Three patients have a CTLA4 mutation, and 1 patient has an STAT3 mutation. These mutations were previously reported in association with inflammatory bowel disease, but a specific connection with lymphocytic colitis has not been described. This report investigates the histopathology of such lesions in children and adolescents.

Lymphocytic and collagenous colitis in children and adolescents: Comprehensive clinicopathologic analysis with long-term follow-up.

Microscopic colitis (MC) is characterized by chronic watery diarrhea, endoscopically normal findings, and abnormal histology. While mostly encountered in adults, pediatric cases are rare and may show varying presentations. Our pathology data system was searched from 1984 to 2019 for patients ≤18 years of age with a lymphocytic colitis (LC) or collagenous colitis (CC) pattern of injury. Twenty-seven cases (23 LC and 4 CC) were retrieved. LC was more prevalent than CC (85% vs 15%, respectively) and affected slightly younger individuals (mean, 9.8 years versus 12.25 years). Immune dysregulation was documented in 11 (41%) patients. Most patients presented with watery diarrhea (n = 26, 96%) and either abdominal pain (n = 18, 67%), nausea/vomiting (n = 5, 19%), flatulence (n = 6, 22%), and/or weight loss (n = 1, 4%). A subset of patients (n = 10, 37%) demonstrated endoscopic abnormalities. Histologically, some patients with LC and CC displayed focal cryptitis or crypt abscess formation (n = 7, 26%) and focally increased crypt apoptosis (n = 9, 33%) in the absence of chronic injury. Clinical follow-up data were available for 23 (85%) patients with variable clinical responses recorded. Only 8 patients experienced complete symptom resolution. Twelve patients (11 LC and 1 CC) had subsequent biopsy material; of which, one developed histologic features of inflammatory bowel disease and another was found to have a CTLA-4 deficiency. Our study shows that pediatric patients with MC may have atypical clinical, histologic, and endoscopic findings and variable clinical responses. Underlying inflammatory and/or genetic conditions may be eventually unmasked, and genetic testing may be helpful in a small subset of patients.

Aquaporin-5 Expression Is Reduced in Lymphocytic Colitis.

OBJECTIVE: Aquaporin-5 (AQP5) is a member of a family of water channel proteins involved in the bidirectional transfer of water across cell membranes. Lymphocytic colitis (LC) and collagenous colitis (CC) are clinically similar diseases characterized by chronic watery diarrhea in patients with usually unremarkable colonic mucosa on colonoscopy. The aim of this study was to determine whether AQP5 expression in colonic epithelium is altered in LC and CC. METHODS: Sections of formalin-fixed and paraffin-embedded colorectal biopsies from three control patients (CTL), 8 patients with chronic non-bloody diarrhea with biopsies negative for active inflammation or significant distortion (CTL-D), 8 patients with LC, and 5 with CC were stained for AQP5 using immunohistochemistry. The staining intensity was scored as 3 (strong), 2 (intermediate), 1 (weak), or 0 (no staining). Statistical analysis was performed using Prism 7 Statistical Soft-ware. RESULTS: AQP5 was strongly expressed (score 3) in the epithelial cells in all three CTL cases and all 8 CTL-D cases. In the 5 cases of CC, 3(60%) had score 3 and 2(40%) had score 2, but none had a score of 1 or 0. Of the 8 LC cases, 2(25%) had score 3, 3 had score 2(37.5%), and 3 had score 1(37.5%) (p=0.0031). In the three cases of LC with markedly reduced AQP5 (score 1), enteric steroid treatment did not lead to significant improvement in diarrhea. CONCLUSIONS: Colorectal AQP5 expression is reduced in most cases of LC. Markedly reduced AQP5 expression in LC may identify a subset of patients with suboptimal response to enteric steroid treatment. Additional larger studies are needed to confirm these findings.This abstract was presented in part at Digestive Diseases Week in San Diego, CA, May 2019.

Family history, comorbidity, smoking and other risk factors in microscopic colitis: a case-control study.

OBJECTIVES: Data on heredity, risk factors and comorbidity in microscopic colitis, encompassing collagenous colitis (CC) and lymphocytic colitis (LC), are limited. AIM: The aim was to carry out a case-control study of family history, childhood circumstances, educational level, marital status, smoking and comorbidity in microscopic colitis. METHODS: A postal questionnaire was sent in 2008-2009 to microscopic colitis patients resident in Sweden and three population-based controls per patient, matched for age, sex and municipality. RESULTS: Some 212 patients and 627 controls participated in the study. There was an association with a family history of microscopic colitis in both CC [odds ratio (OR): 10.3; 95% confidence interval (CI): 2.1-50.4, P=0.004] and LC (OR not estimated, P=0.008). Current smoking was associated with CC [OR: 4.7; 95% CI: 2.4-9.2, P<0.001) and LC (OR: 3.2; 95% CI: 1.6-6.7, P=0.002). The median age at diagnosis was around 10 years earlier in ever-smokers compared with never-smokers.CC was associated with a history of ulcerative colitis (UC) (OR: 8.7, 95% CI: 2.2-33.7, P=0.002), thyroid disease (OR: 2.3; 95% CI: 1.1-4.5, P=0.02), coeliac disease (OR: 13.1; 95% CI: 2.7-62.7, P=0.001), rheumatic disease (OR 1.9; 95% CI: 1.0-3.5, P=0.042) and previous appendicectomy (OR: 2.2; 95% CI: 1.3-3.8, P=0.003), and LC with UC (OR: 6.8; 95% CI: 1.7-28.0, P=0.008), thyroid disease (OR: 2.4; 95% CI: 1.1-5.4, P=0.037) and coeliac disease (OR: 8.7; 95% CI: 2.8-26.7, P<0.001). CONCLUSION: Association with a family history of microscopic colitis indicates that familial factors may be important. The association with a history of UC should be studied further as it may present new insights into the pathogenesis of microscopic colitis and UC.

ENaC Dysregulation Through Activation of MEK1/2 Contributes to Impaired Na+ Absorption in Lymphocytic Colitis.

BACKGROUND: Lymphocytic colitis (LC) causes watery diarrhea. We aimed to identify mechanisms of altered Na absorption and regulatory inputs in patients with LC by examining the epithelial Na channel (ENaC) function as the predominant Na transport system in human distal colon. METHODS: Epithelial Na channel function and regulation was analyzed in biopsies from sigmoid colon of patients with LC and in rat distal colon in Ussing chambers. ENaC-subunit expression was measured by real-time PCR and RNA sequencing. Correction factors for subepithelial resistance contributions were determined by impedance spectroscopy. Upstream regulators in LC were determined by RNA sequencing. RESULTS: Epithelial Na channel-mediated electrogenic Na transport was inhibited despite aldosterone stimulation in human sigmoid colon of patients with LC. The increase in γ-ENaC mRNA expression in response to aldosterone was MEK1/2-dependently reduced in LC, since it could be restored toward normal by MEK1/2 inhibition through U0126. Parallel experiments for identification of signaling in rat distal colon established MEK1/2 to be activated by a cytokine cocktail of TNFα, IFNγ, and IL-15, which were identified as the most important regulators in the upstream regulator analysis in LC. CONCLUSIONS: In the sigmoid colon of patients with LC, the key effector cytokines TNFα, IFNγ, and IL-15 inhibited γ-ENaC upregulation in response to aldosterone through a MEK1/2-mediated pathway. This prevents ENaC to reach its maximum transport capacity and results in Na malabsorption which contributes to diarrhea.

Differential expression of interleukin-1/Toll-like receptor signaling regulators in microscopic and ulcerative colitis.

AIM: To investigate Toll-like receptor (TLR) signaling regulators in microscopic and ulcerative colitis patients. METHODS: Total RNA and microRNA were isolated from fresh frozen colonic biopsies of non-inflamed controls and patients with active or in-remission collagenous colitis (CC), lymphocytic colitis (LC), or ulcerative colitis (UC). We compared expressions of interleukin-1 receptor-associated kinase (IRAK)-2, IRAK-M, interleukin (IL)-37, microRNA (miR)-146a, miR-155, and miR-21 using quantitative real time reverse transcription polymerase chain reaction. RESULTS: IRAK-M expression was increased in LC patients with active disease in histopathological remission (LC-HR; P = 0.02) and UC patients (P = 0.01), but no differences in IRAK-2 expression were detected compared to controls. miR-146a, -155 and -21 expressions were increased in LC-HR (P = 0.04, 0.07, and 0.004) and UC (P = 0.02, 0.04 and 0.03) patients. miR-146a and miR-21 expressions were significantly enhanced in UC patients compared to UC remission (UC-R; P = 0.01 and 0.04). Likewise, active CC patients showed significantly increased expression of miR-155 (P = 0.003) and miR-21 (P = 0.006). IL-37 expression was decreased in both CC (P = 0.03) and LC (P = 0.04) patients with a similar trend in UC patients but not statistically significant, whilst it was increased in UC-R patients compared to controls (P = 0.02) and active UC (P = 0.001). CONCLUSION: The identification of differentially expressed miRNAs, IL-37, and IRAK-M suggests different pathophysiologic mechanisms in various disease stages in LC, CC, and UC.

Lymphocytic colitis is associated with increased pro-inflammatory cytokine profile and up regulation of prostaglandin receptor EP4.

Microscopic colitis (MC) is comprised of two entities, lymphocytic (LC) and collagenous colitis. Up to 20% of patients with chronic diarrhea that have a normal appearing colonoscopy will be diagnosed with MC. Since MC was first described less than 40 years ago, little is known about the mechanisms involved in disease pathogenesis. Nonsteroidal anti-inflammatory drugs are associated with an increased risk of MC and some reports suggest a dysregulation in prostaglandin production. Recent genome wide screens have found an association between prostaglandin receptor EP4 expression and inflammatory bowel disease; however, EP4 expression has never been studied in MC. The aim of this study was to assess colonic mucosal inflammatory cytokine profiles in patients with LC and to assess expression of the prostaglandin receptor EP4. Colonic mucosal biopsies were obtained from patients undergoing colonoscopy for investigation of diarrhea and in those undergoing colon cancer screening. Following histological assessment, expression of cytokines and the prostaglandin receptor EP4 was analyzed using real-time reverse transcriptase-PCR and immunohistochemistry. Patients with LC had markedly increased mRNA expression for TNF-α, IFN-γ and IL-8 compared to normal controls (p<0.001). No significant differences were noted for IL-1ß, IL-4, IL-10 or IL-12/23. Interestingly, those with LC had increased EP4 receptor expression, which positively correlated with increased TNF-α expression. This is the first report to demonstrate that LC is associated with increased TNF-α, INF-γ and IL-8 concurrent with a marked up-regulation of EP4. These findings add to our knowledge on the pathogenesis of LC and may give rise to possible new therapeutic and/or diagnostic tools in the management of MC.

Microscopic colitis patients demonstrate a mixed Th17/Tc17 and Th1/Tc1 mucosal cytokine profile.

BACKGROUND: Microscopic colitis (MC) is a chronic inflammatory bowel disorder of unknown aetiology comprising collagenous colitis (CC) and lymphocytic colitis (LC). Data on the local cytokine profile in MC is limited. This study investigated the T helper (Th) cell and cytotoxic T lymphocyte (CTL) mucosal cytokine profile at messenger and protein levels in MC patients. METHODS: Mucosal biopsies from CC (n=10), LC (n=5), and CC or LC patients in histopathological remission (CC-HR, n=4), (LC-HR, n=6), ulcerative colitis (UC, n=3) and controls (n=10) were analysed by real-time PCR and Luminex for expression/production of IL-1ß, -4, -5, -6, -10, -12, -17, -21, -22, -23, IFN-γ, TNF-α, T-bet and RORC2. RESULTS: Mucosal mRNA but not protein levels of IFN-γ and IL-12 were significantly up regulated in CC, LC as well as UC patients compared to controls. Transcription of the Th1 transcription factor T-bet was significantly enhanced in CC but not LC patients. mRNA levels for IL-17A, IL-21, IL-22 and IL-6 were significantly up regulated in CC and LC patients compared to controls, albeit less than in UC patients. Significantly enhanced IL-21 protein levels were noted in both CC and LC patients. IL-6 protein and IL-1ß mRNA levels were increased in CC and UC but not LC patients. Increased mucosal mRNA levels of IFN-γ, IL-21 and IL-22 were correlated with higher clinical activity, recorded as the number of bowel movements per day, in MC patients. Although at lower magnitude, IL-23A mRNA was upregulated in CC and LC, whereas TNF-α protein was increased in CC, LC as well as in UC patients. Neither mRNA nor protein levels of IL-4, IL-5 or IL-10 were significantly changed in any of the colitis groups. LC-HR and especially CC-HR patients had normalized mRNA and protein levels of the above cytokines compared to LC and CC patients. No significant differences were found between LC and CC in cytokine expression/production. CONCLUSION: LC and CC patients demonstrate a mixed Th17/Tc17 and Th1/Tc1 mucosal cytokine profile.

The behavior of matrix metalloproteinase-9 in lymphocytic colitis, collagenous colitis and ulcerative colitis.

Matrix metalloproteinases play an important role in extracellular matrix remodelling. It has been proposed that matrix metalloproteinase-9 (MMP-9) is involved in epithelial damage in ulcerative colitis (UC). However, to our knowledge, no data are available in terms of MMP-9 expression in microscopic colitis. Determination of mucosal protein expression levels of MMP-9 in lymphocytic colitis (LC), collagenous colitis (CC) and UC. MMP-9 immunohistochemical expressions were analyzed in paraffin-embedded tissue samples by immunohistochemistry including patients with LC, CC, UC, active diverticulitis, inactive diverticular disease and healthy control subjects. UC was also subgrouped according to the severity of inflammation. Immunostaining was determined semiquantitatively. Independent colonic biopsies from healthy and severe UC cases were used for gene expression analyses. For further comparison MMP-9 serum antigen levels were also determined in patients with UC and control patients without macroscopic or microscopic changes during colonoscopy. MMP-9 mucosal expression was significantly higher in UC (26.7 ± 19.5%) compared to LC (6.6 ± 9.3%), CC (6.4 ± 7.6%), active diverticulitis (5.33 ± 2.4%), inactive diverticular disease (5.0 ± 2.2%) and controls (6.3 ± 2.6%) (P < 0.001). The immunohistochemical expression of MMP-9 in LC and CC was similar as compared to controls. MMP-9 expression was significantly higher in each inflammatory group of UC compared to controls (mild: 11.0 ± 2.8%, moderate: 23.9 ± 3.7%, severe UC: 52.6 ± 3.9% and 6.3 ± 2.6%, respectively, P < 0.005). The gene expression microarray data and RT-PCR results demonstrated a significantly higher expression of MMP-9 in severely active UC compared to healthy controls (P < 0.001). Significantly higher MMP-9 serum antigen concentrations were observed in UC patients compared with the control group (P < 0.05). MMP-9 seems to play no role in the inflammatory process of LC and CC. In contrast, the mucosal up-regulation of MMP-9 correlated with the severity of inflammation in UC. The increased MMP-9 expression could contribute to the severity of mucosal damage in active UC.