Toll-like receptor-3 (TLR3) priming may enhance mesenchymal stem cell (MSC) immunosuppressive activity, but this mechanism has not been investigated in the context of inflammatory bowel disease. Thus, we assessed the immunosuppressive properties of TLR3-primed MSCs using a trinitrobenzene sulfonate (TNBS)-induced mouse model of colitis. Intraperitoneally injected polyribocytidylic acid (poly (I:C)- (a ligand of TLR3) primed human umbilical cord-derived MSCs (hUC-MSCs) migrated to the inflamed colon and effectively improved clinical and pathological manifestations in colitic mice compared with mice treated with unstimulated hUC-MSCs (UCMs). Poly (I:C)-MSCs decreased a wide range of inflammatory cytokines and increased systemic interleukin-10 (IL-10) levels in colonic tissues. Poly (I:C)-MSCs also impaired T-helper type 1/17 (Th1/17) cell expansion and enhanced the suppressive effects of regulatory T cells (Treg) in vitro and in vivo. Poly (I:C)-MSCs suppressed the proliferation of activated mesenteric lymph node (MLN) cells via the overproduction of prostaglandin E (PGE) and upregulation of Jagged-1. PGE produced by hUC-MSCs in response to poly (I:C) increased the production of IL-10 and promoted the differentiation of Treg, which could be reversed by inhibition of Notch-1. Collectively, preconditioning MSCs with poly (I:C) enhanced the therapeutic effects of hUC-MSCs in TNBS-induced colitis, and TLR3-activated Notch-1 signaling regulated the immune suppression of hUC-MSCs through the production of PGE.

Background Small bowel fibrostenotic strictures are common in patients with Crohn’s disease (CD). No global consensus recommendations on definitions, diagnosis and clinical management are available. Methods Several systematic reviews followed by a RAND/University of California Los Angeles appropriateness study on the definitions, diagnosis and clinical management of fibrostenosing CD in clinical practice were performed. A panel of 28 global experts and a patient representative were convened. They assessed a total of 152 candidate items. The items were subsequently evaluated for appropriateness. Results No accurate predictive biomarkers are available for naïve or anastomotic fibrostenosing strictures. Accurate diagnosis of fibrostenosing CD requires cross-sectional imaging which should evaluate bowel wall thickness, luminal narrowing and prestenotic dilatation. A potential inflammatory component should be assessed. Abdominal cross-sectional imaging was considered necessary prior to any treatment decision. The panel proposed an approach to medical, endoscopic, and surgical therapies (Figure 1 and Table 1). Technical characteristics for endoscopic balloon dilation and follow up strategies after successful dilation therapy were identified. Appropriateness, types and performance of different surgical approaches in various settings were evaluated. Conclusion This global consensus provides clinical guidance for the diagnostic and therapeutic management of patients with fibrostenotic CD.

Background Intestinal fibrosis is considered an inevitable consequence of chronic inflammatory bowel disease (IBD), leading to stricture formation and need for surgery. During the process of fibrogenesis, extracellular matrix (ECM) components critically regulate the function of mesenchymal cells. We characterized the composition and function of the ECM in fibrostenosing Crohn’s disease (CD) and control tissues. Methods Decellularized full thickness intestinal tissue platforms were tested using three different protocols and ECM composition in different tissue phenotypes was explored by proteomics and validated by quantitative polymerase chain reaction (qPCR) and immunohistochemistry. Primary human intestinal myofibroblasts (HIMF) treated with Milk Fat Globule-Epidermal growth factor 8 (MFGE8) were evaluated regarding mechanism of their anti-fibrotic response, and the action of MFGE8 was tested in experimental intestinal fibrosis. Results We established and validated an optimal decellularization protocol for intestinal IBD tissues. Matrisome analysis revealed elevated MFGE8 expression in CD strictured tissue (CDs), which was confirmed at the mRNA and protein levels. Treatment with MFGE8 inhibited ECM production in normal control (NL) HIMF but not CDs HIMF. Next generation sequencing (NGS) uncovered functionally relevant integrin-mediated signaling pathways, and blockade of integrin αvβ5 and focal adhesion kinase rendered HIMF non-responsive to MFGE8. MFGE8 prevented and reversed experimental intestinal fibrosis in vitro and in vivo. Conclusion MFGE8 displays anti-fibrotic effects, and its administration may represent a future therapeutic approach for prevention of IBD-induced intestinal strictures.

Background Mesenteric fat wrapping around the intestinal wall, so called ‘creeping fat’ (CF), is spatially linked with stricture formation in Crohn’s disease (CD). Intestinal muscularis propria (MP) smooth muscle cell (HIMC) hyperplasia is a major contributor to luminal narrowing in stricturing CD. We investigated CF derived factors and their effect on HIMC hyperplasia in vitro and in vivo using human tissues, primary human cells and a colitis model. Methods Secretion of free fatty acids (FFA) by mesenteric fat (MF) or CF organ cultures was determined via lipidomic mass spectrometry. Effects of different length and types of FFA as well as CF and MF conditioned medium on proliferation of primary HIMF was assessed. Next generation sequencing (NGS) and lipidomics on HIMF was performed and relevant pathways inhibited with small molecules or siRNA knockdown. In the dextrane sodium sulfate (DSS) induced colitis model CPT-1 blockade was achieved via the small molecule etomoxir. Results Histopathology analysis of intestinal resection tissues revealed CD CF being located in the subserosa and its presence was linked with dramatic thickening of the MP. CF conditioned medium markedly upregulated HIMC proliferation compared to mesenteric fat from CD, UC and normal controls. CF released higher amounts of total, saturated and poly-unsaturated FFA with elevated levels of five long-chain (LC-)FFA, including palmitate. LC, but not medium or short chain FFA selectively increased proliferation in HIMC and fibroblasts, but not other intestinal cell types. NGS revealed gene regulation suggesting LC-FFA transport as a putative mechanism. Lipidomic analysis indicated the majority of palmitate being converted into phospholipids, predominantly phosphatidylcholine. Inhibition of LC-FFA uptake into cells via CD36, metabolism through inhibition of acyl-CoA synthetase, choline-phosphate cytidylyltransferase & choline kinase or blockade of LC-FFA uptake into mitochondria through CPT-1A reduced palmitate and CF conditioned medium induced HIMC proliferation. MP thickness increased in acute DSS colitis. Prophylactic inhibition of CPT-1 with etomoxir in acute DSS colitis did not reduce histopathologic inflammation or inflammatory cytokine gene expression, but reduced MP thickness and gene expression of the smooth muscle cell genes desmin and Sm22. Conclusion Subserosal CF releases LC-FFA inducing a selective proliferative response by HIMC. This effect was dependent on CD36, acyl-CoA synthetase and CPT-1. LC-FFA in HIMC are converted into phospholipids. Inhibtion of CPT-1 in DSS colitis reduced the increased MP thickness. These results point to CF as a novel contributor to stricture formation in CD.

Background Efforts to delineate factors linked with rural to urban transition, paralleling the staggering rise in CD incidence, have been hampered by the universal modernization in the west. China is ongoing transition from a predominantly traditional rural to an urban and industrialized society. We aimed to study the effect of changes and their relationship to CD. Methods SOURCE characterized environmental exposures, diet, and microbiome in rural and urban genetically similar Chinese controls and newly diagnosed CD in Guangdong province, and in Israeli CD and controls Results 380 subjects (median age 34years, 51% males); 40 CD and 162 rural controls and 121 urban controls from China, and 25 CD and 32 controls from Israel. Significant factors affecting the gut microbial composition were identified using PERMANOVA including the amount of time spent in urban area, dietary consumption of fat, fruits, iron, and added sugar, and exposures to farm animals. As the amount of time spent in urban area was linked with microbial composition, we stratified subjects living in rural area to those spending<50% of their time in the last year in urban areas ('rural', n=88) and those living in rural area but spending³50% of their time in urban environment ('rural-urban’, n=74) and compared then to the city-dwellers ('urban', n=121). Environmental exposures differed (Fig. 1) whereby flush toilet availability was reported in 5% rural, 35% rural-urban, and 61% urban, and 100% of Israelis. Having farm animals was noted in 60% rural, 24% rural-urban, and 3% of urban. Dietary habits also differed substantially; drinking at least weekly soft drink was reported in 5% rural and 27% of urban Chinese subjects, and 60% of the Israeli controls, while coffee was reported in 1% of rural, 15% of rural-urban, 32% urban, and 77% of Israelis. UniFrac-based PCoA indicated separation between rural and rural-urban control samples, with differences in β diversity (Fig. 2). Additionally, a-diversity was lower in rural-urban vs. rural controls, and in CD vs. urban cases. Composing a rural index based on an independent cohort (PMID:28915922) also indicated reduced rural index in rural-urban vs. rural controls, and in CD vs. urban cases. Using another independent health microbial index (PMID:35197084) also indicated reduced health index in rural-urban vs. rural controls, and in CD vs. urban cases (Fig. 2). Conclusion Trajectory of rural-urban transition is continuous and not dichotomous, whereby time spent in urban area associates with diet, exposures, and gut microbial composition. Living in rural area but spending ³50% of daily-life in urban environment (rural-urban) is linked with reduction in a-diversity, reduced rural index, and reduced microbial health index mirroring CD profile.