Society: AGA

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.

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 was performed on HIMF and relevant pathways were inhibited with small molecules or siRNA knockdown. In the dextran sodium sulfate (DSS) induced colitis model fat deletion was accomplished using the transgenic FAT-ATTAC mouse and CPT-1 blockade was achieved via the small molecule etomoxir.

Results: Histopathology of intestinal resection tissues revealed CD CF being located in the subserosa and its presence was linked with dramatic thickening of the MP. MP thickening was also observed in the acute DSS colitis model compared to controls and the thickening was reduced by fat deletion in the FAT-ATTAC mouse. 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. Prophylactic inhibition of CPT-1 with etomoxir in acute DSS colitis did not reduce histopathologic inflammation or inflammatory cytokines 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 taken up by HIMC are converted into phospholipids. Deletion of fat or inhibition of CPT-1 in acute DSS colitis reduced the increased MP thickness in proximity to mesenteric fat. These results point to CF as a novel contributor to stricture formation in CD.

Claudio Fiocchi

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