REG4 induces angiogenesis in ulcerative colitis by upregulating MMP-7 expression and downregulating sVEGFR1 level.

BACKGROUND: Angiogenesis is suggested as pathogenesis of inflammatory bowel disease (IBD). However, how angiogenesis mediates pathogenesis of ulcerative colitis (UC), a common form of IBD, remains elusive. METHODS: Dextran sulfate sodium salt (DSS)-induced UC mouse models were established. Then, these mice were injected with short hairpin RNA against regenerating family member4 (shREG4), followed by evaluation of weight and disease activity index (DAI). Collected colon tissues underwent hematoxylin and eosin staining and immunohistochemistry to detect morphological changes and microvascular density (MVD). Inflammation cytokines, soluble vascular endothelial growth factor receptor1 (sVEGFR1) and VEGF levels were measured using enzyme-linked immunosorbent (ELISA) assay. Matrix metallopeptidase7 (MMP-7) was measured using western blot. REG4 overexpression plasmid-transfected NCM460 cells experienced quantitative real-time reverse transcription polymerase chain reaction and western blot. Human intestinal microvascular endothelial cells (HIMECs) were co-cultured with NCM460 cells that had been transfected with REG overexpression plasmid and treated with MMP-7 mAb and/or sVEGFR1. MMP-7 or sVEGFR1 levels in both culture medium were tested using ELISA. HIMECs angiogenesis was measured by Angiogenesis kit. RESULTS: In vivo, DSS-induced mice exhibited lower weight and higher DAI. Collected mice colon tissues showed enhanced inflammation, morphological changes, MVD, and expressions of VEGF, REG4 and MMP-7, which were all reversed after DSS-induced mice injected with shREG4. In vitro, REG4 overexpression-transfected NCM460 cells and its culture medium presented higher MMP-7 level. After co-culture with REG4 overexpression-transfected NCM460 cells, HIMECs culture medium demonstrated lower level of sVEGFR1 and higher angiogenesis, and the latter was reversed by MMP-7 mAb and/or sVEGFR1 treatment in NCM460 cells. CONCLUSION: REG4 induces angiogenesis in UC by upregulating MMP-7 and downregulating VEGFR1.