Astragalus mongholicus altered the secretion and expression of transforming growth factor-?1 and basic fibroblast growth factor in cultured human peritoneal mesothelial cells / 医学研究生学报

Objective: To investigate the effect of Astragalus mongholicus injection(AM) on the secretion and mRNA expression of transforming growth factor((TGF-?1)) and basic fibroblast growth factor(bFGF) in cultured human peritoneal mesothelial cells((HPMC).) Methods: HPMC got from patients underwent surgical operation were cultured in vitro.At first,cells from the third passage were incubated with RPMI1640 culture medium containing 0.1% FBS for 24 hours.Then,they were divided into control group,PDS group,AM group 1,AM group 2 and AM group 3 for testing.Each group was supplemented with equal volum of RPMI1640 culture medium containing 20% FBS.After 24 hour incubating,mitochondrial dehydrogenases activity(MTT assay),the levels of TGF-?1 and bFGF in supernatant of the cell culture and the mRNA expression of TGF-?1 and bFGF were detected.Results: Significant decrease of mitochondrial dehydrogenase activities were observed in PDS group as compared with those in control group and AM groups(P0.05).The levels of TGF-?1 and bFGF in supernatant of the cell culture were significantly lower in control group than those in PDS group.Marked lower levels of TGF-?1 and bFGF were found in AM group 1,AM group 2 and AM group 3 as compared with those in PDS group(P0.05).The mRNA expressions of TGF-?1 and bFGF in PDS group increased significantly as compared with those in control group.And significant mRNA expressions of TGF-?1 and bFGF were found in PDS group when compared with those in AM groups(P

Effect of High Glucose on the Expression of MCP-1 and VCAM-1 in Cultured Human Peritoneal Mesothelial Cells / 대한신장학회잡지

BACKGROUND: High glucose in peritoneal dialysis solution has been implicated in the pathogenesis of peritoneal fibrosis. Macrophages in peritoneal cavity seem to participate in the process of peritoneal fibrosis through the production of various cytokines and growth factors. Monocyte chemoattractant protein-1(MCP-1) plays a key role in the recruitment of monocytes toward the peritoneal cavity. Vascular cell adhesion molecule-1(VCAM-1) is assumed to be important in the transmigration of monocytes. MCP-1 and VCAM-1 can be induced by various cytokines and growth factors in human peritoneal mesothelial cells(HPMC). However, effect of high glucose on the expression of MCP-1 and VCAM-1 in HPMC has not been known well. METHODS: Cultured HPMC were conditioned with glucose(5-100mM) or mannitol for varying periods up to 7 days. Cell proliferation and mRNA expression of MCP-1 and VCAM-1 were assessed by MTT assay and Northern blot analysis respectively. MCP-1 protein was measured using ELISA. Chemotactic activity of high glucose-conditioned culture supernant were evaluated by chemotactic assay. Effect of protein tyrosine kinase(PTK) inhibitor on the high glucose-induced MCP-1 mRNA expression was examined. RESULTS: Glucose inhibited the cell proliferation in a time and dose dependent manner. Northern blot analysis showed that high glucose increased the MCP-1 mRNA expression in a time(2-7days) and dose(15-100mM) dependent manner, but not VCAM-1 mRNA expression. MCP-1 protein in cell culture supernant was also increased. Equivalent osmotic concentration of mannitol had no significant effect. High glucose-conditioned supernant had an increased chemotactic activity for monocyte, which was neutralized by specific anti-MCP-1 antibody. PTK inhibitors such as genistein and herbimycin A suppressed the high glucose-induced MCP-1 mRNA expression in a dose dependent manner. CONCLUSION: High glucose induced MCP-1 expression in HPMC partly via pathways involving PTK.

Nitric Oxide Inhibits VCAM-1 Expression in Human Peritoneal Mesothelial Cells: Possible Role of Cyclic GMP and NF-kappa B / 대한신장학회잡지

Leukocyte adhesion to mesothelium is an important step during peritonitis, which may be mediated by adhesion molecules including vascular cell adhesion molecule-1(VCAM-1). Nitric oxide(NO) is known to be an endogenous inhibitor of leukocyte adhesion. We investigated the effect of NO on VCAM-1 expression in cultured human peritoneal mesothelial cells and the possible role of cGMP and NF-kappa B. Cells were exposed to tumor necrosis factor-alpha(TNF-alpha) for the indicated periods in the presence or absence of NO donors, 3-morpholino-sydnonimine (SIN-1) or nitroprusside(NP). The expression of VCAM-1 mRNA and cell surface VCAM-1 molecule was measured by Northern blot analysis and flow cytometry. To detect NF-kappa B binding activity, electrophoretic mobility shift assay(EMSA) was performed. To determine the role of guanylate cyclase or cGMP, inhibitor of guanylate cyclase, 1H-[1, 2, 4] oxadiazolo [4, 3-a] quinoxalin-1-one(ODQ) or analogue of cGMP, 8-bromo-cGMP was used. Both SIN-1 and NP inhibited the TNF-alpha-induced VCAM-1 mRNA expression in a dose dependent manner. SIN-1 also inhibited the expression of cell surface VCAM-1 molecule. Furthermore, SIN-1 and NP inhibited the expression of VCAM-1 mRNA induced by interleukin-1(IL-1beta) or lipopolysaccharide(LPS) as well. By EMSA, SIN-1 inhibited the TNF-alpha-induced NF-kappa B activity. The 8-bromo-cGMP had no significant effect on TNF-alpha-induced VCAM-1 mRNA expression and ODQ also had no significant influence on the inhibitory effect of SIN-1. In conclusion, NO may play an important role in mediating the inflammatory process during peritonitis by down-regulating the mesothelial VCAM-1 expression via suppression of NF-kappa B activity through cGMP- independent pathway.