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目的探讨高糖对人肾小球系膜细胞(human mesangial cells,HMCs)单核细胞趋化蛋白-1(monocyte chemoat-tractant protein-1,MCP-1)及细胞外基质的主要成分(extracellular matrix,ECM)纤维连接蛋白(fibronectin,FN)表达的影响及己酮可可碱(pentoxifylline,PTX)联合霉酚酸酯(mycophenolate,MMF)对上述指标的干预作用。方法将培养的HMCs分为8组:正常对照组(NG,5 mmol/L葡萄糖);高糖组(HG,30 mmol/L葡萄糖);甘露醇渗透压对照组(Man,5 mmol/L葡萄糖+25 mmol/L甘露醇);高糖+PTX-0.03组(P1,30 mmol/L葡萄糖+0.03 mg/mL PTX);高糖+PTX-0.3组(P2,30 mmol/L葡萄糖+0.3 mg/mL PTX);高糖+MMF-10组(M1,30 mmol/L葡萄糖+10μg/mLMMF);高糖+MMF-100组(M2,30 mmol/L葡萄糖+100μg/mL MMF);高糖+PTX+MMF组(P+M,30 mmol/L葡萄糖+0.3 mg/mL PTX+100μg/mL MMF),分别在24、48、72 h采用RT-PCR法和ELISA法检测PTX、MMF及PTX+MMF对高糖条件下MCs内MCP-1 mRNA及蛋白、FN表达的影响。结果高糖组HMCs MCP-1 mRNA、蛋白的表达及FN的分泌较正常对照组显著增加(P<0.01),且48 h表达最高;不同浓度的PTX、MMF均能下调MCP-1 mRNA、蛋白及FN的表达(P<0.01);不同浓度的MMF对MCP-1 mRNA、蛋白的表达及FN分泌的抑制程度不同,呈时间剂量依赖性(P<0.05);不同浓度的PTX则随着时间的延长对其抑制作用无明显差别;PTX联合MMF组比单独PTX各组在各时间点的抑制程度更明显(P<0.01),比单独MMF各组在各时间点抑制作用增强,(24 h,P<0.05),但随着时间延长,差异无统计学意义。结论 PTX及MMF可能通过阻抑MCP-1的表达及FN的分泌延缓糖尿病肾病(diabetic nephropathy,DN)中肾小球硬化及间质纤维化的进展,联合给药组作用优于单药,可能从改善血流动力及抗炎角度解释了PTX联合MMF对DN肾脏的协同保护作用。
Objective To investigate the effects of high glucose on the expression of monocyte chemoat-tractant protein-1 (MCP-1) and the extracellular matrix (MMP) in human mesangial cells (HMCs) (ECM) fibronectin (FN) expression and pentoxifylline (PTX) combined with mycophenolate (MMF) on the above indicators. Methods The cultured HMCs were divided into 8 groups: normal control group (NG, 5 mmol / L glucose); high glucose group (HG, 30 mmol / L glucose); mannitol osmotic pressure control group +25 mmol / L mannitol); high glucose + PTX-0.03 group (P1, 30 mmol / L glucose + 0.03 mg / mL PTX); high glucose + PTX-0.3 group / mL PTX); high glucose + MMF-10 group (M1, 30 mmol / L glucose + 10μg / mLMMF); high glucose + MMF-100 group (M2, 30mmol / L glucose + 100μg / mL MMF) PTX + MMF group (P + M, 30 mmol / L glucose + 0.3 mg / mL PTX + 100 μg / mL MMF) were used to detect PTX, MMF and PTX at 24,48 and 72 h + MMF on the expression of MCP-1 mRNA, protein and FN in MCs under high glucose conditions. Results The expression of MCP-1 mRNA and protein and the secretion of FN in HMCs of high glucose group were significantly higher than those of normal control group (P <0.01), and were highest at 48 h. The expressions of MCP-1 mRNA and protein (P <0.01). The different concentrations of MMF inhibited the expression of MCP-1 mRNA and protein and the secretion of FN in a time and dose-dependent manner (P <0.05) (P <0.01). Compared with MMF group, the inhibitory effect of PTX group and MMF group was better than that of PTX alone group at each time point , P <0.05), but with the extension of time, the difference was not statistically significant. Conclusions PTX and MMF may delay the progression of glomerulosclerosis and interstitial fibrosis in diabetic nephropathy (DN) by inhibiting the expression of MCP-1 and FN secretion. The combination of PTX and MMF may be superior to monotherapy in the treatment of diabetic nephropathy (DN) From the perspective of improving hemodynamics and anti-inflammatory, the synergistic protective effect of PTX combined with MMF on DN kidneys was explained.