低分子肝素抑制肾小球系膜细胞增生机制的研究

Mechanism of Inhibiting Rat Glomerular Mesangial Cells Proliferation by Low-Molecu- lar-Weight Heparins

目的低分子肝素是肾脏疾病治疗中常用药物,能抑制肾小球系膜细胞的增生,其机制常不清楚,本研究探讨低分子肝素(LMWH)对大鼠肾小球系膜细胞(GMCs)增生水平的影响及其机制。方法GMCs细胞共设3组:①GMCs组;②LPS组:即GMCs+LPS;③LMWH组:即GMCs+LPS+LMWH,而LMWH终浓度分别为2.5 IU/ml,25 IU/ml和250 IU/ml。采用MTT法于培养24 h和48 h观察各组中GMCs增生水平,选择药物的最佳GMCs增生抑制浓度及时间;采用免疫细胞化学方法观察培养48 h后3组(LMWH终浓度为250 IU/ml)GMCs涂片中单核细胞趋化蛋白-1(MCP-1)及核转录因子-κB(NF-κB)的表达;采用ELISA方法观察培养的上清液中MCP-1浓度。结果LMWH 250 IU/ml组对GMCs增生的抑制最为显著,其GMCs增生水平低于LPS组和LMWH 2.5 IU/ml,25 IU/ml组(P<0.05);各浓度LMWH对GMCs增生的抑制作用48 h强于24 h(P<0.05)。LMWH 250 IU/ml组48 h的MCP-1阳性率明显低于LPS组(P<0.01),而与GMCs组差异无显著性(P>0.05);LPS组48 h的MCP-1阳性率明显高于GMCs组(P<0.01)。LMWH 250 IU/ml组48 h的NF-κB表达阳性率与LPS组和GMCs组比较差异均无显著性(P>0.05);LPS组的NF-κB表达阳性率明显高于GMCs组(P<0.01)。LMWH 250 IU/ml组GMCs培养上清液中MCP-1浓度明显低于LPS组(P<0.01),与GMCS组差异?

Objective Low-molecular-weight heparins (LMWH) can inhibit proliferation of glomerular mesangial cells (GMCs) and it is a major and effective drug in the treatment of renal diseases but the mechanism has still not been explained. This paper studies the mechanism by which LMWH inhibits monocyte chemoaltractant protein-1 (MCP-1) expression, secretion and regulation in rat GMCs. Methods Three groups were established; GMCs group, LPS group (GMCs+ LPS) and LMWH group (GMCs + LPS + LMWH). GMCs proliferation was detected by MIT at 24 and 48 hs after culture. MCP-1 and nuclear transcriptional factor of Kappa B ( NF-κB) expressions were assayed by immunohistochemistry at 48 h after curture. MCP-1 concentration was determined by EL1SA. Results In the LMWH group with the terminal concentration of 250 IU/ml, the proliferative rate of GMCs was lower than that in the LPS group and was also lower than those in the LMWH groups with the terminal concentration of 2. 5 IU/ml and 25 IU/ml respectively ( P < 0.05). The proliferative inhibition of GMCs induced by LMWH of three different terminal concentrations was more significant at 48 h than at 24 h after culture. The positive rate of MCP-1 expression of GMCs in the LMWH group with the terminal concentration of 250 IU/ml was obviously lower than that in the LPS group at 48 h afterculture ( P <0.01), but there was no statistical difference when compared with the concentration in the GMCs group. The pasitive rale of MCP-1 expression of GMCs in the LPS group was obviously higher than that in the GMCs group ( P < 0.01). There was no statistical difference in the NF-κB expression at 48 h after culture between the LMWH group and LPS group as well as the GMCs group, while it was obviously higher in the LPS group than that in the GMCs group ( P < 0.01). The MCP-1 concentration in the LMWH group with the terminal concentration of 250 IU/ml was significantly lower than that in the LPS group ( P < 0.01) and did not differ from that in the GMCs group. Conclusions LMWH can obviously inhibit proliferation of GMGs, down-regulate the abnormal expression and secretion of MCP-1, but can not inhibit the activity of NF-icB.