| Objective To observe the effects of atorvastatin on monocyte chemoattractant protein-1(MCP-1) expression induced by advanced glycation end products (AGEs) in human umbilical vein endothelial cells(HUVECs)and to investigate the role of atorvastatin on peroxisome proliferator-activated receptor-γ(PPAR-γ)and nuclear factor-κB(NF-κB)in the anti-inflammation.Methods 1 Collagenase was used to isolate endothelial cells from human umbilical vein; The third to the fifth passage cells were used in following experiments.2 Human umbilical vein endothelial cells were identified by immunocellochemistry and morphology.3 Grouping:①Blank control group;②BSA group:controled with AGEs group;③AGEs group : cells were incubated with different concentration of AGEs(10-4-10-1mg/ml)for 24 hours;④Atorvastatin group: cells were incubated with different concentration of atorvastatin(0.1,1,10μmol/L)for 1 hour,then incubated with AGEs (10-1mg/ml) together for 24 hours;⑤15d-PGJ2(PPAR-γactivtor)group: cells were incubated with 15d-PGJ2(10μmol/L)for 1 hour,then incubated with AGEs (10-1mg/ml) for 24 hours;⑥GW9662(PPAR-γinhibitor)group:cells were incubated with GW9662(5000nmol/L)for 1 hour,then incubated with atorvastatin (1μmol/L)and AGEs (10-1mg/ml) together for 24 hours.4 RT-PCR was used to examine MCP-1 and PPAR-γmRNA expression in HUVECs.5 Western-blot was performed to detect the level of NF-κB p65 in cellular nucleusResults 1 We found the cultured cells were oval or polygon and retained cobblestone appearance through inverted phase contrast microscope. Brown particles could be observed in cytoplasm (CD31 or CD34 positive cells) after immunocytochemical staining with CD31 or CD34.2 In comparison with control group, AGEs(10-4-10-1 mg/ml)promoted the expression of MCP-1 mRNA to 1.53 times,2.12 times,2.56 times and 4.71 times respectively; The expression of MCP-1 mRNA was elevated significantly by AGEs(10-4mg/ml)(0.26±0.02 vs 0.17±0.04, P<0.01).3 Compared with AGEs group, atorvastatin (0.1,1,10μmol/L) diminished MCP-1 mRNA expression induced by AGEs in HUVECs in a concentration-dependent manner; Atorvastatin( 1μmol/L) remarkably decreased the expression of MCP-1 mRNA(0.63±0.11 vs 1.03±0.07, P<0.01).4 Comparaed with control group, AGEs decreased the expression of PPAR-γmRNA in HUVECs(0.22±0.08 vs 0.69±0.09, P<0.01)and increased the level of NF-κB p65 in cellular nucleus (0.78±0.06 vs 0.31±0.01, P<0.01); By compared with AGEs group,atorvastatin obviously enhanced the expression of PPAR-γmRNA in HUVECs(0.59±0.02 vs 0.22±0.08, P<0.01)and reduced the level of NF-κB p65 in cellular nucleus(0.40±0.03 vs 0.78±0.06, P<0.01).5 15d-PGJ2 obviously decreased the level of NF-κB p65 in cellular nucleus (0.21±0.01 vs 0.78±0.06, P<0.01)and the expression of MCP-1 mRNA in HUVECs(0.17±0.02 vs 0.93±0.12,P<0.01) comparaed with AGEs group; GW9662 remarkably antagonized the effect of atorvastatin on the level of MCP-1 mRNA and NF-κB p65 induced by AGEs in HUVECs (P<0.01).Conclusion1 High purity HUVECs were obtained by collagenase, which provids an ideal experimental model for pathophysiology studying of vascular endothelial cells in vitro;2 AGEs down-regulates PPAR-γmRNA expression ,enhance NF-κB activity and MCP-1 mRNA expression in HUVECs;3 Atorvastatin attenuates the down-regulation of PPAR-γmRNA , and reduce NF-κB activity and MCP-1 mRNA expression induced by AGEs in HUVECs;4 Activation of PPAR-γinhibits NF-κB transactivation , decrease MCP-1 mRNA expression;5 Atorvastatin plays an anti-in?ammatory role in the prevention and treatment of atherosclerosis, probably associated with the modulation of PPAR-γand the inhibition of NF-κB activity.
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