Effect Of Lovastatin On MRNA And Protein Expression Of Matrix Metal-Loproteinase-2 Secreted By Macrophage

Objective To study the effect of lovastatin on mRNA and protein expression of Matrix Metalloproteinase-2 secreted by mouse macrophage and to investigate the non-lipid mechanisms on antiatherosclerosis of lovastatin.Methods (1)The conditioned media was generated by incubating the macrophage for 24 hours at 37 ℃ with RPMI1640 supplemented with 10% FBS and different concentrations of lovastatin, MTT assay was adopted to evaluate the inhibiting effect of lovastatin on macrophage. (2)In situ hybridization was used to detect mRNA expression of MMP-2 in macrophage and integral optical density of MMP-2mRNA was detected by image analysis system to evaluate the effect of different concentrations of lovastatin on mRNA expression of MMP-2. (3)Immunocytochemistry was used to detect protein expression of MMP-2 in macrophage and integral optical density of MMP-2 protein was detected by image analysis system to evaluate the effect of different concentrations of lovastatin on protein expression of MMP-2. (4)The identity of MMPs was characterized by Western Blot after SDS-Polyacry lamide gel electrophoresis. (5)The data were analysed by using random blocks design variance analysis, muLtiple comparison q test and linear correlation analysis. P<0. 05 meaned that there was difference in statistics. P<0. 01 meaned that there was significant difference in statistics.ResuLts (1)MTT assay indicated that lovastatin had obvious inhibiting effect on mouse macrophage at 10μmol/L(TCL<90%) and no inhibiting effect on it at 0.01, 0.1, 1 μnol/L (TCL>90%). (2)In situ hybridization showed mRNA expression of MMP-2 decreased gradually with the increase of lovastatin concentration. There was significant difference in groups(P<0. 01). The concentration of lovastatin correlated with MMP-2mRNA integral optical density negatively(r=-0. 774, P<0. 01). It indicated that lovastatin could inhibit mRNA expression of MMP-2 in a dose-dependent manner. (3)Immunocytochemistry confirmed expression of MMP-2 decreased graduallywith the increase of lovastatin concentration. There was significant difference in groups (P<0. 01). The concentration of lovastatin correlated with MMP-2 protein integral optical density negatively(r=-0. 734, P<0. 01). It indicated that lovastatin could inhibit protein expression of MMP-2 in a dose-dependent manner. (4) As shown by Western Blot,black protein ladders were found at 72KDa and 62KDa in control group, and they were MMP-2 zymogen and active type. Black ladders were found only at 72KDa in lovastatin groups.Conclusion Lovastatin have direct inhibiting effect on macrophages and reduce their quantity; it also can inhibit mRNA and protein expression of MMP-2 secreted by macrophage and may be have inhibiting effect on zymogen activation. All above can enhance the stability of atherosclerotic plaque.