Curcumin Inhibits Vascular Smooth Muscle Cells Proliferation By Inhibition Ras Signaling Cascade

Objective: To study the effects of curcumin on the proliferation of cultured rat aortic vascular smooth muscle cells(VSMCs) and to screen a safe,effective and cheap drug for prevention and treatment of vascular proliferative disorders.Methods: Rat aortic VSMCs were cultured in vitro with different concentration of curcumin. There were three drug groups: low(7.5μM),moderate(15μM),high dos(e30μM) and three control groups: solvent control,blank control,positive drug(Rapamycin, 10nM) control. The effects of curcumin on the growth and proliferation of VSMCs were investigated by light microscopy, cell counting and MTT assay in different time. Western Blot was used to detect p-ERK1/2, p-Akt and Caspase-9 in Ras signaling pathway.Results: 1. The effects of curcumin with different dose or in different time on the growth of VSMCs observed by the light microscope: With the increase of concentration and time, curcumin caused a marked dose-dependent and time-dependent proliferation inhibitory effect on VSMCs in moderate and high dose groups. In addition to this, cell death was observed in the high-dose group obviously. 2. The effects of curcumin with different dose or in different time on the proliferation of VSMCs confirmed by the cell counting: solvent group and blank group showed no statistic difference in cell number in different time(p﹥0.05).Compared with the solvent and blank control groups, moderate,high dose drug groups and the positive drug control group showed significantly proliferation inhibitory effects. The proliferation inhibition rate in positive drug group of VSMCs at 24h,48h,72h was 31.8%,91.8%,79.6%respectively. The proliferation inhibition rate in low-dose group of VSMCs at 24h,48h,72h was 19.9%,13.2%,26.8%respectively, all have a remarkable reduction compared with the positive control groups(p<0.05). The proliferation inhibition rate in moderate-dose group of VSMCs at 24h,48h and 72h was 48.7%,76.6%,69.2%respectively, there was a marked reduction at 48h and 72h compared with the positive control groups(p<0.05).The proliferation inhibition rate in high-dose group of VSMCs at 24h,48h and 72h was 58.8%,91.1%,98.3%. There was no significant difference between high-dose group and the positive control in inhibition rate at 48h. The proliferation inhibition rate of VSMCs in high-dose group at 24h and 72h are higher than positive control(p<0.05).3 The effects of curcumin with different dose or in different time on the proliferation of VSMCs confirmed by MTT assay: solvent group and blank group showed no statistic difference in OD570 in different time(p﹥0.05). In accordance with the result of the cell counting, moderate,high dose drug groups and the positive group also showed significantly proliferation inhibitory effects. The inhibition rate in positive drug group of VSMCs at 24h,48h,72h was 21.8%,57%,66.2%respectively. All have a notable reduction compared with solvent group and blank group. The inhibition rate in low-dose group of VSMCs at 24h,48h,72h was -2.9%,22.8%,15.0%respectively; there was a marked reduction at 48h and 72h compared with the positive control groups(p<0.01). The inhibition rate in moderate-dose group at 24h,48h,72h was 16.3%,52.3%,60.0%; there was no statistic difference in different time compared with positive drug group(p>0.05). The inhibition rate in high-dose group was 28.9%,67.8%,87.2%at 24h,48h,72h ; there was no statistic difference in different time compared with positive drug group at 24h,48h(p>0.05); the inhibition rate was higher than positive drug group at 72h(p<0.05).4. p-ERK1/2, p-Akt and Caspase-9 in Ras signaling pathway tested by western blot: curcumin may inhibit the activation of ERK1/2, Akt and elevate Caspase-9 in a dose-dependent manner.Conclusion: 1.Curcumin effectively inhibits the proliferation of rat aortic VSMCs in vitro with dose and time dependency.2. The curcumin antiproliferative effect potential mechanism is to inhibit the activation of Ras-Raf-ERK1/2 cell signaling pathway.3. The curcumin antiproliferation mechanism is partly dependent in inhibition Ras-PI3K-Akt cell signaling pathway and may startup mitochondrial apoptosis pathway.