Expression Of Osteopontin In Proliferated Vasular Smooth Muscle Cells Induced By Endothelin-1

Backgrouds:Osteopontin(OPN) is a kind of secreted noncollagenous phosphated glucoprotein. Uninjured arteries have undetectable level of OPN, but it is much higher in the arteries with atherosclerosis(AS). The migration and proliferation of vascular smooth muscle cell(VSMC) can be promoted by OPN, and it has been proved to participate in the phenotypic switch of VSMC(from contractile phenotype to synthetic phenotype). In the blood plasma of patients with AS, the level of endothelin-1 (ET-1) increases obviously, and is closely related to the severity and extent of AS. Previous studies have shown that ET-1 could stimulate VSMC proliferation, but the mechanisms of its effect are not clear yet. Besides, as to whether OPN is expressed in the proliferated VSMC induced by ET-1, and whether OPN plays a role in the effect of ET-1 and how OPN plays its role, no report has been published.Objectives:To find out the effect of ET-1 on VSMC proliferation and OPN expression, analyze the relationship between them, study the role of OPN in the process of VSMC proliferation induced by ET-1, and so to provide a new way of monitoring VSMC proliferation and a new idea of retarding or preventing the progression of AS.Methods:1. Human umbilical artery smooth muscle cells(HUASMCs) were cultured by explant method; 2. Cultured HUASMCs were randomly divided into 5 groups. Group 1, without ET-1, was control group. Group2, group3, and group4 were respectively treated with 1nmol/L, 10nmol/L, and 100nmol/L ET-1 for 48 hours. Group5 was pretreated with 10μmol/L BQ123(a type of specific antagonist of ET receptor A) for 2 hours, and then cultured in 10nmol/L ET-1 for 48 hours. The level of optical density(OD) was evaluated by MTT assay (λ=490nm); 3. With the method of immunocytology, proliferating cell nuclear antigen(PCNA) of groupl, group3 and group5 were stained and compared; 4. OPN expression was also measured by immunocytology, and the positive rate was compared among groups; 5. Analyze the relationship between OPN expression and HUASMC proliferation.Results:1. After treated with 1nmol/L, 10nmol/L and 100nmol/L ET-1, levels of OD were significantly higher than that of control group (P<0.01);2. After pretreated with BQ123, levels of OD and PCNA expression were significantly lower than those of group3 (P<0.01);3. ET-1 increased OPN expression dose-dependently (P<0.01), and BQ123 could reduce its effect;4. OPN expression was correlated with VSMC proliferation (r=0.963,P<0.01).Conclusions:1. ET-1 could promote VSMC proliferation dose-dependently;2. OPN expression increased significantly after treated with ET-1, and was positively correlated with VSMC proliferation. OPN expression was both the cause and the result of the phenotypic switch of VSMC, and the examination of OPN might indicate the early proliferation of VSMC;3. BQ123 could reduce the effect of ET-1 and might retard or prevent the progression of AS.