【摘要】 目的 通过体外培养的方法，建立原代心肌细胞柯萨奇病毒感染性病毒性心肌炎模型，用中药虎杖苷(PD)处理该疾病模型，观察对该疾病的干预作用。方法 体外培养心肌细胞，用100 TCID50柯萨奇B3病毒(CVB3)感染心肌细胞，建立病毒性心肌炎实验模型，然后分别用0.02mmol/L、0.2mmol/L、2mmol/L三种不同浓度的PD处理感染CVB3病毒的心肌细胞，观察心肌细胞的形态和搏动，用细胞免疫荧光技术标记纤维状肌动蛋白(F-actin)和VEGF蛋白，四唑蓝比色法(MTT Assay)测定心肌细胞活性和病毒抑制率，流式细胞仪检测各组心肌细胞VEGF蛋白和F-actin平均荧光强度。结果 0.02mmol/L和0.2 mmol/L PD对心肌细胞无毒性作用。2 mmol/L PD对心肌细胞有一定毒性作用；病毒对照组细胞病变明显，0.02 mmol/L和0.2 mmol/L PD处理组均呈现明显的抗病毒作用；免疫荧光显微镜下观察，病毒对照组心肌细胞骨架破坏、VEGF蛋白解聚、重构；0.02 mmol/L和0.2 mmol/L PD处理后F-actin和VEGF蛋白分布均有所恢复；流式细胞仪测得各浓度PD处理组F-actin和VEGF蛋白表达量均明显高于病毒对照组(P<0.05)。结论 柯萨奇B3病毒对新生大鼠原代心肌细胞有直接损伤性作用，并可导致心肌细胞F-actin骨架蛋白解聚和弥散性分布，影响心肌收缩力；PD可以明显抑制柯萨奇B3病毒对新生大鼠原代心肌细胞的致病变效应，维持心肌细胞骨架正常形态和功能，尤以0.2mmol/L浓度的PD效果最佳。

    【关键词】 虎杖苷；柯萨奇B3病毒；心肌细胞；纤维状肌动蛋白；细胞骨架

    Antiviral effect of polydatin in vivo on viral myocarditis

    SONG Jun-hua,WANG Shu-yun,MOU Dao-yu,et al.Qingdao Healthy School,Qingdao 266071,China

    【Abstract】 Objective To study the antiviral and protective effect of polydatin (PD) on the cultured rat neonatal cardiomyocytes of viral myocarditis (VMC) induced by coxsackievirus B3 (CVB3).Methods Primary cultures of Sprague-Dawley (SD) rat neonatal cardiac myocytes were prepared and inoculated with 100 TCID50 CVB3 as experimental model of viral myocarditis. Different concentration of PD was added into cardiomyocytes infected by CVB3. The cardiomyocyte viability were detected by MTT assay for calculating the survival rate and the inhibitory rate of the virus. Myocardial F-actin and VEGF were analyzed by immunofluorescent staining with confocal microscopy. Mean fluorescent intensity of F-actin and VEGF were determined by flow cytometry.Results There is no toxicity on cardiomyocytes when PD is at concentration of 0.02 mmol/L and 0.2 mmol/L while there is toxicity at concentration of 2 mmol/L. CVB3 could reduce the viability of cardiomyocytes, depolymerize F-actin cytoskeleton and reorganize VEGF protein. Different concentration of PD could effectively lessen the occurrence of cytopathic effect and increase the survival rate of CVB3 infected cardiomyocytes significantly. PD could also inhibit the replication of CVB3 and attenuate the CVB3-induced changes of myocardial cytoskeleton and VEGF organization, especially at dose of 0.2 mmol/L. The results of flow cytometry showed the expression of F-actin and VEGF were significantly increased in PD treated group compared with viral control group and the level of VEGF was increased in dose-dependent manner.Conclusion CVB3 could cause damage of rat neonatal cardiomyocytes directly, induce F-actin cytoskeleton depolymerization and VEGF protein reorganization and then decline the contractility of cardiac myocytes;PD could effectively lessen the occurrence of cytopathic effect induced by CVB3 and maintain normal morphology and function of cardiomyocytes, especially at a dose of 0.2mmol/L. ......