内科

目的探讨乙酰紫草素对HCV复制的影响及其作用机制。方法（1）体外培养Huh7细胞，用不同浓度乙酰紫草素溶液与Huh7细胞进行共同培养，采用MTS法检测细胞的相对活力，观察乙酰紫草素对Huh7细胞的毒性作用。（2）采用JFH-1毒株感染Huh7细胞后，用不同浓度（0、0.1、0.2、0.4、0.8 μM）的乙酰紫草素溶液进行处理，检测HCV RNA以及HCV core蛋白的表达水平，观察乙酰紫草素对Huh7细胞中HCV JFH-1复制的影响。（3）取JFH-1感染的Huh7细胞分为HCV组和HCV+乙酰紫草素组，HCV+乙酰紫草素组用0.8 μM乙酰紫草素处理，HCV组不处理；取未感染JFH-1的Huh7细胞，分为乙酰紫草素组和对照组，乙酰紫草素组用0.8 μM乙酰紫草素处理，对照组不处理。通过检测LC3 mRNA以及LC3B蛋白的表达水平，观察乙酰紫草素对细胞自噬标志因子LC3表达的影响。结果（1）乙酰紫草素浓度在1.0 μM或更低时对Huh7细胞生长影响很小或没有影响，在2.50 μM以上时表现出明显的细胞毒性作用。（2）不同浓度乙酰紫草素（0.1、0.2、0.4、0.8 μM）处理组细胞内HCV RNA水平均显著低于未用乙酰紫草素处理组，差异有统计学意义（P＜0.05），HCV core蛋白的表达量均显著低于未用乙酰紫草素处理组。（3）在基因水平上，LC3 mRNA的表达水平表现为HCV组＞对照组＞HCV+乙酰紫草素组＞乙酰紫草素组，差异有统计学意义（P＜0.05）；在蛋白水平上，LC3B蛋白的表达水平表现为HCV组＞对照组＞HCV+乙酰紫草素组＞乙酰紫草素组。结论乙酰紫草素可以抑制HCV复制，同时下调自噬标志因子LC3的表达，可通过调节细胞自噬途径影响HCV病毒的复制。

ObjectiveTo investigate the effects of acetyl shikonin on HCV replication and its mechanism(s). Methods(1) Huh7 cells were cultured in vitro and co-culture with different concentrations of acetyl shikonin solution. MTS cell proliferation assay was used to detect the relative viability of the cells and the cytotoxic effect of acetyl shikonin on Huh7 cells was observed. (2)After infected with JFH-1 strain, Huh7 cells were treated with different concentrations (0, 0.1, 0.2, 0.4, 0.8 μM) of acetyl shikonin solution. The expression levels of HCV RNA and HCV core protein were detected and the effect of acetyl shikonin on HCV JFH-1 replication in Huh7 cells was observed. (3)Huh7 cells infected with JFH-1 were divided into HCV group and HCV+acetyl shikonin group. The HCV+acetyl shikonin group was treated with 0.8 μM acetyl shikonin, while the HCV group was not treated with acetyl shikonin. Huh7 cells noninfected with JFH-1 were divided into acetyl shikonin group and control group. The acetyl shikonin group was treated with 0.8 μM acetyl shikonin, and the control group was not treated with acetyl shikonin. The effect of acetyl shikonin on the expression of autophagy marker LC3 was observed by detecting the expression levels of LC3 mRNA and LC3B protein. Results(1)Acetyl shikonin had little or no effect on the growth of Huh7 cells at the concentration ≤1.0 μM, and showed a significant cytotoxic effect at the concentration≥2.50 μM. (2)The levels of HCV RNA in the groups treated with different concentrations of acetyl shikonin (0.1, 0.2, 0.4, 0.8 μM) were significantly lower than those in the group untreated with acetyl shikonin, the difference was statistically significant (P＜0.05), the expression level of HCV core protein was significantly lower than that in the group untreated with acetyl shikonin. (3)At the gene level, the mRNA expression level of LC3 was HCV group ＞control group＞HCV+acetyl shikonin group＞acetyl shikonin group, the difference was statistically significant (P＜0.05). At the protein level, the protein expression level of LC3B was HCV group＞control group＞HCV+acetyl shikonin group＞acetyl shikonin group. ConclusionAcetyl shikonin can inhibit HCV replication and down-regulate the expression of the autophagy marker LC3, acetyl shikonin can affect the replication of HCV virus by regulating the autophagy pathway.