穿心莲内酯靶向Akt信号干预HIV感染T细胞的效率研究

Exploring the efficiency of andrographolide targeting Akt signal to interfere with HIV-infected T cells

目的探讨穿心莲内酯靶向Akt信号干预人类免疫缺陷病毒(HIV)感染T细胞的效率。方法培养Jurkat T细胞,用WST-1试剂检测穿心莲内酯干预对Jurkat T细胞活性的影响。将Jurkat T细胞分为空白对照组、穿心莲内酯组、LY294002组(PI3K抑制剂)及穿心莲内酯联合LY294002组。各组药物分别干预24 h后,采用实时荧光定量聚合酶链反应(qRT-PCR)检测各组细胞PI3K、Akt mRNA相对表达量,采用Western blotting检测各组细胞p-PI3K/PI3K、p-Akt/Akt蛋白相对表达量。制备HIV假病毒,用假病毒感染各组细胞,萤光素酶系统检测假病毒感染Jurkat T细胞的效率。结果WST-1试剂检测结果显示,随着穿心莲内酯浓度增加,Jurkat T细胞活力降低(P<0.05)。qRT-PCR和Western blotting检测结果显示,穿心莲内酯降低Jurkat T细胞PI3K及其下游Akt mRNA相对表达量及p-PI3K/PI3K、p-Akt/Akt蛋白相对表达量(P<0.05)。HIV假病毒感染实验结果显示,穿心莲内酯降低HIV假病毒感染Jurkat T细胞的效率,联合组比单独组的抑制效果更好(P<0.05)。结论穿心莲内酯可能通过PI3K降低其下游Akt的活化,从而减少HIV假病毒在Jurkat T细胞间的传播及进入细胞的机会,降低其感染Jurkat T细胞的概率。

Objective This study aims to investigate the efficiency of puerarin targeting the Akt signaling pathway in intervening with Human Immunodeficiency Virus(HIV)infection in T cells.Methods Jurkat T cells were cultured,and the impact of puerarin intervention on Jurkat T cell activity was assessed using WST-1 reagent.Jurkat T cells were divided into a blank control group,puerarin group,LY294002 group(PI3K inhibitor),and a combined puerarin-LY294002 group.After 24 hours of drug intervention,real-time quantitative polymerase chain reaction(qRT-PCR)was employed to measure the relative expression of PI3K and Akt mRNA in each group,and Western blotting was used to detect the relative expression of p-PI3K/PI3K and p-Akt/Akt proteins.HIV pseudovirus was prepared to infect cells in each group,and the efficiency of pseudovirus infection in Jurkat T cells was assessed using a luciferase system.Results WST-1 assay results revealed a dose-dependent reduction in Jurkat T cell viability with increasing concentrations of puerarin(P<0.05).qRT-PCR and Western blotting results indicated that puerarin decreased the relative expression of PI3K and downstream Akt mRNA,as well as p-PI3K/PI3K and p-Akt/Akt proteins in Jurkat T cells(P<0.05).Results from the HIV pseudovirus infection experiment demonstrated that puerarin reduced the efficiency of HIV pseudovirus infection in Jurkat T cells,with the combined treatment showing superior inhibitory effects compared to the individual treatment(P<0.05).Conclusion Puerarin may reduce the activation of Akt downstream of PI3K,thereby decreasing the opportunity for HIV pseudovirus spread and entry into Jurkat T cells,ultimately lowering the probability of infection.