改良三甲散含药脑脊液对脂多糖诱导的BV2小胶质细胞炎症模型相关因子及蛋白表达的影响

目的探讨改良三甲散含药脑脊液对脂多糖(LPS)所诱导的BV2小胶质细胞炎性反应的影响。方法将BV2细胞分为空白对照组,模型组,10.0%、5.0%、2.5%改良三甲散组,石杉碱甲组。细胞培养贴壁后,改良三甲散各组及石杉碱甲组分别加入10.0%、5.0%、2.5%、10.0%含药脑脊液,37℃孵育2 h后除空白对照组均加入LPS(终浓度为1 mg/L),继续孵育24 h后收集培养上清,提取蛋白样品。应用硝酸还原酶法检测各组细胞培养上清中一氧化氮(NO)的含量;ELISA法检测各组细胞培养上清中诱导型一氧化氮合酶(i NOS)、白细胞介素1β(IL-1β)、肿瘤坏死因子α(TNF-α)的含量;Western blot法检测各组细胞c-Jun氨基末端激酶(JNK)、磷酸化c-Jun氨基末端激酶(p-JNK)的表达。结果模型组培养上清中NO、i NOS、IL-1β、TNF-α的含量明显升高,p-JNK表达上调;10%、5%改良三甲散组培养上清中NO、i NOS、IL-1β、TNF-α的含量明显下降,p-JNK表达下调。结论改良三甲散含药脑脊液对LPS所致的BV2小胶质细胞的炎性反应具有调节作用,其作用机制可能与调控p-JNK的表达、减少炎性介质的产生有关。

Influence of medicated cerebrospinal fluid of modified Sanjia San on relative factors of BV2 microglia cell inflammatory model and protein expressions induced by lipopolysaccharide

Objective To investigate the influence of medicated cerebrospinal fluid of modified Sanjia San on inflammatory reactions of BV2 microglia cells induced by lipopolysaccharide (LPS).Methods BV2 microglia cells were divided into blank control group, model group, three modified Sanjia San groups and huperzine group.After the cells cultured and adhered to wall, modified Sanjia San groups and huperzine group were, respectively, given medicated cerebrospinal fluid (10.0%, 5.0%, 2.5%, 10.0%) and incubated at 37 ℃ for 2 h.Except of blank control group, all groups were given LPS (1 mg/L as final concentration) and incubated continuously for 24 h, and then supernatant was collected for extracting protein samples.The content of nitric oxide (NO) in supernatant was detected by using nitrate reductase method in all groups.The content of inducible nitric oxide synthase (NOS), interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in supernatant were detected by using enzyme-linked immunosorbent assay (ELISA) in all groups.The expressions of c-Jun N-terminal kinase (JNK) and phospho-c-Jun N-terminal kinase (p-JNK) were detected by using Western blot test in all groups.Results The content of supernatant NO, iNOS, IL-1β and TNF-α increased significantly, and expression of p-JNK was up-regulated in model group.The content of supernatant NO, iNOS, IL-1β and TNF-α decreased significantly, and expression of p-JNK was down-regulated in 10.0% and 5.0% modified Sanjia San groups.Conclusion The medicated cerebrospinal fluid of modified Sanjia San has the regulative effect on inflammatory reactions of BV2 microglia cells induced by LPS, and the mechanism may be related to the controlling of p-JNK expression and reducing of generation of inflammatory media.