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目的探讨金银花抗氧化作用分子学机理。方法采用过氧化氢(H2O2)作用于人正常RBL肝细胞,制作氧化应激损伤的细胞模型。将培养的细胞分为四个实验组:正常对照组(C),H2O2损伤组(H2),金银花前保护组(Jb),金银花后保护组(Ja)。采用TUNEL和DNALadder法,检测各组细胞凋亡情况。应用免疫组化和Westernblot观察各组细胞的HSP-70、NF-kB、Bcl-2、Bax及Caspase-3的表达及免疫反应活性。应用MTT实验检测RBL细胞的生存率。结果Jb组的细胞凋亡率明显低于H2和Ja组,Bcl-2表达增高,HSP-70、NF-kB、Bax和Caspase-3的表达降低。结论金银花对RBL细胞氧化应激损伤具有一定保护作用,且前保护作用效果好于后保护作用,其保护作用机理可能是通过抑制HSP-70和NF-kB的表达,阻抑NF-kB信号传导途径并提高细胞内抗氧化防御酶系水平。
Objective To explore the molecular mechanism of antioxidant effect of honeysuckle. Methods Human normal RBL hepatocytes were treated with hydrogen peroxide (H2O2) to prepare cell models of oxidative stress injury. The cultured cells were divided into four experimental groups: normal control group (C), H2O2 injury group (H2), pre-honeysuckle protection group (Jb), and honeysuckle post-protection group (Ja). TUNEL and DNALadder methods were used to detect the apoptosis in each group. Immunohistochemistry and Western blot were used to observe the expression and immunoreactivity of HSP-70, NF-kB, Bcl-2, Bax and Caspase-3 in each group of cells. MTT assay was used to detect the survival rate of RBL cells. Results The apoptosis rate of Jb group was significantly lower than that of H2 and Ja groups. The expression of Bcl-2 was increased, and the expression of HSP-70, NF-kB, Bax and Caspase-3 were decreased. Conclusion Honeysuckle has a protective effect on oxidative stress injury in RBL cells, and the former protective effect is better than the latter. The protective mechanism may be through inhibiting the expression of HSP-70 and NF-kB and blocking NF-kB signaling. Route and increase the level of antioxidant defense enzymes in cells.