Résumé
Objective To explore effects of ginsenosides Rg1 on the expression of poly(ADP-ribose) polymerase-1 (PARP-1) and tumor necrosis factor receptor (TNFR) 1 in cortex cells after focal cerebral ischemia in rats. Methods Ninety healthy rats were randomly divided into sham-operative group, focal cerebral ischemia group, ginsenoside Rg 1groups (low, medium and high concentrations) and drug control group. Rats were intraperitoneally injected saline 45 mg/kg, saline 45 mg/kg+ginsenosides Rg1 10, 20 and 40 mg/kg, nimodipine 1 mg/kg 5 d before surgery, respectively. Focal cerebral isch?emia model was made by middle cerebral artery occluding in rats. The neurological deficit score and TTC staining were used to verify the success of the rat model. The expressions of PARP-1 and TNFR1 were evaluated by immunohistochemical meth?od and Western blot technique. Results There were obvious symptoms of neurological deficit and large pale infarct area in focal cerebral ischemia group compared with those of sham-operative group. There were higher percentages of neurological deficit score and infarct area in ginsenosides Rg1 groups and positive control group than those of sham-operative group, but which were lower than those of ischemia group (P<0.05). There were no significant differences between ginsenosides Rg1 groups and positive control group. The positive cells of PARP-1 and TNFR1 were higher in ginsenosides Rg1 low-dose group than those of sham-operative group and positive control group, while ones of medium and high-dose Rg1 group were higher than those of sham-operative group, and were lower than those of ischemia group (P<0.05). Compared with sham-op?erative group, PARP-1 and TNFR1 expression strips were significantly enhanced in ischemia group. Expression strips were higher in ginsenosides Rg1 low-dose group than those of sham-operative group. Expression strips were higher in ginsen?osides Rg1 medium-dose group than those of sham-operative group, but which were lower than those of ischemia group, and ones of high-dose group were lower than ischemia group (P<0.05). Conclusion Ginsenoside Rg1 shows protective effects on focal ischemia injury, which may be related with down-regulation of the expression of PARP-1 and TNFR1.
Résumé
Objective To explore a novel and highly specific small-molecule TNFβinhibitor by using computer-aid?ed virtual screening and cell-based assays in vitro. Methods Computer-aided drug design and virtual screening were used to design and identify chemical compounds that targeted TNFβbased on the crystal structure of the TNFβ-TNFR1 com?plex. The effect of the small-molecule compound against TNFβ-induced cytotoxicity of L929 cells was detected by MTT as?say, and the efficacy of the compound to inhibit TNFβ-induced apoptosis of L929 cells was determined by flow cytometry as?say. The impact of the compound on L929 cell cycle was examined by Propidium Iodide (PI) staining and flow cytometry, and the influence of the compound on TNFβ-triggered signal pathway was analyzed by Western blot assay and Ultra VIEW VOX 3D Live Cell Imaging System. Results No.35 compound (named as C35 thereafter) could effectively inhibit TNFβ-induced cell death in a dose dependent manner, and the half-maximum inhibition concentration (IC50) was 8.19μmol/L. Furthermore, C35 had lower cytotoxicity and minimal effect on L929 proliferation. Here we further revealed that C35 could affect TNFβ-induced apoptotic pathway by blocking the activation of Caspase 3, and markedly reduce L929 cell apoptosis induced by TNFβ. Conclusion A novel TNFβsmall-molecule inhibitor was identified by combining computer-aided virtual screening with functional assays, and which could block TNFβ-triggered apoptotic pathway and efficiently inhibit the cell death in?duced by TNFβ.