In order to study the effects and the possible mechanisms of Daxx overexpressed in HepG2 to hydrogen peroxide treatment, and to search new targets for cancer chemotherapy, HepG2cells were transfected using lipofectamine 2000, and selected by treatment with G418. Stable cell lines were confirmed by reverse transeriptase polymerase chain reaction (RT-PCR) targeting vector gene. Experiments include the following groups: (1) control group (non-transfected cells); (2) transfected with empty vector (HepG2/GFP cells); and (3) transfected with pEGFP-C1-Daxx (HepG2/GFP-Daxx cells). After incubation with hydrogen peroxide (H2O2) for 24 h, cellular viability was analyzed by MTT, and cellular apoptosis was measured by flow cytometric analysis. Gene expression at protein level was detected by Western blot. The RT-PCR results showed that Daxx RNA in cells transfected with pEGFP-C1-Daxx was increased significantly compared with that in the HepG2/GFP cells. Fluorescence microscopy revealed that Daxx protein was localized in the nuclei. Hydrogen peroxide was used to induce apoptosis of HepG2 cells and observed that the hydrogen peroxide decreased the viability of HepG2 cells in concentration-dependent pattern. The IC50 values in three groups (Normal cells, HepG2/GFP cells and HepG2/GFP-Daxx cells) were 0.72, 0.76, and 0.49 mmol/L respectively. The apoptotic ratio was significantly higher in HepG2/GFP-Daxx cells as compared to the other two groups. HepG2/GFP-Daxx cell incubated with hydrogen peroxide, showed a significant increase in the activation of caspase-3 and JNK as compare with the other groups. Over-expression of Daxx facilitated HepG2 cells apoptosis induced by hydrogen peroxide. Furthermore, there may be a synergetic relation with apoptosis and increase of JNK activity.

Objective To investigate the effect of caveolin-1 on rabbit carotid artery anastomotic stenosis and its relationship with TNF-α.Methods 40 New Zealand white rabbits were randomly divided into normal group,surgical group,empty vector group and caveolin-1 transfecting group.Carotid artery end-to-end anastomosis was done in the rabbits except these in normal group.Five specimens were randomly taken on day 7 after surgery for Westem blot to detect the expression of caveolin-1 and TNF-α; The rest specimens were taken for HE staining at fourth week.The ratio of intima/media area were measured by Image-Pro Plus 6.0 software in order to observe the proliferation of intima.Results Compared with normal group,in surgical group intimal proliferation was significant,the intima/media ratio was significantly higher (P ＜ 0.05) ; Compared with surgical group,in caveolin-1 transfected group neointimal proliferation was not obvious,the intima/media ratio decreased (P ＜ 0.05).Western blot showed that:compared with the surgical group,caveolin-1 expression in transfected group was significantly higher (P ＜ 0.05) ; compared with normal group,the TNF-α expression in surgical group increased (t:41.28,P ＜ 0.05) ; Compared with surgical group,TNF-α expression in transfected group decreased significantly (t:36.37,P ＜ 0.05).Conclusions Caveolin-1 inhibits vascular anastomotic stenosis,possibly by down-regulating TNF-α expression.

Objective To investigate the ischemic injury of hepatic cell caused by hepatic artery occlusion.Methods The hepatic artery was occluded in 20 dogs via operation,while the portal vein remained patent.Specimens were gained from the right liver at four time points:before occlusion of the hepatic artery,20(minutes),40 minutes and 60 minutes after artery occlusion.Each specimen was examined by HE and BCL-2 by immunohistochemistry.The gray scale of BCL-2 in HE sections was detected.Results Hepatic cellular injury was obvious 20 minutes after occlusion of the hepatic artery.Irreversible hepatic cellular injury was(observed) 60 minutes after hepatic artery occlusion.The results showed that the gray scale of BCL-2 at every time point after hepatic artery occlusion were significantly different from that before hepatic artery occlusion(P

Objective To investigate the effect of Caveolin-1 on extracellular regulated protein kinases of rabbit carotid artery anastomotic restenosis.Methods 40 New Zealand white rabbits were randomly divided into normal control group,carotid artery end to end anastomosis surgical group,empty vector transfection on the site of anastomosis group and Caveolin-1 transfected group.Left carotid artery endto-end anastomosis was performed,and the mixture of Caveolin-1 plasmid and liposome lipofectin 2000 (transfected group) or empty plasmid and lipofectin 2000 mixture (empty vector group) were transfected on anastomosis.Specimens were taken at 7 d after surgery for Western blot and RT-PCR to detect the expression of protein and mRNA.Specimens were taken for HE staining at 28 d to observe the proliferation of intima,and measured the ratio of intima/media area by Image-Pro Plus 6.0 software.Results Compared with surgical group,the ratio of intima/media area in Caveolin-1 transfected group decreased by about 50％.Compared with surgical group and empty vector group,the Caveolin-1 mRNA expression and protein activity significantly increased (t =36.59,P ＜ 0.01) ; the ERK1/2 mRNA expression and protein activity significantly decreased on rabbit carotid artery anastomotic site in Caveolin-1 transfected group (t =32.64and 7.27,P ＜ 0.01).Conclusions Caveolin-1 inhibits anastomotic restenosis possibly by regulating the activation of ERK.

Aim To explore the anti-proliferation effects of curcumin trinicotinate ( CurTn ) on vascular smooth muscle cell ( VSMC ) and its mechanism. Methods The cells were cultured in DMEM supple-mented with 10% fetal bovine serum. MTT assay was used to examine cell proliferation. FCM was used to observe cell cycle. The expressions of PCNA, Cy-clinD1 and p-ERK1/2 were analyzed using Western blot. Results CurTn could inhibit the proliferation of VSMC and showed a certain amount-time relationship. What’ s more, CurTn could increase the G1 phase pro-portion of cell, decrease the S phase proportion and the expression level of PCNA protein. It was also found that CurTn significantly inhibit the protein expression of p-ERK1/2 and Cyclin D1 . Conclusion CurTn may inhibit the proliferation of VSMC via downregulating the expression of CyclinD1 and p-ERK1/2 .