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Objective To explore influence of JAK/STAT3 signaling pathway on the apoptosis of HCC cells.Methods DNA-vector-based RNAi approach silence was used to down-regulate STAT3 expression in Bel-7402 cells.According to the STAT3 cDNA sequence in the GeneBank database,the plasmid pGCsi.U6/neoRFP-STAT3 that was designed for expression of STAT3 siRNA was constructed and synthesized,and then transfected into the Bel-7402 cells with iipofectamine 2000.The apoptotic rate was measured with flow cytometry (FCM) and annexinV/PI apoptosis detection kit staining.The mitochondrial membrane potential (BΨm) was visualized by the JC-1 fluorescence staining and the inverted fluorescence microscope.Moreover,the expression of caspase-3 protein was analyzed by Western blotting.Results The apoptotic ratio of STAT3-siRNA group was (38.82 ± 0.88) ％,which was significantly higher than that in other group [control group (9.22 ± 0.38) ％,scramble-siRNA group (16.47 ± 1.04) ％,P ＜ 0.05].The mitochondrial membrane potential of STAT3-siRNA group observed by the JC-1 fluorescence staining was decreased significantly[(91.33 ± 1.78) ％] and [(89.90 ± 1.92) ％ vs (59.06 ± 1.89) ％,P ＜ 0.05].The Western blot results showed that the protein expression of active caspase-3 in STAT3-siRNA group was significantly higher than other groups (0.48 ± 0.05 vs 0.22 ± 0.04 and 0.26 ± 0.06,P ＜ 0.05).Conclusions STAT3 gene silencing significantly improves the apoptotic effect in the Bel-7402 ceils.
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Objective To explore the role of phosphatidylinositol 3-kinase (PI3K) signal pathway in interleukin-8 (IL-8)/chemokine (C-X-C Motif) receptor 2 (CXCR2)-mediated growth and metastasis of liver cancer.Methods Eighty-two cases of liver cancer tissues and serum samples,and 21 cases of normal tissues and serum samples were collected.The level of serum IL-8 was measured by enzyme-linked immunesorbent assay (ELISA).Expression of CXCR2 was detected by immunohistochemical method,and its correlation with clinicopathological parameters was also analyzed.Different stages of liver cancer fresh tissues were selected,and expressions of CXCR2,PI3Kγ,AKT,and NF-κB were detected by Westeru blotting analyses.Results The levels of serum IL-8 in liver cancer patients (stages Ⅰ,Ⅱ,Ⅲ,Ⅳ) were (120.34 ±15.52) pg/ml,(221.60 ±23.10) pg/ml,(279.50 ±24.25) pg/ml,(324.81 ±22.50) pg/ml,respectively.With the increasing degree of liver cancers,levels of IL-8 were increased,most significant was in the Ⅳ period of liver cancer; the expression of CXCR2 was significantly correlated with pathological stages,degree of differentiation,lymph node and distant metastases of liver cancer (P ＜ 0.05) ; serum IL-8 levels were consistent with expression of CXCR2 in liver cancer tissues,which had a strong correlation with activation of PI3K γ,AKT,and NF-κ B proteins.Conclusions IL-8/CXCR2 was able to mediate the growth and metastasis of liver cancers,which might be through the activation of PI3K signaling pathway.
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Objective To evaluate the effects of remifentanil on the proliferation,the cell cycle and apoptosis of human liver carcinoma cell line HepG2 in vitro.Methods Human liver carcinoma cells HepG2 were cultured in vitro.The HepG2 cells of the test group were incubated in the RPMI-1640 medium with remifentanil at different concentration(0.001,0.01,0.1,1,10,100,200 μmol/L).The HepG2 cells of the control group were incubated in the RPMI-1640 medium for 48 hours.The level of the cell proliferation was evaluated with methylthiazolyldiphenyl-tetrazolium bromide (MTT) method.The cell cycle was detected with flow cytometry (FCM).The morphological change of apoptosis cell was observed by fluorescence microscopy after staining by Hoechst33258.Results Remifentanil inhibited the proliferation of the HepG2 cells with a dose-dependent effect.Compared with control group,the cell proliferation capability was apparently decreased in the test group (P ＜ 0.05) when the concentration of remifentanil was over 1 μmol/L (P ＜0.05).However,no significant difference in cell proliferation was found when remifentanil was 100 and 200 μmol/ L.The ratio of G0/G1 phase of HepG2 cells was significantly enhanced and the ratio of S phase of HepG2 cells was significantly decreased when remifentanil was over 1 μmol/L.The fluorescent microscopy stained by the Hoechst33258 showed part of HcpG2 cells apoptosis in test group,and the apoptotic rate was significantly increased when remifentanil was over 1 μmol/L (P ＜ 0.05).Conclusions The data suggest that remifentanil would inhibit the proliferation of HepG2 cells and induce apoptosis when remifentanil was over 1 μmol/L.
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Objective:To compare the killing effects on HCC SMMC-7721 cells of immunoliposome PE38、immunotoxin PE38 and liposome PE38. Methods:Dissolute PC,CHOL and CHS in chloroform in proportion.Dry membrane was formed after chloroform was removed.Add the protein solution of PE38 to dissolute the dry membrane.Then pass the solution over a Sephadex G-50 column after ultrasoned and filtrated to detect the encapsulation efficiency of the liposome.The solution reacted in EDC,SSNHS and MES for 30 minutes.Then add Ab to the solution in 4℃ over night.MTT method was used to detect the killing effects on HCC cells of immunoliposome PE38、immunotoxin PE38 and liposome PE38 in vitro. Results:The killing effects on HCC cells of immunoliposome PE38 is the best,and that of liposome PE38 is the worst.The respective IC_(50) s are:0.59 ?g/ml、6.04 ?g/ml and 92.07 ?g/ml. Conclusion:Immunoliposome PE38 may be a protential durg in the treatment of hepatocarinoma.