AIM: To evaluate the effects of NS-398, a cyclooxygenase-2 (COX-2) inhibitor, on the proliferation and apoptosis of HepG2 cells. METHODS: The effects of NS-398 on the proliferation of HepG2 cells were evaluated by MTT...AIM: To evaluate the effects of NS-398, a cyclooxygenase-2 (COX-2) inhibitor, on the proliferation and apoptosis of HepG2 cells. METHODS: The effects of NS-398 on the proliferation of HepG2 cells were evaluated by MTT. DNA fragmentation gel analysis was used to analyze the apoptotic cells. DNA ploidy and apoptotic cell percentage were calculated by flow cytornetry. The expression of COX-2 and Bcl-2 mRNA was identified by competitive RT-PCR. Furthermore, expression level of Bcl-2 was detected using Western blot in HepG2 after treated with NS-398. RESULTS: NS-398 inhibited cell proliferation and induced apoptosis of HepG2 cells in a concentration-dependent manner. DNA ploidy analysis showed that S phase cells were significantly decreased with increase of NS-398 concentration. The quiescent GO/G1 phase was accumulated with decrease of Bcl-2 mRNA. Whereas NS-398 had no effect on the expression of COX-2 mRNA, and no correlations were found between COX-2 mRNA and HepG2 cell proliferation and apoptosis induced by NS-398 (r=0.056 and r=0.119, respectively). Bcl-2 protein level was inhibited after treated with NS-398. CONCLUSION: NS-398 significantly inhibits the proliferation and induces apoptosis of HepG2 cells. Mechanisms involved may be accumulation of quiescent GO/G1 phase and decrease of Bcl-2 expression.展开更多
AIM: To study the cell cycle alterations of human hepatoma cell line HepG2 in vitro after ^60Co γ-irradiation and further to examine the mechanisms underlying the enhancement of radiosensitivity to γ-irradiation in ...AIM: To study the cell cycle alterations of human hepatoma cell line HepG2 in vitro after ^60Co γ-irradiation and further to examine the mechanisms underlying the enhancement of radiosensitivity to γ-irradiation in HepG2 transiently transfected with wild type p27^kip1. METHODS: The proliferation of HepG2 cells was evaluated with MTT assay, and the cell cycle profile and apoptosis were assessed by cell morphology, DNA fragmentation analysis and flow cytometry. HepG2 cells were transfectedwith p27^kip1 wild type by using Lipofectamine (LF2000), and the expression and subcellular localization of p27^kip1 in HepG2 were detected by immunocytochemistry.RESULTS: ^60Co γ-irradiation inhibited the growth of HepG2 cells in a dose-dependent manner. Apoptosis of HepG2 cells was induced 48 h after ~, ray exposure. Furthermore research was carried out to induce exogenous expression of p27^kip1 in HepG2. The expression of p27^kip1 induced G0/G1 phase arrest in HepG2 cells. The overexpression of p27^kip1 enhanced ^60Co γ-irradiation-induced radiosensitivity in HepG2 cells. CONCLUSION: Overexpression of p27^kip1 is a rational approach to improve conventional radiotherapy outcomes, which may be a possible strategy for human hepatoma therapy.展开更多
AIM： To analyze the neutralizing activity of antibodies against E1 region of hepatitis C virus （HCV）. Specific polydonal antibody was raised via immunization of New Zealand rabbits with a synthetic peptide that had...AIM： To analyze the neutralizing activity of antibodies against E1 region of hepatitis C virus （HCV）. Specific polydonal antibody was raised via immunization of New Zealand rabbits with a synthetic peptide that had been derived from the E1 region of HCV and was shown to be highly conserved among HCV published genotypes. METHODS： Hyper-immune HCV E1 antibodies were incubated over night at 4 ℃ with serum samples positive for HCV RNA, with viral loads ranging from 615 to 3.2 million IU/mL. Treated sera were incubated with HepG2 cells for 90 min. Blocking of viral binding and entry into cells by anti E1 antibody were tested by means of RTPCR and flow cytometry. RESULTS： Direct immunostaining using FITC conjugated E1 antibody followed by Flow cytometric analysis showed reduced mean fluorescence intensity in samples pre-incubated with E1 antibody compared with untreated samples. Furthermore, 13 out of 18 positive sera （72%） showed complete inhibition of infectivity as detected by RT-PCR. CONCLUSION： In house produced E1 antibody, blocks binding and entry of HCV virion infection to target cells suggesting the involvement of this epitope in virus binding and entry. Isolation of these antibodies that block virus attachment to human cells are useful as therapeutic reagents.展开更多
文摘AIM: To evaluate the effects of NS-398, a cyclooxygenase-2 (COX-2) inhibitor, on the proliferation and apoptosis of HepG2 cells. METHODS: The effects of NS-398 on the proliferation of HepG2 cells were evaluated by MTT. DNA fragmentation gel analysis was used to analyze the apoptotic cells. DNA ploidy and apoptotic cell percentage were calculated by flow cytornetry. The expression of COX-2 and Bcl-2 mRNA was identified by competitive RT-PCR. Furthermore, expression level of Bcl-2 was detected using Western blot in HepG2 after treated with NS-398. RESULTS: NS-398 inhibited cell proliferation and induced apoptosis of HepG2 cells in a concentration-dependent manner. DNA ploidy analysis showed that S phase cells were significantly decreased with increase of NS-398 concentration. The quiescent GO/G1 phase was accumulated with decrease of Bcl-2 mRNA. Whereas NS-398 had no effect on the expression of COX-2 mRNA, and no correlations were found between COX-2 mRNA and HepG2 cell proliferation and apoptosis induced by NS-398 (r=0.056 and r=0.119, respectively). Bcl-2 protein level was inhibited after treated with NS-398. CONCLUSION: NS-398 significantly inhibits the proliferation and induces apoptosis of HepG2 cells. Mechanisms involved may be accumulation of quiescent GO/G1 phase and decrease of Bcl-2 expression.
基金Supported by the National Postdoctor Research Foundation of China,No.2003034383
文摘AIM: To study the cell cycle alterations of human hepatoma cell line HepG2 in vitro after ^60Co γ-irradiation and further to examine the mechanisms underlying the enhancement of radiosensitivity to γ-irradiation in HepG2 transiently transfected with wild type p27^kip1. METHODS: The proliferation of HepG2 cells was evaluated with MTT assay, and the cell cycle profile and apoptosis were assessed by cell morphology, DNA fragmentation analysis and flow cytometry. HepG2 cells were transfectedwith p27^kip1 wild type by using Lipofectamine (LF2000), and the expression and subcellular localization of p27^kip1 in HepG2 were detected by immunocytochemistry.RESULTS: ^60Co γ-irradiation inhibited the growth of HepG2 cells in a dose-dependent manner. Apoptosis of HepG2 cells was induced 48 h after ~, ray exposure. Furthermore research was carried out to induce exogenous expression of p27^kip1 in HepG2. The expression of p27^kip1 induced G0/G1 phase arrest in HepG2 cells. The overexpression of p27^kip1 enhanced ^60Co γ-irradiation-induced radiosensitivity in HepG2 cells. CONCLUSION: Overexpression of p27^kip1 is a rational approach to improve conventional radiotherapy outcomes, which may be a possible strategy for human hepatoma therapy.
基金Supported by the Ministry of Scientific Research, Academy of Scientific Research and Technology, Medical Research Council Code: P5-MED-030-01US-Egypt joint project BIO7-002-011
文摘AIM： To analyze the neutralizing activity of antibodies against E1 region of hepatitis C virus （HCV）. Specific polydonal antibody was raised via immunization of New Zealand rabbits with a synthetic peptide that had been derived from the E1 region of HCV and was shown to be highly conserved among HCV published genotypes. METHODS： Hyper-immune HCV E1 antibodies were incubated over night at 4 ℃ with serum samples positive for HCV RNA, with viral loads ranging from 615 to 3.2 million IU/mL. Treated sera were incubated with HepG2 cells for 90 min. Blocking of viral binding and entry into cells by anti E1 antibody were tested by means of RTPCR and flow cytometry. RESULTS： Direct immunostaining using FITC conjugated E1 antibody followed by Flow cytometric analysis showed reduced mean fluorescence intensity in samples pre-incubated with E1 antibody compared with untreated samples. Furthermore, 13 out of 18 positive sera （72%） showed complete inhibition of infectivity as detected by RT-PCR. CONCLUSION： In house produced E1 antibody, blocks binding and entry of HCV virion infection to target cells suggesting the involvement of this epitope in virus binding and entry. Isolation of these antibodies that block virus attachment to human cells are useful as therapeutic reagents.
