AIM: To investigate the inhibitory effect of Chinese herbal medicine on the transcription of hepatitis C virus (HCV) structural gene in Hela D cells.METHODS: Hela cell line was transfected with recombinant pBK-CMV-HCV...AIM: To investigate the inhibitory effect of Chinese herbal medicine on the transcription of hepatitis C virus (HCV) structural gene in Hela D cells.METHODS: Hela cell line was transfected with recombinant pBK-CMV-HCV containing HCV structural gene by Lipofectamine. RT-nested-PCR and Western blot assay were used to testify the HCV gene expression in Hela cells. The Hela cells expressing HCV structural protein were named Hela D cells. Prescriptions of Xiao chaihu Decoction (XCHD),Fufang Huangqi (FFHQ) and Bingganling (BGL) wererespectively added to Hela D cells in various concentrations. Semi-quantitative RT-nested-PCR product analysis was performed according to the fluorescent density between HCV DNA band and GAPDH DNA band in gel electrophoresisafter screened. RESULTS: Recombinant pBK-CMV-HCV could correctly express the HCV structural gene in Hela D cells. After coculture of Hela D cells with three prescriptional different concentrations for 48 h respectively, the transcription of HCVgene decreased with increasing of the concentration of each prescription. The lightness ratio of HCV product bands to GAPDH product bands was 0.24, 0.10 and 0.12 in Hela D cells incubated with 0.1 g/mL of XCHD, FFHQand BGL respectively and the lightness ratio HCV product bands to GAPDH product bands was 0.75, 0.67 and 0.61respectively in the control cells. CONCLUSION: The prescriptions of XCHD, FFHQ and BGL partly inhibit the transcription of HCV structural gene inHela D cells.展开更多
lnterleukin-21(IL-21)is a recently characterized T cell-derived cytokine with a significant homology to IL-2, IL-4 and IL-15.To determine whether IL-21 has broad immunoregulatory activity and can stimulate durable ant...lnterleukin-21(IL-21)is a recently characterized T cell-derived cytokine with a significant homology to IL-2, IL-4 and IL-15.To determine whether IL-21 has broad immunoregulatory activity and can stimulate durable antitumour responses,we constructed mouse IL-21(mIL-21) recombinant plasmid and evaluated its antitumor efficacy.Mouse IL-21 cDNA was amplified from Con A-activated mouse T cells by RT-PCR.Recombinant pcDNA3.1/mIL-21 was constructed and transfected into Sp2/0 cells.Mouse IL-21 expression was analyzed by Western blotting and its activities were detected by ~3H-TdR incorporation and MTT assay.The recombinant pcDNA3.1/mIL-21 was injected s.c.into tumor lump.Tumor size,weight,the activities of CTLs,NK cells and LAK cells and serum IFN-γ,level were measured for evaluating mIL-21 mediated antitumor responses.The results indicated that mIL-21 was correctly expressed in Sp2/0 cells and it can improve the proliferation of T cells and B cells,and enhance NK cytotoxic activity in vitro.The activities of CTL and NK cells,and serum IFN-γlevel were significantly improved,furthermore the tumor growth was obviously suppressed in pcDNA3.1/mIL-21 treated mice.However,the LAK activity did not alter significantly.Taken together,this study suggests that the injection with recombinant plasmid containing mIL-21 is a potential approach for tumor gene therapy.Cellular & Molecular Immunology.2004;1(6):461-466.展开更多
A novel tuberculosis(TB)gene vaccine containing mouse granulocyte macrophage-colony stimulating factor (mGM-CSF)and a TB antigen(Ag85A)was developed in this study.The genes encoding Ag85A and mGM-CSF were amplified by...A novel tuberculosis(TB)gene vaccine containing mouse granulocyte macrophage-colony stimulating factor (mGM-CSF)and a TB antigen(Ag85A)was developed in this study.The genes encoding Ag85A and mGM-CSF were amplified by PCR respectively from the Ag85A-containing pBSby5 and pC-mGM-CSF.The genes were then cloned into two different polylinker sites of plasmid pIRES,forming a novel TB gene vaccine construct pI85AGM. Following transfection of pI85AGM plasmid into 7721 cell line by Lipofectamine^(TM),the expression of Ag85A and GM-CSF proteins was identified by Western blotting or RT-PCR.Then Balb/c mice were inoculated with the recombinant pI85AGM,pI85A,pIGM or plasmid alone,respectively.The activities of CTL,NK cells and the Ag85A-stimulated proliferation of spleen cells were measured by MTT method.The serum antibody against Ag85A was detected by ELISA.The results showed that the Ag85A and GM-CSF proteins could be expressed in 7721 cell line and the activity of CTLs and the proliferation of spleen cells were significantly increased in the pI85AGM-immunized mice,indicating that the pI85AGM-immunized mice could generate specific immune responses to Ag85A.This study might provide possibility for developing novel anti-TB gene vaccine.Cellular & Molecular Immunology.2005;2(1):57-62.Cellular & Molecular Immunology.2005;2(1):57-62.展开更多
基金Supported by the Chinese medicine and pharmacology bureau of Jiangsu Province in China
文摘AIM: To investigate the inhibitory effect of Chinese herbal medicine on the transcription of hepatitis C virus (HCV) structural gene in Hela D cells.METHODS: Hela cell line was transfected with recombinant pBK-CMV-HCV containing HCV structural gene by Lipofectamine. RT-nested-PCR and Western blot assay were used to testify the HCV gene expression in Hela cells. The Hela cells expressing HCV structural protein were named Hela D cells. Prescriptions of Xiao chaihu Decoction (XCHD),Fufang Huangqi (FFHQ) and Bingganling (BGL) wererespectively added to Hela D cells in various concentrations. Semi-quantitative RT-nested-PCR product analysis was performed according to the fluorescent density between HCV DNA band and GAPDH DNA band in gel electrophoresisafter screened. RESULTS: Recombinant pBK-CMV-HCV could correctly express the HCV structural gene in Hela D cells. After coculture of Hela D cells with three prescriptional different concentrations for 48 h respectively, the transcription of HCVgene decreased with increasing of the concentration of each prescription. The lightness ratio of HCV product bands to GAPDH product bands was 0.24, 0.10 and 0.12 in Hela D cells incubated with 0.1 g/mL of XCHD, FFHQand BGL respectively and the lightness ratio HCV product bands to GAPDH product bands was 0.75, 0.67 and 0.61respectively in the control cells. CONCLUSION: The prescriptions of XCHD, FFHQ and BGL partly inhibit the transcription of HCV structural gene inHela D cells.
文摘lnterleukin-21(IL-21)is a recently characterized T cell-derived cytokine with a significant homology to IL-2, IL-4 and IL-15.To determine whether IL-21 has broad immunoregulatory activity and can stimulate durable antitumour responses,we constructed mouse IL-21(mIL-21) recombinant plasmid and evaluated its antitumor efficacy.Mouse IL-21 cDNA was amplified from Con A-activated mouse T cells by RT-PCR.Recombinant pcDNA3.1/mIL-21 was constructed and transfected into Sp2/0 cells.Mouse IL-21 expression was analyzed by Western blotting and its activities were detected by ~3H-TdR incorporation and MTT assay.The recombinant pcDNA3.1/mIL-21 was injected s.c.into tumor lump.Tumor size,weight,the activities of CTLs,NK cells and LAK cells and serum IFN-γ,level were measured for evaluating mIL-21 mediated antitumor responses.The results indicated that mIL-21 was correctly expressed in Sp2/0 cells and it can improve the proliferation of T cells and B cells,and enhance NK cytotoxic activity in vitro.The activities of CTL and NK cells,and serum IFN-γlevel were significantly improved,furthermore the tumor growth was obviously suppressed in pcDNA3.1/mIL-21 treated mice.However,the LAK activity did not alter significantly.Taken together,this study suggests that the injection with recombinant plasmid containing mIL-21 is a potential approach for tumor gene therapy.Cellular & Molecular Immunology.2004;1(6):461-466.
文摘A novel tuberculosis(TB)gene vaccine containing mouse granulocyte macrophage-colony stimulating factor (mGM-CSF)and a TB antigen(Ag85A)was developed in this study.The genes encoding Ag85A and mGM-CSF were amplified by PCR respectively from the Ag85A-containing pBSby5 and pC-mGM-CSF.The genes were then cloned into two different polylinker sites of plasmid pIRES,forming a novel TB gene vaccine construct pI85AGM. Following transfection of pI85AGM plasmid into 7721 cell line by Lipofectamine^(TM),the expression of Ag85A and GM-CSF proteins was identified by Western blotting or RT-PCR.Then Balb/c mice were inoculated with the recombinant pI85AGM,pI85A,pIGM or plasmid alone,respectively.The activities of CTL,NK cells and the Ag85A-stimulated proliferation of spleen cells were measured by MTT method.The serum antibody against Ag85A was detected by ELISA.The results showed that the Ag85A and GM-CSF proteins could be expressed in 7721 cell line and the activity of CTLs and the proliferation of spleen cells were significantly increased in the pI85AGM-immunized mice,indicating that the pI85AGM-immunized mice could generate specific immune responses to Ag85A.This study might provide possibility for developing novel anti-TB gene vaccine.Cellular & Molecular Immunology.2005;2(1):57-62.Cellular & Molecular Immunology.2005;2(1):57-62.
