To investigate the phenotypic knockout of HIV-1 chemokine coreceptor CXCR4 and CCR5 by intrakines and its inhibitory effect on HIV-1 infection. Primary human PBLs were transduced with the recombinant vector pLNCX-R-K-...To investigate the phenotypic knockout of HIV-1 chemokine coreceptor CXCR4 and CCR5 by intrakines and its inhibitory effect on HIV-1 infection. Primary human PBLs were transduced with the recombinant vector pLNCX-R-K-S-K followed by anti-NGFR/anti-IgG-magnetic bead method selection and FCM detection. The transduced PBI.S were infected with DP1 HFV-1 virus thereafter envelope-mediated syncytium formation and p24 detection were carried out to study the blockage of HIV-1 infection by co-inactivation of CCR5 and CXCR4. pLNCX-R-K-S-K -transduced PBLs were isolated with an anti-NGFR/anti-IgG-magnetic bead method. After isolation, about 70% of the PBI.S were posi- tive for the NGFR marker. When the transduced PBLs were infected with DP1 HIV-1 virus, envelop-mediated syncytium for- mation was almost completely inhibited by pLNCX-R-K-S-K transfection. Also, p24 antigen was very low in the cultures of pLNCX-R-K-S-K transduced PBLs. pLNCX-R-K-S-K transduction inhibited the produc- tion of DP1 p24 antigen by 15%, 43% and 19% on days 4, 7 and 10 respectively. The lymphocytes with the phenotypic knockout of CCR5 and CXCR4 could protect primary human PBLs from DP1 HIV-1 virus infection.展开更多
Objective: To evaluate the humoral immune induction in rats of a candidate AIDS vaccine expressing the gag p24 gene froma subtype B HIV-1 isolate. Methods: The amplified p24 gene was inserted into aneukaryotic express...Objective: To evaluate the humoral immune induction in rats of a candidate AIDS vaccine expressing the gag p24 gene froma subtype B HIV-1 isolate. Methods: The amplified p24 gene was inserted into aneukaryotic expression vector to form the supercoiled DNAvaccine. The linearized expressed DNA vaccine was preparedfrom the expression plasmid by polymerase chain reaction(PCR). The antigen gene expression in rats of the linearizedand supercoiled DNA vaccines were in vitro and in vivodetected. Results: In vitro transcription and Northern hybridizationshowed that the linearized DNA vaccine could synthesizeamounts or p24 mRNA similar to the supercoiled DNA vaccine.Antibody assays of inoculated rats confirmed that thelinearized expression DNA could induce a slightly higherantibody titer than the expression plasmid, while the highestautibody titer had been induced by plasmid plus adjuvantinoculation. Conclusion: The construction of a candidate AIDS vaccinebased on the p24 gene could shed light on a potential IV vaccine, meriting evaluation in a rhesus macaque SHIV-AIDSmodel.展开更多
文摘To investigate the phenotypic knockout of HIV-1 chemokine coreceptor CXCR4 and CCR5 by intrakines and its inhibitory effect on HIV-1 infection. Primary human PBLs were transduced with the recombinant vector pLNCX-R-K-S-K followed by anti-NGFR/anti-IgG-magnetic bead method selection and FCM detection. The transduced PBI.S were infected with DP1 HFV-1 virus thereafter envelope-mediated syncytium formation and p24 detection were carried out to study the blockage of HIV-1 infection by co-inactivation of CCR5 and CXCR4. pLNCX-R-K-S-K -transduced PBLs were isolated with an anti-NGFR/anti-IgG-magnetic bead method. After isolation, about 70% of the PBI.S were posi- tive for the NGFR marker. When the transduced PBLs were infected with DP1 HIV-1 virus, envelop-mediated syncytium for- mation was almost completely inhibited by pLNCX-R-K-S-K transfection. Also, p24 antigen was very low in the cultures of pLNCX-R-K-S-K transduced PBLs. pLNCX-R-K-S-K transduction inhibited the produc- tion of DP1 p24 antigen by 15%, 43% and 19% on days 4, 7 and 10 respectively. The lymphocytes with the phenotypic knockout of CCR5 and CXCR4 could protect primary human PBLs from DP1 HIV-1 virus infection.
基金Financially supported by Natural Science Foundation of China(No 39870725)Natural Science Foundation of Guangdong(No 980642)Research Foundation of Guangdong Education Buresu(No.20032).
文摘Objective: To evaluate the humoral immune induction in rats of a candidate AIDS vaccine expressing the gag p24 gene froma subtype B HIV-1 isolate. Methods: The amplified p24 gene was inserted into aneukaryotic expression vector to form the supercoiled DNAvaccine. The linearized expressed DNA vaccine was preparedfrom the expression plasmid by polymerase chain reaction(PCR). The antigen gene expression in rats of the linearizedand supercoiled DNA vaccines were in vitro and in vivodetected. Results: In vitro transcription and Northern hybridizationshowed that the linearized DNA vaccine could synthesizeamounts or p24 mRNA similar to the supercoiled DNA vaccine.Antibody assays of inoculated rats confirmed that thelinearized expression DNA could induce a slightly higherantibody titer than the expression plasmid, while the highestautibody titer had been induced by plasmid plus adjuvantinoculation. Conclusion: The construction of a candidate AIDS vaccinebased on the p24 gene could shed light on a potential IV vaccine, meriting evaluation in a rhesus macaque SHIV-AIDSmodel.