Human immunodeficiency virus type 1(HIV-1)V2 L175P mutation increases the binding capability of neutralizing antibodies to HIV-1 / 中华皮肤科杂志

Objective To study the impact of HIV-1 V2 L175P mutation on the binding capability of anti-V3 neutralizing antibodies to HIV-1.Methods A series of eukaryotic cell expression plasmids were used to concatenate wild type and mutant env gene of HIV-1 and green fluorescent protein(GFP)gene.The recombinant plasmids were transfected into 293T cells to express HIV-1 gp120 protein on the surface of cells.The successfully transfected cells were screened by GFP florescence marker.Immunostaining and dual fluorescence flow cytometry were performed to test the binding affinity of several common V3 region specific neutralizing antibodies to wild type or mutant gp120 proteins.Results The mean fluorescence intensity(MFI)of mutant gp120-expressing 293T cells were significantly higher than that of negative control cells(expressing GFP).Flow cytometry showed that the curve for mutant gp120-expressing 293T cells was obviously different in shape and peak from that for the negative control,while most parts of the curve for the wild type gp120-expressing 293T cells overlapped with those for the negative control.Conclusion The V2 region mutation may increase the sensitivity of HIV-1 to the neutralization by V3 region specific antibodies.

Efficiency of Recombinant Bacille Calmette-Guerin in Inducing Humoral and Cell Mediated Immunities against Human Immunodeficiency Virus Type 1 Third Variable Domain in Immunized Mice

PURPOSE: The third variable (V3) loop of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein has been intensively studied for AIDS vaccine development. Bacille Calmette-Guerin (BCG) is widely used to immunize against tuberculosis and has many advantages as a vaccine vehicle, such as low toxicity, adjuvant potential, low cost, and long-lasting immune-inducing capacity. This work was initiated to investigate the immunogenicity of recombinant BCG (rBCG-mV3) designed to express trimeric HIV-1 V3 loop (mV3) in rBCG-mV3-immunized animals. MATERIALS AND METHODS: HIV-1 V3-concatamer was cloned into pMV261, a BCG-expression vector, and then rBCG-mV3 was constructed by introducing the recombinant plasmid (pMV-V3). The recombinant BCG was examined with regard to its expression of V3-concatamer and the genetic stability in vivo and in vitro. The immune responses induced by recombinant BCG were tested in immunized mice and guinea pigs. RESULTS: The rBCG-mV3 expressed detectable amounts of V3-concatamer when induced by single heat-shock. The recombinant BCG was genetically stable and maintained the introduced mV3 gene for several weeks. V3-specific antibodies were clearly detected 6 weeks after inoculation. The antibody titer rapidly increased after immunization up to 10 weeks, and then maintained for over 4 weeks. IgG2a was prevalent in the V3-specific antiserum. The recombinant BCG was also effective in inducing delayed-type hypersensitivity responses in the immunized guinea pigs. rBCG-immunized mice retained substantial amounts of V3-specific T cells in the spleen, even 5 months after the first immunization. CONCLUSION: Recombinant BCG-mV3 is very efficient in inducing humoral and long-lasting cell-mediated immunity against HIV-1 V3 in the immunized animals.

Efficiency of Recombinant Bacille Calmette-Guerin in Inducing Humoral and Cell Mediated Immunities against Human Immunodeficiency Virus Type 1 Third Variable Domain in Immunized Mice

PURPOSE: The third variable (V3) loop of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein has been intensively studied for AIDS vaccine development. Bacille Calmette-Guerin (BCG) is widely used to immunize against tuberculosis and has many advantages as a vaccine vehicle, such as low toxicity, adjuvant potential, low cost, and long-lasting immune-inducing capacity. This work was initiated to investigate the immunogenicity of recombinant BCG (rBCG-mV3) designed to express trimeric HIV-1 V3 loop (mV3) in rBCG-mV3-immunized animals. MATERIALS AND METHODS: HIV-1 V3-concatamer was cloned into pMV261, a BCG-expression vector, and then rBCG-mV3 was constructed by introducing the recombinant plasmid (pMV-V3). The recombinant BCG was examined with regard to its expression of V3-concatamer and the genetic stability in vivo and in vitro. The immune responses induced by recombinant BCG were tested in immunized mice and guinea pigs. RESULTS: The rBCG-mV3 expressed detectable amounts of V3-concatamer when induced by single heat-shock. The recombinant BCG was genetically stable and maintained the introduced mV3 gene for several weeks. V3-specific antibodies were clearly detected 6 weeks after inoculation. The antibody titer rapidly increased after immunization up to 10 weeks, and then maintained for over 4 weeks. IgG2a was prevalent in the V3-specific antiserum. The recombinant BCG was also effective in inducing delayed-type hypersensitivity responses in the immunized guinea pigs. rBCG-immunized mice retained substantial amounts of V3-specific T cells in the spleen, even 5 months after the first immunization. CONCLUSION: Recombinant BCG-mV3 is very efficient in inducing humoral and long-lasting cell-mediated immunity against HIV-1 V3 in the immunized animals.