Molecular approaches to diagnostic and clinical problems in AIDS patients
Description
Human immunodeficiency virus (HIV) infection causes a decrease in CD4 cell number and an increase in CD8 cell count. These cell numbers are used to assess disease progression in HIV-infected patients. In children, frequent invasive blood draws are undesirable. We therefore developed the technique of quantitative competitive reverse transcriptase polymerase chain reaction (QC-RT-PCR) to quantitate CD4 and CD8 mRNA levels from heelsticks or fingerpricks. This technique is extremely sensitive and can be applied to frozen samples. We found no correlation between CD4 and CD8 cell counts and mRNA levels of CD4 and CD8 as determined by QC-RT-PCR CD8+ cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are the major cytotoxic components of the antiviral immune response. The major pathway used by these cells in the antiviral response includes perforin and granzymes. We assessed the mRNA levels of granzyme B in peripheral blood mononuclear cells of IRV-infected versus noninfected individuals by QC-RT-PCR. There were significantly fewer HIV patients with detectable granzyme B levels than controls. This implies that cytotoxicity may be impaired due a deficient quantity of cytotoxic granules The in utero HIV transmission rate should theoretically be much higher than currently estimated. Protective mechanisms may exist to prevent in utero transmission. Immunoglobulin G (IgG) crosses the placenta, and neutralizing antiviral antibody may prevent in utero transmission. To study this hypothesis, the quantity and quality of the antiviral IgG in amniotic fluid of infected mother-infant pairs was assessed. Using p24 antigen capture ELISA, detectable levels of HIV p24 antigen were found in the amniotic fluid of infected mothers. By western blot, significantly high levels of antiviral IgG were found in amniotic fluid samples versus plasma. Infectivity reduction assays were performed, and extremely dilute human and macaque amniotic fluids were able to neutralize their respective viruses' infectivity, except for the samples from an SIV-positive mother whose infant was infected in utero. These results show that HIV-1 and SIV neutralizing antibodies may play a crucial role in protecting the fetus from in utero infection