Sequential analysis of apoptosis induction in peripheral blood mononuclear cells and lymph nodes in the early phase of pathogenic and nonpathogenic SIVmac infection.

To investigate the role of apoptosis in the early phase of HIV infection, we used macaques infected with simian immunodeficiency virus strain mac (SIVmac) as a primate model and examined sequentially the characteristics of apoptosis of lymphocytes in peripheral blood mononuclear cells (PBMCs) and lymph nodes in the early phase of SIVmac infection. Five macaques infected with a pathogenic strain of SIV, SIVmac239, were analyzed during the first 4 weeks after infection. Peripheral CD4+ and CD8+ cells transiently decreased at 1 week postinfection. The percentage of apoptotic cells in cultured PBMCs increased from about 2 weeks postinfection. The number of apoptotic cells in lymph node sections was higher on days 13 and 28 postinfection than before infection and on day 5 postinfection. Fas antigen expression on peripheral lymphocytes was upregulated from day 8 postinfection. These results indicate that apoptosis is induced about 2 weeks after SIVmac239 infection, following the upregulation of Fas antigen expression on lymphocytes. Since apoptosis was induced about 1 week after the decrease in peripheral CD4+ and CD8+ cell counts, it appears that the apoptosis induction does not play an important role in the transient lymphopenia in the early phase of SIVmac infection. In macaques infected with a nonpathogenic derivative of SIVmac239, SIVmac delta nef, apoptosis of lymphocytes was induced as it was in SIVmac239-infected macaques, but to a lesser degree, suggesting a correlation between the extent of apoptosis induction in lymphocytes in the early phase of SIVmac infection and the pathogenicity of SIVmac.

Analysis of evolutionary conservation in CD1d molecules among primates.

The hereditary conservation in the genetically encoded CD1D sequences of various primates was analyzed. Genomic CD1D sequences of 17 rhesus macaques with distinct origins, eight Indian and nine Chinese, were examined and differences of only one or two nucleotides were detected and the consensus sequence of rhesus CD1D was determined. CD1D consensus sequences of three African green monkeys (AGMs) and the rhesus monkeys were then compared to study the evolutionary differences among interspecies. The CD1D consensus sequence determined from AGMs apparently differed by seven nucleotides from the rhesus consensus sequence, and nucleotide difference induced only three amino acid changes within Exon3, corresponding to the alpha2 domain of CD1d having a hydrophobic ligand-binding pocket. Such changes in the alpha2 domain may alter the characteristics of the SIV-derived glycolipid/lipid antigens presented by each CD1d molecule to innate natural killer T cells. In addition, the CD1D genomic sequences of three chimpanzees (chimps) were determined. To our surprise, although Exon2 and Exon3 reflecting antigen-binding alpha1 and alpha2 domains in chimps' CD1D were identical to that in humans except one amino acid, three amino acids within Exon4, reflecting alpha3 domain, were distinct from humans, and one of them was identical to those in rhesus and AGM CD1D. On the basis of the findings, the evolutionary relationship of the CD1d molecules among the various primates and their HIV-1/SIV susceptibility will be discussed.

DNA vaccination of macaques by a full genome HIV-1 plasmid which produces noninfectious virus particles.

In this study, we tried a DNA vaccination regime in rhesus macaques using a full genome HIV-1 plasmid. The HIV-1 genome is under the control of its original LTR promoter, but has a mutated zinc finger motif gene in the nucleocapsid region. Due to the lack of genomic RNA packaging, the plasmid produces only noninfectious viral particles. We repeatedly injected four macaque monkeys intramuscularly with the naked DNA over a period of 40 weeks. To evaluate the humoral and cell-mediated immunity provided by this DNA vaccination, no other booster or other recombinant viral vectors were used. Immunological responses against HIV-1 were elicited in all of the vaccinated monkeys: stable anti-HIV-1 Env antibodies were raised in two monkeys and CTL activities were induced in the other monkeys. The macaques were intravenously challenged at 54 weeks with 100 TCID(50) of SHIV-NM-3rN, which possesses an envelope gene homologous to the one in the vaccinated plasmid. In all of the vaccinated macaques, the peak plasma viral loads induced by the challenge virus were two to three orders of magnitude lower than those of the naive controls. These results suggest that a DNA vaccination regime with a full genome plasmid alone is potentially efficacious and provides a new possibility for the development of an AIDS vaccine.