Frequency of provirus-bearing CD4+ cells in HIV type 1 infection correlates with extent of in vitro apoptosis of CD8+ but not of CD4+ cells.

Lymphocytes from HIV-1-infected subjects undergo massive apoptosis when cultured in vitro, and this phenomenon might reflect pathogenetic mechanisms leading to immune dysfunction in vivo. However, (1) lymphocyte death is not restricted to CD4+ cells but seems to involve predominantly CD8+ cells, and (2) the same phenomenon occurs in other viral infections. Furthermore, it is not known whether a relationship exists between the HIV-1 burden and this type of cell death. In this work we sought to determine whether the HIV-1 provirus load correlates with the propensity to apoptosis of CD4+ and CD8+ cells. We studied 10 HIV-1-infected patients with CD4+ cell counts above 500/mm3 and free of concomitant infections. We correlated the frequency of HIV-1-infected CD4+ cells with the extent of culture-induced apoptosis as well as with the phenotype of the apoptotic lymphocytes. We found that the magnitude of apoptosis correlated with the frequency of HIV-1-infected CD4+ cells (p = 0.0007), and that increasing viral load and apoptosis were associated with a shift to the selective death of CD8+ cells. Our data support the view that, in addition to CD4+ cell killing, another immunopathogenic effect of HIV might be that of priming CD8+ cells to apoptosis. In vivo, this could eventually lead to the exhaustion of the cytotoxic T cell compartment.

Detection and characterization of apoptotic peripheral blood lymphocytes in human immunodeficiency virus infection and cancer chemotherapy by a novel flow immunocytometric method.

We have developed a quantitative and sensitive flow cytometric method for the detection of human apoptotic lymphocytes that, unlike previously described assays, allows their identification in mixed populations of peripheral blood leukocytes as well as their immunophenotyping. Apoptotic lymphocytes are identified on the basis of peculiar light scatter changes, reflecting their smaller size and their modified nucleus/cytoplasm organization, and of the decreased expression of surface CD45 molecules. Based on these criteria, apoptotic lymphocytes generated by exposure to ionizing radiation can be easily distinguished from viable cells and from necrotic lymphocytes generated by treatment with antibody and complement. Using this assay, we reappraised the phenomenon of the in vitro apoptosis of lymphocytes from patients with human immunodeficiency virus (HIV) infection. Lymphocytes from HIV patients, unlike those from normal HIV-negative subjects, undergo apoptosis upon simple in vitro culture. We found that the percentages of lymphocytes undergoing apoptosis were significantly higher in patients with low CD4 cell counts (< 400/microL) than in patients at earlier stages (> 400 CD4 cells/microL). However, phenotypic analysis disclosed that apoptotic lymphocytes generated in these cultures were mostly CD8+ T cells and CD19+ B cells. Thus, in contrast to what has been previously suggested, the phenomenon of in vitro lymphocyte apoptosis might not be pathogenetically related to the depletion of CD4+ T cells in acquired immunodeficiency syndrome. Nevertheless, it might represent an useful marker of disease progression. Our assay allows the analysis of unfractionated peripheral blood leukocytes and thus the identification of apoptotic lymphocytes circulating in vivo. Apoptotic lymphocytes could indeed be detected in the circulation of a patient with cancer shortly after high-dose cytotoxic chemotherapy. By contrast, no apoptotic lymphocytes could be detected in vivo in patients with early or advanced HIV infection.

Absence of antibodies to human herpesvirus-6 in patients with slowly-progressive human immunodeficiency virus type 1 infection.

To evaluate a possible role for Human Herpesvirus-type 6 (HHV-6) coinfection as a co-factor in the progression of HIV-1 disease, we investigated the prevalence of seropositivity for HHV-6 in a cohort of HIV-1 infected patients. These patients were retrospectively divided into two groups according to the decline of CD4+ T cells during the follow up: 11 were classified as rapid decliners (less than 400 CD4+/cmm within 1 year), and 38 as slow decliners (greater than 400 CD4+/cmm after at least 4 years' follow up). HHV-6 antibodies were detected by a commercial immunofluorescence assay and by a Western blotting assay developed in our laboratory. Our results show that Western blot appears to provide results satisfactorily free of false positivities. We found that the frequency of HHV-6 seropositivity was significantly lower in the group of slow decliners, compared both to rapid decliners and to the general population. These data suggest a role for HHV-6 co-infection in the progression of HIV-1 disease.