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Detection and characterization of apoptotic peripheral blood lymphocytes in
human immunodeficiency virus infection and cancer chemotherapy by a novel
flow immunocytometric method
M Carbonari, M Cibati, M Cherchi, D Sbarigia, AM Pesce, L Dell'Anna, A Modica and M Fiorilli
Department of Clinical Immunology, University of Rome La Sapienza, Italy.
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.
Volume 83,
Issue 5,
pp. 1268-1277,
03/01/1994
Copyright © 1994 by The American Society of Hematology
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