Mitochondrial dysfunctions in circulating T lymphocytes from human immunodeficiency virus-1 carriers

In several models of lymphocyte apoptosis, two alterations of mitochondrial function precede advanced DNA fragmentation: (1) a reduction of mitochondrial transmembrane potential (ΔΨ(m)) and (2) an increase in mitochondrial generation of superoxide anion. Here we show that two fluorochromes allow for the identification of analogous mitochondrial perturbations in circulating T lymphocytes from human immunodeficiency virus (HIV)-1⁺ donors. The first among these fluorochromes, the cationic lipophilic dye DiOC₆(3), measures ΔΨ(m); the second marker, hydroethidine (HE), is nonfluorescent, unless it is oxidized by superoxide anions to the product athidium (Eth). CD4⁺ or CD8⁺ cells from clinically asymptomatic HIV-1 carriers contain a significantly elevated percentage of cells endowed with enhanced HE → Eth conversion and/or reduced DiOC₆(3) uptake as compared with normal controls. Phenotypic characterization of (HE → Eth)(high) cells from HIV⁺ donors shows that these cells possess a low ΔΨ(m), thus demonstrating a functional alteration of mitochondria. In addition, (HE → Eth)(high) cells display a reduced incorporation of the cardiolipin-specific dye nonyl-acridine orange (NAO), showing a structural defect of the cardiolipin-containing inner mitochondrial membrane. Control experiments involving rotenone, an inhibitor of the respiratory chain complex I, indicate that the reactive oxygen species responsible for HE → Eth conversion is generated during mitochondrial electron transport. In synthesis, it appears that mitochondrial alterations occur in a significant percentage of circulating T lymphocytes from HIV-1 carriers. The extent of ΔΨ(m) reduction, as determined ex vivo, correlates with the frequency of cella undergoing DNA fragmentation after overnight in vitro culture. These observations may be important for the understanding and for the direct ex vivo quantitation of HIV-triggered lymphocyte destruction.