Biology Reference
In-Depth Information
damages (Cossarizza et al., 1997). Signi®cant Dc alterations and a high ten-
dency to undergo spontaneous apoptosis were present in PBL from the sub-
jects studied, but the presence of NAC, NAM, or LAC was able to rescue most
cells from apoptosis. The changes in Dc and apoptosis observed in lymphocytes
collected in the earliest phases of the acute viral syndrome decreased sig-
ni®cantly after a few days, even in the absence of antiretroviral therapy. Sig-
ni®cant correlation was found between spontaneous apoptosis and TNF-a or
p24 plasma levels, as well as between apoptosis and the percentages of circu-
lating CD4
or CD8
T cells. It was suggested that NAC, NAM, or LAC
could rescue cells through a protective e¨ect on mitochondria, a well-known
target for the action of TNF-a and ROS, the production of which is strongly
induced by this cytokine.
In a second paper, we described the role of the CD95/CD95L system during
primary infection, analyzing the functional characteristics of lymphocytes in 12
patients during the acute phase of viral infection and in the following months
(Cossarizza et al., 2000). Almost all cells, including CD8
T cells with a virgin
phenotype, B lymphocytes, and natural killer (NK) cells, displayed CD95
molecules on the plasma membrane. Activation of CD95 on the surface of iso-
lated lymphocytes by anti-CD95 monoclonal antibodies or binding to CD95L
induced rapid apoptosis that was accompanied by relevant changes in Dc.We
showed that during acute, primary infection, a prolonged deregulation of the
CD95/CD95L system may exist, which provokes mitochondrial damages and
relevant apoptotic phenomena. Such deregulation is likely not entirely related
to virus production, but may contribute to the pathogenesis of the disease. It
was pointed out that mitochondria are involved in the complex balance existing
between proapoptotic events (increase in CD95 expression), likely triggered by
the host as a method to limit viral production, and antiapoptotic events (de-
crease in CD95L expression), likely triggered by the virus as a way to increase
its production and survival.
Concerning chronic HIV infection, we have focused our attention on a cel-
lular model of in vitro infection. In this chapter, we show some cyto¯uorimetric
studies on mitochondrial membrane potential and apoptosis in a cell line
chronically infected with HIV-1, U1, used for many years as a model for studies
on the physiopathology of infected cells (Gri½n et al., 1989; Kalebic et al.,
1991; Poli et al., 1994; Pomerantz et al., 1990). We have analyzed the capacity
of such cells to undergo apoptosis after treatment with TNF-a and cyclo-
hexamide (CHX), two classical agents used to induce cell death in the U937 cell
line (Cossarizza et al., 1995b).
DYES, CHEMICALS, AND CELL CULTURES
JC-1 and MitoTracker
TM
Red CMXRos (MT ) were from Molecular Probes
( Eugene, OR). JC-1 was dissolved in N,N-dimethylformamide ( DMF ) at a
concentration of 1 mg/mL and stored at ÿ20
C ; MT was dissolved in
dimethylsulfoxide (DMSO) at a concentration of 1 mM and stored at ÿ20
C.
