Biology Reference
In-Depth Information
Recognition of Lymphocytic Markers
HIV-1 disease pathogenesis has been characterized by marked alterations in the
physiology of lymphocytic cells. These alterations are either a consequence of
the massive immune responses that are induced by the chronic viral infection
or direct e¨ects of viral products on cellular proteins. The continuous presence
of virus constitutively stimulates the cells of the immune system and thereby
results in phenotypic changes in the cells. For instance, CD8
T lymphocytes
express enhanced cell-surface levels of the T-cell activation markers CD38 and
CD95. Conversely, it has been reported that circulating CD8
T cells have de-
creased levels of CD3z and CD28 (Trimble et al., 2000). Intracellular molecules
also ¯uctuate during HIV-1 infection. For instance, levels of bcl-2 and thio-
redoxin in monocytes are decreased in HIV-1 seropositive individuals with
CD4
T cell counts above 500 ml/microliter. However, in patients with low
levels of CD4
T cells, monocytic bcl-2 is expressed in amounts commensurate
with monocytes from uninfected persons and thioredoxin levels in these cells
are elevated (Elbim et al., 1999).
Besides the widespread consequence of the presence of large amounts of
foreign molecules, cellular infection with the virus also has marked, direct
e¨ects on host protein expression. For instance, HIV-1 Tat has been shown to
increase metalloproteinase production ( Lafrenie et al., 1996), to increase CXC-
chemokine receptor 4 expression (Secchiero et al., 1999), to induce CD69 sur-
face membrane display ( Blazquez et al., 1999), to enhance the level of a hema-
topoietic cell-speci®c transcription factor and down-regulate the chemokine
MIP-1a (Sharma et al., 1996), to increase Bcl-2 expression (Wang et al., 1999),
and to up-regulate IL-4 receptors (Husain et al., 1996). Both HIV-1 Nef and
Vpu inhibit the surface expression of CD4 and MHC class I molecules (Aiken
et al., 1994; Collins et al., 1998; Kerkau et al., 1997; Schubert et al., 1998;
Schwartz et al., 1996). Nef also inhibits CD25 and c-myb (Greenway et al.,
1995). HIV-1 Vpr acts to induce I-kB and Bcl-2 while inhibiting the expres-
sion of Bax (Ayyavoo et al., 1997; Conti et al., 1998). Besides these de®nitive
changes in the expression of cellular proteins attributable to speci®c viral pro-
teins, there are many other changes that have been demonstrated that have not
been clearly assigned to speci®c viral gene product. HIV-1 infection causes sig-
ni®cant, widespread changes in the expression of cellular proteins, and these
changes are important in HIV-1 disease pathogenesis.
The analysis of phenotypic changes in lymphocytes and other blood cells
from patients with HIV-1 disease have been necessarily limited to the detection
levels of the available technologies. Flow cytometric analysis is particularly
useful for analyzing these changes because it provides a cell-by-cell assessment.
Because these changes are often seen in subsets of the total cells, a cell-by-cell
analysis, which can also provide a more extended phenotypic analysis, is desir-
able. EAS is able to provide ¯ow cytometry with a considerably greater sensi-
tivity and thereby greater resolving power. Consequently, enzymatic ampli®ca-
