SDF-1alpha is a potent inducer of HIV-1-Specific CD8+ T-cell chemotaxis, but migration of CD8+ T cells is impaired at high viral loads.

Multiple impairments in HIV-1-specific cytotoxic T cells (CTL) have been reported, but derangements in HIV-1-specific CD8+ T-cell chemotaxis have not been described previously. We assessed migration to SDF-1alpha (stromal cell-derived factor 1-alpha) and CX3CL1 in vitro and expression of cognate receptors, CXCR4 and CX3CR1, by flow cytometry in peripheral blood and lymph node CD8+ T cells from HIV-1-seropositive and -seronegative individuals. Compared with seronegative individuals, percentages of CXCR4+CD8+ T cells were reduced (median, 26% versus 74%, p < 0.001) and percentages of CX3CR1+CD8+ T cells were increased (median, 33% versus 15%, p = 0.03) in seropositive individuals. Robust migration of peripheral blood mononuclear cell (PBMC) CD8+ T cells to SDF-1alpha (1 alphag/ml) was observed in both HIV-1-seropositive (median chemotactic index [CI] 4.9) and -seronegative (median CI 2.8) subjects (p = 0.46). CI to SDF-1alpha was not significantly related to percentage of CXCR4+CD8+ T cells or density of CXCR4, but correlated inversely with plasma HIV-1 RNA concentration (r = -0.82, p = 0.03). Little chemotaxis was observed in response to CX3CL1 and it was unrelated to CX3CR1 expression. Lymph node CD8+ T-cell chemotaxis to SDF-1alpha and CX3CL1 in four subjects demonstrated the same patterns observed in PBMC. HIV-1-specific tetramer-staining CD8+ T cells exhibited chemotaxis of similar magnitude as PBMC CD8+ T cells in a subset of subjects. These data suggest that SDF-1alpha is a potent chemoattractant for HIV-1-specific CTL, but that impairments in migration of HIV-1-specific CTL may exist at high viral loads. Improved understanding of the determinants of CTL localization may provide insight into novel therapies to enhance delivery of CTL to sites of HIV-1 replication.

Effect of cryopreservation on measurement of cytomegalovirus-specific cellular immune responses in HIV-infected patients.

To determine the feasibility of cytomegalovirus (CMV)-specific cell-mediated immunity (CMI) studies using cryopreserved cells, we compared lymphocyte proliferation assays (LPA), responder cell frequency (RCF), interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) production using fresh and cryopreserved peripheral blood mononuclear cells (PBMCs) from 53 HIV-infected patients and 15 uninfected controls. Qualitative CMV LPA results were concordant in >/=84% of the specimens from either HIV-infected patients or controls. Proliferation-based RCF, IL-2, and IFN-gamma comparisons showed that cryopreservation reduces the number of CMV-specific responders and decreases cytokine secretion, without changing the rank order of the results (p <.01). In contrast, the number of flow cytometry-enumerated IFN-gamma-producing CD4+ cells was not significantly changed by cryopreservation. In HIV-infected patients, the differences between fresh and frozen cell assays were not influenced by CD4 cell numbers or HIV viral load. These data indicate that cryopreserved cells are suitable for longitudinal studies of the CMV-specific immune response in HIV-infected patients and uninfected controls.

Morphology and mobility of oyster hemocytes: evidence for seasonal variations.

Hemocytes of Crassostrea virginica were video recorded and tracked to determine their locomotive rates and to assign these rates to Wright-stained morphological variants. From 24 oysters examined in January, February, March, and May, 1571 hemocytes were video recorded, identified, and their rate of locomotion (ROL) measured. Granulocytes (three types) and agranulocytes (one lymphoid and three nonlymphoid types) were recognized. Focusing on 15 oysters in March and May, 20,318 hemocytes were counted from duplicate slides to verify the classification and to show that predominant hemocytes vary greatly between samples and among individual oysters, yet population differences can be detected. Measured rates of locomotion indicate that the granulocyte subpopulation moved significantly faster (3.3 microns/min) than the agranulocyte subpopulation (0.7 microns/min) because most (81%) agranulocytes were not mobile. Of the mobile hemocytes, granulocytes were also significantly faster (4.8 microns/min vs. 3.5 microns/min, P less than 0.0001), and basophilic granulocytes (BASOs) were the most active and abundant cell type. Examination of monthly percentages of cells and ROL indicates, however, that granulocyte dominance and ROL are not invariable. Granulocyte percentages of more than 60% in January, February, and March decreased to 32% in May, and BASO dominance was reduced to 15%. Further, percentages of mobile granulocytes decreased from greater than 65% in January, February, and March to 50% in May. ROL for all cells decreased from greater than 2.3 microns/min in these months to 1.0 microns/min in May. The fewer mobile hemocytes tracked in May had significantly (P less than .05) lower average ROL (4.0 microns/min) than those in January and March (4.7 microns/min each). Agranulocytes increased in May due to an increase in nonlymphoid cells.