CD49a Identifies Polyfunctional Memory CD8 T Cell Subsets that Persist in the Lungs After Influenza Infection.

CD8 T cell memory offers critical antiviral protection, even in the absence of neutralizing antibodies. The paradigm is that CD8 T cell memory within the lung tissue consists of a mix of circulating TEM cells and non-circulating TRM cells. However, based on our analysis, the heterogeneity within the tissue is much higher, identifying TCM, TEM, TRM, and a multitude of populations which do not perfectly fit these classifications. Further interrogation of the populations shows that TRM cells that express CD49a, both with and without CD103, have increased and diverse effector potential compared with CD49a negative populations. These populations function as a one-man band, displaying antiviral activity, chemokine production, release of GM-CSF, and the ability to kill specific targets in vitro with delayed kinetics compared with effector CD8 T cells. Together, this study establishes that CD49a defines multiple polyfunctional CD8 memory subsets after clearance of influenza infection, which act to eliminate virus in the absence of direct killing, recruit and mature innate immune cells, and destroy infected cells if the virus persists.

Electron transport complex I is required for CD8+ T cell function.

After Ag encounter, CD8+ T cells become activated and begin to proliferate. Early during infection, when Ag-specific effector CD8+ T cells are proliferating, producing cytokines, and lysing infected cells in vivo, their mitochondrial potential is increased. The purpose of the experiments presented here was to determine whether mitochondrial function was required for CD8+ T cell function. To block mitochondrial function, transgenic CD8+ T cells were incubated with increasing doses of rotenone, an inhibitor of electron transport complex I. Within minutes of T cell activation, rotenone incubation decreased the production of H(2)O(2), calcium flux, and ERK1/2 phosphorylation. Failure to undergo signal transduction resulted in a decrease in T cell division initiated by peptide-coated cells, CD3/CD28 Abs, and PMA/ionomycin stimulation. Decreased function following rotenone incubation was not restricted to naive cells, as effector and memory CD8+ T cells isolated directly ex vivo from lymphocytic choriomeningitis virus-infected mice displayed decreased production of IFN-gamma and TNF-alpha production after peptide stimulation. Furthermore, incubation with rotenone decreased degranulation of effector and memory cells, a critical step in the cytolysis of infected cells. These data suggest that electron transport complex I is required for CD8+ T cell signal transduction, proliferation, cytokine production, and degranulation.

Partial reconstitution of virus-specific memory CD8+ T cells following whole body gamma-irradiation.

CD8+ memory T cells are critical in providing immunity to viral infection. Previous studies documented that antigen-specific CD8+ memory T cells are more resistant to radiation-induced apoptosis than naive T cells. Here, we determined the number and in vivo function of memory CD8+ T cells as immune reconstitution progressed following irradiation. Immediately following irradiation, the number of memory CD8+ T cells declined 80%. As reconstitution progressed, the number of memory cells reached a zenith at 33% of pre-irradiation levels, and was maintained for 120 days post-irradiation. In vitro, memory CD8+ T cells were able to produce cytokines at all times post-irradiation, but when adoptively transferred, they were not able to expand upon rechallenge immediately following irradiation, but regained this ability as reconstitution progressed. When proliferation was examined in vitro, irradiated memory CD8+ T cells were able to respond to mitogenic growth but were unable to divide.

Antioxidant treatment reduces expansion and contraction of antigen-specific CD8+ T cells during primary but not secondary viral infection.

During many viral infections, antigen-specific CD8(+) T cells undergo large-scale expansion. After viral clearance, the vast majority of effector CD8(+) T cells undergo apoptosis. Previous studies have implicated reactive oxygen intermediates (ROI) in lymphocyte apoptosis. The purpose of the experiments presented here was to determine the role of ROI in the expansion and contraction of CD8(+) T cells in vivo during a physiological response such as viral infection. Mice were infected with lymphocytic choriomeningitis virus (LCMV) and treated with Mn(III)tetrakis(4-benzoic acid)porphyrin chloride (MnTBAP), a metalloporphyrin-mimetic compound with superoxide dismutase activity, from days 0 to 8 postinfection. At the peak of CD8(+)-T-cell response, on day 8 postinfection, the numbers of antigen-specific cells were 10-fold lower in MnTBAP-treated mice than in control mice. From days 8 to 30, a contraction phase ensued where the numbers of antigen-specific CD8(+) T cells declined 25-fold in vehicle-treated mice compared to a 3.5-fold decrease in MnTBAP-treated mice. Differences in contraction appeared to be due to greater proliferation in drug-treated mice. By day 38, the numbers of antigen-specific CD8(+) memory T cells were equivalent for the two groups. The administration of MnTBAP during secondary viral infection had no effect on the expansion of antigen-specific CD8(+) secondary effector T cells. These data suggest that ROI production is critical for the massive expansion and contraction of antigen-specific CD8(+) T cells during primary, but not secondary, viral infection.

Mitochondrial potential and reactive oxygen intermediates in antigen-specific CD8+ T cells during viral infection.

Following many viral infections, there are large expansions of Ag-specific CD8(+) T cells. After viral clearance, mechanisms exist to ensure that the vast majority of effector cells undergo apoptosis. In studies of thymocyte apoptosis, loss of mitochondrial potential (deltapsi(m)) and excess production of reactive oxygen intermediates have been implicated as key events in cellular apoptosis. The purpose of the experiments presented in this work was to determine these parameters in Ag-specific CD8(+) T cells during a physiological response such as viral infection. Using lymphocytic choriomeningitis virus infection of mice, we found that Ag-specific CD8(+) effector T cells that had undergone recent TCR stimulation had an increased deltapsi(m). These cells also had increased levels of superoxide. As these cells progressed through the contraction of the immune response, their potential decreased, but superoxide levels remained similar to naive cells. One of the consequences of reduced mitochondrial potential is membrane permeability and subsequent caspase activation. We examined both the enzymatic activity and levels of cleaved caspase 3, an effector caspase, and could only detect increased levels in Ag-specific CD8(+) T cells on day 5 postinfection, a time point in which virus was still present. This contrasts with Ag-specific effector cells examined during the contraction phase that had no detectable caspase activity directly ex vivo. These data suggest that the apoptotic program begins earlier than previously expected on day 5, during the expansion phase.