Interleukin-4 induced down-regulation of skin homing receptor expression by human viral-specific CD8 T cells may contribute to atopic risk of cutaneous infection.

Factors controlling the expression of cutaneous lymphocyte-associated antigen (CLA) by T cells are poorly understood, but data from murine and human CD4(+) T cell systems have suggested that cytokines play an important role. However, there are no data examining the influence of cytokines on the expression of CLA by human antigen-specific CD8(+) T cells. Peripheral blood mononuclear cells (PBMC) were isolated from 10 HLA-A*0201-positive healthy individuals. Using HLA-peptide tetrameric complexes refolded with immunodominant peptides from Epstein-Barr virus (EBV), cytomegalovirus (CMV) and influenza A virus, we investigated the temporal associations of CLA expression by viral-specific CD8(+) T cells following stimulation with antigen. Ex vivo influenza matrix-specific CD8(+) T cells expressed significantly (P < 0.05) greater levels of CLA than EBV BMLF1 and CMV pp65-specific CD8(+) T cells (mean 9.7% influenza matrix versus 1.4% BMLF1 versus 1.1% pp65) and these differences were sustained on culture. However, regardless of viral specificity, interleukin (IL)-12 and IL-4 induced significant (P < 0.05) dose-dependent up-regulation and down-regulation of CLA expression, respectively, with IL-4 showing a dominant negative effect. In many cases, IL-4 resulted in complete abrogation of detectable CLA expression by the viral-specific CD8(+) T cells. Overall these data demonstrate that CLA expression by human viral-specific CD8(+) T cells is highly dynamic and that IL-4 causes significant down-regulation. Disorders associated with a type 2 cytokine shift may reduce the efficiency of skin homing by viral-specific CD8(+) T cells. Furthermore, the ability to modify the local and systemic microenvironment may offer novel therapeutic strategies that influence tissue-specific T cell homing.

Direct measurement of CD8+ T cell responses in macaques infected with simian immunodeficiency virus.

The simian immunodeficiency virus (SIV) macaque model system has been used extensively to study AIDS pathogenesis and to test candidate vaccines for their ability to protect against homologous or heterologous challenge with pathogenic SIV or SHIV. Recent studies suggest that stimulation of HIV-1-specific CTL responses is important for effective vaccination against HIV-1. While quantitative measurements of SIV-specific cytotoxic T lymphocyte (CTL) responses have been facilitated by the use of tetrameric peptide complexes, this technique is currently limited to the study of Mamu-A*01-positive rhesus macaques. Furthermore, very few SIV-specific CTL epitopes have been identified, and there is limited identification of other MHC alleles in macaques. In this study, cytokine flow cytometry (CFC) was used to quantify SIV-specific CD8+ antigen-reactive T cells in macaques infected with SIV. We found a strong correlation (r = 0.96, P < 0.001) between CD8+ antigen-reactive T cells stained with the Mamu-A*01 p11C, C-M tetramer and production of intracellular TNF-alpha in the CFC assay. Furthermore, the CFC assay was used to identify a novel SIV-specific CTL epitope in Envelope (SIV Env, a.a. 486-494, sequence AEVAELYRL). The use of the CFC assay facilitates the study of antigen-reactive T cell responses in SIV infection and vaccination.

Simian immunodeficiency virus-specific cytotoxic T lymphocytes and protection against challenge in rhesus macaques immunized with a live attenuated simian immunodeficiency virus vaccine.

In this study, we examined the role of simian immunodeficiency virus (SIV)-specific cytotoxic T lymphocytes (CTLs) in macaques immunized with an attenuated strain of simian immunodeficiency virus (SIVmac239Deltanef) in protection against pathogenic challenge with SIVmac251. Our results indicate that attenuated SIVmac239Deltanef can elicit specific CTL precursor cells (CTLp), but no correlation was observed between breadth or strength of CTLp response to structural proteins SIV-Env, -Gamg or -Pol (as measured by limiting dilution assay) and protection against infection. In one animal, we longitudinally followed the SIV-Gag-specific response to an MHC class I Mamu-A*01-restricted epitope p11C, C-M using a tetrameric MHC/peptide complex reagent. A low frequency of SIV p11C, C-M peptide-specific tetramer-reactive cells was present at the time of challenge but could be expanded in vitro. Surprisingly, the low level of Mamu-A*01/p11C, C-M-specific CTLs induced through attenuated SIVmac239Deltanef vaccination increased in the absence of detectable SIVmac251 or SIVmac239Deltanef proviral DNA. Overall, our results suggest that protection against infection in this model can be achieved through more than one mechanism, with SIV-specific CTLs being important in controlling SIVmac239Deltanef viral replication postchallenge.

Effective induction of simian immunodeficiency virus-specific cytotoxic T lymphocytes in macaques by using a multiepitope gene and DNA prime-modified vaccinia virus Ankara boost vaccination regimen.

DNA and modified vaccinia virus Ankara (MVA) are vaccine vehicles suitable and safe for use in humans. Here, by using a multicytotoxic T-lymphocyte (CTL) epitope gene and a DNA prime-MVA boost vaccination regimen, high levels of CTLs specific for a single simian immunodeficiency virus (SIV) gag-derived epitope were elicited in rhesus macaques. These vaccine-induced CTLs were capable of killing SIV-infected cells in vitro. Fluorescence-activated cell sorter analysis using soluble tetrameric major histocompatibility complex-peptide complexes showed that the vaccinated animals had 1 to 5% circulating CD8(+) lymphocytes specific for the vaccine epitope, frequencies comparable to those in SIV-infected monkeys. Upon intrarectal challenge with pathogenic SIVmac251, no evidence for protection was observed in at least two of the three vaccinated animals. This study does not attempt to define correlates of protective immunity nor design a protective vaccine against immunodeficiency viruses, but it demonstrates clearly that the DNA prime-MVA boost regimen is an effective protocol for induction of CTLs in macaques. It also shows that powerful tools for studying the role of CTLs in the control of SIV and human immunodeficiency virus infections are now available: epitope-based vaccines, a protocol for an effective induction of CTLs in primates, and a simple and sensitive method for quantitation of epitope-specific T cells. The advantages of the DNA prime-MVA boost regimen as well as the correlations of tetramer staining of peripheral blood lymphocytes with CTL killing in vitro and postchallenge control of viremia are discussed.