Lymphoid tissue immunopathology is a characteristic feature of chronic HIV/SIV infection in AIDS-susceptible species, but is absent in SIV-infected natural hosts. To investigate factors contributing ...to this difference, we compared germinal center development and SIV RNA distribution in peripheral lymph nodes during primary SIV infection of the natural host sooty mangabey and the non-natural host pig-tailed macaque. Although SIV-infected cells were detected in the lymph node of both species at two weeks post infection, they were confined to the lymph node paracortex in immune-competent mangabeys but were seen in both the paracortex and the germinal center of SIV-infected macaques. By six weeks post infection, SIV-infected cells were no longer detected in the lymph node of sooty mangabeys. The difference in localization and rate of disappearance of SIV-infected cells between the two species was associated with trapping of cell-free virus on follicular dendritic cells and higher numbers of germinal center CD4+ T lymphocytes in macaques post SIV infection. Our data suggests that fundamental differences in the germinal center microenvironment prevent productive SIV infection within the lymph node germinal centers of natural hosts contributing to sustained immune competency.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Standard proliferation assays used for analysis of CD4
+ T cell function have significant shortcomings, including limited sensitivity, lack of truly quantitative readouts and significant variability. ...We have optimized an intracellular cytokine staining (ICS) assay in rhesus macaques which allows us to identify virus-specific CD4
+ T cells at the single-cell level with high sensitivity while reducing background staining to a minimum. A variety of parameters were tested to determine the optimal experimental conditions necessary for the detection of antigen-specific CD4
+ T cells in macaques. Central to our optimized protocol was the addition of cross-linked costimulatory anti-CD28 and anti-CD49d Mabs, a modification which resulted in up to threefold enhancement of the frequency of TNF-α-secreting CD4
+ T cells following superantigen- or antigen-specific stimulation. The ICS protocol was also optimized with respect to antigen concentration and duration of antigenic stimulation. These modifications resulted in a convenient and highly reproducible assay with intra- and inter-assay variability of less than 10%. Although cryopreservation of PBMC generally led to a 40% to 80% decrease in the frequency of antigen-specific CD4
+ T cells detected by ICS using stimulation with viral proteins, the use of overlapping peptide pools minimized the effects of cryopreservation on ICS responses. The use of more sensitive techniques such as ICS permits delineation of antigen-specific cells at the single cell level and should provide new insights into pathogen-specific immune responses in the rhesus macaque model.
Retroviral restriction factor TRIM5alpha exhibits a high degree of sequence variation among primate species. It has been proposed that this diversity is the cumulative result of ancient, ...lineage-specific episodes of positive selection. Here, we describe the contribution of within-species variation to the evolution of TRIM5alpha. Sampling within two geographically distinct Old World monkey species revealed extensive polymorphism, including individual polymorphisms that predate speciation (shared polymorphism). In some instances, alleles were more closely related to orthologues of other species than to one another. Both silent and nonsynonymous changes clustered in two domains. Functional assays revealed consequences of polymorphism, including differential restriction of a small panel of retroviruses by very similar alleles. Together, these features indicate that the primate TRIM5alpha locus has evolved under balancing selection. Except for the MHC there are few, if any, examples of long-term balancing selection in primates. Our results suggest a complex evolutionary scenario, in which fixation of lineage-specific adaptations is superimposed on a subset of critical polymorphisms that predate speciation events and have been maintained by balancing selection for millions of years.
Although CTL escape has been well documented in pathogenic simian immunodeficiency virus (SIV) infection, there is no information on CTL escape in nonpathogenic SIV infection in nonhuman primate ...hosts like the sooty mangabeys. CTL responses and sequence variation in the SIV nef gene were evaluated in one sooty mangabey and one rhesus macaque inoculated together with the same stock of cloned SIVmac239. Each animal developed an immunodominant response to a distinct CTL epitope in Nef, aa 157–167 in the macaque and aa 20–28 in the mangabey. Nonsynonymous mutations in their respective epitopes were observed in both animals and resulted in loss of CTL recognition. These mutations were present in the majority of proviral DNA sequences at 16 weeks post infection in the macaque and >2 years post infection in the mangabey. These results document the occurrence of CTL escape in a host that does not develop AIDS, and adds to the growing body of evidence that CTL exert significant selective pressure in SIV infection.
The characterization of antiviral CTL responses has largely been limited to assessing Ag-specific immune responses in the peripheral blood. Consequently, there is an incomplete understanding of the ...cellular immune responses at mucosal sites where many viruses enter and initially replicate and how the Ag specificity and activation status of CTL derived from these mucosal sites may differ from that of blood-derived CTL. In this study, we show that EBV-specific CTL responses in the tonsils are of comparable specificity and breadth but of a significantly higher magnitude compared with responses in the peripheral blood. EBV-specific, tonsil-resident, but not PBMC-derived, T cells expressed the integrin/activation marker CD103 (alphaEbeta7), consistent with the detection of its ligand, E-cadherin, on tonsillar squamous cells. These CD8-positive, CD103-positive, tonsil-derived CTL were largely CCR7- and CD45RA- negative effector-memory cells and responded to lower Ag concentrations in in vitro assays than their CD103-negative PBMC-derived counterparts. Thus, EBV-specific CTL in the tonsil, a crucial site for EBV entry and replication, are of greater magnitude and phenotypically distinct from CTL in the peripheral blood and may be important for effective control of this orally transmitted virus.
PDF
