Cervicovaginal lamina propria lymphocytes: phenotypic characterization and their importance in cytotoxic T-lymphocyte responses to simian immunodeficiency virus SIVmac251

Abstract

Most human immunodeficiency virus (HIV) type 1 infections occur by the mucosal route. Thus, it is important to assess the immune responses to HIV in the vaginal, cervical, and rectal compartments. Here we quantitated the virus-specific CD8+ T-cell response and characterized the phenotype of lymphocytes in the genital tracts of naive macaques, macaques acutely or chronically infected with simian immunodeficiency virus SIVmac251, and macaques chronically infected with chimeric simian/human immunodeficiency virus SHIV(KU2.) Vaginal biopsy samples or samples obtained at the time of euthanasia were used in this analysis. The percentage of Gag-specific, tetramer-positive T cells was as high as 13 to 14% of the CD3+ CD8+ T-cell population in the vaginal and cervical laminae propriae of both SIVmac251 and SHIV(KU2) chronically infected macaques. In most cases, the frequency of this response in the cervicovaginal compartment far exceeded the frequency in the blood or the draining iliac lymph node. Vaginal laminae propriae of naive macaques contained 55 to 65% CD3+ CD8+ cells and 28 to 34% CD3+ CD4+ cells, while the majority of intraepithelial cells were CD8+ T cells (75 to 85%). For the same cells, the surface expression of CD62L was low whereas that of alphaEbeta7 was high. No difference in the expression of CD45RA on CD8+ T cells was observed in the chronic stage of SIVmac251 infection. Although no decrease in the percentage of CD4+ cells in the genital tract was observed within the first 12 days of infection, by 6 weeks from SIVmac251 infection and thereafter the percentage of CD4+ T cells was decreased in the laminae propriae of the vagina and cervix. Expression of CD45RA did not differ in naive and acutely SIVmac251 infected macaques. Information on the quality and quantity of local immune responses may help in the design of vaccine strategies aimed at containing viral replication at the site of viral encounter.

FIG. 1.

Enumeration of T cells in vagina laminae propriae of naive and SIV_mac251-infected macaques. Shown are percentages of CD3⁺ CD4⁺ and CD3⁺ CD8⁺ cells in vaginal laminae propriae of naïve macaques (a) and of SIV_mac251-infected macaques during primary (b) or chronic (c) infection. Cells were gated through the CD3⁺ population (4 × 10⁴ cells were acquired through this gate for each sample analyzed), and quadrant analysis for CD4-PE- and CD8-PerCP-positive cells was used to generate the percentages shown in the histograms. LP, lamina propria.

FIG. 2.

CD3⁺ CD8⁺ CD45RA⁺ frequency in the vaginal tissues of naive and chronically infected macaques. Shown are the percentages of CD3⁺ CD8⁺ cells of the vaginal lamina propria that express the CD45RA marker in naive (b) or chronically infected (c) macaques. Tricolor staining for CD3-FITC, CD8-PerCP, and CD45RA-HRD was used to generate these data. Cells were gated through the CD3⁺ population (4 × 10⁴ cells were acquired through this gate for each sample analyzed) and then analyzed for expression of CD8 and CD45RA using quadrant analysis (a).

FIG. 3.

CD8⁺ CD103⁺ T cells in the cervicovaginal and rectal mucosae of naive macaques. Bar graphs show CD103 (αEβ7) expression in intraepithelial and lamina propria CD8⁺ T lymphocytes of vaginal (a) and rectal (b) tissues of naive macaques. As shown in the top two panels, two-color staining was done for CD8 and CD103. Cells were gated on the CD8⁺ population (10⁴ cells were acquired through this gate), and expression of CD103 on this population was determined by histogram analysis as shown.

FIG. 4.

Staining of control tissues with the Gag181 tetramer. Vaginal biopsy specimens obtained from naive Mamu-A*0-positive macaques and from infected Mamu-A*01-negative macaques were used as negative controls for the Gag181 tetramer staining.

FIG. 5.

Frequency of Gag181 tetramer-specific CD8⁺ T cells in the systemic and genital compartments of chronically SIV_mac251 infected macaques. The percentage in the upper right portion of each panel represents tetramer-positive CD3⁺ CD8⁺ T cells. Cells were gated through the CD3⁺ population, and 10⁴ CD3⁺ CD8⁺ cells were acquired for analysis of each sample where possible. Staining was done on lymphocytes from the blood, vaginal and cervical laminae propriae (LP), and iliac lymph nodes (LN), as indicated.

FIG. 6.

Frequency of Gag181 tetramer-positive CD3⁺ CD8⁺ T cells in tissues of chronically SHIV_KU2 infected macaques. Flow charts show the proportions of Gag181 tetramer-positive lymphocytes isolated from the blood, iliac lymph nodes, and vaginal laminae propriae of SHIV_KU2-infected macaques. Cells were analyzed as described in the legend to Fig. 5 and in Materials and Methods.