Depot Medroxyprogesterone Acetate Use Is Associated With Elevated Innate Immune Effector Molecules in Cervicovaginal Secretions of HIV-1-Uninfected Women.

OBJECTIVE: The effects of sex hormones on the immune defenses of the female genital mucosa and its susceptibility to infections are poorly understood. The injectable hormonal contraceptive depot medroxyprogesterone acetate (DMPA) may increase the risk for HIV-1 acquisition. We assessed the local concentration in the female genital mucosa of cationic polypeptides with reported antiviral activity in relation to DMPA use. METHODS: HIV-1-uninfected women were recruited from among couples testing for HIV in Nairobi, Kenya. Cervicovaginal secretion samples were collected, and the concentrations of HNP1-3, LL-37, lactoferrin, HBD-2, and SLPI were measured by enzyme-linked immunosorbent assays. Levels of cationic polypeptides in cervicovaginal secretions were compared between women who were not using hormonal contraception and those using DMPA, oral, or implantable contraception. RESULTS: Among 228 women, 165 (72%) reported not using hormonal contraception at enrollment, 41 (18%) used DMPA, 16 (7%) used an oral contraceptive, and 6 (3%) used a contraceptive implant. Compared with nonusers of hormonal contraception, DMPA users had significantly higher mean levels of HNP1-3 (2.38 vs. 2.04 log10 ng/mL; P = 0.024), LL-37 (0.81 vs. 0.40 log10 ng/mL; P = 0.027), and lactoferrin (3.03 vs. 2.60 log10 ng/mL; P = 0.002), whereas SLPI and HBD-2 were similar. CONCLUSIONS: Although all analyzed cationic polypeptides have intrinsic antiviral capacity, their interaction and cumulative effect on female genital mucosa susceptibility to infections in vivo has yet to be unraveled. This study suggests a potential mechanism underlying the effect of DMPA on the innate immune defenses, providing a rationale to investigate its effect on HIV-1 acquisition risk.

HIV Infection in Uncircumcised Men Is Associated With Altered CD8 T-cell Function But Normal CD4 T-cell Numbers in the Foreskin.

BACKGROUND: Human immunodeficiency virus (HIV)-infected (HIV+) men are more susceptible to sexually transmitted infections, and may be superinfected by HIV. We hypothesized that HIV induces immune alterations in the foreskin that may impact the subsequent acquisition/clearance of genital coinfections. METHODS: Foreskin tissue and blood were obtained from 70 HIV-uninfected and 20 HIV+ men undergoing circumcision. T cells were characterized by flow cytometry, immunohistochemistry, and polymerase chain reaction. RESULTS: There was substantial influx of CD8 T-cells into the foreskins of HIV+ men (108.8 vs 23.1 cells/mm(2); P < .001); but foreskin CD4 T-cell density was unchanged (43.0 vs 33.7/mm(2); P = .67), despite substantial blood depletion (409.0 vs 877.8 cells/µL; P < .001). While frequencies of foreskin C-C chemokine receptor type 5(+) (CCR5(+)) T cells, T regulatory cells, and T-helper 17 cells were unaltered in HIV+ men, CD8 T-cell production of tumor necrosis factor α (TNFα) was decreased. HIV-specific CD8 T cells were present in the foreskins of HIV+ men, although their frequency and function was reduced compared to the blood. CONCLUSIONS: Foreskin CD4 T-cell density and CCR5 expression were not reduced during HIV infection, perhaps explaining susceptibility to HIV superinfection. Foreskin CD8 T-cell density was increased, but decreased production of TNFα may enhance susceptibility to genital coinfections in HIV+ men.

Feasibility and safety of cervical biopsy sampling for mucosal immune studies in female sex workers from Nairobi, Kenya.

BACKGROUND: There is an urgent need to improve our understanding of the mucosal immuno-pathogenesis of HIV acquisition in the female genital tract, particularly in high-risk women such as female sex workers (FSWs). Cervical biopsy samples offer technical advantages over cytobrush sampling, but there are concerns that this might increase HIV acquisition, particularly if healing is slow and/or women do not abstain from sex during healing. METHODOLOGY/PRINCIPAL FINDINGS: Cervical biopsy samples and cervico-vaginal swabs for co-infection diagnostics, prostate specific antigen (PSA) and immune studies were collected from 59 women, including HIV seropositive and HIV-exposed seronegative (HESN) FSWs as well as lower risk women from Nairobi, Kenya. A clinical-demographic questionnaire was administered and women were instructed to avoid sexual intercourse, douching and the insertion of tampons for 14 days. All participants underwent a repeat exam to assess healing within the 14 days, and had HIV diagnostics at six months. Cervical sampling was well tolerated, and 82% of participants had healed macroscopically by 5 days. Both self-report and PSA screening suggested high levels of compliance with pre- and post-procedure abstinence. Delayed healing was associated with vulvovaginal candidiasis (VVC) and HESN status. At six-month follow up all low-risk and HESN participants remained HIV seronegative. CONCLUSION: Cervical biopsy sampling is a safe and well-tolerated method to obtain cervical biopsies in this context, particularly if participants with VVC are excluded. As healing could be delayed up to 11 days, it is important to support (both financially and with rigorous counseling) a period of post-procedure abstinence to minimize HIV risk.

HIV-neutralizing activity of cationic polypeptides in cervicovaginal secretions of women in HIV-serodiscordant relationships.

BACKGROUND: HIV exposed seronegative (HESN) women represent the population most in need of a prophylactic antiviral strategy. Mucosal cationic polypeptides can potentially be regulated for this purpose and we here aimed to determine their endogenous expression and HIV neutralizing activity in genital secretions of women at risk of HIV infection. METHODOLOGY/PRINCIPAL FINDINGS: Cervicovaginal secretions (CVS) of Kenyan women in HIV-serodiscordant relationships (HESN, n = 164; HIV seropositive, n = 60) and low-risk controls (n = 72) were assessed for the cationic polypeptides HNP1-3, LL-37 and SLPI by ELISA and for HIV neutralizing activity by a PBMC-based assay using an HIV primary isolate. Median levels of HNP1-3 and LL-37 in CVS were similar across study groups. Neither HSV-2 serostatus, nor presence of bacterial vaginosis, correlated with levels of HNP1-3 or LL-37 in the HESN women. However, an association with their partner's viral load was observed. High viral load (>10,000 HIV RNA copies/ml plasma) correlated with higher levels of HNP1-3 and LL-37 (p = 0.04 and 0.03, respectively). SLPI was most abundant in the low-risk group and did not correlate with male partner's viral load in the HESN women. HIV neutralizing activity was found in CVS of all study groups. In experimental studies, selective depletion of cationic polypeptides from CVS rendered the remaining CVS fraction non-neutralizing, whereas the cationic polypeptide fraction retained the activity. Furthermore, recombinant HNP1-3 and LL-37 could induce neutralizing activity when added to CVS lacking intrinsic activity. CONCLUSIONS/SIGNIFICANCE: These findings show that CVS from HESN, low-risk, and HIV seropositive women contain HIV neutralizing activity. Although several innate immune proteins, including HNP1-3 and LL-37, contribute to this activity these molecules can also have inflammatory properties. This balance is influenced by hormonal and environmental factors and in the present HIV serodiscordant couple cohort study we show that a partner's viral load is associated with levels of such molecules.

Detection of intraepithelial and stromal Langerin and CCR5 positive cells in the human endometrium: potential targets for HIV infection.

Both the upper (endocervix and uterus) and lower (ectocervix and vagina) female genital tract mucosa are considered to be target sites for sexual transmission of HIV. There are a few reports on the T cell and antigen-presenting cell distribution in human endometrial tissue however, there is little known about the expression of the HIV co-receptor CCR5 and HIV-binding C-type lectin receptors on endometrial cell subsets. We therefore assessed endometrial tissue sections from HIV seronegative women undergoing hysterectomy of a benign and non-inflammatory cause for phenotypic characterization of potential HIV target cells and receptors by immunohistochemistry. Langerin was expressed on intraepithelial CD1a+CD4+ and CD11c+CD4+ Langerhans cells. Furthermore, CCR5+CD4+CD3+ T cells, DC-SIGN+MR+CD11c+ myeloid dendritic cells and MR+CD68+ macrophages were found within or adjacent to the epithelium of the uterine lumen. In addition, occasional CD123+ BDCA-2+ plasmacytoid dendritic cells were detected deep in the endometrial stroma. Both T cells and several antigen-presenting cells were detected in lymphoid aggregate formations in close proximity to the epithelial lining. The finding of intraepithelial and stromal Langerin+ cells as well as CCR5+ CD4+ T cells is novel for human endometrium.

Slumbering mucosal immune response in the cervix of human papillomavirus DNA-positive and -negative women.

Persistent human papillomavirus (HPV) infection is a prerequisite for cervical cancer and results from bypassing the local immune response. Twenty-four volunteers underwent an ectocervical biopsy, Pap smear, tests for sexually transmitted infections including HIV and HPV genotyping. All answered a questionnaire regarding medical history. Repeat Pap smear and HPV genotyping was performed 9-26 months later. Quantitative reverse transcriptase (qRT-)PCR was used to assess expression of CD3, CD4, CD8, CD19, CD27, IL-2, IL-12, IL-4, IL-10, IL-17, HLA-DRα, TGFß, IFNγ, PD-1, PD-L1, CTLA-4, LAG3, IgA, IgG, CCR5, CCL5/RANTES and the IL-7 receptor in the biopsies. Eleven of 24 volunteers were HPV DNA-positive at baseline. Four of 10 were infected with a persistent HPV genotype at follow-up. All target molecules were successfully amplified and quantified except for IL-4. We found no difference in mRNA expression of these molecules when comparing HPV DNA-positive and -negative women, neither when comparing persistently infected individuals or those who cleared the infection. However, mRNA expression of the B cell phenotypic marker CD19 was higher in women using hormonal contraception than those not (p<0.05). HPV infection does not evoke a local inflammatory immune response in the ectocervix measurable with qRT-PCR. Hormonal contraception may influence B cell activity in the cervix.

Upregulation of interferon-alpha and RANTES in the cervix of HIV-1-seronegative women with high-risk behavior.

OBJECTIVE: The expression of innate immune molecules associated with potential blocking activity of HIV-1 propagation was analyzed in the cervical tissue of a group of African HIV-1 IgG-negative commercial sex workers (CSWs) with an HIV-1-encountering risk behavior. METHODS: Cervical biopsies from the superior portion of the ectocervix were assessed for innate immune molecules and evaluated in situ by computerized image analysis at the single-cell level. RESULTS: A higher expression of interferon-alpha (IFNalpha) and RANTES was detected in CSWs and HIV-1-infected individuals as compared to low-risk HIV-1-uninfected controls (Neg Ctrls). Most (>90%) of RANTES-expressing cells were CD8 cells as determined by confocal microscopy. In contrast, the expression of leukemia inhibitory factor (LIF) and secretory leukocyte protease inhibitor (SLPI) was comparable between the groups. The expression of beta-defensin 2 was highest in HIV-1-infected individuals. CONCLUSIONS: Induction of IFNalpha and RANTES expression in cervical mucosa may contribute to protection of sexual HIV-1 transmission in subjects with a higher risk behavior.

Aberrant cellular immune responses in humans infected persistently with parvovirus B19.

A subset of parvovirus B19 (B19) infected patients retains the infection for years, as defined by detection of B19 DNA in bone marrow. Thus far, analysis of B19-specific humoral immune responses and viral genome variations has not revealed a mechanism for the absent viral clearance. In this study, ex-vivo cellular immune responses were assessed by enzyme linked immunospot assay mounted against the majority of the translated viral genome. Compared to seropositive healthy individuals, individuals with B19 persistence (2-8 years) showed larger number of responses to the structural proteins (P = 0.0022), whereas responses to the non-structural protein were of lower magnitude (P = 0.012). These observations provide the first findings of immunological discrepancies between individuals with B19 persistence and healthy individuals, findings that may reflect both failed immunity and antigenic exhaustion.