Our earlier studies in pig-tailed macaques demonstrated varying SHIV susceptibility during the menstrual cycle, likely caused by cyclic variations in immune responses in the female genital tract. There is concern that high-dose, long-lasting, injectable progestin-based contraception could mimic the high-progesterone luteal phase and predispose women to HIV-1 acquisition and transmission. In this study, we adopted a systems biology approach employing proteomics (tandem mass spectrometry), transcriptomics (RNA microarray hybridization), and other specific protein assays (enzyme-linked immunosorbent assays and multiplex chemokine-cytokine measurements) to characterize the effects of hormonal changes on the expression of innate factors and secreted proteins in the macaque vagina. Several antiviral factors and pathways (including acute phase response signaling and complement system) were overexpressed in the follicular phase. Conversely, during the luteal phase there were factors overexpressed (including moesins, syndecans, integrins, among others) that could play direct or indirect roles in enhancing HIV-1 infection. Thus, our study showed that specific pathways and proteins/genes might be working in tandem to regulate innate immunity, thus fostering further investigation and future design of approaches to help counter HIV-1 acquisition in the female genital tract. IMPORTANCE: HIV infection in women is poorly understood. High levels of the hormone progesterone may make women more vulnerable to infection. This could be the case during the menstrual cycle, when using hormone-based birth control, or during pregnancy. The biological basis for increased HIV vulnerability is not known. We used an animal model with high risk for infection during periods of high progesterone. Genital secretions and tissues were studied during the menstrual cycle. Our goal was to identify biological factors upregulated at high progesterone levels, and we indeed show an upregulation of genes and proteins which enhance the ability of HIV to infect when progesterone is high. In contrast, during low progesterone periods, we find more HIV inhibitory factors. This basic research study contributes to our understanding of mechanisms that may regulate HIV infection in females under hormonal influences. Such knowledge is needed for the development of novel prevention strategies.

Fluctuations in susceptibility to HIV or SHIV during the menstrual cycle are currently not fully documented. To address this, time point of infection was determined in 19 adult female pigtail macaques vaginally challenged during their undisturbed menstrual cycles with repeated, low-dose SHIVSF162P3 exposures. Eighteen macaques (95%) first displayed viremia in the follicular phase, as compared to 1 macaque (5%) in the luteal phase (p<0.0001). Due to a viral eclipse phase, we estimated a window of most frequent virus transmission between days 24-31 of the menstrual cycle, in the late luteal phase. Thus, susceptibility to vaginal SHIV infection is significantly elevated in the second half of the menstrual cycle when progesterone levels are high, and when local immunity may be low. Such susceptibility windows have been postulated before but not definitively documented. Our data support findings of higher susceptibility to HIV in women during progesterone-dominated periods including pregnancy and contraceptive use.

Animal models for research on susceptibility to HIV are currently not available. Here we explore whether a macaque model of repeated low-dose rectal or vaginal virus challenges could be employed. We tested the hypothesis that susceptibility to Simian HIV is not merely stochastic in this model but rather is associated with identifiable host factors. Forty macaques required a median of 3.5 SHIVSF162P3 challenges for infection. We studied the association of their susceptibility with 13 predisposing plasma cytokines/chemokines (RANTES, Eotaxin, monocyte chemoattractant protein (MCP)-1, IL-7, MIP-1beta, TNF-alpha, MIP-1alpha, granulocyte colony-stimulating factor, IL-8, interferon-gamma, IL-17, IL-1beta, IL-6). Higher plasma RANTES, IL-8, and Eotaxin were associated with lower susceptibility, that is, higher resistance to infection. In a group of macaques with low IL-8 and RANTES, a median 3 exposures were required to infect; whereas, when either IL-8 or RANTES were high, a median 12 exposures were required. Thus, susceptibility was associated with identifiable discrete host factors and was not stochastic. In addition, the macaque model identified key human resistance factors (RANTES, Eotaxin), but also revealed a novel association with resistance (IL-8). Future direct evaluation of these or other factors in the animal model may be beneficial for developing new immunomodulation strategies for HIV prevention.