This proposal aims to improve our understanding of how variations in hormone levels and reproductive tract microbes contribute to HIV acquisition risk in women. These are significant areas of research because more than half of new HIV infections are acquired by women, predominantly through heterosexual transmission. The goal of the proposed research project is to define how hormonal variation during the normal menstrual cycle (endogenous hormones) and with DMPA contraceptives (synthetic progestin) and bacterial vaginosis affect protein expression in ectocervical CD4+ T lymphocytes and their susceptibility to fusion and productive infection with HIV. These studies are powered to allow determination of hormonal effects, microbiota effects and interaction effects between hormones and vaginal microbes. We have found that bacterial metabolites called short chain fatty acids (SFCAs) that are present in healthy cervicovaginal fluid and increase dramatically during BV, strongly heighten the susceptibility of CD4+ T cells to productive HIV infection through the inhibition of histone deacetylases (HDACs). Finally, we will investigate a novel strategy to reduce HIV transmission by making cells more resistant to HIV infection. The three overlapping areas to be investigated in this project are: 1. Determine the effects of hormone levels and BV on susceptibility and activation state of CD4+ T lymphocytes from ectocervical biopsies. Ectocervical CD4+ T cell susceptibility to fusion and productive infection by HIV will be directly measured using our combination reporter virus system by flow cytometry. Activation states of CD4+ T cells will be quantified by flow cytometry and label-free proteomics, an unbiased and powerful strategy for identifying specific activation pathways. Advanced bioinformatics will be used to identify specific activation pathways that contribute to HIV replication in ectocervical CD4+ T cells. 2. Quantify bacterial species and metabolites present in cervicovaginal lavage (CVL) samples and correlate metabolites with specific species or groups of bacteria. CVL samples will be analyzed for bacterial species and for bacterial metabolites-including short chain fatty acids (SCFAs) that are present in the healthy female reproductive tract and elevated in BV. We have recently found that SCFAs enhance HIV infection of CD4+ T cells via a mechanism involving inhibition of HDACs. Advanced bioinformatics will be used to identify correlations between bacteria and their products and ectocervical CD4+ T cell protein expression and susceptibility to HIV. 3. Determine the ability of garcinol to reduce the susceptibility of ectocervical CD4+ T cells to HIV ex vivo in women with different hormone patterns and cervicovaginal microbiota. Garcinol, a histone acetyltransferase inhibitor, reverses the effects of SCFA-mediated HDAC inhibition of HIV susceptibility in vitro, and actually reduces infection even in unstimulated cells. These findings suggest that garcinol, or similar compounds, may represent a novel strategy to reduce heterosexual transmission of HIV by inhibiting productive infection of ectocervical CD4+ T cells.
This project addresses several issues of central importance to heterosexual HIV transmission. First, we will address how variation in endogenous (from two stages of the menstrual cycle) and synthetic (from women on Depo-Provera contraception) hormones affects the composition of the cervicovaginal microbiota and their metabolites. Second, we will use sophisticated new technologies to evaluate the susceptibility of ectocervical CD4+ T cells to HIV, determine how cells have been activated in vivo, and use sophisticated bioinformatics strategies to determine which activation pathways increase HIV replication. Third, we will determine if bacteria or their metabolites affect HIV replication, including determining whether short chain fatty acids (SCFAs) are a mechanistic link between BV and HIV acquisition risk. Fourth, we will investigate the potential of a novel therapeutic strategy to reduce heterosexual HIV transmission.
|Lucera, Mark B; Fleissner, Zach; Tabler, Caroline O et al. (2017) HIV signaling through CD4 and CCR5 activates Rho family GTPases that are required for optimal infection of primary CD4+ T cells. Retrovirology 14:4|
|Lucera, Mark B; Tilton, Carisa A; Mao, Hongxia et al. (2014) The histone deacetylase inhibitor vorinostat (SAHA) increases the susceptibility of uninfected CD4+ T cells to HIV by increasing the kinetics and efficiency of postentry viral events. J Virol 88:10803-12|