Objective:
The rectal mucosa is a critical site of HIV vulnerability. We sought to identify transcriptomic features of rectal mucosal tissue prior to exposure associated with support or restriction of HIV replication.
Design:
Rectal tissue from HIV-negative cisgender men (n=57) underwent concurrent i) RNAseq transcriptomic analyses (2 biopsies/participant) and ii) challenge with HIV in the ex vivo explant model of infection (3 biopsies challenged/participant) as part of a larger cohort study to understand the rectal mucosal immune environment among men who have sex with men.
Methods:
P24 was quantified in the explant supernatants over a culture period of 18 days via ELISA. Participant median p24 log Area Under the Curve was correlated with bulk transcriptomic data (Illumina HiSeq3000) to identify associations between gene expression and p24 production. Significant differentially expressed genes (DEG) were identified via DESeq2 analysis, and analyzed with Reactome to identify pathways of interest.
Results:
183 DEG (181 upregulated, 2 downregulated) were associated with higher p24 accumulation in the ex vivo challenge model, including T cell activation, B cell function, and chemokine DEG. Reactome analysis of the upregulated genes identified ‘Adaptive Immune System’, ‘Cytokine Signaling in Immune System’, and ‘Innate Immune System’ as significant upregulated pathways.
Conclusions:
For the first time, we identified rectal tissue transcriptomic signatures associated with increased p24 production utilizing an ex vivo model. Our findings are highly relevant to HIV transmission and the early establishment of HIV reservoirs in humans, and future studies should examine the identified pathways as targets for new or improved biomedical prevention or treatment interventions.
Approximately half of the people with HIV (PWH) in the United States are retained in HIV care and only 57% have achieved viral suppression, due to barriers including transportation access, stigma, poor mental health, substance use, and medical mistrust. Community-based HIV care models have potential to address the diverse needs of patients and to improve retention in care, but their success is contingent on acceptance by patients and key community stakeholders. Recognizing that the preferences of PWH who are out-of-care (PWH-OOC) likely differ from those retained in care, we conducted a mixed-methods study from June 2019 to May 2021 composed of surveys with PWH-OOC (n = 50) and in-depth interviews with key clinic and community stakeholders (n = 41) to examine the relative preference and perceived advantages and disadvantages for six different community-based HIV care models versus the traditional fixed-clinic model. Survey data was analyzed to assess average rank preference for each care model and interview transcripts were thematically coded to examine factors influencing model acceptance. The highest preference for care delivery was via a mobile clinic, followed by community-based peer navigation, primary care clinics, telemedicine, traditional HIV subspeciality clinic, homeless shelter, and drug treatment center. Common factors influencing preference included convenience, accessibility, potential to preserve confidentiality, quality of care assurance, opportunity to develop rapport with their HIV care provider, access to a smart device, and potential to alleviate versus exacerbate HIV stigma. Participants discussed need for integration of care models and for individuals to choose different care models at different times. Providers and patients differed in preference for care model and weighting of relative advantages and disadvantages of each. Findings highlight the need to integrate alternative, community-based care models into the national plan to end the HIV epidemic and to allow for PWH-OOC to choose the model most fitting based on individual circumstances.
Young men who have sex with men (YMSM) represent a particularly high-risk group for HIV acquisition in the US, despite similarly reported rates of sexual activity as older, adult MSM (AMSM). Increased rates of HIV infection among YMSM compared to AMSM could be partially attributable to differences within the rectal mucosal (RM) immune environment associated with earlier sexual debut and less lifetime exposure to receptive anal intercourse. Using an ex vivo explant HIV challenge model, we found that rectal tissues from YMSM supported higher levels of p24 at peak viral replication timepoints compared to AMSM. Among YMSM, the RM was characterized by increased CD4+ T cell proliferation, as well as lower frequencies of tissue resident CD8+ T cells and pro-inflammatory cytokine producing CD4+ and CD8+ T cells. In addition, the microbiome composition of YMSM was enriched for anaerobic taxa that have previously been associated with HIV acquisition risk, including Prevotella, Peptostreptococcus, and Peptoniphilus. These distinct immunologic and microbiome characteristics were found to be associated with higher HIV replication following ex vivo challenge of rectal explants, suggesting the RM microenvironment of YMSM may be uniquely conducive to HIV infection.
Introduction: We previously showed that the rectal mucosal immune environment among men who have sex with men (MSM) engaging in condomless receptive anal intercourse (CRAI) is immunologically distinct from that of men who do not engage in anal intercourse (AI). Here, we further examined these differences with quantitative immunohistochemistry to better understand the geographic distribution of immune markers of interest. Methods: We enrolled a cohort of MSM engaging in CRAI (n = 41) and men who do not engage in AI (n = 21) between October 2013 and April 2015. Participants were healthy, HIV-negative men aged 18–45 from the metro Atlanta area. We performed rectal mucosal sampling via rigid sigmoidoscopy during two study visits separated by a median of nine weeks and timed with sexual activity for MSM engaging in CRAI. We used standardized, automated immunohistochemistry and quantitative image analysis to investigate the rectal mucosal distribution of neutrophils (MPO), IL-17-producing cells (IL-17) and Tregs (FOXP3) in the lamina propria, and cellular proliferation (Ki67) and adherens junction protein (E-cadherin) in the epithelium. We examined associations between biomarker expression and the rectal mucosal microbiota composition by 16s rRNA sequencing. Results: Relative to the colonic crypt base, IL-17, FOXP3, and MPO expression increased towards the rectal lumen, while Ki67 decreased and E-cadherin was more uniformly distributed. Throughout the rectal mucosa distribution examined, MSM engaging in CRAI had higher mean lamina propria MPO expression (p = 0.04) and epithelial Ki67 (p = 0.04) compared to controls. There were no significant differences in IL-17, FOXP3 or E-cadherin expression. We found no significant associations of the five biomarkers with the global rectal microbiota composition or the individual taxa examined. Conclusions: Understanding the mucosal distribution of inflammatory mediators can enhance our knowledge of the earliest events in HIV transmission. Neutrophil enrichment and crypt epithelial cell proliferation likely represent sub-clinical inflammation in response to CRAI in the rectal mucosa of MSM, which could increase the risk for HIV acquisition. However, the contributory role of the microbiota in mucosal inflammation among MSM remains unclear. HIV prevention may be enhanced by interventions that reduce inflammation or capitalize on the presence of specific inflammatory mechanisms during HIV exposure.
Our understanding of innate immune responses in human rectal mucosal tissues (RM) and their contributions to promoting or restricting HIV transmission is limited. We defined the RM composition of innate and innate-like cell subsets, including plasmacytoid dendritic cells; CD1c + myeloid DCs; neutrophils; macrophages; natural killer cells (NK); Marginal Zone-like B cells (MZB); γδ T cells; and mucosal-associated invariant T cells in RM from 69 HIV-negative men by flow cytometry. Associations between these cell subsets and HIV-1 replication in ex vivo RM explant challenge experiments revealed an inverse correlation between RM-NK and p24 production, in contrast to a positive association between RM-MZB and HIV replication. Comparison of RM and blood-derived MZB and NK illustrated qualitative and quantitative differences between tissue compartments. Additionally, 22 soluble molecules were measured in a subset of explant cultures (n = 26). Higher production of IL-17A, IFN-γ, IL-10, IP-10, GM-CSF, sFasL, Granzyme A, Granzyme B, Granulysin, and Perforin following infection positively correlated with HIV replication. These data show novel associations between MZB and NK cells and p24 production in RM and underscore the importance of inflammatory cytokines in mucosal HIV infection, demonstrating the likely critical role these innate immune responses play in early mucosal HIV replication in humans.