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Heterogeneity of HIV-1 Replication in Ectocervical and Vaginal Tissue Ex Vivo.

ABSTRACT: In clinical trials evaluating HIV-1 prevention products, ex vivo exposure of mucosal tissue to HIV-1 is performed to inform drug levels needed to suppress viral infection. Understanding assay and participant variables that influence HIV-1 replication will help with assay implementation. Demographic and behavioral data were obtained from 61 healthy women aged 21-45. Paired cervical tissue (CT) and vaginal tissue (VT) biopsies were collected and treated with HIV-1_BaL or HIV-1_JR-CSF, washed, and cultured. On days 3, 7, and/or 11, culture supernatant was collected, and viral replication was monitored by p24 ELISA. Tissue was extracted at study end, and HIV-1 relative RNA copies were determined by polymerase chain reaction. Cumulative p24 and RNA were log-transformed and analyzed using a linear mixed model, t-test, and an intraclass correlation coefficient (ICC). HIV replication was similar between CT and VT for each virus, but HIV-1_BaL had 1.5 log₁₀ and 0.9 log₁₀ higher levels of p24 than HIV-1_JR-CSF in CT and VT, respectively (p < .001), which correlated with HIV-1 relative RNA copies. Cumulative p24 and RNA copies in both tissues demonstrated low intraperson correlation for both viruses (ICC ≤0.513 HIV-1_BaL; ICC ≤0.419 HIV-1_JR-CSF). Enrollment into previous clinical studies in which genital biopsies were collected modestly decreased the HIV-1_BaL cumulative p24 for CT, but not for VT. To improve the ex vivo challenge assay, viruses should be evaluated for replication in mucosal tissue before study implementation, baseline mucosal tissue is not needed if a placebo/no treatment group is included within the clinical trial, and previous biopsy sites should be avoided.

SUBMITTER: Dezzutti CS

PROVIDER: S-EPMC8104025 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The human vaginal microbiome (VMB) is a complex bacterial community that interacts closely with vaginal epithelial cells (VECs) impacting the mucosal phenotype and its responses to pathogenic insults. The VMB and VEC relationship includes nutrient exchange and regulation of signaling molecules that controls numerous host functions and defends against invading pathogens. To better understand infection and replication of sexually transmitted viral pathogens in the human vaginal mucosa we used our ex vivo VEC multilayer culture system. We tested the hypothesis that selected VMB communities could be identified that alter the replication of sexually transmitted viruses consistent with reported clinical associations. Sterile VEC multilayer cultures or those colonized with VMB dominated by specific Lactobacillus spp., or VMB lacking lactobacilli, were infected with Zika virus, (ZIKV) a single stranded RNA virus, or Herpes Simplex Virus type 2 (HSV-2), a double stranded DNA virus. The virus was added to the apical surface of the cultured VEC multilayer to model transmission during vaginal intercourse. Viral replication was measured 48 h later by qPCR. The results indicated that VEC cultures colonized by VMB containing Staphylococcus spp., previously reported as inflammatory, significantly reduced the quantity of viral genomes produced by ZIKV. HSV-2 titers were decreased by nearly every VMB tested relative to the sterile control, although Lactobacillus spp.-dominated VMBs caused the greatest reduction in HSV-2 titer consistent with clinical observations. To explore the mechanism for reduced ZIKV titers, we investigated inflammation created by ZIKV infection, VMB colonization or pre-exposure to selected TLR agonists. Finally, expression levels of human beta defensins 1-3 were quantified in cultures infected by ZIKV and those colonized by VMBs that impacted ZIKV titers. Human beta defensins 1-3 produced by the VEC showed no association with ZIKV titers. The data presented expands the utility of this ex vivo model system providing controlled and reproducible methods to study the VMB impact on STIs and indicated an association between viral replication and specific bacterial species within the VMB.

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Heterogeneity of HIV-1 Replication in Ectocervical and Vaginal Tissue Ex Vivo.

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