An ex vivo Model of HIV-1 Infection in Human Lymphoid Tissue and Cervico-vaginal Tissue

Introini, Andrea; Vanpouille, Christophe; Grivel, Jean‐Charles; Margolis, Leonid

doi:10.21769/bioprotoc.1047

Cited by 16 publications

(11 citation statements)

References 9 publications

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“…The explants were maintained at the liquid-air interface for 18 days, as previously described [79,80]. Culture supernatant was sampled every 3 days before replacing it with fresh medium, and stored at -80°C.…”

Section: Methodsmentioning

confidence: 99%

Seminal plasma induces inflammation and enhances HIV-1 replication in human cervical tissue explants

et al. 2017

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The most immediate and evident effect of mucosal exposure to semen in vivo is a local release of proinflammatory mediators accompanied by an influx of leukocytes into the female genital mucosa (FGM). The implication of such response in HIV-1 transmission has never been addressed due to limitations of currently available experimental models. Using human tissue explants from the uterine cervix, we developed a system of mucosal exposure to seminal plasma (SP) that supports HIV-1 replication. Treatment of ectocervical explants with SP resulted in the upregulation of inflammatory and growth factors, including IL-6, TNF, CCL5, CCL20, CXCL1, and CXCL8, and IL1A, CSF2, IL7, PTGS2, as evaluated by measuring protein levels in explant conditioned medium (ECM) and gene expression in tissue. SP treatment was also associated with increased recruitment of monocytes and neutrophils, as observed upon incubation of peripheral blood leukocytes with ECM in a transwell system. To evaluate the impact of the SP-mediated response on local susceptibility to HIV-1, we infected ectocervical explants with the CCR5-tropic variant HIV-1BaL either in the presence of SP, or after explant pre-incubation with SP. In both experimental settings SP enhanced virus replication as evaluated by HIV-1 p24gag released in explant culture medium over time, as well as by HIV-1 DNA quantification in explants infected in the presence of SP. These results suggest that a sustained inflammatory response elicited by SP soon after coitus may promote HIV-1 transmission to the FGM. Nevertheless, ectocervical tissue explants did not support the replication of transmitted/founder HIV-1 molecular clones, regardless of SP treatment. Our system offers experimental and analytical advantages over traditional models of HIV-1 transmission for the study of SP immunoregulatory effect on the FGM, and may provide a useful platform to ultimately identify new determinants of HIV-1 infection at this site.

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Section: Methodsmentioning

confidence: 99%

Seminal plasma induces inflammation and enhances HIV-1 replication in human cervical tissue explants

et al. 2017

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“…For each specimen, the mucosa was separated from any residual underlying stromal or muscular tissue and dissected into approximately 8 mm 3 blocks using a scalpel and tweezers, as previously described. 46 One or two tissue blocks were snap-frozen immediately after dissection and cryopreserved at -80°C for in situ staining. The remaining tissue blocks were digested with 0.1 mg/ml of DNase I (Roche, Basel, Switzerland) and 0.5 mg/ml of Collagenase II (Sigma-Aldrich, St. Louis, MO, USA) in a thermoshaker (Labnet International, Edison, NJ, USA) rocking at 1400 rpm for 30 min at 37°C.…”

Section: Methodsmentioning

confidence: 99%

MAIT cells reside in the female genital mucosa and are biased towards IL-17 and IL-22 production in response to bacterial stimulation

et al. 2017

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The female genital tract (FGT) mucosa is a critically important site for immune defense against microbes. Mucosal-associated invariant T (MAIT) cells are an innate-like T cell population that recognizes microbial riboflavin metabolite antigens in an MR1-dependent manner. The role of MAIT cells in the FGT mucosa is unknown. Here, we found that MAIT cells and MR1+ antigen-presenting cells were present in the upper and lower FGT, with distinct tissue localization of MAIT cells in endometrium versus cervix. MAIT cells from the FGT and blood displayed a distinct phenotype with expression of IL-18Rα, CD127, α4β7, PD-1, as well as the transcription factors PLZF, RORγt, Helios, Eomes and T-bet. Their expression levels of PLZF and Eomes were lower in the FGT compared to blood. When stimulated with Escherichia coli, MAIT cells from the FGT displayed a bias towards IL-17 and IL-22 expression, whereas blood MAIT cells produced primarily IFN-γ, TNF, and Granzyme B. Furthermore, both FGT- and blood-derived MAIT cells were polyfunctional and contributed to the T cell-mediated response to E. coli. Thus, MAIT cells in the genital mucosa have a distinct IL-17/IL-22 profile and may play an important role in immunological homeostasis and control of microbes at this site.

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“…Tissue samples were maintained in ice-cold RPMI 1640 and processed within 24 hours of surgery. For each specimen, the mucosa was separated from underlying stroma and dissected into blocks as previously described (43). Tissue blocks were snap-frozen immediately after dissection and stored at −80°C for in situ staining, or digested with 0.1 mg/ml of DNase I (Roche, Basel, Switzerland) and 0.5 mg/ml of Collagenase II (Sigma-Aldrich, St. Louis, MO, USA) for 30 min at 37°C.…”

Section: Methodsmentioning

confidence: 99%

Innate Invariant NKT Cell Recognition of HIV-1–Infected Dendritic Cells Is an Early Detection Mechanism Targeted by Viral Immune Evasion

Paquin‐Proulx

Gibbs

Bächle

et al. 2016

The Journal of Immunology

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Invariant natural killer T (iNKT) cells are innate-like T cells that respond rapidly with a broad range of effector functions upon recognition of glycolipid antigens presented by CD1d. HIV-1 carries Nef- and Vpu-dependent mechanisms to interfere with CD1d surface expression, indirectly suggesting a role for iNKT cells in control of HIV-1 infection. Here, we investigated if iNKT cells can participate in the innate cell-mediated immune response to HIV-1. Infection of dendritic cells (DCs) with Nef- and Vpu-deficient HIV-1 induced upregulation of CD1d in a TLR7-dependent manner. Infection of DCs caused modulation of enzymes in the sphingolipid pathway and enhanced expression of the endogenous glucocylceramide antigen. Importantly, iNKT cells responded specifically to rare DCs productively infected with Nef- and Vpu-defective HIV-1. Transmitted founder viral isolates differed in their CD1d down-regulation capacity, suggesting that diverse strains may be differentially successful in inhibiting this pathway. Furthermore, both iNKT cells and DCs expressing CD1d and HIV-receptors resided in the female genital mucosa, a site where HIV-1 transmission occurs. Together, these findings suggest that innate iNKT cell sensing of HIV-1 infection in DCs is an early immune detection mechanism, which is independent of priming and adaptive recognition of viral antigen, and is actively targeted by Nef- and Vpu-dependent viral immune evasion mechanisms.

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An ex vivo Model of HIV-1 Infection in Human Lymphoid Tissue and Cervico-vaginal Tissue

Abstract: Human tissue explants are

Cited by 16 publications

References 9 publications

Seminal plasma induces inflammation and enhances HIV-1 replication in human cervical tissue explants

Seminal plasma induces inflammation and enhances HIV-1 replication in human cervical tissue explants

MAIT cells reside in the female genital mucosa and are biased towards IL-17 and IL-22 production in response to bacterial stimulation

Innate Invariant NKT Cell Recognition of HIV-1–Infected Dendritic Cells Is an Early Detection Mechanism Targeted by Viral Immune Evasion

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