|Call||Career Development Fellowship (CDF)|
Mucosal type I IFN desensitization and the risk of HIV acquisition
Aim 1a. To establish longitudinal patterns of IFNa2 expression in the FRT, and correlate these patterns to epidemiological variables. Aim 1b. To correlate high/low IFN status defined in Aim 1a with IFN responsiveness in mucosal HIV target cells ex vivo. Aim 1c. To correlate high/low IFN status defined in Aim 1a with altered HIV susceptibility of mucosal target cells in vitro.
|Centre for the Aids Programmes of Research in South Africa (CAPRISA)||South Africa|
|Durban, South Africa|
|Type||Name||Title||University||Start Date||End Date|
|Honours||Nokwanda Masondo||Characterizing Type I IFN expression in mucosal samples from CAPRISA 082 study||UKZN||2018||2019|
|Honours||Nobuhle Dlamini||Inflammatory plasma cytokine differences between sexes during active TB in IMPRESS||UKZN||2019||2020|
This proposal aims to answer important questions regarding the role of antiviral interferon responses during HIV infection, characterizing both the causes and consequences of increased IFN production in the genital tract. In the first reporting period we have characterized the longitudinal expression of IFNs in mucosal specimens from the CAPRISA 082 cohort and correlated interferon expression with a number of epidemiological and biological variables in order to characterize the drivers of increased IFN expression. Additionally, this project provides a training platform for a number of undergraduate and graduate students from University of KwaZulu-Natal. This study will contribute toward an understanding of how basic immunological mechanisms are exploited by HIV. Interferon therapy and blockade has been unsuccessfully tested in various clinical trials for HIV treatment further highlighting the need to better understand the IFN-mediated antiviral immune responses at the initial site of infection in humans as this is crucial if antiviral properties of IFNs will be successfully exploited in HIV prevention strategies.
CAPRISA/University of Manitoba
|University of Manitoba, Canada||PhD||2014-10-20|
Human Immuno-deficiency Virus (HIV) Tuberculosis (TB)
Several clinical trials have demonstrated that antiretroviral (ARV) drugs taken as pre-exposure prophylaxis (PrEP) can prevent HIV infection, with the magnitude of protection ranging from -49 to 86% (refs. ). Although these divergent outcomes are thought to be due primarily to differences in product adherence, biological factors likely contribute. Despite selective recruitment of higher-risk participants for prevention trials, HIV risk is heterogeneous even within higher-risk groups. To determine whether this heterogeneity could influence patient outcomes following PrEP, we undertook a post hoc prospective analysis of results from the CAPRISA 004 trial for 1% tenofovir gel (n = 774 patients), one of the first trials to demonstrate protection against HIV infection. Concentrations of nine proinflammatory cytokines were measured in cervicovaginal lavages at >2,000 visits, and a graduated cytokine score was used to define genital inflammation. In women without genital inflammation, tenofovir was 57% protective against HIV (95% confidence interval (CI): 7-80%) but was 3% protective (95% CI: -104-54%) if genital inflammation was present. Among women who highly adhered to the gel, tenofovir protection was 75% (95% CI: 25-92%) in women without inflammation compared to -10% (95% CI: -184-57%) in women with inflammation. Immunological predictors of HIV risk may modify the effectiveness of tools for HIV prevention; reducing genital inflammation in women may augment HIV prevention efforts.
Interferons, induced early during viral infections, represent important regulators of both innate and adaptive immune responses, and provide protective effects against a wide range of pathogens, including HIV. Several in vitro studies and some in vivo data from HIV-exposed seronegative cohorts indicate that interferons and interferon-mediated immune responses are crucial in preventing early HIV replication. Following establishment of HIV infection, the uncontrolled (aberrant) activation of the immune system, in part regulated by interferon levels, contributes to HIV-1-induced immune activation and disease progression. Modulation of interferon responses prior to and during HIV infection shows promise for development of novel therapeutics to prevent HIV transmission, clear HIV infection, and dampen chronic immune activation. In this review we discuss the role that interferons play in protection from HIV infection, acute infection, and their role in HIV pathogenesis and disease progression. Lastly, we review recent advances in modulating interferon responses for purposes of developing novel HIV therapeutic approaches.
Members of the interferon regulatory factor (IRF) family control the expression of numerous proteins, many of which are central to regulating host immune responses. IRF1 is one of the central mediators of the innate and adaptive immune responses required for antigen processing and presentation, Th1/Th2 differentiation, and natural killer (NK) cell and macrophage function. Many viruses have evolved mechanisms to target the IRF1 pathway in order to promote viral pathogenesis. During early HIV infection, IRF1 acts as a double-edged sword, critical for driving viral replication as well as eliciting antiviral responses. In this review, we describe the strategies that HIV-1 has evolved to modulate IRF1 in order to enhance viral replication and to disarm the host immune system. IRF1 has been shown to be an important factor in natural protection against HIV in highly exposed seronegative (HESN) individuals and is crucial in regulating the initial stages of HIV replication and HIV disease progression, as well as the establishment of latency. An understanding of how the protective effects of IRF1 responses are controlled in HESN individuals, naturally resistant to HIV infection, may provide important clues on how to regain control of HIV and tip the balance of immunity in favor of the host, or provide new opportunities to eliminate HIV in its host altogether.
The gastrointestinal (GI) mucosa is central to HIV pathogenesis, and the integrin a4b7 promotes the homing of immune cells to this site, including those that serve as viral targets. Data from simian immunodeficiency virus (SIV) animal models suggest that a4b7 blockade provides prophylactic and therapeutic benefits. We show that pre-HIV infection frequencies of a4b7+ peripheral blood CD4+ T cells, independent of other T cell phenotypes and genital inflammation, were associated with increased rates of HIV acquisition in South African women. A similar acquisition effect was observed in a Kenyan cohort and in nonhuman primates (NHPs) after intravaginal SIV chal- lenge. This association was stronger when infection was caused by HIV strains containing V2 envelope motifs with a preference for a4b7 binding. In addition, pre-HIV a4b7+ CD4+ T cells predicted a higher set-point viral load and a greater than twofold increased rate of CD4+ T cell decline. These results were confirmed in SIV-infected NHPs. Increased frequencies of pre-HIV a4b7+ CD4+ T cells were also associated with higher postinfection expression of lipopolysaccharide binding protein, a microbial translocation marker, suggestive of more extensive gut damage. CD4+ T cells expressing a4b7 were rapidly depleted very early in HIV infection, particularly from the GI mucosa, and were not restored by early antiretroviral therapy. This study provides a link between a4b7 expression and HIV clinical outcomes in humans, in line with observations made in NHPs. Given the availability of a clinically approved anti-a4b7 monoclonal antibody for treatment of inflammatory bowel disease, these data support fur- ther evaluation of targeting a4b7 integrin as a clinical intervention during HIV infection.
Background: While non-optimal vaginal bacteria and inflammation have been associated with increased HIV risk, the upstream drivers of these phenotypes are poorly defined in young African women.
Setting: Mombasa, Kenya.
Methods: We characterized vaginal microbiome and cytokine profiles of sexually active young women aged 14-24 years (n=168) in three study groups: those engaging in formal sex work, in transactional sex, and non-sex workers. Vaginal secretions were collected via self-inserted SoftCup, and assayed for cytokines and vaginal microbiome via multiplex ELISA and 16S rRNA sequencing, respectively. Epidemiological data were captured using a validated questionnaire.
Results: The median age of participants was 20 (IQR: 18-22). Approximately two-thirds of young women (105/168) had vaginal microbial communities characterized by Gardnerella and/or Prevotella spp.-dominance; a further 29% (49/168) were predominantly Lactobacillus iners. Microbiome clustering explained a large proportion of cytokine variation (>50% by the first 2 principle components). Age was not associated with vaginal microbial profiles in bivariable or multivariable analyses. Women self-identifying as sex workers had increased alpha (intra-individual) diversity, independent of age, recent sexual activity, HIV and other STIs (beta = 0.47, 95% CI: 0.05 - 0.90, p = 0.03). Recent sex (number of partners or sex acts last week, time since last vaginal sex) correlated with increased alpha diversity, particularly in participants who were not involved in sex work.
Conclusion: Non-optimal vaginal microbiomes were common in young Kenyan women and associated with sex work and recent sexual activity, but independent of age. Restoring optimal vaginal microflora may represent a useful HIV prevention strategy.
Not all individuals exposed to HIV become infected. Understanding why these HIV-exposed seronegative individuals remain uninfected will help inform the development of preventative measures against HIV infection. Interferon regulatory factor-1 (IRF1) plays a critical role both in host antiviral immunity and in HIV-1 replication. This study examined IRF1 expression regulation in the ex vivo peripheral blood mononuclear cells of HIV-exposed seronegative commercial sex workers who can be epidemiologically defined as relatively resistant to HIV infection (HIV-R), versus HIV-uninfected, susceptible controls (HIV-S). Whereas HIV-susceptible individuals demonstrated a biphasic, prolonged increase in IRF1 expression after interferon-γ stimulation, HIV-R individuals demonstrated a robust, but transient response. We also found that the IRF1 promoter in HIV-R was primed by increased basal histone deacetylase-2 binding, independently of transcription regulators, STAT1 and nuclear factor-κB/p65, implicating an epigenetic silencing mechanism. Interestingly, the transitory IRF1 response in HIV-R was sufficient in comparable regulation of interleukin-12 and interleukin-4 expression compared with the HIV-susceptible controls. This is the first study characterizing IRF1 responsiveness in individuals who demonstrate altered susceptibility to HIV infection. These data suggest that transitory IRF1 responsiveness in HIV-R may be one of the key contributors to the altered susceptibility to HIV infection during the early stages of primary HIV infection.
Cells from women who are epidemiologically deemed resistant to HIV infection exhibit a 40-60% reduction in endogenous IRF-1 (interferon regulatory factor-1), an essential regulator of host antiviral immunity and the early HIV replication. This study examined the functional consequences of reducing endogenous IRF-1 on HIV-1 replication and immune response to HIV in natural HIV target cells. IRF-1 knockdown was achieved in ex vivo CD4(+) T cells and monocytes with siRNA. IRF-1 level was assessed using flow cytometry, prior to infection with HIV-Bal, HIV-IIIB, or HIV-VSV-G. Transactivation of HIV long terminal repeats was assessed by p24 secretion (ELISA) and Gag expression (reverse transcription-polymerase chain reaction (RT-PCR)). The expression of IRF-1-regulated antiviral genes was quantitated with RT-PCR. A modest 20-40% reduction in endogenous IRF-1 was achieved in >87% of ex vivo-derived peripheral CD4(+) T cells and monocytes, resulted in >90% reduction in the transactivation of the HIV-1 genes (Gag, p24) and, hence, HIV replication. Curiously, these HIV-resistant women demonstrated normal immune responses, nor an increased susceptibility to other infection. Similarly, modest IRF-1 knockdown had limited impact on the magnitude of HIV-1-elicited activation of IRF-1-regulated host immunologic genes but resulted in lessened duration of these responses. These data suggest that early expression of HIV-1 genes requires a higher IRF-1 level, compared to the host antiviral genes. Together, these provide one key mechanism underlying the natural resistance against HIV infection and further suggest that modest IRF-1 reduction could effectively limit productive HIV infection yet remain sufficient to activate a robust but transient immune response.
Burkholderia cenocepacia is a member of the Burkholderia cepacia complex, a group of metabolically versatile bacteria that have emerged as opportunistic pathogens in cystic fibrosis and immunocompromised patients. Previously a screen of transposon mutants in a rat pulmonary infection model identified an attenuated mutant with an insertion in paaE, a gene related to the phenylacetic acid (PA) catabolic pathway. In this study, we characterized gene clusters involved in the PA degradation pathway of B. cenocepacia K56-2 in relation to its pathogenicity in the Caenorhabditis elegans model of infection. We demonstrated that targeted-insertion mutagenesis of paaA and paaE, which encode part of the putative PA-coenzyme A (CoA) ring hydroxylation system, paaZ, coding for a putative ring opening enzyme, and paaF, encoding part of the putative beta-oxidation system, severely reduces growth on PA as a sole carbon source. paaA and paaE insertional mutants were attenuated for virulence, and expression of paaE in trans restored pathogenicity of the paaE mutant to wild-type levels. Interruption of paaZ and paaF slightly increased virulence. Using gene interference by ingested double-stranded RNA, we showed that the attenuated phenotype of the paaA and paaE mutants is dependent on a functional p38 mitogen-activated protein kinase pathway in C. elegans. Taken together, our results demonstrate that B. cenocepacia possesses a functional PA degradation pathway and that the putative PA-CoA ring hydroxylation system is required for full pathogenicity in C. elegans.