Adaptive NK Cells Rapidly Expand during Acute HIV Infection and Persist Despite Early Initiation of Antiretroviral Therapy.

HIV is associated with NK cell dysfunction and expansion of adaptive-like NK cells that persist despite antiretroviral therapy (ART). We investigated the timing of NK cell perturbations during acute HIV infection and the impact of early ART initiation. PBMCs and plasma were obtained from people with HIV (PWH; all men who have sex with men; median age, 26.0 y) diagnosed during Fiebig stages I, II, III, or IV/V. Participants initiated ART a median of 3 d after diagnosis, and immunophenotyping was performed at diagnosis and longitudinally after ART. Anti-CMV Abs were assessed by ELISA. Samples from matched HIV-uninfected males were also analyzed. Proportions of adaptive NK cells (A-NKs; defined as Fcε-Receptor-1γ-) were expanded at HIV diagnosis at all Fiebig stages (pooled median 66% versus 25% for controls; p < 0.001) and were not altered by early ART initiation. Abs to CMV immediate early protein were elevated in PWH diagnosed in Fiebig stages III and IV/V (p < 0.03 for both). Proportions of A-NKs defined as either Fcε-Receptor-1γ- or NKG2C+/CD57+ were significantly associated with HIV DNA levels at diagnosis (p = 0.046 and 0.029, respectively) and trended toward an association after 48 wk of ART. Proportions of activated HLA-DR+/CD38+ NK cells remained elevated in PWH despite early ART initiation. NK cell activation and A-NK expansion occur very early after HIV transmission, before T cell activation, and are not altered by ART initiation during acute infection. A-NKs may contribute to HIV control and thus be useful for HIV cure.

Injecting drug use and hepatitis C virus infection independently increase biomarkers of inflammatory disease risk which are incompletely restored by curative direct-acting antiviral therapy.

Background: Hepatitis C virus (HCV) infections are more prevalent in people who inject drugs (PWID) who often experience additional health risks. HCV induces inflammation and immune alterations that contribute to hepatic and non-hepatic morbidities. It remains unclear whether curative direct acting antiviral (DAA) therapy completely reverses immune alterations in PWID. Methods: Plasma biomarkers of immune activation associated with chronic disease risk were measured in HCV-seronegative (n=24) and HCV RNA+ (n=32) PWID at baseline and longitudinally after DAA therapy. Adjusted generalised estimating equations were used to assess longitudinal changes in biomarker levels. Comparisons between community controls (n=29) and HCV-seronegative PWID were made using adjusted multiple regression modelling. Results: HCV-seronegative PWID exhibited significantly increased levels of inflammatory biomarkers including soluble (s) TNF-RII, IL-6, sCD14 and sCD163 and the diabetes index HbA1c as compared to community controls. CXCL10, sTNF-RII, vascular cell adhesion molecule-1 and lipopolysaccharide binding protein (LBP) were additionally elevated in PWID with viremic HCV infection as compared to HCV- PWID. Whilst curative DAA therapy reversed some biomarkers, others including LBP and sTNF-RII remained elevated 48 weeks after HCV cure. Conclusion: Elevated levels of inflammatory and chronic disease biomarkers in PWID suggest an increased risk of chronic morbidities such as diabetes and cardiovascular disease. HCV infection in PWID poses an additional disease burden, amplified by the incomplete reversal of immune dysfunction following DAA therapy. These findings highlight the need for heightened clinical surveillance of PWID for chronic inflammatory diseases, particularly those with a history of HCV infection.

Factors associated with the development of coronary artery disease in people with HIV.

BACKGROUND: People living with HIV (PLHIV) are at increased risk for coronary artery disease (CAD). This study aimed to describe the features associated with CAD in PLHIV. METHODS: A case ([n =160] PLHIV with CAD) control ([n =317] PLHIV matched by age and sex without CAD) study was performed at the Alfred Hospital, Melbourne, Australia (January 1996 and December 2018). Data collected included CAD risk factors, duration of HIV infection, nadir and at-event CD4+ T-cell counts, CD4:CD8 ratio, HIV viral load, and antiretroviral therapy exposure. RESULTS: Participants were predominantly male (n =465 [97.4%]), with a mean age of 53years. Traditional risk factors associated with CAD in univariate analysis included hypertension (OR 11.4 [95%CI 5.01, 26.33], P <0.001), current cigarette smoking (OR 2.5 [95% CI 1.22, 5.09], P =0.012), and lower high-density lipoprotein cholesterol (OR 0.14 [95%CI 0.05, 0.37], P <0.001). There was no association between duration of HIV infection, nadir or current CD4 cell count. However, current and ever exposure to abacavir (cases: 55 [34.4%]; controls: 79 [24.9%], P =0.023 and cases: 92 [57.5%]; controls: 154 [48.6%], P =0.048, respectively) was associated with CAD. In conditional logistic regression analysis, current abacavir use, current smoking, and hypertension remained significantly associated (aOR=1.87 [CI=1.14, 3.07], aOR=2.31 [1.32, 4.04], and aOR=10.30 [5.25, 20.20] respectively). CONCLUSION: Traditional cardiovascular risk factors and exposure to abacavir were associated with CAD in PLHIV. This study highlights that aggressive management of cardiovascular risk factors remains critical for reducing risk in PLHIV.

Regional Analysis of Intact and Defective HIV Proviruses in the Brain of Viremic and Virally Suppressed People with HIV.

Here, we provide the first regional analysis of intact and defective HIV reservoirs within the brain. Brain tissue from both viremic and virally suppressed people with HIV (PWH) harbored HIV pol DNA in all regions tested, with lower levels present in basal ganglia and cerebellum relative to frontal white matter. Intact proviruses were primarily found in the frontal white matter but also detected in other brain regions of PWH, demonstrating frontal white matter as a major brain reservoir of intact, potentially replication competent HIV DNA that persists despite antiretroviral therapy. ANN NEUROL 2023;94:798-802.

Favipiravir in early symptomatic COVID-19, a randomised placebo-controlled trial.

Background: Well tolerated antivirals administered early in the course of COVID-19 infection when the viremia is highest could prevent progression to severe disease. Favipiravir inhibits SARS-CoV-2 viral replication in vitro with evidence of clinical benefit in open label trials. Placebo controlled studies of people with early symptomatic COVID-19 with regular assessments of SARS-CoV-2 viral load can determine if it has an antiviral effect and improves clinical outcomes. Methods: People with PCR-confirmed COVID-19 and 5 days or less of symptoms were randomised 1:1 to favipiravir 1800 mg on day 1, then 800 mg twice daily or matched placebo for 14 days. SARS-CoV-2 viral load was quantitated from second daily self-collected nose-throat swabs while receiving study drug. The primary endpoint was time to virological cure defined as 2 successive swabs negative for SARS-CoV-2 by PCR and secondary outcomes were progression of disease severity, symptom resolution and safety. Findings: Between 31 July 2020 and 19 September 2021, 200 people were enrolled (199 in the community, 1 in hospital) with 190 receiving one or more doses of drug (modified intention to treat [mITT] population). There was no difference in time to virological cure (Log-rank p=0.6 comparing Kaplan Meier curves), progression to hospitalisation (14 favipiravir, 9 placebo; p=0.38), time to symptom resolution (cough, fever, sore throat) and there were no deaths. 51 people related an adverse event that was possibly drug related, but these were evenly distributed (n=24 favipiravir, n=27 placebo). Sensitivity analyses where the definition of virological cure was changed to: a single negative PCR, exclude datapoints based on the presence or absence of human DNA in the swab, a SARS-CoV-2 viral load < 300 copies/mL being considered negative all demonstrated no difference between arms. Interpretation: Favipiravir does not improve the time to virological cure or clinical outcomes and shows no evidence of an antiviral effect when treating early symptomatic COVID-19 infection. Funding: The study was supported in part by grants from the Commonwealth Bank Australia, the Lord Mayor's Charitable Foundation, Melbourne Australia and the Orloff Family Charitable Trust, Melbourne, Australia. JHM is supported by the Medical Research Future Fund, AYP, JT are supported by the Australian National Health and Medical Research Council.

The Impact of Over-The-Counter Lactic Acid Containing Vaginal Gels on the Integrity and Inflammatory State of the Vaginal Epithelium in vitro.

The vaginal microbiome influences a wide range of health outcomes in women, where a microbiome dominated by Lactobacillus spp. is considered optimal and associated with reduced risk of pre-term birth and acquisition of sexually transmitted infections including HIV. Conversely, replacement of lactobacilli by non-optimal bacteria leads to the development of bacterial vaginosis, which is associated with increased risk of these outcomes. Lactobacilli produce the metabolite lactic acid (LA) which is a potent antibacterial and antiviral agent. The potential therapeutic benefits of LA have prompted the development of numerous over-the-counter LA-containing gels for use in the vagina, although a comprehensive analysis of the impact of these formulations on the cervicovaginal epithelium and pro-inflammatory cytokine/chemokine responses, has not been assessed. Here, we evaluated the properties of 11 over-the-counter gels, including 9 containing LA, marketed for use in the vagina. Ten of the 11 gels had an osmolality greater than vaginal fluid from women with Lactobacillus-dominated microbiota (370 ± 40 mOsmol/kg in women with Nugent score 0-3), with six gels that were hyperosmolal >2,000 mOsmol/kg. Using a reconstructed primary cell model of the vaginal epithelium, we found hyperosmolal gels had a detrimental impact on epithelial barrier integrity, resulting in substantial cellular toxicity (<10% viability as compared to untreated cells) and reduced epithelial barrier integrity [≈30% of untreated cells, assessed by transepithelial electrical resistance (TEER)]. Treatment of vaginal tissues with most of the gels elicited the production of pro-inflammatory factors including IL-1α (8 of 11) and IL-1ß (10 of 11) which are associated with heightened risk of HIV acquisition in vivo. The majority of the OTC gels elicited moderate tissue damage as determined by histology. The detrimental effects of these gels on the human vaginal epithelium in vitro may predict compromised epithelial barrier integrity and genital inflammation in vivo, which has implications for sexual and reproductive health. This study highlights the importance of evaluating the impact of intravaginal products on the integrity and inflammatory status of the mucosal epithelium to avoid unfavorable off target effects.

Effects and safety of Chinese herbal medicine on inflammatory biomarkers in cardiovascular diseases: A systematic review and meta-analysis of randomized controlled trials.

Inflammation drives cardiovascular disease (CVD) in individuals with underlying chronic inflammatory diseases, including People with HIV (PWH), independently of dyslipidemia. Adjunctive treatments that lower inflammation may be useful to lower CVD risk in such populations. There is very little data on the efficacy of Chinese herbal medicine (CHM) in reducing inflammation in PWH to address its potential in reducing this CVD risk factor, therefore we evaluated its impact on inflammatory biomarkers relevant to CVD risk in the general population. Six English and Chinese databases were searched for studies investigating CHM's effects on inflammatory biomarkers relevant to CVD from respective inceptions to February 2022. A systematic review and meta-analysis of randomized controlled trials (RCTs) were conducted and the most-frequently prescribed herbs were identified. Thirty-eight RCTs involving 4,047 participants were included. Greater than or equal to 50% of included studies had a low risk of bias in five domains (random sequence generation, detection, attrition, reporting and other bias) and 97% had a high risk of performance bias. CHM provided significant additive effects on attenuating relevant inflammatory indices including hs-CRP (SMD -2.05, 95% CI -2.55 to -1.54), IL-6 (SMD -1.14, 95% CI -1.63 to -0.66) and TNF-α levels (SMD -0.88, 95% CI -1.35 to -0.41), but no significant effects on hs-CRP were found between CHM and placebo when co-treating with Western drugs (MD 0.04, 95% CI -1.66 to 1.74). No severe adverse events were reported in CHM groups. The two most prevalent herbs present in formulae demonstrating reduction of at least one inflammatory biomarker were Dan shen (Salviae Miltiorrhizae Radix et Rhizoma) and Huang qi (Astragali Radix). CHM, in combination with standard anti-inflammatory medications, may depress inflammation and reduce the risk of inflammatory conditions such as CVD. Rigorously-conducted trials and adequate reporting are needed to provide more robust evidence supporting the use of CHM to reduce CVD risk in people with underlying chronic inflammation such as PWH.

Lactic acid from vaginal microbiota enhances cervicovaginal epithelial barrier integrity by promoting tight junction protein expression.

BACKGROUND: Women with a cervicovaginal microbiota dominated by Lactobacillus spp. are at reduced risk of acquiring sexually transmitted infections including HIV, but the biological mechanisms involved remain poorly defined. Here, we performed metaproteomics on vaginal swab samples from young South African women (n = 113) and transcriptomics analysis of cervicovaginal epithelial cell cultures to examine the ability of lactic acid, a metabolite produced by cervicovaginal lactobacilli, to modulate genital epithelial barrier function. RESULTS: Compared to women with Lactobacillus-depleted microbiota, women dominated by vaginal lactobacilli exhibit higher abundance of bacterial lactate dehydrogenase, a key enzyme responsible for lactic acid production, which is independently associated with an increased abundance of epithelial barrier proteins. Physiological concentrations of lactic acid enhance epithelial cell culture barrier integrity and increase intercellular junctional molecule expression. CONCLUSIONS: These findings reveal a novel ability of vaginal lactic acid to enhance genital epithelial barrier integrity that may help prevent invasion by sexually transmitted pathogens. Video abstract.

Effect of Rosuvastatin Therapy on Biomarkers of Inflammation and Immune Activation in People With Human Immunodeficiency Virus at Intermediate Cardiovascular Risk.

BACKGROUND: Statins may help prevent cardiovascular disease (CVD) in people with human immunodeficiency virus (PWH) with chronic inflammation owing to their pleotropic lipid-lowering and anti-inflammatory properties. METHODS: The impact of 48 weeks of rosuvastatin therapy on inflammation and immune activation in a double-blind, placebo-controlled trial in PWH at moderate cardiovascular disease risk was assessed. RESULTS: Rosuvastatin did not alter plasma levels of interleukin 6, soluble tumor necrosis factor receptor type 2, CXCL10, soluble CD14, or soluble vascular cellular adhesion molecule 1 (P ≥ .1 for all). Proportions of CD16+ monocyte subsets were increased in PWH receiving rosuvastatin. CONCLUSIONS: The potential benefits of statin use in PWH with normal lipid levels requires further clinical outcome research.

Development of a Novel In Vitro Primary Human Monocyte-Derived Macrophage Model To Study Reactivation of HIV-1 Transcription.

Latent HIV reservoirs persist in people living with HIV despite effective antiretroviral therapy and contribute to rebound viremia upon treatment interruption. Macrophages are an important reservoir cell type, but analysis of agents that modulate latency in macrophages is limited by lack of appropriate in vitro models. We therefore generated an experimental system to investigate this by purifying nonproductively infected human monocyte-derived macrophages (MDM) following in vitro infection with an M-tropic enhanced green fluorescent protein reporter HIV clone and quantified activation of HIV transcription using live-cell fluorescence microscopy. The proportion of HIV-infected MDM was quantified by qPCR detection of HIV DNA, and GFP expression was validated as a marker of productive HIV infection by colabeling of HIV Gag protein. HIV transcription spontaneously reactivated in latently infected MDM at a rate of 0.22% ± 0.04% cells per day (mean ± the standard error of the mean, n = 10 independent donors), producing infectious virions able to infect heterologous T cells in coculture experiments, and both T cells and TZM-bl cells in a cell-free infection system using MDM culture supernatants. Polarization to an M1 phenotype with gamma interferon plus tumor necrosis factor resulted in a 2.3-fold decrease in initial HIV infection of MDM (P < 0.001, n = 8) and a 1.4-fold decrease in spontaneous reactivation (P = 0.025, n = 6), whereas M2 polarization using interleukin-4 prior to infection led to a 1.6-fold decrease in HIV infectivity (P = 0.028, n = 8) but a 2.0-fold increase in the rate of HIV reactivation in latently infected MDM (P = 0.023, n = 6). The latency reversing agents bryostatin and vorinostat, but not panobinostat, significantly induced HIV reactivation in latently infected MDM (P = 0.031 and P = 0.038, respectively, n = 6). IMPORTANCE Agents which modulate latent HIV reservoirs in infected cells are of considerable interest to HIV cure strategies. The present study characterizes a robust, reproducible model enabling quantification of HIV reactivation in primary HIV-infected human MDM which is relatively insensitive to the monocyte donor source and hence suitable for evaluating latency modifiers in MDM. The rate of initial viral infection was greater than the rate of HIV reactivation, suggesting that different mechanisms regulate these processes. HIV reactivation was sensitive to macrophage polarization, suggesting that cellular and tissue environments influence HIV reactivation in different macrophage populations. Importantly, latently infected MDM showed different susceptibilities to certain latency-reversing agents known to be effective in T cells, indicating that dedicated strategies may be required to target latently infected macrophage populations in vivo.

Differential expression of HIV envelope epitopes on the surface of HIV-Infected macrophages and CD4+ T cells.

HIV-infected macrophages contribute to persistence of HIV reservoirs in people living with HIV receiving antiretroviral therapy. A potential strategy to eliminate reservoirs is the use of antibody-dependent cellular cytotoxicity (ADCC) against infected cells expressing the HIV envelope (Env) protein on their surface. Designing ADCC strategies requires knowledge of exposed Env epitopes on the cell surface and identifying antibodies capable of opsonising infected cells, yet little is known regarding the ability of HIV-infected macrophages to be targeted with such strategies. Using a panel of neutralising and poorly-neutralising anti-Env antibodies we compared Env epitopes expressed on infected monocyte-derived macrophages (MDM) and autologous T cells. Our results reveal potential differences in epitope expression on macrophage- and T cell-expressed Env. Notably, HIVBaL-infected macrophages were more susceptible to opsonisation by NIH45-46 (median = 40.4%) compared to infected T cells (13.6%; p = 0.002), which were more susceptible to opsonisation by 17b and 447.52D (88.6% and 45.6% respectively) compared to MDM (30% and 6.7%, p = 0.002 and 0.004 respectively). Furthermore, neutralising antibodies 10E8 and PGT145 were relatively ineffective at opsonising Env expressed on the surface of infected T cells or macrophages, indicating that the context in which Env is presented on infected cells may differ to that of cell-free virions.

HERV-K Gag RNA and Protein Levels Are Elevated in Malignant Regions of the Prostate in Males with Prostate Cancer.

Heightened expression of human endogenous retrovirus (HERV) sequences has been associated with a range of malignancies, including prostate cancer, suggesting that they may serve as useful diagnostic or prognostic cancer biomarkers. We analysed the expression of HERV-K (Gag and Env/Np9 regions), HERV-E 4.1 (Pol and Env regions), HERV-H (Pol) and HERV-W (Gag) sequences in prostate cancer cells lines and normal prostate epithelial cells using qRT-PCR. HERV expression was also analysed in matched malignant and benign prostate tissue samples from men with prostate cancer (n = 27, median age 65.2 years (range 47-70)) and compared to prostate cancer-free male controls (n = 11). Prostate cancer epithelial cell lines exhibited a signature of HERV RNA overexpression, with all HERVs analysed, except HERV-E Pol, showing heightened expression in at least two, but more commonly all, cell lines analysed. Analysis of primary prostate material indicated increased expression of HERV-E Pol but decreased expression of HERV-E Env in both malignant and benign regions of the prostate in men with prostate cancer as compared to those without. Expression of HERV-K Gag was significantly higher in malignant regions of the prostate in men with prostate cancer as compared to matched benign regions and prostate cancer-free men (p < 0.001 for both), with 85.2% of prostate cancers donors showing malignancy-associated upregulation of HERV-K Gag RNA. HERV-K Gag protein was detected in 12/18 (66.7%) malignant tissues using immunohistochemistry, but only 1/18 (5.6%) benign tissue sections. Heightened expression of HERV-K Gag RNA and protein appears to be a sensitive and specific biomarker of prostate malignancy in this cohort of men with prostate carcinoma, supporting its potential utility as a non-invasive, adjunct clinical biomarker.

Monocytes from men living with HIV exhibit heightened atherogenic potential despite long-term viral suppression with antiretroviral therapy.

OBJECTIVE: People living with HIV have an increased risk of cardiovascular disease (CVD) despite effective antiretroviral therapy (ART). Monocytes play a key role in the early stages of atherosclerosis-driven CVD by forming lipid-laden foam cells within artery walls. HIV infection potentiates foam cell formation ex vivo, but the mechanisms contributing to this are not known. METHODS: We investigated the atherosclerosis-promoting potential of monocytes from 39 virologically suppressed men living with HIV (MLHIV) on ART and no evidence of CVD, and 25 HIV-uninfected controls of comparable age, sex, smoking status and CVD risk. RESULTS: Despite absence of clinical atherosclerosis in both MLHIV and uninfected cohorts (evidenced by a carotid intima-media thickness of 0.6 mm for both groups; P = 0.254), monocytes from MLHIV showed increased potential to form atherosclerosis-promoting foam cells compared with controls in an ex-vivo assay (36.6% vs. 27.6%, respectively, P = 0.003). Consistent with observations of persistent inflammation and immune/endothelial activation in ART-treated HIV infection, levels of soluble tumour necrosis factor receptor II, CXCL10 and soluble VCAM-1 were elevated in MLHIV (P ≤ 0.005 for all), but were not significantly associated with foam cell formation. Foam cell formation was associated with an impaired ability of monocytes to undergo reverse transmigration, and a reduced ability to efflux cholesterol ex vivo (P < 0.05 for both). Importantly, foam cell formation declined significantly with duration of viral suppression (P = 0.004). CONCLUSION: These findings highlight the persistence of HIV-related changes to the atherogenic potential of monocytes despite long-term viral suppression, and provide insights into mechanisms potentially driving increased CVD in ART-treated HIV infection.

Corrigendum: The HIV Reservoir in Monocytes and Macrophages.

[This corrects the article DOI: 10.3389/fimmu.2019.01435.].

How Monocytes Contribute to Increased Risk of Atherosclerosis in Virologically-Suppressed HIV-Positive Individuals Receiving Combination Antiretroviral Therapy.

Combination antiretroviral therapy (ART) is effective at suppressing HIV viremia to achieve persistently undetectable levels in peripheral blood in the majority of individuals with access and ability to maintain adherence to treatment. However, evidence suggests that ART is less effective at eliminating HIV-associated inflammation and innate immune activation. To the extent that residual inflammation and immune activation persist, virologically suppressed people living with HIV (PLWH) may have increased risk of inflammatory co-morbidities, and adjunctive therapies may need to be considered to reduce HIV-related inflammation and fully restore the health of virologically suppressed HIV+ individuals. Cardiovascular disease (CVD) is the single leading cause of death in the developed world and is becoming more important in PLWH with access to ART. Arterial disease due to atherosclerosis, leading to acute myocardial infarction (AMI) and stroke, is a major component of CVD. Atherosclerosis is an inflammatory disease, and epidemiological comparisons of atherosclerosis and AMI show a higher prevalence and suggest a greater risk in PLWH compared to the general population. The reasons for greater prevalence of CVD in PLWH can be broadly grouped into four categories: (a) the higher prevalence of traditional risk factors e.g., smoking and hypertension (b) dyslipidemia (also a traditional risk factor) caused by off-target effects of ART drugs (c) HIV-related inflammation and immune activation and (d) other undefined HIV-related factors. Management strategies aimed at reducing the impact of traditional risk factors in PLWH are similar to those for the general population and their effectiveness is currently being evaluated. Together with improvements in ART regimens and guidelines for treatment, and a greater awareness of its impact on CVD, the HIV-related risk of AMI and stroke is decreasing but remains elevated compared to the general community. Monocytes are key effector cells which initiate the formation of atherosclerotic plaques by migrating into the intima of coronary arteries and accumulating as foam cells full of lipid droplets. This review considers the specific role of monocytes as effector cells in atherosclerosis which progresses to AMI and stroke, and explores mechanisms by which HIV may promote an atherogenic phenotype and function independent of traditional risk factors. Altered monocyte function may represent a distinct HIV-related factor which increases risk of CVD in PLWH.

The HIV Reservoir in Monocytes and Macrophages.

In people living with HIV (PLWH) who are failing or unable to access combination antiretroviral therapy (cART), monocytes and macrophages are important drivers of pathogenesis and progression to AIDS. The relevance of the monocyte/macrophage reservoir in PLWH receiving cART is debatable as in vivo evidence for infected cells is limited and suggests the reservoir is small. Macrophages were assumed to have a moderate life span and lack self-renewing potential, but recent discoveries challenge this dogma and suggest a potentially important role of these cells as long-lived HIV reservoirs. This, combined with new HIV infection animal models, has led to a resurgence of interest in monocyte/macrophage reservoirs. Infection of non-human primates with myeloid-tropic SIV implicates monocyte/macrophage activation and infection in the brain with neurocognitive disorders, and infection of myeloid-only humanized mouse models are consistent with the potential of the monocyte/macrophage reservoir to sustain infection and be a source of rebound viremia following cART cessation. An increased resistance to HIV-induced cytopathic effects and a reduced susceptibility to some antiretroviral drugs implies macrophages may be relevant to residual replication under cART and to rebound viremia. With a reappraisal of monocyte circulation dynamics, and the development of techniques to differentiate between self-renewing tissue-resident, and monocyte-derived macrophages in different tissues, a new framework exists to contextualize and evaluate the significance and relevance of the monocyte/macrophage HIV reservoir. In this review, we discuss recent developments in monocyte and macrophage biology and appraise current and emerging techniques to quantify the reservoir. We discuss how this knowledge influences our evaluation of the myeloid HIV reservoir, the implications for HIV pathogenesis in both viremic and virologically-suppressed PLWH and the need to address the myeloid reservoir in future treatment and cure strategies.

Molecular dissection of an inhibitor targeting the HIV integrase dependent preintegration complex nuclear import.

Human immunodeficiency virus (HIV) continues to be a major contributor to morbidity and mortality worldwide, particularly in developing nations where high cost and logistical issues severely limit the use of current HIV therapeutics. This, combined HIV's high propensity to develop resistance, means that new antiviral agents against novel targets are still urgently required. We previously identified novel anti-HIV agents directed against the nuclear import of the HIV integrase (IN) protein, which plays critical roles in the HIV lifecycle inside the cell nucleus, as well as in transporting the HIV preintegration complex (PIC) into the nucleus. Here we investigate the structure activity relationship of a series of these compounds for the first time, including a newly identified anti-IN compound, budesonide, showing that the extent of binding to the IN core domain correlates directly with the ability of the compound to inhibit IN nuclear transport in a permeabilised cell system. Importantly, compounds that inhibited the nuclear transport of IN were found to significantly decrease HIV viral replication, even in a dividing cell system. Significantly, budesonide or its analogue flunisolide, were able to effect a significant reduction in the presence of specific nuclear forms of the HIV DNA (2-LTR circles), suggesting that the inhibitors work though blocking IN, and potentially PIC, nuclear import. The work presented here represents a platform for further development of these specific inhibitors of HIV replication with therapeutic and prophylactic potential.

Distinct Immune Responses Elicited From Cervicovaginal Epithelial Cells by Lactic Acid and Short Chain Fatty Acids Associated With Optimal and Non-optimal Vaginal Microbiota.

Non-optimal vaginal microbiota, as observed in bacterial vaginosis (BV), is typically characterized by a depletion of beneficial lactobacilli and an abundance of numerous anaerobes. These non-optimal conditions are associated with subclinical cervicovaginal inflammation and an increased risk of HIV infection compared to women colonized with optimal vaginal microbiota dominated by lactobacilli. Lactic acid (LA) is a major organic acid metabolite produced by vaginal lactobacilli that elicits anti-inflammatory effects from cervicovaginal epithelial cells and is dramatically depleted during BV. However, it is unclear if LA retains its anti-inflammatory activity in the presence of vaginal microbiota metabolites comprising short chain fatty acids (SCFAs) and succinic acid, which are also produced by an optimal vaginal microbiota. Furthermore, the immunomodulatory effect of SCFAs and succinic acid on cervicovaginal epithelial cells at higher concentrations present during BV is unknown. Here we report that in the presence of physiologically relevant concentrations of SCFAs and succinic acid at pH 3.9 (as found in women with lactobacillus-dominated microbiota) LA induced an anti-inflammatory state in cervicovaginal epithelial cells and inhibited inflammation elicited by the toll-like receptor (TLR) agonists polyinosinic:polycytidylic acid and Pam3CSK4. When cervicovaginal epithelial cells were treated with a vaginal microbiota metabolite mixture representative of BV, containing a lower concentration of LA but higher concentrations of SCFA/succinic acid at pH 7, no anti-inflammatory was observed. Rather, the vaginal microbiota metabolite mixture representative of BV dysregulated the immune response of cervicovaginal epithelial cells during prolonged and sustained treatments. This was evidenced by increased basal and TLR-induced production of pro-inflammatory cytokines including tumor necrosis factor-α, but decreased basal production of chemokines including RANTES and IP-10. Further characterization of individual components of the BV vaginal microbiota mixture suggested that acetic acid is an important vaginal microbiota metabolite capable of eliciting diverse immunomodulatory effects on a range of cervicovaginal epithelial cell targets. These findings indicate that elevated levels of SCFAs are a potential source of cervicovaginal inflammation in women experiencing BV, and support the unique anti-inflammatory properties of LA on cervicovaginal epithelial cells as well as a role for LA or LA-producing lactobacilli to reverse genital inflammation associated with increased HIV risk.