CXCR3 and CXCR5 are highly expressed in HIV-1-specific CD8 central memory T cells from infected patients.

New ways of characterizing CD8+ memory T cell responses in chronic infections are based on the measurement of chemokine receptor expression (CXCR3, CXCR5, and CX3CR1). We applied these novel phenotyping strategies to chronic HIV infection by comparing healthy donors (HDs), HIV-infected patients receiving antiretroviral therapy (ART), and spontaneous HIV controllers (HICs). In all groups, the memory cells exhibited high proportion of CXCR3+ cells. Proportions of CXCR5+ and CX3CR1+ cells were preferentially observed among central memory cells (Tcm) and effector memory cells (Tem) respectively. Chronic controlled HIV infection impacted the chemokine receptor profile of both HIV-specific and nonspecific CD8+ T cells. In total CD8+ T cells, the proportions of CXCR3- CXCR5- CX3CR1- Tcm and Tem were lower in HIV-infected patients than in HDs with subtle differences between ART and HICs. Such phenotyping strategy also revealed differences in exhaustion and senescence phenotypes, the CXCR3+ CXCR5+ CX3CR1- being more exhausted and senescent than the CXCR3+ CXCR5- CX3CR1- Tcm fraction. Among HIV-specific CD8+ T cells, the vast majority of Tcm cells were CXCR3+ and CXCR5+ cells in contrast with their nonspecific counterparts. In conclusion, the addition of migration markers contributes to better characterize Tcm/Tem compartment.

Pharmacokinetics and tissue distribution of tenofovir, emtricitabine and dolutegravir in mice.

BACKGROUND: Studies of antiretroviral drug (ARV) tissue distribution in preclinical models, such as mice, are key to understanding viral persistence. OBJECTIVES: To determine the plasma and tissue pharmacokinetics and tissue distributions of tenofovir, emtricitabine and dolutegravir in mice. METHODS: ARVs were simultaneously administered to two different strains, and their levels in plasma and tissue samples were determined by a validated LC-MS/MS method. A non-compartmental analysis was performed to estimate the main pharmacokinetic parameters. A tissue penetration factor (TPF) was calculated as the ratio of the concentration in the tissue concerned to that in plasma. RESULTS: ARV plasma pharmacokinetic parameters in both strains were similar to those estimated in the clinical context. Tissue concentrations were highest in the digestive tract, followed by the liver and kidneys, lymphatic system, pancreas, adipose tissue and lungs. Tissue concentrations were lowest in the brain. Triple therapy could not be considered effective in any of the tissues considered. The TPF values obtained showed that tenofovir diffused widely, especially in the digestive tract, liver and kidneys. Emtricitabine had a TPF above 100% in two-thirds of the tissues. Dolutegravir was poorly distributed to all tissues. CONCLUSIONS: Drug specificity was observed, with higher levels of exposure to tenofovir than to emtricitabine or dolutegravir. Tissue specificity was also observed, with strong penetration of the digestive tract and weak penetration of the brain. These data have important implications for future preclinical and clinical studies for developing new HIV therapies with the goal of an HIV cure.

The Integrase Inhibitors Dolutegravir and Raltegravir Exert Proadipogenic and Profibrotic Effects and Induce Insulin Resistance in Human/Simian Adipose Tissue and Human Adipocytes.

BACKGROUND: Although some integrase strand transfer inhibitors (INSTIs) promote peripheral and central adipose tissue/weight gain in people with human immunodeficiency virus (PHIV), the underlying mechanism has not been identified. Here, we used human and simian models to assess the impact of INSTIs on adipose tissue phenotype and function. METHODS: Adipocyte size and fibrosis were determined in biopsies of subcutaneous and visceral adipose tissue (SCAT and VAT, respectively) from 14 noninfected macaques and 19 PHIV treated or not treated with an INSTI. Fibrosis, adipogenesis, oxidative stress, mitochondrial function, and insulin sensitivity were assessed in human proliferating or adipocyte-differentiated adipose stem cells after long-term exposure to dolutegravir or raltegravir. RESULTS: We observed elevated fibrosis, adipocyte size, and adipogenic marker expression in SCAT and VAT from INSTI-treated noninfected macaques. Adiponectin expression was low in SCAT. Accordingly, SCAT and VAT samples from INSTI-exposed patients displayed higher levels of fibrosis than those from nonexposed patients. In vitro, dolutegravir and, to a lesser extent, raltegravir were associated with greater extracellular matrix production and lipid accumulation in adipose stem cells and/or adipocytes as observed in vivo. Despite the INSTIs' proadipogenic and prolipogenic effects, these drugs promoted oxidative stress, mitochondrial dysfunction, and insulin resistance. CONCLUSIONS: Dolutegravir and raltegravir can directly impact adipocytes and adipose tissue. These INSTIs induced adipogenesis, lipogenesis, oxidative stress, fibrosis, and insulin resistance. The present study is the first to shed light on the fat modifications observed in INSTI-treated PHIV.

Early SIV and HIV infection promotes the LILRB2/MHC-I inhibitory axis in cDCs.

Classical dendritic cells (cDCs) play a pivotal role in the early events that tip the immune response toward persistence or viral control. In vitro studies indicate that HIV infection induces the dysregulation of cDCs through binding of the LILRB2 inhibitory receptor to its MHC-I ligands and the strength of this interaction was proposed to drive disease progression. However, the dynamics of the LILRB2/MHC-I inhibitory axis in cDCs during early immune responses against HIV are yet unknown. Here, we show that early HIV-1 infection induces a strong and simultaneous increase of LILRB2 and MHC-I expression on the surface of blood cDCs. We further characterized the early dynamics of LILRB2 and MHC-I expression by showing that SIVmac251 infection of macaques promotes coordinated up-regulation of LILRB2 and MHC-I on cDCs and monocytes/macrophages, from blood and lymph nodes. Orientation towards the LILRB2/MHC-I inhibitory axis starts from the first days of infection and is transiently induced in the entire cDC population in acute phase. Analysis of the factors involved indicates that HIV-1 replication, TLR7/8 triggering, and treatment by IL-10 or type I IFNs increase LILRB2 expression. Finally, enhancement of the LILRB2/MHC-I inhibitory axis is specific to HIV-1 and SIVmac251 infections, as expression of LILRB2 on cDCs decreased in naturally controlled chikungunya virus infection of macaques. Altogether, our data reveal a unique up-regulation of LILRB2 and its MHC-I ligands on cDCs in the early phase of SIV/HIV infection, which may account for immune dysregulation at a critical stage of the anti-viral response.

High proportion of PD-1-expressing CD4+ T cells in adipose tissue constitutes an immunomodulatory microenvironment that may support HIV persistence.

We and others have demonstrated that adipose tissue is a reservoir for HIV. Evaluation of the mechanisms responsible for viral persistence may lead to ways of reducing these reservoirs. Here, we evaluated the immune characteristics of adipose tissue in HIV-infected patients receiving antiretroviral therapy (ART) and in non-HIV-infected patients. We notably sought to determine whether adipose tissue's intrinsic properties and/or HIV induced alteration of the tissue environment may favour viral persistence. ART-controlled HIV infection was associated with a difference in the CD4/CD8 T-cell ratio and an elevated proportion of Treg cells in subcutaneous adipose tissue. No changes in Th1, Th2 and Th17 cell proportions or activation markers expression on T cell (Ki-67, HLA-DR) could be detected, and the percentage of CD69-expressing resident memory CD4+ T cells was not affected. Overall, our results indicate that adipose-tissue-resident CD4+ T cells are not extensively activated during HIV infection. PD-1 was expressed by a high proportion of tissue-resident memory CD4+ T cells in both HIV-infected patients and non-HIV-infected patients. Our findings suggest that adipose tissue's intrinsic immunomodulatory properties may limit immune activation and thus may strongly contribute to viral persistence.

Adipose Tissue Is a Neglected Viral Reservoir and an Inflammatory Site during Chronic HIV and SIV Infection.

Two of the crucial aspects of human immunodeficiency virus (HIV) infection are (i) viral persistence in reservoirs (precluding viral eradication) and (ii) chronic inflammation (directly associated with all-cause morbidities in antiretroviral therapy (ART)-controlled HIV-infected patients). The objective of the present study was to assess the potential involvement of adipose tissue in these two aspects. Adipose tissue is composed of adipocytes and the stromal vascular fraction (SVF); the latter comprises immune cells such as CD4+ T cells and macrophages (both of which are important target cells for HIV). The inflammatory potential of adipose tissue has been extensively described in the context of obesity. During HIV infection, the inflammatory profile of adipose tissue has been revealed by the occurrence of lipodystrophies (primarily related to ART). Data on the impact of HIV on the SVF (especially in individuals not receiving ART) are scarce. We first analyzed the impact of simian immunodeficiency virus (SIV) infection on abdominal subcutaneous and visceral adipose tissues in SIVmac251 infected macaques and found that both adipocytes and adipose tissue immune cells were affected. The adipocyte density was elevated, and adipose tissue immune cells presented enhanced immune activation and/or inflammatory profiles. We detected cell-associated SIV DNA and RNA in the SVF and in sorted CD4+ T cells and macrophages from adipose tissue. We demonstrated that SVF cells (including CD4+ T cells) are infected in ART-controlled HIV-infected patients. Importantly, the production of HIV RNA was detected by in situ hybridization, and after the in vitro reactivation of sorted CD4+ T cells from adipose tissue. We thus identified adipose tissue as a crucial cofactor in both viral persistence and chronic immune activation/inflammation during HIV infection. These observations open up new therapeutic strategies for limiting the size of the viral reservoir and decreasing low-grade chronic inflammation via the modulation of adipose tissue-related pathways.

High eomesodermin expression among CD57+ CD8+ T cells identifies a CD8+ T cell subset associated with viral control during chronic human immunodeficiency virus infection.

During HIV infection, increased CD57 expression among CD8(+) T cells has been associated with immune senescence and defective immune responses. Interestingly, CD57-expressing CD8(+) T cells exhibit a dual profile, being simultaneously highly cytotoxic (terminally differentiated effectors) and poorly proliferative (replicative senescent). Recent publications point toward a positive role of CD57-expressing CD8(+) T cell subsets, presumably due to their high cytolytic activity. We further investigated the phenotype of CD57-expressing CD8(+) T cells in healthy donors and during HIV infection combining CD57 expression to Eomesodermin (EOMES), a T box transcription factor which determines, coordinately with T-bet, effector and memory CD8(+) T cell differentiation. We defined in healthy donors two functionally distinct CD57-expressing CD8(+) T cell subsets exhibiting different levels of EOMES expression: EOMES(hi) CD57(+) and EOMES(int) CD57(+) CD8(+) T cells. EOMES(hi) CD57(+) cells exhibited low cytotoxic activity but preserved proliferative capacity and interleukin 7 (IL-7) receptor expression, whereas EOMES(int) CD57(+) cells exhibited obvious cytotoxic functions and a more terminally differentiated phenotype. We next performed a similar analysis in different contexts of HIV infection: primary infected patients, long-term viremic patients, aviremic patients treated with antiretroviral therapy, and HIV controllers; we demonstrated a higher percentage of CD57-expressing cells in all HIV-infected patients regardless of virological status. When heterogeneity in EOMES expression among CD57 cells was taken into account, we detected significantly higher proportions of EOMES(hi) CD57(+) cells among HIV-specific and nonspecific CD8(+) T cells from HIV controllers than in aviremic antiretroviral-treated patients and viremic patients. Importantly, such a peculiar non-terminally differentiated EOMES(hi) CD57(+) phenotypic profile was associated with viral control. Importance: This study demonstrates that functional heterogeneity exists among CD57-expressing CD8 T cells, which include both terminally differentiated, highly cytotoxic EOMES(int) CD57(+) CD8(+) T cells and less differentiated EOMES(hi) CD57(+) CD8 T cells, which do not exhibit immediate cytotoxic functions but present high proliferative capacity. Interestingly, HIV controllers present a high proportion of EOMES(hi) CD57 cells among CD57-expressing HIV-specific CD8 T cells compared to both long-term viremic and aviremic antiretroviral therapy (ART)-treated patients, suggesting a beneficial role for this cell subset in viral control.

Paradoxical resistance to high-fat diet-induced obesity and altered macrophage polarization in mineralocorticoid receptor-overexpressing mice.

The mineralocorticoid receptor (MR) exerts proadipogenic and antithermogenic effects in vitro, yet its in vivo metabolic impact remains elusive. Wild type (WT) and transgenic (Tg) mice overexpressing human MR were subjected to standard chow (SC) or high-fat diet (HFD) for 16 wk. Tg mice had a lower body weight gain than WT animals and exhibited a relative resistance to HFD-induced obesity. This was associated with a decrease in fat mass, an increased population of smaller adipocytes, and an improved glucose tolerance compared with WT animals. Quantitative RT-PCR studies revealed decreased expression of PPARγ2, a master adipogenic gene, and of glucocorticoid receptor and 11ß-hydroxysteroid dehydrogenase type 1, consistent with an impaired local glucocorticoid signaling in adipose tissues (AT). This paradoxical resistance to HFD-induced obesity was not related to an adipogenesis defect since differentiation capacity of Tg preadipocytes isolated from stroma-vascular fractions was unaltered, suggesting that other nonadipocyte factors might compromise AT development. Although AT macrophage infiltration was not different between genotypes, Tg mice exhibited a distinct macrophage polarization, as revealed by FACS analysis and CD11c/CD206 expression studies. We further demonstrated that Tg macrophage-conditioned medium partially impaired preadipocyte differentiation. Therefore, we propose that modification of M1/M2 polarization of hMR-overexpressing macrophages could account in part for the metabolic phenotype of Tg mice. Collectively, our results provide evidence that MR exerts a pivotal immunometabolic role by controlling adipocyte differentiation processes directly but also indirectly through macrophage polarization regulation. Our findings should be taken into account for the pharmacological treatment of metabolic disorders.

Ageing combines CD4 T cell lymphopenia in secondary lymphoid organs and T cell accumulation in gut associated lymphoid tissue.

BACKGROUND: CD4 T cell lymphopenia is an important T cell defect associated to ageing. Higher susceptibility to infections, cancer, or autoimmune pathologies described in aged individuals is thought to partly rely on T cell lymphopenia. We hypothesize that such diverse effects may reflect anatomical heterogeneity of age related T cell lymphopenia. Indeed, no data are currently available on the impact of ageing on T cell pool recovered from gut associated lymphoid tissue (GALT), a crucial site of CD4 T cell accumulation. RESULTS: Primary, secondary and tertiary lymphoid organs of C57BL/6 animals were analysed at three intervals of ages: 2 to 6 months (young), 10 to 14 months (middle-aged) and 22 to 26 months (old). We confirmed that ageing preferentially impacted CD4 T cell compartment in secondary lymphoid organs. Importantly, a different picture emerged from gut associated mucosal sites: during ageing, CD4 T cell accumulation was progressively developing in colon and small intestine lamina propria and Peyer's patches. Similar trend was also observed in middle-aged SJL/B6 F1 mice. Interestingly, an inverse correlation was detected between CD4 T cell numbers in secondary lymphoid organs and colonic lamina propria of C57BL/6 mice whereas no increase in proliferation rate of GALT CD4 T cells was detected. In contrast to GALT, no CD4 T cell accumulation was detected in lungs and liver in middle-aged animals. Finally, the concomitant accumulation of CD4 T cell in GALT and depletion in secondary lymphoid organs during ageing was detected both in male and female animals. CONCLUSIONS: Our data thus demonstrate that T cell lymphopenia in secondary lymphoid organs currently associated to ageing is not sustained in gut or lung mucosa associated lymphoid tissues or non-lymphoid sites such as the liver. The inverse correlation between CD4 T cell numbers in secondary lymphoid organs and colonic lamina propria and the absence of overt proliferation in GALT suggest that marked CD4 T cell decay in secondary lymphoid organs during ageing reflect redistribution of CD4 T cells rather than generalized CD4 T cell decay. Such anatomical heterogeneity may provide an important rationale for the diversity of immune defects observed during ageing.

Identification of macaque dendritic cell precursors in blood and tissue reveals their dysregulation in early SIV infection.

Distinct dendritic cell (DC) subsets play important roles in shaping immune responses. Circulating DC precursors (pre-DCs) are more susceptible to HIV infection in vitro, which may explain the inefficiency of immune responses against HIV. However, the interplay between HIV and pre-DC is not defined in vivo. We identify human pre-DC equivalents in the cynomolgus macaque and then analyze their dynamics during simian immunodeficiency virus (SIV) infection to illustrate a sharp decrease of blood pre-DCs in early SIV infection and accumulation in lymph nodes (LNs), where they neglect to upregulate CD83/CD86 or MHC-II. Additionally, SIV infection attenuates the capacity of stimulated LN pre-DCs to produce IL-12p40. Analysis of HIV cohorts provides correlation between costimulatory molecule expression on pre-DCs and T cell activation in spontaneous HIV controllers. These findings pinpoint certain dynamics and functional changes of pre-DCs during SIV infection, providing a deeper understanding of immune dysregulation mechanisms elicited in people living with HIV.

Early and long-lasting alteration of effector CD45RA(-)Foxp3(high) regulatory T-cell homeostasis during HIV infection.

Regulatory T-cell (Treg) quantification in human immunodeficiency virus (HIV) infection remains ill defined because of the lack of reliable specific markers to identify human Tregs and the diversity of clinical stages of HIV infection. Using a recently described Treg identification strategy based on CD45RA and Foxp3 expression, we performed an extensive quantification of total, naive (CD45RA(+)Foxp3(low)), and effector (CD45RA(-)Foxp3(high)) Tregs in different contexts of HIV infection: primary HIV infection, long-term viremic patients, aviremic patients treated with highly active antiretroviral therapy, and HIV controllers. We showed that although total Treg percentages were mildly affected by HIV infection, Treg absolute numbers were significantly reduced in all groups studied. We demonstrated that although naive Treg numbers were essentially preserved, effector Tregs were consistently affected during HIV infection. Finally, we demonstrated that effector but not total or naive Treg numbers were negatively correlated with the magnitude of HIV-specific CD8 T-cell responses.

Expansion of Immature Neutrophils During SIV Infection Is Associated With Their Capacity to Modulate T-Cell Function.

In spite of the efficacy of combinational antiretroviral treatment (cART), HIV-1 persists in the host and infection is associated with chronic inflammation, leading to an increased risk of comorbidities, such as cardiovascular diseases, neurocognitive disorders, and cancer. Myeloid cells, mainly monocytes and macrophages, have been shown to be involved in the immune activation observed in HIV-1 infection. However, less attention has been paid to neutrophils, the most abundant circulating myeloid cell, even though neutrophils are strongly involved in tissue damage and inflammation in several chronic diseases, in particular, autoimmune diseases. Herein, we performed a longitudinal characterization of neutrophil phenotype and we evaluated the interplay between neutrophils and T cells in the model of pathogenic SIVmac251 experimental infection of cynomolgus macaques. We report that circulating granulocytes consists mainly of immature CD10- neutrophils exhibiting a prime phenotype during primary and chronic infection. We found that neutrophil priming correlates with CD8+ T cell activation. Moreover, we provide the evidence that neutrophils are capable of modulating CD4+ and CD8+ T-cell proliferation and IFN-γ production in different ways depending on the time of infection. Thus, our study emphasizes the role of primed immature neutrophils in the modulation of T-cell responses in SIV infection.

Detection of SARS-CoV-2 in subcutaneous fat but not visceral fat, and the disruption of fat lymphocyte homeostasis in both fat tissues in the macaque.

The well documented association between obesity and the severity of SARS-CoV-2 infection raises the question of whether adipose tissue (AT) is impacted during this infection. Using a model of SARS-CoV-2 infection in cynomolgus macaques, we detected the virus within subcutaneous AT (SCAT) but not in visceral AT (VAT) or epicardial AT on day 7 post-infection. We sought to determine the mechanisms responsible for this selective detection and observed higher levels of angiotensin-converting-enzyme-2 mRNA expression in SCAT than in VAT. Lastly, we evaluated the immunological consequences of SARS-CoV-2 infection on AT: both SCAT and VAT T cells showed a drastic reduction in CD69 expression, a standard marker of resident memory T cell in tissue, that is also involved in the migratory and metabolic properties of T cells. Our results demonstrate that in a model of mild infection, SCAT is selectively infected by SARS-CoV-2 although changes in the immune properties of AT are observed in both SCAT and VAT.

Reprogramming dysfunctional CD8+ T cells to promote properties associated with natural HIV control.

Virus-specific CD8+ T cells play a central role in HIV-1 natural controllers to maintain suppressed viremia in the absence of antiretroviral therapy. These cells display a memory program that confers them stemness properties, high survival, polyfunctionality, proliferative capacity, metabolic plasticity, and antiviral potential. The development and maintenance of such qualities by memory CD8+ T cells appear crucial to achieving natural HIV-1 control. Here, we show that targeting the signaling pathways Wnt/transcription factor T cell factor 1 (Wnt/TCF-1) and mTORC through GSK3 inhibition to reprogram HIV-specific CD8+ T cells from noncontrollers promoted functional capacities associated with natural control of infection. Features of such reprogrammed cells included enrichment in TCF-1+ less-differentiated subsets, a superior response to antigen, enhanced survival, polyfunctionality, metabolic plasticity, less mTORC1 dependency, an improved response to γ-chain cytokines, and a stronger HIV-suppressive capacity. Thus, such CD8+ T cell reprogramming, combined with other available immunomodulators, might represent a promising strategy for adoptive cell therapy in the search for an HIV-1 cure.

Prolonged Antiretroviral Treatment Induces Adipose Tissue Remodelling Associated with Mild Inflammation in SIV-Infected Macaques.

During chronic SIV/HIV infection, adipose tissue (AT) is the target of both antiretroviral treatment (ART) and the virus. AT might subsequently contribute to the low-grade systemic inflammation observed in patients on ART. To evaluate the inflammatory profile of AT during chronic SIV/HIV infection, we assayed subcutaneous and visceral abdominal AT from non-infected (SIV-, control), ART-naïve SIV-infected (SIV+) and ART-controlled SIV-infected (SIV+ART+) cynomolgus macaques for the mRNA expression of genes coding for factors related to inflammation. Significant differences were observed only when comparing the SIV+ART+ group with the SIV+ and/or SIV- groups. ART-treated infection impacted the metabolic fraction (with elevated expression of PPARγ and CEBPα), the extracellular matrix (with elevated expression of COL1A2 and HIF-1α), and the inflammatory profile. Both pro- and anti-inflammatory signatures were detected in AT, with greater mRNA expression of anti-inflammatory markers (adiponectin and CD163) and markers associated with inflammation (TNF-α, Mx1, CCL5 and CX3CL1). There were no intergroup differences in other markers (IL-6 and MCP-1). In conclusion, we observed marked differences in the immune and metabolic profiles of AT in the context of an ART-treated, chronic SIV infection; these differences were related more to ART than to SIV infection per se.

CIITA promoter polymorphism impairs monocytes HLA-DR expression in patients with septic shock.

Low monocyte (m)HLA-DR expression is associated with mortality in sepsis. G-286A∗rs3087456 polymorphism in promoter III of HLA class II transactivator (CIITA), the master regulator of HLA, has been associated with autoimmune diseases but its role in sepsis has never been demonstrated. In 203 patients in septic shock, GG genotype was associated with 28-day mortality and mHLA-DR remained low whereas it increased in patients with AA or AG genotype. In ex vivo cells, mHLA-DR failed to augment in GG in comparison with AG or AA genotype on exposure to IFN-γ. Promoter III transcript levels were similar in control monocytes regardless of genotype and exposure to IFN-γ. Promoter III activity was decreased in GG genotype in monocyte cell line but restored after stimulation with IFN-γ. Hereby, we demonstrated that G-286A∗rs3087456 significantly impact mHLA-DR expression in patients with septic shock in part through CIITA promoter III activity, that can be rescued using IFN-γ.

Inhibition of Adipose Tissue Beiging by HIV Integrase Inhibitors, Dolutegravir and Bictegravir, Is Associated with Adipocyte Hypertrophy, Hypoxia, Elevated Fibrosis, and Insulin Resistance in Simian Adipose Tissue and Human Adipocytes.

For people living with HIV, treatment with integrase-strand-transfer-inhibitors (INSTIs) can promote adipose tissue (AT) gain. We previously demonstrated that INSTIs can induce hypertrophy and fibrosis in AT of macaques and humans. By promoting energy expenditure, the emergence of beige adipocytes in white AT (beiging) could play an important role by limiting excess lipid storage and associated adipocyte dysfunction. We hypothesized that INSTIs could alter AT via beiging inhibition. Fibrosis and gene expression were measured in subcutaneous (SCAT) and visceral AT (VAT) from SIV-infected, dolutegravir-treated (SIVART) macaques. Beiging capacity was assessed in human adipose stromal cells (ASCs) undergoing differentiation and being exposed to dolutegravir, bictegravir, or raltegravir. Expression of beige markers, such as positive-regulatory-domain-containing-16 (PRDM16), were lower in AT of SIVART as compared to control macaques, whereas fibrosis-related genes were higher. Dolutegravir and bictegravir inhibited beige differentiation in ASCs, as shown by lower expression of beige markers and lower cell respiration. INSTIs also induced a hypertrophic insulin-resistant state associated with a pro-fibrotic phenotype. Our results indicate that adipocyte hypertrophy induced by INSTIs is involved via hypoxia (revealed by a greater hypoxia-inducible-factor-1-alpha gene expression) in fat fibrosis, beiging inhibition, and thus (via positive feedback), probably, further hypertrophy and associated insulin resistance.

Increased CD127 expression on activated FOXP3+CD4+ regulatory T cells.

Regulatory T cells (Treg) are commonly identified by CD25 (IL-2R alpha) surface expression and/or intracellular expression of the FOXP3 transcription factor. In addition, Treg are also characterized by low CD127 (IL-7R alpha) expression when compared to conventional T cells and their biology in the periphery is considered essentially independent of IL-7. We further investigated CD127 expression on Treg and we demonstrated differential CD127 expression depending on Treg subsets considered. Notably, we observed high CD127 expression on inducible costimulatory molecule (ICOS)- and CD103-expressing Treg subsets. Since these two markers reflect activation status, we addressed whether Treg activation modulated CD127 expression. We demonstrated that in contrast to conventional T cells, Treg significantly upregulated CD127 expression during in vitro and in vivo activation using adoptive transfer and contact dermatitis models. High CD127 expression on Treg was also predominantly detected ex vivo in some specific sites, notably bone marrow and skin. Importantly, higher CD127 expression on Treg correlated with higher phosphorylation of STAT5 upon IL-7 exposure. High CD127 expression on Treg also provided survival advantage upon in vitro incubation with IL-7. We thus demonstrated that low CD127 expression is not an intrinsic characteristic of Treg and we identified activated Treg as a potential target of endogenous or therapeutic IL-7.

Ablation of thymic export causes accelerated decay of naive CD4 T cells in the periphery because of activation by environmental antigen.

A model of chemical thymectomy by inducible Rag ablation was used to study peripheral T cell homeostasis. Induction of Rag ablation was efficient and complete, leading to cessation of thymic T cell production within 3-4 weeks. The decay of peripheral T cells became apparent with a delay of an additional 2-3 weeks and was entirely accounted for by loss of naïve T cells, whereas numbers of memory phenotype and regulatory T cells were not decreased. Naïve CD4 T cells decayed with an average half-life of 50 days, whereas naïve CD8 T cells exhibited a considerably longer half-life. The rapid decay of naïve CD4 T cells was not caused by intrinsic survival differences compared with naïve CD8 T cells, but was caused by changes in the lymphopenic environment resulting in higher microbial load and consequential activation. This finding suggests that in lymphopenic conditions involving compromised thymic function replenishment and survival of a naïve CD4 T cell repertoire may be severely curtailed because of chronic activation. Such a scenario might play a role in the aging immune system and chronic viral infection, such as HIV infection, and contribute to loss of CD4 T cells and impaired immune function. As our data show, continued replenishment with cells from the thymus seems to be required to maintain efficient gut mucosal defense.