Cellular HIV-1 reservoirs that persist despite antiretroviral treatment are incompletely defined. We show that during suppressive antiretroviral therapy, CD4+ T memory stem cells (TSCM) harbor high per-cell levels of HIV-1 DNA, and make increasing contributions to the total viral CD4+ T cell reservoir over time. Moreover, phylogenetic studies suggested long-term persistence of viral quasispecies in CD4+ TSCM cells. Thus, HIV-1 may exploit stem cell characteristics of cellular immune memory to promote long-term viral persistence.

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Initiation of antiretroviral therapy during the earliest stages of HIV-1 infection may limit the seeding of a long-lasting viral reservoir, but long-term effects of early antiretroviral treatment initiation remain unknown. Here, we analyzed immunological and virological characteristics of nine patients who started antiretroviral therapy at primary HIV-1 infection and remained on suppressive treatment for >10 years; patients with similar treatment duration but initiation of suppressive therapy during chronic HIV-1 infection served as controls. We observed that independently of the timing of treatment initiation, HIV-1 DNA in CD4 T cells decayed primarily during the initial 3 to 4 years of treatment. However, in patients who started antiretroviral therapy in early infection, this decay occurred faster and was more pronounced, leading to substantially lower levels of cell-associated HIV-1 DNA after long-term treatment. Despite this smaller size, the viral CD4 T cell reservoir in persons with early treatment initiation consisted more dominantly of the long-lasting central-memory and T memory stem cells. HIV-1-specific T cell responses remained continuously detectable during antiretroviral therapy, independently of the timing of treatment initiation. Together, these data suggest that early HIV-1 treatment initiation, even when continued for >10 years, is unlikely to lead to viral eradication, but the presence of low viral reservoirs and durable HIV-1 T cell responses may make such patients good candidates for future interventional studies aiming at HIV-1 eradication and cure. IMPORTANCEAntiretroviral therapy can effectively suppress HIV-1 replication to undetectable levels; however, HIV-1 can persist despite treatment, and viral replication rapidly rebounds when treatment is discontinued. This is mainly due to the presence of latently infected CD4 T cells, which are not susceptible to antiretroviral drugs. Starting treatment in the earliest stages of HIV-1 infection can limit the number of these latently infected cells, raising the possibility that these viral reservoirs are naturally eliminated if suppressive antiretroviral treatment is continued for extremely long periods of time. Here, we analyzed nine patients who started on antiretroviral therapy within the earliest weeks of the disease and continued treatment for more than 10 years. Our data show that early treatment accelerated the decay of infected CD4 T cells and led to very low residual levels of detectable HIV-1 after long-term therapy, levels that were otherwise detectable in patients who are able to maintain a spontaneous, drug-free control of HIV-1 replication. Thus, long-term antiretroviral treatment started during early infection cannot eliminate HIV-1, but the reduced reservoirs of HIV-1 infected cells in such patients may increase their chances to respond to clinical interventions aiming at inducing a drug-free remission of HIV-1 infection.

Background COVID-19 patients can develop a cytokine release syndrome that eventually leads to acute respiratory distress syndrome (ARDS) requiring invasive mechanical ventilation (IMV). Since interleukin-6 (IL-6) is a relevant cytokine in ARDS, the blockade of its receptor with Tocilizumab (TCZ) could reduce mortality and/or morbidity in severe COVID-19. Objective To determine whether baseline IL-6 serum levels can predict the need for IMV and the response to TCZ. Methods Retrospective observational study performed in hospitalized patients diagnosed of COVID-19. Clinical information and laboratory findings, including IL-6 levels, were collected approximately 3 and 9 days after admission to be matched with pre- and post-administration of TCZ. Multivariable logistic and linear regressions, and survival analysis were performed depending on outcomes: need for IMV, evolution of arterial oxygen tension/fraction of inspired oxygen ratio (PaO 2 /FiO 2 ) or mortality. Results One hundred and forty-six patients were studied, predominantly male (66%); median age was 63 years. Forty-four patients (30%) required IMV, and 58 patients (40%) received treatment with TCZ. IL-6 levels>30 pg/ml was the best predictor for IMV (OR:7.1; p<0.001). Early administration of TCZ was associated with improvement of oxygenation (PaO 2 /FiO 2 ) in patients with high IL-6 (p=0.048). Patients with high IL-6 not treated with TCZ showed high mortality (HR: 4.6; p=0.003), as well as those with low IL-6 treated with TCZ (HR: 3.6; p=0.016). No relevant serious adverse events were observed in TCZ-treated patients. Conclusion Baseline IL-6>30 pg/ml predicts IMV requirement in patients with COVID-19 and contributes to establish an adequate indication for TCZ administration.

The generation of natural regulatory T cells (nTregs) is crucial for the establishment of immunologic self-tolerance and the prevention of autoimmunity. Still, the origin of nTregs and the mechanisms governing their differentiation within the thymus are poorly understood, particularly in humans. It was recently shown that conventional dendritic cells (cDCs) in human thymus were capable of inducing nTreg differentiation. However, the function of plasmacytoid DCs (pDCs), the other major subset of thymic DCs, remains unknown. Here we report that pDCs resident in the human thymus, when activated with CD40 ligand (CD40L) plus interleukin-3, efficiently promoted the generation of CD4 ؉ CD25 ؉ Foxp3 ؉ nTregs from autologous thymocytes. The progenitors of these nTregs were selectively found within CD4 ؉ CD8 ؉ thymocytes that had accomplished positive selection, as judged by their CD69 hi TCR hi phenotype. Supporting the involvement of the CD40-CD40L pathway in pDC-induced nTreg generation, we show that positively selected CD4 ؉ CD8 ؉ progenitors specifically transcribed CD40L in vivo and upregulated CD40L expression on T-cell receptor engagement, thereby promoting the activation of pDCs. Finally, evidence is provided that nTregs primed by pDCs displayed reciprocal interleukin-10/transforming growth factor-␤ cytokine expression profiles compared with nTregs primed by cDCs. This functional diversity further supports a nonredundant tolerogenic role for thymic pDCs in the human thymus. (Blood. 2010;115(26):5366-5375) IntroductionDendritic cells (DCs) are highly specialized antigen-presenting cells (APCs) that are crucial in the regulation of innate and adaptive immunity. DCs respond to danger signals by antigen uptake, maturation, and induction of antigen-specific immune responses in secondary lymphoid organs. 1,2 In addition, tissue DCs are able to present antigen in a tolerogenic fashion, leading to anergy or deletion of potentially self-reactive T cells, or inducing the generation of regulatory T cells (Tregs). [3][4][5][6] Tregs play a key role in the control of immune homeostasis and immunologic tolerance and are crucial to protect against fatal autoimmunity throughout life. Tregs also regulate or suppress other classes of immune responses, such as allograft rejection, allergy, tumor immunity, and responses to microbes. 7,8 Thus, there is great interest in understanding how do Tregs develop.During thymopoiesis, nearly all positively selected CD4 ϩ CD8 ϩ double-positive (DP) thymocytes carrying high-affinity T-cell receptors (TCRs) for self-peptide-major histocompatibility complexes are eliminated by a mechanism of clonal deletion that ensures central tolerance. 6 Self-reactive thymocytes that escape clonal deletion undergo an alternative process of nondeletional tolerance that involves their positive selection and further differentiation into immunosuppressive CD4 ϩ T cells, termed natural Tregs (nTregs). [8][9][10][11] In addition, Tregs can be induced in the periphery (iTregs) from CD4 ϩ T lymphocytes. 4 Generation o...

IntroductionThe identification of the lectin gene cluster at chromosome 19p13.2 1 has led to the realization that some C-type lectins are capable of mediating intercellular adhesion, pathogen-binding, and antigen internalization for induction of T cell responses. 2 The paradigmatic example of this type of lectin is dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN), which efficiently internalizes antigens, 3 mediates dendritic cell intercellular adhesions, 4 and recognizes a wide range of microorganisms through binding to mannose-and Lewis-containing glycans. 5 C-type lectins on dendritic cells enhance their ability for pathogen recognition 6 and contribute to modulation of toll-like receptor (TLR)-initiated signals. 7 Consequently, the definition of the range of dendritic cell lectins and their binding specificities might provide adequate targets for immune intervention and prevention of pathogen entrance and spreading.The lectin gene cluster at chromosome 19p13.2 includes the genes encoding for the type II C-type lectins DC-SIGN, liver/lymph node-specific intercellular adhesion molecule-3-grabbing integrin (L-SIGN), CD23, and liver and lymph node sinusoidal endothelial cell C-type lectin (LSECtin). 1,4,8,9 DC-SIGN is expressed on myeloid dendritic cells, 4,10 and alternatively activated in vitro on macrophages. 11 In vivo it is found on interstitial dendritic cells, 12 a subset of CD14ϩ peripheral blood DC, 13 human microvascular endothelial cells, 8 and on synovial, placenta, lymph node, and alveolar macrophages. 14-16 By contrast, L-SIGN is exclusively expressed on endothelial cells of the liver, lymph nodes, and placenta, 17,18 but not on myeloid cells.The LSECtin (CLEC4G) gene is located between the CD23 and DC-SIGN genes with the three genes arranged in the same orientation. 9 LSECtin encodes a protein with a lectin domain followed by a 110-residue stalk region, a transmembrane domain, and a 31-residue cytoplasmic domain. 9 LSECtin has been previously detected on liver and lymph node sinusoidal endothelial cells at the protein and RNA level. 9 LSECtin functions as an attachment factor for Ebola virus and SARS, but it does not bind HIV or hepatitis C virus. 19 We now describe the expression of LSECtin isoforms in ex vivo isolated human peripheral blood and thymic dendritic cells as well as in dendritic cells and macrophages generated in vitro. LSECtin exhibits ligand-induced internalization, and its sugar recognition specificity differs from that of DC-SIGN. The presence of LSECtin on myeloid cells should therefore contribute to expanding their antigen-capture and pathogen-recognition capabilities. Materials and methodsThe study described was approved by the Centro de Investigaciones Biologicas (CSIC) Institutional Review Board. The study did not involve any direct contact with human subjects. Cell cultureHuman peripheral blood mononuclear cells were isolated from buffy coats from normal donors over a Lymphoprep (Nycomed Pharma, Oslo, Norway) gradient according to sta...

The majority of HIV-1 elite controllers (EC) restrict HIV-1 replication through highly functional HIV-1-specific T cell responses, but mechanisms supporting the evolution of effective HIV-1-specific T cell immunity in these patients remain undefined. Cytosolic immune recognition of HIV-1 in conventional dendritic cells (cDC) can facilitate priming and expansion of HIV-1-specific T cells; however, HIV-1 seems to be able to avoid intracellular immune recognition in cDCs in most infected individuals. Here, we show that exposure of cDCs from EC to HIV-1 leads to a rapid and sustained production of type I interferons and upregulation of several interferon-stimulated effector genes. Emergence of these cell-intrinsic immune responses was associated with a reduced induction of SAMHD1 and LEDGF/p75, and an accumulation of viral reverse transcripts, but inhibited by pharmacological blockade of viral reverse transcription or siRNA-mediated silencing of the cytosolic DNA sensor cGAS. Importantly, improved cell-intrinsic immune recognition of HIV-1 in cDCs from elite controllers translated into stronger abilities to stimulate and expand HIV-1-specific CD8 T cell responses. These data suggest an important role of cell-intrinsic type I interferon secretion in dendritic cells for the induction of effective HIV-1-specific CD8 T cells, and may be helpful for eliciting functional T cell immunity against HIV-1 for preventative or therapeutic clinical purposes.

Natural progression of HIV-1 infection depends on genetic variation in the human major histocompatibility complex (MHC) class I locus, and the CD8+ T cell response is thought to be a primary mechanism of this effect. However, polymorphism within the MHC may also alter innate immune activity against human immunodeficiency virus type 1 (HIV-1) by changing interactions of human leukocyte antigen (HLA) class I molecules with leukocyte immunoglobulin-like receptors (LILR), a group of immunoregulatory receptors mainly expressed on myelomonocytic cells including dendritic cells (DCs). We used previously characterized HLA allotype-specific binding capacities of LILRB1 and LILRB2 as well as data from a large cohort of HIV-1-infected individuals (N = 5126) to test whether LILR-HLA class I interactions influence viral load in HIV-1 infection. Our analyses in persons of European descent, the largest ethnic group examined, show that the effect of HLA-B alleles on HIV-1 control correlates with the binding strength between corresponding HLA-B allotypes and LILRB2 (p = 10−2). Moreover, overall binding strength of LILRB2 to classical HLA class I allotypes, defined by the HLA-A/B/C genotypes in each patient, positively associates with viral replication in the absence of therapy in patients of both European (p = 10−11–10−9) and African (p = 10−5–10−3) descent. This effect appears to be driven by variations in LILRB2 binding affinities to HLA-B and is independent of individual class I allelic effects that are not related to the LILRB2 function. Correspondingly, in vitro experiments suggest that strong LILRB2-HLA binding negatively affects antigen-presenting properties of DCs. Thus, we propose an impact of LILRB2 on HIV-1 disease outcomes through altered regulation of DCs by LILRB2-HLA engagement.