Immunological signature in human cases of monkeypox infection in 2022 outbreak: an observational study.

BACKGROUND: An unprecedented global monkeypox outbreak started in May, 2022. No data are yet available about the dynamics of the immune response against monkeypox virus. The aim of this study was to describe kinetics of T-cell response, inflammatory profile, and pox-specific T-cell induction in patients with laboratory-confirmed monkeypox. METHODS: 17 patients with laboratory-confirmed monkeypox admitted at the Lazzaro Spallanzani National Institute for Infectious Diseases (Rome, Italy), from May 19, to July 7, 2022, were tested for differentiation and activation profile of CD4 and CD8 T (expression of CD38, PD-1, and CD57 assessed by flow cytometry), frequency of pox-specific T cells (by standard interferon-γ ELISpot), and release of interleukin (IL)-1ß, IL-6, IL-8, and tumour necrosis factor (TNF) in plasma (by ELISA). All patients were tested 10-12 days after symptoms onset. In a subgroup of nine patients with a laboratory-confirmed monkeypox, the kinetics of the immune response were analysed longitudinally according to timing from symptoms onset and compared with ten healthy donors (ie, health-care workers recruited from the same institution). FINDINGS: Among the 17 patients, ten were HIV negative and seven HIV positive, all with good viro-immunological status. On days 0-3 from symptom onset, patients with laboratory-confirmed monkeypox were characterised by a statistically significant reduction in CD4+ T cells (p=0·0011) and a concurrent increase of CD8+ T cells (p=0·0057) compared with healthy donors. A lower proportion of naive (CD45RA+CD27+) CD4+ T cells was observed in six (67%) of nine patients and a concomitant higher proportion of effector memory (CD45RA-CD27-) CD4+ T cells in all patients; this skewed immune profile tended to normalise over time. A similar differentiated profile was also observed in CD8+ T cells with a consistent expansion of terminally differentiated CD8+ T cells. Patients with monkeypox had a higher proportion of CD4+CD38+ and CD38+CD8+ T-cells than healthy donors, which normalised after 12-20 days from symptom onset. The expression of PD-1 and CD57 on CD4+ and CD8+ T-cells showed kinetics similar to that observed for CD38. Furthermore, the inflammatory cytokines (IL-1ß, IL-6, IL-8, and TNF) were higher in patients with monkeypox than in healthy donors and, although they decreased over time, they remained elevated after recovery. Almost all patients (15 [94%] of 16) developed a pox-specific Th1 response. No differences in immune cells profile were observed between patients with and without HIV, whereas paucysimptomatic patients (without systemic symptoms, with less than five skin lesions, and no other mucosal localisation of monkeypox) showed a less perturbed immune profile early after symptom onset. INTERPRETATION: Our data showed the immunological signature of monkeypox virus infection, characterised by an early expansion of activated effector CD4+ and CD8+ T cells that persisted over time. Almost all patients, even regardless of HIV infection, developed a poxvirus-specific Th1 cell response. These results might have implications on the expected immunogenicity of monkeypox vaccination, suggesting that it might not be necessary to vaccinate people who have already been infected. FUNDING: Italian Ministry of Health. TRANSLATION: For the Italian translation of the abstract see Supplementary Materials section.

Safety and immune response kinetics of GRAd-COV2 vaccine: phase 1 clinical trial results.

Despite the successful deployment of efficacious vaccines and therapeutics, the development of novel vaccines for SARS-CoV-2 remains a major goal to increase vaccine doses availability and accessibility for lower income setting. We report here on the kinetics of Spike-specific humoral and T-cell response in young and old volunteers over 6 months follow-up after a single intramuscular administration of GRAd-COV2, a gorilla adenoviral vector-based vaccine candidate currently in phase-2 of clinical development. At all three tested vaccine dosages, Spike binding and neutralizing antibodies were induced and substantially maintained up to 3 months, to then contract at 6 months. Potent T-cell responses were readily induced and sustained throughout the study period, with only minor decline. No major differences in immune response to GRAd-COV2 vaccination were observed in the two age cohorts. In light of its favorable safety and immunogenicity, GRAd-COV2 is a valuable candidate for further clinical development and potential addition to the COVID-19 vaccine toolbox to help fighting SARS-CoV-2 pandemic.

Use of Pembrolizumab for Treatment of Progressive Multifocal Leukoencephalopathy in People Living with HIV.

Progressive Multifocal Leukoencephalopathy (PML) is a demyelinating disease occurring in advanced HIV infection, caused by the reactivation of poliomavirus JC (JCV). The use of pembrolizumab for treatment is based on the inhibition of programmed cell death protein 1 (PD-1), potentially improving the anti JCV-specific response. We used pembrolizumab with combined antiretroviral treatment (cART) on a compassionate-use basis. At each administration, clinical evaluation, MRI and laboratory testing, including CD3, CD4, CD8, PD-1 markers, HIV-RNA and JCV-DNA in cerebrospinal fluid (CSF)/plasma pairs, were performed. The JCV-specific T cell response was analysed by Elispot assay. This study included five HIV patients: four male, median age 43 years (29-52), median CD4 and CD8 count 150 (15-158) and 973 (354-1250) cell/mm3, respectively; median JCV-DNA and HIV-RNA in CSF/plasma pairs 9.540/1.503 cps/mL and 2.230/619 cp/mL, respectively. Overall, patients received between two and seven doses of pembrolizumab. After treatment, we observed JCV-DNA reduction and PD-1 down-regulation both in CSF and in plasma (high in circulating CD4 and CD8 at baseline), which remained stable at low levels in all patients. Three out of five patients showed stability of clinical picture and neuroimaging, while two others died. More data are needed in order to identify predictors of response to therapy.

Myeloid-Derived Suppressor Cells in COVID-19: The Paradox of Good.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Viral replication in the respiratory tract induces the death of infected cells and the release of pathogen- associated molecular patterns (PAMPs). PAMPs give rise to local inflammation, increasing the secretion of pro- inflammatory cytokines and chemokines, which attract immune cells from the blood into the infected lung. In most individuals, lung-recruited cells clear the infection, and the immune response retreats. However, in some cases, a dysfunctional immune response occurs, which triggers a cytokine storm in the lung, leading to acute respiratory distress syndrome (ARDS). Severe COVID-19 is characterized by an impaired innate and adaptive immune response and by a massive expansion of myeloid-derived suppressor cells (MDSCs). MDSCs function as protective regulators of the immune response, protecting the host from over-immunoreactivity and hyper-inflammation. However, under certain conditions, such as chronic inflammation and cancer, MDSCs could exert a detrimental role. Accordingly, the early expansion of MDSCs in COVID-19 is able to predict the fatal outcome of the infection. Here, we review recent data on MDSCs during COVID-19, discussing how they can influence the course of the disease and whether they could be considered as biomarker and possible targets for new therapeutic approaches.

Kinetics of the B- and T-Cell Immune Responses After 6 Months From SARS-CoV-2 mRNA Vaccination in Patients With Rheumatoid Arthritis.

Objective: To assess the kinetics of the humoral and cell-mediated responses after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination in rheumatoid arthritis (RA) patients treated with different immunosuppressive therapies. Methods: Following vaccine completed schedule, health care workers (HCWs, n = 49) and RA patients (n = 35) were enrolled at 5 weeks (T1) and 6 months (T6) after the first dose of BNT162b2-mRNA vaccination. Serological response was assessed by quantifying anti-receptor-binding domain (RBD)-specific immunoglobulin G (IgG) and SARS-CoV-2 neutralizing antibodies, while cell-mediated response was assessed by a whole-blood test quantifying the interferon (IFN)-γ response to spike peptides. B-cell phenotype and IFN-γ-specific T-cell responses were evaluated by flow cytometry. Results: After 6 months, anti-RBD antibodies were still detectable in 91.4% of RA patients, although we observed a significant reduction of the titer in patients under Cytotoxic T-Lymphocyte Antigen 4 (CTLA-4)-Ig [median: 16.4 binding antibody units (BAU)/ml, interquartile range (IQR): 11.3-44.3, p < 0.0001] or tumor necrosis factor (TNF)-α inhibitors (median: 26.5 BAU/ml, IQR: 14.9-108.8, p = 0.0034) compared to controls (median: 152.7 BAU/ml, IQR: 89.3-260.3). All peripheral memory B-cell (MBC) subpopulations, in particular, the switched IgG+ MBCs (CD19+CD27+IgD-IgM-IgG+), were significantly reduced in RA subjects under CTLA-4-Ig compared to those in HCWs (p = 0.0012). In RA patients, a significantly reduced anti-RBD IgG titer was observed at T6 vs. T1, mainly in those treated with CTLA-4-Ig (p = 0.002), interleukin (IL)-6 inhibitors (p = 0.015), and disease-modifying antirheumatic drugs (DMARDs) ± corticosteroids (CCSs) (p = 0.015). In contrast, a weak nonsignificant reduction of the T-cell response was reported at T6 vs. T1. T-cell response was found in 65.7% of the RA patients at T6, with lower significant magnitude in patients under CTLA-4-Ig compared to HCWs (p < 0.0001). The SARS-CoV-2 IFN-γ-S-specific T-cell response was mainly detected in the CD4+ T-cell compartment. Conclusions: In this study, in RA patients after 6 months from COVID-19 vaccination, we show the kinetics, waning, and impairment of the humoral and, to a less extent, of the T-cell response. Similarly, a reduction of the specific response was also observed in the controls. Therefore, based on these results, a booster dose of the vaccine is crucial to increase the specific immune response regardless of the immunosuppressive therapy.

In Acute Dengue Infection, High TIM-3 Expression May Contribute to the Impairment of IFNγ Production by Circulating Vδ2 T Cells.

γδ T cells are innate cells able to quickly eliminate pathogens or infected/tumoral cells by their antiviral and adjuvancy activities. The role of γδ T cells during Dengue Viral Infection (DENV) infection is not fully elucidated. Nevertheless, human primary γδ T cells have been shown to kill in vitro DENV-infected cells, thus highlighting their possible antiviral function. The aim of this work was to characterize the phenotype and function of Vδ2 T cells in DENV patients. Fifteen DENV patients were enrolled for this study and peripheral blood mononuclear cells (PBMC) were used to analyze Vδ2-T-cell frequency, differentiation profile, activation/exhaustion status, and functionality by multiparametric flow cytometry. Our data demonstrated that DENV infection was able to significantly reduce Vδ2-T-cell frequency and to increase their activation (CD38 and HLA-DR) and exhaustion markers (PD-1 and TIM-3). Furthermore, Vδ2 T cells showed a reduced capability to produce IFN-γ after phosphoantigenic stimulation that can be associated to TIM-3 expression. Several studies are needed to depict the possible clinical impact of γδ-T-cell impairment on disease severity and to define the antiviral and immunoregulatory activities of γδ T cells in the first phases of infection.

The unbalanced p53/SIRT1 axis may impact lymphocyte homeostasis in COVID-19 patients.

BACKGROUND/OBJECTIVES: A dysregulated inflammatory profile plays an important role in coronavirus disease-2019 (COVID-19) pathogenesis. Moreover, the depletion of lymphocytes is typically associated with an unfavourable disease course. We studied the role and impact of p53 and deacetylase Sirtuin 1 (SIRT1) on lymph-monocyte homeostasis and their possible effect on T and B cell signalling. METHODS: Gene expression analysis and flow cytometry were performed on peripheral blood mononuclear cells (PBMC) of 35 COVID-19 patients and 10 healthy donors (HD). Inflammatory cytokines, the frequency of Annexin+ cells among CD3+ T cells and CD19+ B cell subsets were quantified. RESULTS: PBMC from COVID-19 patients had a higher p53 expression, and higher concentrations of plasma proinflammatory cytokines (IL1ß, TNF-α, IL8, and IL6) than HD. Deacetylase Sirtuin 1 (SIRT1) expression was significantly decreased in COVID-19 patients and was negatively correlated with p53 (p = 0.003 and r = -0.48). A lower expression of IL-7R and B Cell linker (BLNK), key genes for lymphocyte homeostasis and function, was observed in COVID-19 than in HD. The reduction of IgK and IgL chains was seen in lymphopenic COVID-19 patients. A significant increase in both apoptotic B and T cells were observed. Inflammatory cytokines correlated positively with p53 (IL-1ß: r = 0.5 and p = 0.05; IL-8: r = 0.5 and p = 0.05) and negatively with SIRT1 (IL1-ß: r = -0.5 and p = 0.04; TNF-α: r = -0.4 and p = 0.04). CONCLUSIONS: Collectively, our data indicate that the inflammatory environment, the dysregulated p53/SIRT1 axis and low expression of IL7R and BLNK may impact cell survival, B cell signalling and antibody production in COVID-19 patients. Further studies are required to define the functional impact of low BLNK/IL7R expression during severe acute respiratory syndrome coronavirus-2 infection.

Myocarditis-associated necrotizing coronary vasculitis: incidence, cause, and outcome.

AIMS: Necrotizing coronary vasculitis (NCV) is a rare entity usually associated to myocarditis which incidence, cause, and response to therapy is unreported. METHODS AND RESULTS: Among 1916 patients with biopsy-proven myocarditis, 30 had NCV. Endomyocardial samples were retrospectively investigated with immunohistochemistry for toll-like receptor 4 (TLR4) and real-time polymerase chain reaction (PCR) for viral genomes. Serum samples were processed for anti-heart autoantibodies (Abs), IL-1ß, IL-6, IL-8, tumour necrosis factor (TNF)-α. Identification of an immunologic pathway (including virus-negativity, TLR4-, and Ab-positivity) was followed by immunosuppression. Myocarditis-NCV cohort was followed for 6 months with 2D-echo and/or cardiac magnetic resonance and compared with 60 Myocarditis patients and 30 controls. Increase in left ventricular ejection fraction ≥10% was classified as response to therapy. Control endomyocardial biopsy followed the end of treatment. Twenty-six Myocarditis-NCV patients presented with heart failure; four with electrical instability. Cause of Myocarditis-NCV included infectious agents (10%) and immune-mediated causes (chest trauma 3%; drug hypersensitivity 7%; hypereosinophilic syndrome 3%; primary autoimmune diseases 33%, idiopathic 44%). Abs were positive in immune-mediated Myocarditis-NCV and virus-negative Myocarditis; Myocarditis-NCV patients with Ab+ presented autoreactivity in vessel walls. Toll-like receptor 4 was overexpressed in immune-mediated forms and poorly detectable in viral. Interleukin-1ß was significantly higher in Myocarditis-NCV than Myocarditis, the former presenting 24% in-hospital mortality compared with 1.5% of Myocarditis cohort. Immunosuppression induced improvement of cardiac function in 88% of Myocarditis-NCV and 86% of virus-negative Myocarditis patients. CONCLUSION: Necrotizing coronary vasculitis is histologically detectable in 1.5% of Myocarditis. Necrotizing coronary vasculitis includes viral and immune-mediated causes. Intra-hospital mortality is 24%. The immunologic pathway is associated with beneficial response to immunosuppression.

Early expansion of myeloid-derived suppressor cells inhibits SARS-CoV-2 specific T-cell response and may predict fatal COVID-19 outcome.

The immunological mechanisms underlying the clinical presentation of SARS-CoV-2 infection and those influencing the disease outcome remain to be defined. Myeloid-derived suppressor cells (MDSC) have been described to be highly increased during COVID-19, however, their role remains elusive. We performed an in depth analysis of MDSC in 128 SARS-CoV-2 infected patients. Polymorphonuclear (PMN)-MDSC expanded during COVID-19, in particular in patients who required intensive care treatments, and correlated with IL-1ß, IL-6, IL-8, and TNF-α plasma levels. PMN-MDSC inhibited T-cells IFN-γ production upon SARS-CoV-2 peptides stimulation, through TGF-ß- and iNOS-mediated mechanisms, possibly contrasting virus elimination. Accordingly, a multivariate regression analysis found a strong association between PMN-MDSC percentage and fatal outcome of the disease. The PMN-MDSC frequency was higher in non-survivors than survivors at the admission time, followed by a decreasing trend. Interestingly, this trend was associated with IL-6 increase in non-survivors but not in survivors. In conclusion, this study indicates PMN-MDSC as a novel factor in the pathogenesis of SARS-CoV2 infection, and open up to new therapeutic options.

Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring.

BACKGROUND: Personalised medicine in oncology needs standardised immunological assays. Flow cytometry (FCM) methods represent an essential tool for immunomonitoring, and their harmonisation is crucial to obtain comparable data in multicentre clinical trials. The objective of this study was to design a harmonisation workflow able to address the most effective issues contributing to intra- and interoperator variabilities in a multicentre project. METHODS: The Italian National Institute of Health (Istituto Superiore di Sanità, ISS) managed a multiparametric flow cytometric panel harmonisation among thirteen operators belonging to five clinical and research centres of Lazio region (Italy). The panel was based on a backbone mixture of dried antibodies (anti-CD3, anti-CD4, anti-CD8, anti-CD45RA, and anti-CCR7) to detect naïve/memory T cells, recognised as potential prognostic/predictive immunological biomarkers in cancer immunotherapies. The coordinating centre distributed frozen peripheral blood mononuclear cells (PBMCs) and fresh whole blood (WB) samples from healthy donors, reagents, and Standard Operating Procedures (SOPs) to participants who performed experiments by their own equipment, in order to mimic a real-life scenario. Operators returned raw and locally analysed data to ISS for central analysis and statistical elaboration. RESULTS: Harmonised and reproducible results were obtained by sharing experimental set-up and procedures along with centralising data analysis, leading to a reduction of cross-centre variability for naïve/memory subset frequencies particularly in the whole blood setting. CONCLUSION: Our experimental and analytical working process proved to be suitable for the harmonisation of FCM assays in a multicentre setting, where high-quality data are required to evaluate potential immunological markers, which may contribute to select better therapeutic options.

Impact of ART on dynamics of growth factors and cytokines in primary HIV infection.

Antiretroviral treatment (ART) of Primary HIV Infection (PHI) has demonstrated virological and immunological benefits. The effect of early ART during PHI on the level of growth factors and chemokines modulating immune cell functions remains to be established. The aim of our work was to analyze the dynamics of 27 cytokines, chemokines and growth/regulation factors in plasma of HIV infected patients treated during PHI. Patients with PHI (n = 43) were enrolled before, 24 and 48 weeks after therapy initiation. Quantification of soluble immune mediators was performed in plasma from HIV infected patients and healthy donors (HD, n = 7) by Luminex technology. The cytokines profile was strongly perturbed in primary HIV infected patients when compared to healthy donors (HD). After 48 weeks of ART, some of these factors were restored to HD level (IL-2, IL-5, IL-7, IL-9, IL12p70, TNFα) while others persisted higher than HD (IL-6, IL-10, IL-13). Interestingly, a subset of chemokines, such as IL-8, MCP-1, RANTES and CCL27, and growth factors such as HGF, SCF and GM-CSF, increased during ART, reaching values significantly higher than HD after 48 weeks. Moreover, the G-CSF and MIP-1ß soluble mediators were persistently altered and showed an inverse correlation with the CD4/CD8 T cell ratio. The increase of chemokines with antiviral activity and of growth factors with hematopoietic and immunomodulatory properties may have beneficial effects. Other studies are mandatory to evaluate the effects of long lasting levels of these factors to clarify their possible role in the context of protection/pathogenesis.

Myeloid Derived Suppressor Cells Expansion Persists After Early ART and May Affect CD4 T Cell Recovery.

Myeloid-derived suppressor cells (MDSC) are expanded during HIV-1 infection and correlated with disease progression. MDSC expand in the early phase of primary infection depending on TRAIL level. In this study we evaluated the effect of ART on the frequency of MDSC in patients with primary HIV infection (PHI), and their impact on CD4 T cell reconstitution. MDSC frequency was evaluated by flow-cytometry in 60 PHI patients at 12, 24 and 48 weeks after ART initiation. Cytokine plasma levels were evaluated by Luminex technology at the same time points. The capacity of MDSC to modulate hematopoietic early progenitor cells' expansion was evaluated using the OP9/Dl1 in vitro system. As previously described, polymorphonuclear-MDSC (PMN-MDSC) frequency was higher in PHI compared to healthy donors. Interestingly, 48 weeks of successful ART failed to normalize the PMN-MDSC frequency. Moreover, PMN-MDSC frequency was not correlated with residual viral load, suggesting that the persistence of PMN-MDSC was not due to residual viral replication. Interestingly, patients with low PMN-MDSC frequency (<6%) at T0 had a higher HIV DNA at the same time point than individuals with high PMN-MDSC frequency (>6%). We also found an inverse correlation between PMN-MDSC frequency and CD4-T cell count at 48 weeks post-ART, which was confirmed by multivariate analysis adjusting for age and CD4 T cell number at baseline. These data suggest that the persistence of PMN-MDSC may impact CD4 T cell recovery. Indeed, in vitro PMN-MDSC impaired the expansion of CD34+CD38- hematopoietic early progenitors. Further, a balance between TRAIL and GM-CSF may be necessary to maintain a low MDSC level. In conclusion, early ART initiation was not able to normalize PMN-MDSC frequency that might impact the CD4 T cell recovery. These data open new questions regarding the clinical impact of MDSC persistence in HIV+ patients, in particular on non-AIDS related diseases.

In Human Immunodeficiency Virus primary infection, early combined antiretroviral therapy reduced γδ T-cell activation but failed to restore their polyfunctionality.

Primary and chronic human immunodeficiency virus (HIV) infection alters γδ T-cell features. However, there is no evidence about early combined antiretroviral therapy (cART) and γδ T-cell dynamics. In the present study, HIV-positive individuals were divided into those with early primary infection (EPI) and those with late primary infection (LPI). The analysis of γδ T cells was performed by flow cytometry before and after therapy. Polyfunctional profile was assessed after in vitro peripheral blood mononuclear cell (PBMC) exposure to specific antigens. The results show that primary infection induced an expansion of Vδ1 T cells in LPI. Before treatment, a massive activation of γδ T-cell subsets was observed in both groups of patients, that correlated with disease progression and was significantly reduced after cART introduction. Despite this, CD107A-expressing Vδ1 T cells in both groups were significantly fewer than in healthy donors, but were restored by therapy introduction. Polyfunctional analysis of Vδ1 T cells from HIV-positive individuals revealed a lower frequency of CD107A+  CCL-4+ Vδ1 T-cell subsets than healthy donors that persists after therapy. Functional profile of Vδ2 was similar to that in healthy donors before therapy but, at 6 months, a lower frequency of CD107A, interferon-γ- or tumor necrosis factor-α-producing Vδ2 T cells was observed in the EPI group. Finally, individuals with LPI showed a lower frequency of quadruple-functional Vδ2 T-cell subset. In conclusion, during primary HIV infection, the baseline Vδ1 T-cell activation is correlated with immune reconstitution potential. Moreover, an altered γδ polyfunctional profile occurred, persisting after cART. Further studies are needed to understand whether a longer treatment of primary infection may increase γδ T-cell functionality.

Myeloid-Derived Suppressor Cells Specifically Suppress IFN-γ Production and Antitumor Cytotoxic Activity of Vδ2 T Cells.

γδ T cells represent less than 5% of circulating T cells; they exert a potent cytotoxic function against tumor or infected cells and secrete cytokines like conventional αß T cells. As αß T cells γδ T cells reside in the typical T cell compartments (the lymph nodes and spleen), but are more widely distributed in tissues throughout the body. For these reasons, some investigators are exploring the possibility of immunotherapies aimed to expand and activate Vδ2 T cells, or using them as Chimeric Antigen Receptor carriers. However, the role of immunosuppressive microenvironment on Vδ2 T cells during infections and cancers has not been completely elucidated. In particular, the effects of myeloid-derived suppressor cells (MDSC), largely expanded in such pathologies, were not explored. In the present work, we demonstrated that MDSC may inhibit IFN-γ production and degranulation of phosphoantigen-activated Vδ2 T cells. Moreover, the Vδ2 T cells cytotoxic activity against the Burkitt lymphoma cell line Daudi and Jurkat cell line were impaired by MDSC. The Arginase I seems to be involved in the impairment of Vδ2 T cell function induced by both tumor cells and MDSC. These data open a key issue in the context of Vδ2-targeted immunoteraphy, suggesting the need of combined strategies aimed to boost Vδ2 T cells circumventing tumor- and MDSC-induced Vδ2 T cells suppression.

IL-18 and Stem Cell Factor affect hematopoietic progenitor cells in HIV-infected patients treated during primary HIV infection.

The impact of early antiretroviral therapy (ART) during Primary HIV Infection (PHI) on the hematopoietic progenitor cells (HPCs) homeostasis is not available. This study aimed to characterize HPCs and their relationship with cytokines regulating progenitors function in ART-treated patients with PHI. We enrolled HIV infected patients treated with ART during PHI. Circulating HPCs, Lymphoid-HPCs (L-HPCs) frequency and plasmatic concentrations of IL-7, IL-18 and Stem Cell Factor (SCF) were analysed at baseline and after 6 months of therapy. ART introduction during PHI restored the decline of L-HPCs, induced a decrease in the level of pro-inflammatory IL-18 cytokine and a parallel increase of SCF. Moreover, L-HPCs frequency positively correlated with IL-18 at baseline, and with SCF after 6 months of therapy, suggesting that different signals impact L-HPCs expansion and maintenance before and after treatment. Finally, the SCF receptor expression on HPCs decreased after early ART initiation. These insights may open new perspectives for the evaluation of cytokine-driven L-HPCs expansion and their impact on the homeostasis of hematopoietic compartment during HIV infection.

Different features of Vδ2 T and NK cells in fatal and non-fatal human Ebola infections.

BACKGROUND: Human Ebola infection is characterized by a paralysis of the immune system. A signature of αß T cells in fatal Ebola infection has been recently proposed, while the involvement of innate immune cells in the protection/pathogenesis of Ebola infection is unknown. Aim of this study was to analyze γδ T and NK cells in patients from the Ebola outbreak of 2014-2015 occurred in West Africa, and to assess their association with the clinical outcome. METHODOLOGY/PRINCIPAL FINDINGS: Nineteen Ebola-infected patients were enrolled at the time of admission to the Ebola Treatment Centre in Guinea. Patients were divided in two groups on the basis of the clinical outcome. The analysis was performed by using multiparametric flow cytometry established by the European Mobile Laboratory in the field. A low frequency of Vδ2 T-cells was observed during Ebola infection, independently from the clinical outcome. Moreover, Vδ2 T-cells from Ebola patients massively expressed CD95 apoptotic marker, suggesting the involvement of apoptotic mechanisms in Vδ2 T-cell loss. Interestingly, Vδ2 T-cells from survivors expressed an effector phenotype and presented a lower expression of the CTLA-4 exhaustion marker than fatalities, suggesting a role of effector Vδ2 T-cells in the protection. Furthermore, patients with fatal Ebola infection were characterized by a lower NK cell frequency than patients with non fatal infection. In particular, both CD56bright and CD56dim NK frequency were very low both in fatal and non fatal infections, while a higher frequency of CD56neg NK cells was associated to non-fatal infections. Finally, NK activation and expression of NKp46 and CD158a were independent from clinical outcome. CONCLUSIONS/SIGNIFICANCES: Altogether, the data suggest that both effector Vδ2 T-cells and NK cells may play a role in the complex network of protective response to EBOV infection. Further studies are required to characterize the protective effector functions of Vδ2 and NK cells.

Bone Marrow CD34+ Progenitor Cells from HIV-Infected Patients Show an Impaired T Cell Differentiation Potential Related to Proinflammatory Cytokines.

The impact of HIV infection on the frequency and differentiation capability of CD34+ bone marrow hematopoietic progenitor cells (BM-HPCs) is still debated, having a possible primary role in antiretroviral-induced immunoreconstitution. We investigated the influence of HIV replication or proinflammatory cytokines on lymphopoietic capability of BM-HPCs from seven viremic (VR) and five nonviremic (NVR) HIV-infected patients. We found that BM-HPCs from VR patients were unable to differentiate in vitro toward T cells, and produced proinflammatory cytokines in the absence of viral replication. In contrast, the lymphoid differentiation potential of BM-HPCs was partially restored in successfully antiretroviral therapy-treated patients. We also showed that TLR8 triggering induced BM-HPCs from healthy donors to release proinflammatory cytokines affecting T cell differentiation. These data suggest that in HIV-infected patients, the lymphopoiesis capability of BM-HPCs may be modulated by a virus-driven autocrine mechanism involving proinflammatory cytokines.

Granulocytic Myeloid-Derived Suppressor Cells Increased in Early Phases of Primary HIV Infection Depending on TRAIL Plasma Level.

BACKGROUND: It has been demonstrated that myeloid-derived suppressor cells (MDSC) are expanded in HIV-1-infected individuals and correlated with disease progression. The phase of HIV infection during which MDSC expansion occurs, and the mechanisms that regulate this expansion remain to be established. In this study, we evaluated the frequency of MDSC in patients during primary HIV infection (PHI) and factors involved in MDSC control. METHODS: Patients with PHI and chronic HIV infection (CHI) were enrolled. PHI staging was performed according to Fiebig classification, and circulating MDSC frequency and function were evaluated by flow cytometry. Cytokine levels were evaluated by Luminex technology. RESULTS: We found that granulocytic MDSC (Gr-MDSC) frequency was higher in patients with PHI compared with healthy donors, but lower than that in patients with CHI. Interestingly, Gr-MDSC expansion was observed in the early phases of HIV infection (Fiebig II/III), but it was not associated with HIV viral load and CD4 T-cell count. Interestingly, in PHI, Gr-MDSC frequency was inversely correlated with plasmatic level of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), although a direct correlation was observed in CHI. Furthermore, lower level of Granulocyte Macrophage Colony Stimulating Factor (GM-CSF) was observed in PHI compared with that in CHI. In vitro experiments demonstrated that, differently from CHI, recombinant TRAIL-induced apoptosis of Gr-MDSC from PHI, an effect that can be abrogated by GM-CSF. CONCLUSION: We found that Gr-MDSC are expanded early during PHI and may be regulated by TRAIL and GM-CSF levels. These findings shed light on the fine mechanisms regulating the immune system during HIV infection and open new perspectives for immune-based strategies.