[Show abstract][Hide abstract] ABSTRACT: Differences in the expression of Natural Killer cell receptors have been reported to reflect divergent clinical courses in patients with chronic infections or tumors. However, extensive molecular characterization at the transcriptional level to support this view is lacking. The aim of this work was to characterize baseline differences in purified NK cell transcriptional activity stratified by response to treatment with PEG-IFNα/RBV in patients chronically infected with HCV.
To this end we here studied by flow cytometer and gene expression profile, phenotypic and transcriptional characteristics of purified NK cells in patients chronically infected with HCV genotype-1 virus who were subsequently treated with PEG-IFNα/RBV. Results were further correlated with divergent clinical response obtained after treatment.
The pre-treatment transcriptional patterns of purified NK cells from patients subsequently undergoing a sustained virologic response (SVR) clearly segregated from those of non-responder (NR) patients. A set of 476 transcripts, including molecules involved in RNA processing, ubiquitination pathways as well as HLA class II signalling were differently expressed among divergent patients. In addition, treatment outcome was associated with differences in surface expression of NKp30 and NKG2D. A complex relationship was observed that suggested for extensive post-transcriptional editing. Only a small number of the NK cell transcripts identified were correlated with chronic HCV infection/replication indicating that inherent transcriptional activity prevails over environment effects such as viral infection.
Collectively, inherent/genetic modulation of NK cell transcription is involved in setting the path to divergent treatment outcomes and could become useful to therapeutic advantage.
Full-text · Article · Dec 2015 · Journal of Translational Medicine
[Show abstract][Hide abstract] ABSTRACT: During chronic inflammatory disorders, a persistent natural killer (NK) cell derangement is observed. While increased cell turnover is expected, little is known about whether and how NK-cell homeostatic balance is maintained. Here, flow cytometric analysis of peripheral blood mononuclear cells in chronic inflammatory disorders, both infectious and non-infectious, reveals the presence of a CD34(+)CD226(DNAM-1)(bright)CXCR4(+) cell population displaying transcriptional signatures typical of common lymphocyte precursors and giving rise to NK-cell progenies with high expression of activating receptors and mature function and even to α/β T lymphocytes. CD34(+)CD226(bright)CXCR4(+) cells reside in bone marrow, hardly circulate in healthy donors and are absent in cord blood. Their proportion correlates with the degree of inflammation, reflecting lymphoid cell turnover/reconstitution during chronic inflammation. These findings provide insight on intermediate stages of NK-cell development, a view of emergency recruitment of cell precursors, and upgrade our understanding and monitoring of chronic inflammatory conditions.
Full-text · Article · Oct 2015 · Nature Communications
[Show abstract][Hide abstract] ABSTRACT: Introduction
The persistence of immune activation and inflammation in HIV patients with HIV-RNA (VL) undetectable causes many co-morbidities [1–3]. The aim of this study is to correlate monocytes (m) and NK cell activation levels, soluble markers and oxidative stress with clinical, biochemical and metabolic data in HIV-1 infected patients with VL≤50 copies (cp)/mL on antiretroviral therapy.
Materials and Methods
Multicentre, cross-sectional study in patients with VL≤50 cp/mL and on antiretroviral therapy by at least six months. We studied: activation/homing markers (CD38, HLA-DR, CCR-2, PDL-1) on inflammatory, intermediate, proinflammatory m; activatory receptors NKp30, NKp46 and HLA-DR on NK cells; soluble inflammatory (sCD14, adiponectina, MCP-1) and stress oxidative markers (dRoms, antiRoms). Univariate analyses are performed with non-parametric and Pearson tests. The significant correlations were adjusted for possible known confounding factors (smoking, Cytomegalovirus IgG serology, Raltegravir, Protease Inhibitor [PI] therapy and HCV-RNA) with multivariate analysis.
In the 68 patients the positive correlation between age and antiRoms was significant also after adjustment for PI use (p=0.05). The% CD8+T was associated with% proinflammatory m (p=0.043) and with their expression of CCR2 mean fluorescence intensity (MFI) (p=0.012). The% NKp46+ was positively correlated with CD4+T count (p=0.001). The fibrinogen was positively associated with dRoms (p=0.052) and the positive correlation between triglycerides and antiRoms has been confirmed (p<0.001); the impact of antiRoms on HDL/triglycerides ratio (p=0.006) was observed after adjustment for PI use. The BMI was associated with smoking (p=0.011). Only the maraviroc-treated patients showed minimal arterial pressure, fibrinogen and antiRoms lower (p=0.001, 0.004 e 0.006) and sCD14 values higher (p=0.029).
Patients with long history of HIV infection and stable immunological and virological status showed interactions between acquired and innate immunity activation; moreover, the levels of some metabolic and inflammatory parameters correlate with oxidative stress values and innate immunity activation. The age, BMI and smoking impact metabolic and immunological parameters. The correlations between antiretroviral drugs and clinical-immunological parameters need further confirmations.
Full-text · Article · Nov 2014 · Journal of the International AIDS Society
[Show abstract][Hide abstract] ABSTRACT: Natural killer (NK) cell function is regulated by a balance between the triggering of activating and inhibitory receptors expressed on their surface. A relevant effort has been focused so far on the study of KIR carriage/expression setting the basis for NK cell education and self-tolerance. Focus on the evolution and regulation of activating NK receptors has lagged behind so far. Our understanding of activating receptor expression and regulation has recently improved by evidences derived from in vitro and in vivo studies. Virus infection - either acute or chronic - determines preferential expansion of NK cells with specific phenotype, activating receptors, and with recall-like functional activity. Studies on patients with viral infection (HIV and HCV) and specific diverging clinical courses confirm that inter-individual differences may exist in baseline expression of natural cytotoxicity receptors (NKp46 and NKp30). The findings that patients with divergent clinical courses have different kinetics of activating receptor density expression upon NK cell activation in vitro provide an additional, time-dependent, functional parameter. Kinetic changes in receptor expression thus represent an additional parameter to basal receptor density expression. Different expression and inducibilities of activating receptors on NK cells contribute to the high diversity of NK cell populations and may help our understanding of the inter-individual differences in innate responses that underlie divergent disease courses.
Full-text · Article · Jul 2014 · Frontiers in Immunology
[Show abstract][Hide abstract] ABSTRACT: MTB ranks as the first worldwide pathogen latently infecting one third of the population and the second leading cause of death from a single infectious agent, after the human immunodeficiency virus (HIV). The development of vigorous and apparently appropriate immune response upon infection with M. tuberculosis in humans and experimental animals conflict with failure to eradicate the pathogen itself and with its ability to undergo clinical latency from which it may exit. From a clinical standpoint, our views on MTB infection may take advantage from updating the overall perspective, that has quite changed over the last decade, following remarkable advances in our understanding of the manipulation of the immune system by M. tuberculosis and of the role of innate components of the immune response, including macrophages, neutrophils, dendritic cells and NK cells in the initial spread of MTB and its exit from latency. Scope of this review is to highlight the major mechanisms of MTB escape from immune control and to provide a supplementary translational perspective for the interpretation of innate immune mechanisms with particular impact on clinical aspects.
Full-text · Article · Jan 2014 · Mediterranean Journal of Hematology and Infectious Diseases
[Show abstract][Hide abstract] ABSTRACT: Control of HIV replication in elite controller (EC) and long-term nonprogressor (LTNP) patients has been associated with efficient CD8(+)cytotoxic T-lymphocyte function. However, innate immunity may play a role in HIV control. We studied the expression of natural cytotoxicity receptors (NKp46, NKp30, and NKp44) and their induction over a short time frame (2-4 d) on activation of natural killer (NK) cells in 31 HIV controller patients (15 ECs, 16 LTNPs). In EC/LTNP, induction of NKp46 expression was normal but short (2 d), and NKp30 was induced to lower levels vs. healthy donors. Notably, in antiretroviral-treated aviremic progressor patients (TAPPs), no induction of NKp46 or NKp30 expression occurred. More importantly, EC/LTNP failed to induce expression of NKp44, a receptor efficiently induced in activated NK cells in TAPPs. The specific lack of NKp44 expression resulted in sharply decreased capability of killing target cells by NKp44, whereas TAPPs had conserved NKp44-mediated lysis. Importantly, conserved NK cell responses, accompanied by a selective defect in the NKp44-activating pathway, may result in lack of killing of uninfected CD4(+)NKp44Ligand(+) cells when induced by HIVgp41 peptide-S3, representing a relevant mechanism of CD4(+) depletion. In addition, peripheral NK cells from EC/LTNP had increased NKG2D expression, significant HLA-DR up-regulation, and a mature (NKG2A-CD57(+)killer cell Ig-like receptor(+)CD85j(+)) phenotype, with cytolytic function also against immature dendritic cells. Thus, NK cells in EC/LTNP can maintain substantially unchanged functional capabilities, whereas the lack of NKp44 induction may be related to CD4 maintenance, representing a hallmark of these patients.
Full-text · Article · Jul 2013 · Proceedings of the National Academy of Sciences
[Show abstract][Hide abstract] ABSTRACT: Differences in innate immune responses may be associated with different capabilities of controlling HIV infection, not necessarily reflected by CD4(+) T-cell counts alone. We investigated by cytofluorometry the expression of NK cell receptors and ligands in 19 treated HIV-infected patients with CD4+<220/ml at presentation (11 AIDS, 8 non-AIDS) and 10 healthy donors. Expression of NKp46 and NKp30 was significantly higher in non-AIDS vs. AIDS patients. Overall, the level of NKp46 expression directly correlated with the degree of NK cell cytotoxicity. As compared to healthy donors, in both groups, there was a similar increase of CD69 and HLA-DR expression in NK cells that directly correlated with the presence of activation markers (HLA-DR) on CD4(+) and CD8(+) T cells. As compared to AIDS, in non-AIDS patients in vitro activated CD4+ showed higher expression of MIC-A (NKG2D ligand), with significantly higher Nectin-2/DNAM-1 and MIC-A/NKG2D ratios. Thus, NK cell responses in AIDS and non-AIDS patients with similar CD4(+) counts significantly differ despite similar treatment. This suggests an involvement of innate mechanisms, in preventing AIDS-defining opportunistic infections in HIV infection and further suggests, that CD4(+) absolute counts alone, may be inadequate to explain differences in the clinical outcome.
No preview · Article · Mar 2013 · Immunology letters
[Show abstract][Hide abstract] ABSTRACT: Purpose of the study: Promising research suggest that maraviroc (MVC) has favourable clinical outcome in HIV-infected patients (pts). Aim of the study is to assess: durability, safety profile and immunovirological recovery in a large MVC-based cohort. Methods: All HIV pts treated with antiretroviral therapy (ART) containing MVC for at least 6 months (all viruses CCR-5 tropic analyzed by genotypic and phenotypic test) were recruited in an observational multicenter study. Eight Infectious Diseases Centers in Liguria and Piedmont (Italy) collected at baseline and every 3 months demographics, clinical and immunovirological data on a web-based system (by MedinfoDist, University of Genoa). We used SPSS for Pearson Chi square test and for generalized estimating equation (GEE); in this model for longitudinal data and frequency analysis we considered altered: TCD4+≤350/mmc, HIVRNA>50 cp/ml, total cholesterol >200 mg/dl, triglycerides >160 mg/dl, transaminase>40 mg/dl, creatinine>1.3 mg/dl and we divided data in 5 time periods: baseline, 1-6, 9-12, 15-24 and 24-45 months. Summary of results: We enrolled 55 pts: 36 (65%) males, median age 49.6 years (yrs) (range [r] 18.2-76.6, IQR 44.5-53.3), 11 (20%) HCVRNA-positive, 4 (7%) HBsAg-positive, 1 both infection. Twenty-four (44%) pts were classified as CDC C stage, median nadir TCD4+ was 219/mmc (r 11-529, IQR 125-317), median duration of ART was 15.3 yrs (r 1.3-27.3, IQR 12-16.8), median duration of treatment with MVC was 23 months (r 6-47, IQR 14-36). At baseline 42 (76%) pts had HIVRNA >50 cp/ml, 11 (20%) HIVRNA≤50 cp/ml, 2 (4%) pts no data; on treatment at the last examination 53 pts (96%) had HIVRNA≤50 cp/ml, 2 pts still had HIVRNA>50 cp/ml (CCR5 tropic) and median TCD4+ count was 469/mmc (r 73-1802, IQR 302-592). One pt died and only 2 pts shifted to X4. Chi square test at 9-12 months showed p=0.0001 and the 80% of pts had TCD4+>350/mmc; at the same observation time 83.3% of pts had HIVRNA ≤50 cp/ml (p=0.0001). About cholesterol, triglycerides, transaminase and creatinine no significative differences were found and the median value showed no changes. Conclusions: In our study a majority of pts treated with ART containing MVC achieved a count of TCD4 >350/mmc and HIVRNA undetectable within 9-12 months. This regimen is a safe, feasible option and in pts with a poor immunological stage, MVC offered a remarkable TCD4+ count gain with limited X4 strains onset.
Full-text · Article · Nov 2012 · Journal of the International AIDS Society
[Show abstract][Hide abstract] ABSTRACT: Hepatitis C virus (HCV) infection induces the long-term risk of liver cirrhosis or hepatocellular carcinoma and in adults represents the most common cause of liver transplantation. Natural killer (NK) cells participate in innate immune responses with efficient direct antitumor and antiviral defense. Over the years, their complex interaction with downstream adaptive responses and with the regulation of immune responses has been increasingly recognized. Considerable advances have been made particularly in understanding the role of NK cells in the pathophysiology of HCV infection and their possible use as biological markers for clinical purposes. This review summarizes the available data on the role of NK cells in the natural history of HCV infection and their role in the outcome of treatment. The main objective of this review is to summarize recent advancements in the basic understanding of NK cell function highlighting their possible translational use in clinical practice. An integrated practical view on the possible use of currently available predictive immunogenetic and NK cell functional tests is provided, to support clinical management choices for optimal treatment of patients with both standard and new drug regimens.
No preview · Article · Nov 2012 · Expert Review of Clinical Immunology
[Show abstract][Hide abstract] ABSTRACT: It is not yet clear whether immature NK (iNK) cells are bystanders to or rather participate in immune responses to pathogens that may colocalize in areas of NK-cell maturation such as bone marrow or lymph nodes. Mycobacteria, including Bacillus Calmette-Guerin (BCG), have been shown to interact with peripheral NK cells and in vivo may colocalize in areas of iNK-cell development. We studied infection with BCG of human cord blood CD34(+) Lin(-) -derived cultures containing myelomonocytes and iNK cells in vitro. Increased iNK-cell DNAM-1 expression, transient natural cytotoxicity receptor modulation, and production of IFN-γ were observed. Transcriptional receptor modulation was associated to BCG challenge, which determined increased iNK-cell cytotoxic activity against tumor cell lines and also increased killing of immature dendritic cells (iDCs). No requirement for cell contact was recorded for BCG-induced iNK-cell activation, while cytokine production including IL-18, IL-10, GM-CSF, and TGF-β contributed to the observed effects. Thus, iNK cells are affected by mycobacteria in vitro and may contribute to shaping of adaptive mature innate responses through iDC-iNK cross-talk. In addition, iNK-cell activation by BCG may represent a novel additional mechanism contributing to the effects observed upon BCG administration in vivo.
Preview · Article · Sep 2012 · European Journal of Immunology
[Show abstract][Hide abstract] ABSTRACT: Specific NK cell killer inhibitory receptor (KIR):HLA haplotype combinations have been associated with successful clearance of acute and chronic HCV infection. Whether an imbalance of activating NK cell receptors also contributes to the outcome of treatment of chronic HCV infection, however, is not known. We studied peripheral NK cell phenotype and function in 28 chronically viraemic HCV genotype I treatment-naïve patients who underwent treatment with pegylated IFN-α and ribavirin. At baseline, chronically infected patients with sustained virological response (SVR) had reduced CD56(bright) CD16(+/-) cell populations, increased CD56(dull) CD16(+) NK cell proportions, and lower expression of NKp30, DNAM-1, and CD85j. Similarly, reduced NK cell IFN-γ production but increased degranulation was observed among nonresponding (NR) patients. After treatment, CD56(bright) CD16(+/-) NK cell numbers increased in both SVR and NR patients, with a parallel significant increase in activating NKp30 molecule densities in SVR patients only. In vitro experiments using purified NK cells in the presence of rIL-2 and IFN-α confirmed upregulation of NKp30 and also of NKp46 and DNAM-1 in patients with subsequent SVR. Thus, differences in patient NK cell receptor expression and modulation during chronic HCV-1 infection are associated with subsequent outcome of standard treatment. Individual activating receptor expression/function integrates with KIR:HLA genotype carriage to determine the clearance of HCV infection upon treatment.
Preview · Article · Oct 2011 · European Journal of Immunology
[Show abstract][Hide abstract] ABSTRACT: Natural Killer (NK) cells are endowed with cell-structure-sensing receptors providing inhibitory protection from self-destruction (inhibitory NK receptors, iNKRs, including killer inhibitory receptors and other molecules) and rapid triggering potential leading to functional cell activation by Toll-like receptors (TLRs), cytokine receptors, and activating NK cell receptors including natural cytotoxicity receptors (NCRs, i.e., NKp46, NKp46, and NKp44). NCR and NKG2D recognize ligands on infected cells which may be endogenous or may directly bind to some structures derived from invading pathogens. In this paper, we address the known direct or indirect interactions between activating receptors and pathogens and their expression during chronic HIV and HCV infections.
Full-text · Article · Aug 2011 · BioMed Research International
[Show abstract][Hide abstract] ABSTRACT: Long-term side effects may represent a relevant burden of antiretroviral treatment (ART) in HIV-infected patients with good CD4 immune reconstitution over extended time spans. CD4-guided treatment interruption (TI) has been evaluated to address this point and may result in a wide spectrum of time off ART in different patient cohorts. We studied whether differences in innate immune responses, in particular NK cells, are associated to patterns of longer (LoTI) or a shorter (ShTI) TI. Clinical cohort parameters were analyzed on a group of patients widely diverging for TI duration (<9 versus >18 months) on samples before TI, including NK-cell analysis and function by natural cytotoxicity receptor (NCR)-triggered γ-IFN production. Although persistently reduced NCR expression (NKp30) and function were observed in both LoTI and ShTI patients on ART compared with healthy donors, relevant differences were observed at baseline TI in those patients who subsequently developed LoTI course. Lower expression of NKG2D and NKp46 on NK cells. This also translates in reduced γ-IFN production in redirected functional assays. Thus, differences in innate immune balance exist during ART, may be associated to differential control of HIV infection and their understanding could explain clinical differences in individual patients that are not reflected by CD4(+) cell counts alone.
No preview · Article · Feb 2011 · International Immunology