Loss of CCR7 Expression on CD56bright NK Cells Is Associated with a CD56dimCD16+ NK Cell-Like Phenotype and Correlates with HIV Viral Load

et al. (2012) Loss of CCR7 Expression on CD56bright NK Cells Is Associated with a CD56dimCD16+ NK Cell-Like Phenotype and Correlates with HIV Viral Load. PLoS ONE 7(9): e44820. doi:10.1371/journal.pone.0044820 bright Loss of CCR7 Expression on CD56 NK Cells Is dim + Associated with a CD56 CD16 NK Cell-Like Phenotype and Correlates with HIV Viral Load Henoch S. Hong 0 Fareed Ahmad 0 Johanna M. Eberhard 0 Nupur Bhatnagar 0 Benjamin A. Bollmann 0 Phillip Keudel 0 Matthias Ballmaier 0 Margot Zielinska-Skowronek 0 Reinhold E. Schmidt 0 Dirk Meyer- Olson 0 Golo Ahlenstiel, University of Sydney, Australia 0 1 Klinik f u r Immunologie und Rheumatologie, Medizinische Hochschule Hannover, Hannover, Germany, 2 Division of Immunology, New England Primate Research Center, Harvard Medical School , Southborough , Massachusetts, United States of America, 3 Pa diatrische Ha matologie und Onkologie, Medizinische Hochschule Hannover , Hannover , Germany NK cells are pivotal sentinels of the innate immune system and distinct subpopulations in peripheral blood have been described. A number of studies addressed HIV-induced alterations of NK cell phenotype and functionality mainly focusing on CD56dimCD16+ and CD562CD16+ NK cells. However, the impact of HIV-infection on CD56bright NK cells is less well understood. Here we report a rise of CD56bright NK cells in HIV-infected individuals, which lack CCR7-expression and strongly correlate with HIV viral load. CCR72CD56bright NK cells were characterized by increased cytolytic potential, higher activation states and a more differentiated phenotype. These cells thus acquired a number of features of CD56dimCD16+ NK cells. Furthermore, CD56bright NK cells from HIV patients exhibited higher degranulation levels compared to uninfected individuals. Thus, chronic HIV-infection is associated with a phenotypic and functional shift of CD56bright NK cells, which provides a novel aspect of HIV-associated pathogenesis within the NK cell compartment. - Funding: D.M.O. is supported by grants from the Bundesministerium fu r Bildung und Forschung, Stiftung Zukunfts- und Innovationsfonds Niedersachsen, the European AIDS Treatment Network (NEAT) and the Helmholtz-Zentrum fu r Infektionsforschung (IG-SCID-TwinPro02). H.S.H. is supported by a fellowship from the MD/PhD program of the Hannover Biomedical Research School (HBRS) at Hannover Medical School and by a post-doctoral fellowship from the Deutsche Forschungsgemeinschaft (HO 4527/1-1). R.E.S. is supported by grant IND 06/20 from Bundesministerium fu r Bildung und Forschung and NEAT. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. . These authors contributed equally to this work. NK cells are effector cells of the innate immune system, which can spontaneously sense and lyse virus-infected cells [1,2]. Distinct NK cell subpopulations have been described. The majority of human NK cells in peripheral blood are CD56dimCD16+ cells whereas CD56bright cells only constitute approximately 10% of the blood NK cell pool [3]. Among other markers, CD56bright NK cells are characterized by high expression of type II membrane glycoprotein CD94, L-selectin CD62L and lymph-node homing receptor CCR7 [4,5] but low expression of the low affinity IgG-Fcreceptor III (CD16), killer cell immunoglobulin-like receptors (KIRs) and cytolytic molecules such as perforin and granzyme B, which are predominantly features of CD56dimCD16+NK cells [1]. Thus, NK cell subsets seem to have distinct roles in the immune response. Generally, CD56dimCD16+ NK cells are viewed as the cytotoxic NK cell subpopulation whereas CD56bright NK cells were described to have regulatory functions by means of cytokine production, such as IFN-c and TNF among others [1,3]. Recent studies have emphasized the pivotal contributions of NK cells in the host defense against HIV [6,7]. However, a number of defects in NK cell biology caused by HIV-infection have been documented [8]. We have shown an association of chronic HIVinfection with a significant decline of less differentiated and functionally more active CD56dimCD16+ NK cells, which are either CD572 or CD57dim [9]. In addition, we and others characterized an expansion of CD562CD16+ NK cells in HIV infection with a terminally differentiated phenotype [1012], which might reflect an increased turnover of NK cells in chronic HIV infection [13]. Nonetheless, little is known about the impact of HIV viremia and chronic HIV-1 infection on CD56bright NK cells. CD56bright NK cells have been suggested to be less differentiated NK cells, which can give rise to CD56dimCD16+ NK cells [14] and an accumulating body of evidence seems to corroborate this view [5,9,1519]. Enhanced cytolytic activity of these cells has been previously associated with HIV-infection [11,20]. Here we show that high HIV-1 viral load significantly correlates with a loss of CD56bright NK cells expressing CCR7. CCR72CD56bright NK cells exhibited a number of features resembling CD56dimCD16+ NK cells. These results thus present evidence for profound alterations of CD56bright NK cells in HIV-infection. Materials and Methods Ethical Approval The study was performed in strict accordance with the ethical principles as outlined in the WMA Declaration of Helsinki. All study subjects gave written, informed consent prior to their participation. The protocol was approved by the local ethics committee (Votum der Ethikkommission der MHH No. 3150). Study Subjects We obtained peripheral blood samples from 37 untreated and 15 treated HIV-seropositive subjects on highly active antiretroviral therapy (HAART) and 16 uninfected individuals in the HIV outpatient clinic of the Medizinische Hochschule Hannover (MHH). A summary of the demographical data of the studied groups is shown in Table 1 and more detailed information on the HIV-seropositive blood donors are provided in Table S1. Plasma HIV-1 RNA levels were determined using the VERSANT-HIV 1 RNA Assay, version 3.0 (bDNA, Bayer Diagnostics, Berkeley, CA) and absolute lymphocyte counts were routinely determined by differential hemograms. Frequencies of CD4+ T cells and other lymphocyte subpopulations were determined by flow cytometry using a cocktail of diagnostic staining antibodies from Beckman Coulter either directed against CD45, CD3, CD4 and CD8 or CD45, CD56, CD19, CD3 and CD16. Absolute CD4+ T cell counts were calculated by determining their frequency of the total lymphocytes. Isolation of Mononuclear Cells PBMCs were isolated from fresh blood as described previously [12,21]. Aliquots of 107 PBMCs each were cryopreserved in heatinactivated FCS supplemented with 10% dimethyl sulfoxide (DMSO) (Merck). Phenotypic Analysis of NK Cells by Flow Cytometry A list of monoclonal antibodies employed in this study is available upon request. Staining and flow cyto (...truncated)