Natural killer cell education in human health and disease.

Natural killer (NK) cells play an important role in the host response against viral infections and cancer development. They are able to kill virus-infected and tumor cells, and they produce different important cytokines that stimulate the antiviral and antitumor adaptive immune response, particularly interferon gamma. NK cells are of particular importance in herpesvirus infections, which is illustrated by systemic and life-threatening herpesvirus disease symptoms in patients with deficiencies in NK cell activity and by the myriad of reports describing herpesvirus NK cell evasion strategies. The latter is particularly obvious for cytomegaloviruses, but increasing evidence indicates that most, if not all, members of the herpesvirus family suppress NK cell activity to some extent. This review discusses the different NK cell evasion strategies described for herpesviruses and how this knowledge may translate to clinical applications.

Herpesvirus Evasion of Natural Killer Cells

Human cytomegalovirus (HCMV) co-infection is highly prevalent within HIV-1 cohorts and is an important cofactor in driving ongoing immune activation, even during effective antiretroviral treatment. HCMV infection has recently been associated with expansion of adaptive-like natural killer (NK) cells, which harbor epigenetic alterations that impact on their cellular function and phenotype. The influence of HCMV co-infection on the considerable heterogeneity among NK cells and their functional responses to different stimuli was assessed in a cohort of HIV-1-infected individuals sampled during different stages of infection, compared with healthy subjects stratified according to HCMV serostatus. Our data demonstrate a reshaping of the NK cell pool in HIV-1 infection of HCMV-seropositive individuals, with an accentuated peripheral transition of CD56dim NK cells toward a mature CD57+ CD85j+ NKG2C+ NKG2A- phenotype. Lack of PLZF further distinguishes adaptive NK cells from other NK cells expressing CD57 or NKG2C. PLZF- NK cells from HIV-infected individuals had high expression of CD2, were Siglec-7 negative and exhibited downregulation of key signaling molecules, SYK and FceRI-γ, overwhelmingly displaying features of adaptive NK cells that correlated with HCMV serum Ab levels. Notably this adaptive-like signature was detected during early HIV-1 infection and persisted during treatment. Adaptive-like NK cell subsets in HIV-1-infected individuals displayed enhanced IFN-γ production following Fc receptor triggering compared with their conventional NK cell counterparts, and their ability to produce TNF-α and degranulate was preserved. Together, these data suggest that HMCV infection/reactivation, a hallmark of HIV-1 infection, plays a role in driving a relative expansion of NK cells with adaptive features during HIV-1 infection. The identification of selective NK subsets with retained effector activity in HIV-1-infected subjects raises the possibility of developing therapeutic strategies that exploit specific NK subpopulations to achieve better HIV-1 control.

Adaptive Reconfiguration of Natural Killer Cells in HIV-1 Infection.

The interaction of inhibitory killer cell Ig-like receptors (KIRs) with human leukocyte antigen (HLA) class I molecules has been characterized in detail. By contrast, activating members of the KIR family, although closely related to inhibitory KIRs, appear to interact weakly, if at all, with HLA class I. KIR2DS1 is the best studied activating KIR and it interacts with C2 group HLA-C (C2-HLA-C) in some assays, but not as strongly as KIR2DL1. We used a mouse 2B4 cell reporter system, which carries NFAT-green fluorescent protein with KIR2DS1 and a modified DAP12 adaptor protein. KIR2DS1 reporter cells were not activated upon coculture with 721.221 cells transfected with different HLA-C molecules, or with interferon-γ stimulated primary dermal fibroblasts. However, KIR2DS1 reporter cells and KIR2DS1+ primary natural killer (NK) cells were activated by C2-HLA-C homozygous human fetal foreskin fibroblasts (HFFFs) but only after infection with specific clones of a clinical strain of human cytomegalovirus (HCMV). Active viral gene expression was required for activation of both cell types. Primary NKG2A-KIR2DS1+ NK cell subsets degranulated after coculture with HCMV-infected HFFFs. The W6/32 antibody to HLA class I blocked the KIR2DS1 reporter cell interaction with its ligand on HCMV-infected HFFFs but did not block interaction with KIR2DL1. This implies a differential recognition of HLA-C by KIR2DL1 and KIR2DS1. The data suggest that modulation of HLA-C by HCMV is required for a potent KIR2DS1-mediated NK cell activation.

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Modulation of Human Leukocyte Antigen-C by Human Cytomegalovirus Stimulates KIR2DS1 Recognition by Natural Killer Cells.

Understanding how the T cell memory response directed towards human cytomegalovirus (HCMV) develops and changes over time while the virus persists is important. Whilst HCMV primary infection and periodic reactivation is well controlled by T cell responses in healthy people, when the immune system is compromised such as post-transplantation, during pregnancy, or underdeveloped such as in new-born infants and children, CMV disease can be a significant problem. In older people, HCMV infection is associated with increased risk of mortality and despite overt disease rarely being seen there are increases in HCMV-DNA in urine of older people suggesting that there is a change in the efficacy of the T cell response following lifelong infection. Therefore, understanding whether phenomenon such as “memory inflation” of the immune response is occurring in humans and if this is detrimental to the overall health of individuals would enable the development of appropriate treatment strategies for the future. In this review, we present the evidence available from human studies regarding the development and maintenance of memory CD8 + and CD4 + T cell responses to HCMV. We conclude that there is only limited evidence supportive of “memory inflation” occurring in humans and that future studies need to investigate immune cells from a broad range of human tissue sites to fully understand the nature of HCMV T cell memory responses to lytic and latent infection.

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Generation, maintenance and tissue distribution of T cell responses to human cytomegalovirus in lytic and latent infection.

Immune responses of Natural Killer (NK) cell are controlled by the balance between activating and inhibitory receptors, but the expression of these receptors varies between cells within an individual. Although NK cells are a component of the innate immune system, particular NK cell subsets expressing Ly49H are positively selected and increase in frequency in response to cytomegalovirus infection in mice. Recent evidence suggests that in humans certain NK subsets also have an increased frequency in the blood of HCMV infected individuals. However whether these subsets differ in their capacity of direct control of HCMV infected cells remains unclear. In this study we developed a novel in vitro assay to assess whether human NK cells subsets have differential abilities to inhibit HCMV growth and dissemination. NK cells expressing or lacking NKG2C did not display any differences when controlling viral dissemination. However, when in vitro expanded NK cells were used, cells expressing or lacking the inhibitory receptor Leukocyte Immunoglobulin-like receptor 1 (LIR1) were differentially able to control dissemination. Surprisingly, the ability of LIR1+ NK cells to control virus spread differed between HCMV viral strains, and this phenomenon was dependent on amino acid sequences within the viral ligand UL18. Together, the results here outlined an in vitro technique to compare the long-term immune responses of different human NK cell subsets, and suggest, for the first time, phenotypically defined human NK cell subsets may differentially recognise HCMV infected.

IMPORTANCE HCMV infection is ubiquitous in most populations, it is not cleared by the host after primary infection but persists for life. The innate and adaptive immune system controls the spread of virus, of which Natural Killer (NK) cells play a pivotal role. NK cells can respond to HCMV infection by rapid, short-term non-specific innate responses, but evidence from murine studies suggested NK cells may display a long-term, memory like responses to murine cytomegalovirus infection. In this study, we developed a new assay that examines human NK cell subsets that have been suggested to play a long-term memory-like response to HCMV infection. We show that changes in a HCMV viral protein that interacts with an NK cell receptor can change the ability of NK cell subsets to control HCMV while the acquisition odf another receptor has no effect on virus control.

Leukocyte Immunoglobulin-Like Receptor 1-Expressing Human Natural Killer Cell Subsets Differentially Recognize Isolates of Human Cytomegalovirus through the Viral Major Histocompatibility Complex Class I Homolog UL18

Human cytomegalovirus (HCMV) infection is not cleared by the initial immune response but persists for the lifetime of the host, in part due to its ability to establish a latent infection in cells of the myeloid lineage. HCMV has been shown to manipulate the secretion of cellular proteins during both lytic and latent infection; with changes caused by latent infection mainly investigated in CD34+ progenitor cells. Whilst CD34+ cells are generally bone marrow resident, their derivative CD14+ monocytes migrate to the periphery where they briefly circulate until extravasation into tissue sites. We have analyzed the effect of HCMV latent infection on the secretome of CD14+ monocytes, identifying an upregulation of both CCL8 and CXCL10 chemokines in the CD14+ latency-associated secretome. Unlike CD34+ cells, the CD14+ latency-associated secretome did not induce migration of resting immune cell subsets but did induce migration of activated NK and T cells expressing CXCR3 in a CXCL10 dependent manner. As reported in CD34+ latent infection, the CD14+ latency-associated secretome also suppressed the anti-viral activity of stimulated CD4+ T cells. Surprisingly, however, co-culture of activated autologous CD4+ T cells with latently infected monocytes resulted in reactivation of HCMV at levels comparable to those observed using M-CSF and IL-1β cytokines. We propose that these events represent a potential strategy to enable HCMV reactivation and local dissemination of the virus at peripheral tissue sites.

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Kevin C. Chen

Papers

Leukocyte Immunoglobulin-Like Receptor 1-Expressing Human Natural Killer Cell Subsets Differentially Recognize Isolates of Human Cytomegalovirus through the Viral Major Histocompatibility Complex Class I Homolog UL18

Latent Cytomegalovirus-Driven Recruitment of Activated CD4+ T Cells Promotes Virus Reactivation.