Accumulation Of Natural Killer Cells Research Articles

Involvement of natural killer cells in PolyI:C-induced liver injury

The roles of T cells, natural killer T cells (NKT) and macrophages in autoimmune hepatitis have been well documented. However, the roles of natural killer (NK) cells in liver injury remain obscure. Here we examined the effect of Polyinosinic: polycytidylic acid (PolyI:C)-activated NK cells on liver injury. Mice were intraperitoneally injected with PolyI:C at a dose of 20mug/g body wt. The percentage and absolute number of NK cells in the liver were analyzed with flow cytometry. Serum alanine transaminase (ALT) and aspartate aminotransferase (AST) assay and H-E staining were used to evaluate the liver injury. Following PolyI:C injection, NK cells accumulation and activation occurred in the liver. Meanwhile, slight elevation of ALT/AST in the serum, mild inflammation and focal necrosis in the liver were also observed. Depletion of NK cells markedly attenuated PolyI:C-induced liver injury. Neutralization of endogenous Interleukin-12 produced by Kupffer cells abrogated the accumulation of NK cells in the liver and subsequent liver injury. The liver injury was also alleviated by neutralization of vascular cell adhesive molecule-1. These findings suggest that PolyI:C preferentially recruits and activates hepatic NK cells, which may be responsible for the mild hepatitis.

CXC chemokine ligand 10 controls viral infection in the central nervous system: evidence for a role in innate immune response through recruitment and activation of natural killer cells.

How chemokines shape the immune response to viral infection of the central nervous system (CNS) has largely been considered within the context of recruitment and activation of antigen-specific lymphocytes. However, chemokines are expressed early following viral infection, suggesting an important role in coordinating innate immune responses. Herein, we evaluated the contributions of CXC chemokine ligand 10 (CXCL10) in promoting innate defense mechanisms following coronavirus infection of the CNS. Intracerebral infection of RAG1(-/-) mice with a recombinant CXCL10-expressing murine coronavirus (mouse hepatitis virus) resulted in protection from disease and increased survival that correlated with a significant increase in recruitment and activation of natural killer (NK) cells within the CNS. Accumulation of NK cells resulted in a reduction in viral titers that was dependent on gamma interferon secretion. These results indicate that CXCL10 expression plays a pivotal role in defense following coronavirus infection of the CNS by enhancing innate immune responses.

Calling in the troops: regulation of inflammatory cell trafficking through innate cytokine/chemokine networks.

The recruitment of immune effector cells to localized sites of infection is crucial for the effective delivery of innate immune mechanisms. Under the conditions of infections with murine cytomegalovirus (MCMV), a herpesvirus with pathogenic potential, early immune functions are essential in the control of virus replication and virus-induced pathology. Our studies have demonstrated that the chemokine macrophage inflammatory protein-1alpha (MIP-1alpha) is critical for natural killer (NK) cell inflammation and delivery of interferon (IFN)-gamma to mediate downstream protective responses against MCMV infection in liver. Moreover, IFN-alpha/beta-dependent mechanisms promote MIP-1alpha production and subsequently the accumulation of NK cells in liver. Taken together, the studies highlighted in this review define a unique in vivo pathway mediated by innate cytokines in regulating chemokine responses that are essential in the promotion of NK cell inflammation for localized antiviral defense. In addition, the downstream consequences of these events in enhancing endogenous adaptive immune responses will also be discussed. Overall, the innate cytokine/chemokine networks that are described emphasize the emerging importance of chemokine functions for protective immune responses during infection with viruses.

CD4+ T cells initiate pancreatic islet xenograft rejection via an interferon-gamma-dependent recruitment of macrophages and natural killer cells.

In this study, the mechanisms by which CD4+ T cells interact with the innate immune system in xenograft rejection were investigated. Fetal pig pancreas (FPP) grafts were transplanted into female SCID mice. The FPP recipient SCID mice were reconstituted with exogenous leukocytes obtained from male BALB/c mice. Although nonreconstituted SCID recipients or recipients reconstituted with CD4+ T cell-depleted leukocytes showed indefinite FPP graft survival with very few macrophages infiltrating their grafts, reconstitution of SCID recipients with as few as 2x10(5) CD4+ T cells was sufficient to induce rapid xenograft rejection. CD4+ T cells secreted interferon-gamma but not interleukin-4 and initiated the activation and accumulation of macrophages and natural killer cells, that were responsible for the rapid graft destruction. Suppression of interferon-gamma prolonged graft survival and suppressed the macrophages and natural killer cell accumulation and activation. These results demonstrate that CD4+ T cell-dependent cellular xenograft rejection was a result of macrophage and natural killer cell accumulation and activation, but was not mediated by eosinophils. Consistent with this was the finding that interferon-gamma but not interleukin-4 was in part responsible for mediating this effect.

Adenovirus gene transfer vectors inhibit growth of lymphatic tumor metastases independent of a therapeutic transgene.

Adenovirus (Ad) gene transfer vectors traffic to regional lymph nodes (RLNs) after footpad injections in mice, resulting in localized production of interferon gamma (IFN-gamma). With this background, we evaluated the hypothesis that Ad vector administration may inhibit RLN tumor metastasis independent of the transgene in the expression cassette. Tumors of MM48, a cell line with a propensity toward lymphogenous metastasis, were established in the footpads of syngeneic C3H mice, and E1(-)E3(-) Ad vectors encoding no transgene (AdNull) or encoding an irrelevant transgene (AdCD; Escherichia coli cytosine deaminase with no 5-fluorocytosine administration) were administered (10(10) particles) in a peritumoral location. Both vectors suppressed the growth of tumor in the regional (popliteal) lymph node. This effect was localized to the regional, but not distant, lymph nodes (p < 0.05). Heat inactivation of the vector or decreasing the dose of the vector to 10(9) particles did not suppress RLN growth of the tumor when compared with 10(10) particles of active AdNull (p < 0.05 and p < 0.01, respectively). The ability of an E1(-)E4(-) vector expressing beta-galactosidase (AdRSVbetagal.11) to suppress RLN tumor growth showed that the E4 region of the Ad vector was not responsible for the effect. Blocking either IFN-gamma or natural killer (NK) cells with systemic antibody treatment in immunocompetent mice allowed rapid growth of RLN metastases despite Ad vector administration, and Ad vector injection into the footpads of tumor-free mice induced the accumulation of NK cells in the RLN. These data demonstrate that, in a metastatic murine tumor model, a low dose (10(10) particles) of replication-deficient Ad vectors inhibits RLN metastases independent of a therapeutic transgene, an effect that is mediated, at least in part, by IFN-gamma and NK cells.

Chemokine Receptor 1 Knockout Abrogates Natural Killer Cell Recruitment and Impairs Type-1 Cytokines in Lymphoid Tissue during Pulmonary Granuloma Formation

Mice with targeted mutation of chemokine receptor 1 (CCR1) were used to assess the contribution of CCR1 agonists to local, regional, and systemic inflammatory-related events during experimental pulmonary granuloma formation. Models of Th1 (type-1) and Th2 (type-2) cell-mediated lung granulomas were induced in wild-type (CCR+/+) and knockout (CCR1-/-) mice by embolizing Sepharose beads coupled to the purified protein derivative of Mycobacterium bovis or soluble antigens derived from Schistosoma mansoni eggs. Morphometric analysis indicated that granuloma sizes were unchanged in CCR1-/- mice, but flow cytometric analyses of dispersed granulomas revealed that natural killer cell recruitment to type-1 lesions was abrogated by 60%. Analysis of cytokine production by draining lymph node cultures showed altered expression in CCR1-/- mice characterized by reduced interleukin-2 and interferon-gamma in the type-1 response, and enhanced interleukin-5 and interleukin-13 in the type-2 response. Peripheral blood leukocytosis was also enhanced in the type-1 but not the type-2 response. These findings suggest that CCR1 agonists contribute to multiple immunoinflammatory events in the type-1 granulomatous response with natural killer cell accumulation being particularly sensitive to CCR1 disruption. Although functional efficacy of granulomas may be altered, chemokine redundancy and cytokine reserve seem to make the bulk of the exudative response resistant to CCR1 disruption.

Cationic Lipid Gene Transfer of an IL-2 Transgene Leads to Activation of Natural Killer Cells in a SCID Mouse Human Tumor Xenograft

Natural killer (NK) cells play an important role in combating infectious and malignant diseases and interleukin-2 (IL-2) has been shown to promote proliferation and activation of NK cells in vitro and in vivo. Here we investigate the effects of local cationic lipid-mediated IL-2 gene transfer on intratumoral accumulation and activation of NK cells in a SCID mouse tumor model. UM449 human melanoma tumors in SCID mice received intratumoral injections of DMRIE/DOPE admixed with VR1103, a DNA plasmid encoding the gene for human IL-2. Dissagregated tumor cells were tested for IL-2 secretion and were characterized using antibodies to asGM1, MAC-1, and F4/80 antigens. Granzyme A, a proteolytic serine esterase, was also measured in tumor cell lysates. IL-2 secretion from tumors injected with VR1103:DMRIE/DOPE peaked at 48 h after injection and fell to baseline levels on day 8. Intratumoral granzyme A activity was significantly increased in tumors injected with IL-2 plasmid:DMRIE/DOPE complexes, but not by an irrelevant plasmid DNA:DMRIE/DOPE control. Importantly, the growth of UM449 tumors was slowed in VR1103:DMRIE/DOPE-injected tumors. These results indicate that local cationic lipid-mediated gene transfer of IL-2 induces activation of intratumoral NK cells and slows tumor growth.

A chemokine-to-cytokine-to-chemokine cascade critical in antiviral defense.

Macrophage inflammatory protein 1alpha (MIP-1alpha) promotes natural killer (NK) cell inflammation in livers during murine cytomegalovirus (MCMV) infections, and NK cell-produced interferon gamma (IFN-gamma) contributes to defense against MCMV infections. A specific role for local NK cell IFN-gamma production, however, has not been established. The importance of MIP-1alpha and NK cell-produced IFN-gamma in shaping endogenous immune responses and defense in different compartments was examined. MIP-1alpha deficiency profoundly decreased resistance to MCMV and was associated with dramatically reduced NK cell accumulation and IFN-gamma production in liver. MIP-1alpha-independent IFN-gamma responses were observed in serum and spleen, and infection-induced elevations in blood NK cell populations occurred in absence of the factor, but peak liver expression of another chemokine, the monokine induced by IFN-gamma (Mig), depended upon presence of MIP-1alpha, NK cells, and IFN-gamma. The Mig response was also important for viral resistance. Thus, serum cytokine responses are insufficient; MIP-1alpha is critical for NK cell migration and IFN-gamma delivery to mediate protection; and Mig induction in tissues is a downstream protective response resulting from the process. These results define a critical chemokine-to-cytokine-to-chemokine cascade required for defense during a viral infection establishing itself in tissues.

M144, a murine cytomegalovirus (MCMV)-encoded major histocompatibility complex class I homologue, confers tumor resistance to natural killer cell-mediated rejection.

Until now, it has been unclear whether murine cytomegalovirus (MCMV)-encoded protein m144 directly regulates natural killer (NK) cell effector function and whether the effects of m144 are only strictly evident in the context of MCMV infection. We have generated clones of the transporter associated with antigen processing (TAP)-2-deficient RMA-S T lymphoma cell line and its parent cell line, RMA, that stably express significant and equivalent levels of m144. In vivo NK cell-mediated rejection of RMA-S-m144 lymphomas was reduced compared with rejection of parental or mock-transfected RMA-S clones, indicating the ability of m144 to regulate NK cell-mediated responses in vivo. Significantly, the accumulation of NK cells in the peritoneum was reduced in mice challenged with RMA-S-m144, as was the lytic activity of NK cells recovered from the peritoneum. Expression of m144 on RMA-S cells also conferred resistance to cytotoxicity mediated in vitro by interleukin 2-activated adherent spleen NK cells. In summary, the data demonstrate that m144 confers some protection from NK cell effector function mediated in the absence of target cell class I expression, but that in vivo the major effect of m144 is to regulate NK cell accumulation and activation at the site of immune challenge.

Decidual natural killer cells in recurrent spontaneous abortion with normal chromosomal content.

The maternal local immune responses in unexplained recurrent spontaneous abortion (RSA) are not yet well known. Maternal peripheral and decidual natural killer (NK) cells were evaluated in RSA with normal chromosomal content. Maternal peripheral blood, villous trophoblast, and decidua were taken from 15 normal pregnancies and 9 RSA patients with normal chromosomes. The NK cells in decidual lymphocytes were evaluated by flow cytometry using monoclonal antibodies for CD56, CD16, and CD3. The percentages of CD56+ CD16- CD3- cells in decidual lymphocytes in RSA were lower than in normal pregnancies (P < 0.002). The CD56+CD16+/CD56+CD16- cells ratio in RSA was higher than in normal pregnancies (P < 0.02). The lower percentages of CD56+CD16-CD3- cells in RSA cases may show an inappropriate accumulation of NK cells in the decidua, and this finding may be a factor involved in RSA.

Contribution of Natural Killer Cells to Inhibition of Angiogenesis by Interleukin-12

Interleukin-12 (IL-12) inhibits angiogenesis in vivo by inducing interferon-gamma (IFN-gamma) and other downstream mediators. Here, we report that neutralization of natural killer (NK) cell function with antibodies to either asialo GM1 or NK 1.1 reversed IL-12 inhibition of basic fibroblast growth factor (bFGF)-induced angiogenesis in athymic mice. By immunohistochemistry, those sites where bFGF-induced neovascularization was inhibited by IL-12 displayed accumulation of NK cells and the presence of IP-10-positive cells. Based on expression of the cytolytic mediators perforin and granzyme B, the NK cells were locally activated. Experimental Burkitt lymphomas treated locally with IL-12 displayed tumor tissue necrosis, vascular damage, and NK-cell infiltration surrounding small vessels. After activation in vitro with IL-12, NK cells from nude mice became strongly cytotoxic for primary cultures of syngeneic aortic endothelial cells. Cytotoxicity was neutralized by antibodies to IFN-gamma. These results document that NK cells are required mediators of angiogenesis inhibition by IL-12, and provide evidence that NK-cell cytotoxicity of endothelial cells is a potential mechanism by which IL-12 can suppress neovascularization.

Contribution of Natural Killer Cells to Inhibition of Angiogenesis by Interleukin-12

AbstractInterleukin-12 (IL-12) inhibits angiogenesis in vivo by inducing interferon-γ (IFN-γ) and other downstream mediators. Here, we report that neutralization of natural killer (NK) cell function with antibodies to either asialo GM1 or NK 1.1 reversed IL-12 inhibition of basic fibroblast growth factor (bFGF)-induced angiogenesis in athymic mice. By immunohistochemistry, those sites where bFGF-induced neovascularization was inhibited by IL-12 displayed accumulation of NK cells and the presence of IP-10–positive cells. Based on expression of the cytolytic mediators perforin and granzyme B, the NK cells were locally activated. Experimental Burkitt lymphomas treated locally with IL-12 displayed tumor tissue necrosis, vascular damage, and NK-cell infiltration surrounding small vessels. After activation in vitro with IL-12, NK cells from nude mice became strongly cytotoxic for primary cultures of syngeneic aortic endothelial cells. Cytotoxicity was neutralized by antibodies to IFN-γ. These results document that NK cells are required mediators of angiogenesis inhibition by IL-12, and provide evidence that NK-cell cytotoxicity of endothelial cells is a potential mechanism by which IL-12 can suppress neovascularization.

Accumulation of natural killer cells after hepatic artery embolisation in the midgut carcinoid syndrome

Eleven patients with disseminated midgut carcinoid tumour disease were subjected to hepatic artery embolisation. In six patients, lymphocytosis with a predominance of NK cells occurred and the cytotoxic activity of isolated lymphocytes increased. A relation between NK cell accumulation and subsequent radiological and biochemical response was observed, and it is suggested that anti-tumour mechanisms other than ischaemia may contribute to the therapeutic response in these patients.

Sudden tumour regression with enhanced natural killer cell accumulation in a patient with stage IV breast cancer

Spontaneous regression of advanced breast cancer is a rare phenomenon. Efforts have been made in order to explain it by means of immunological mechanisms. Corticosteroids have demonstrated important efficacy in the treatment of breast cancer. We present a patient with stage IV breast cancer in whom large tumour masses dramatically regressed during treatment with dexamethasone alone. In this patient, histological and hormonal findings, with results of analyses on surface and intracellular blood cells markers demonstrated significant redistribution of lymphocytes and accumulation of natural killer cells in tumour masses. It seems that dexamethasone has acted through the hypophyse against cancer.

Substrate-specific proteases (BLT-esterase) are localized predominantly in the natural killer cells of unprimed mice.

In leukocytes isolated from unprimed mice, the levels of extractable N alpha-Cbz-Lys-thiobenzylesteresterase (BLT-esterase) closely correlated with the number of natural killer (NK) cells. The spleens of mice that exhibit severe combined immunodeficiency (SCID) contained much higher levels of this enzyme than other mouse strains. Treatments that resulted in a local accumulation of NK cells (as assessed by lytic activity) produced a concomitant increase in BLT-esterase activity. However, short-term in vitro treatment of spleen cells with interferon (IFN)-alpha/beta indicated that BLT-esterase levels correlated more closely with absolute numbers of NK cells than with their lytic capacity. There was a very good correlation between the numbers of cells bearing the NK phenotype (NK-1.1+) and BLT-esterase levels. Cells positively sorted using the NK-specific antibodies NK-1.1 and LGL-1 had high enzymatic activity. The BLT-esterase levels were high in both the NK-1.1+/LGL-1- and NK-1.1+/LGL-1+ subsets. Highly purified CD4+ and CD8+ T cells and sIg+ B cells demonstrated negligible enzyme, as did populations of cells highly enriched for macrophages or neutrophils. However, it should be stressed that the inbred mice used on this study have been maintained in a pathogen-free facility. It would be anticipated that mice maintained under less stringent conditions could exhibit appreciable levels of BLT-esterase activity in their T cells. Nonetheless, BLT-esterase is present at high levels in NK cells and cannot be regarded as a T cell-specific enzyme.

Distribution of natural killer cells in periodontal diseases: an immunohistochemical study.

We studied the distribution and incidence of natural killer (NK) cells in gingiva affected with adult periodontitis (AP) and rapidly progressive periodontitis (RP) with immunohistochemical methods using antibodies Leu-7 and Leu-11b. The incidence of Leu-7 positive cells revealed no significant difference among mild AP, severe AP, and RP. On the other hand, Leu-11b positive cells were frequently found in severe AP and RP. The accumulation of NK cells reacting with Leu-11b in the infiltrated connective tissue was greater in severe forms of periodontal disease (severe AP and RP) than in mild AP. These results suggest that Leu-11b positive NK cells may play a role in the destruction of tissues in periodontal diseases.