A role for IFN-gamma in primary and secondary immunity generated by NK cell-sensitive tumor-expressing CD80 in vivo

We have investigated the primary and secondary immunity generated in vivo by a MHC class I-deficient tumor cell line that expressed CD80 (B7-1). CD80 expression enhanced primary NK cell-mediated tumor rejection in vivo and T cell immunity against secondary tumor challenge. CD80 expression enhanced primary NK cell-mediated tumor rejection, and both NK cell perforin and IFN-gamma activity were critical for the rejection of MHC class I-deficient RMA-S-CD80 tumor cells. This primary rejection process stimulated the subsequent development of specific CTL and Th1 responses against Ags expressed by the MHC class I-deficient RMA-S tumor cells. The development of effective secondary T cell immunity could be elicited by irradiated RMA-S-CD80 tumor cells and was dependent upon NK cells and IFN-gamma in the priming response. Our findings demonstrate a key role for IFN-gamma in innate and adaptive immunity triggered by CD80 expression on tumor cells.     

Trypanosoma cruzi immune evasion mediated by host cell-derived microvesicles

The innate immune system is the first mechanism of vertebrate defense against pathogen infection. In this study, we present evidence for a novel immune evasion mechanism of Trypanosoma cruzi, mediated by host cell plasma membrane-derived vesicles. We found that T. cruzi metacyclic trypomastigotes induced microvesicle release from blood cells early in infection. Upon their release, microvesicles formed a complex on the T. cruzi surface with the complement C3 convertase, leading to its stabilization and inhibition, and ultimately resulting in increased parasite survival. Furthermore, we found that TGF-β-bearing microvesicles released from monocytes and lymphocytes promoted rapid cell invasion by T. cruzi, which also contributed to parasites escaping the complement attack. In addition, in vivo infection with T. cruzi showed a rapid increase of microvesicle levels in mouse plasma, and infection with exogenous microvesicles resulted in increased T. cruzi parasitemia. Altogether, these data support a role for microvesicles contributing to T. cruzi evasion of innate immunity.     

Donor deficiency of decay-accelerating factor accelerates murine T cell-mediated cardiac allograft rejection

Decay-accelerating factor (DAF) is a cell surface regulator that accelerates the dissociation of C3/C5 convertases and thereby prevents the amplification of complement activation on self cells. In the context of transplantation, DAF has been thought to primarily regulate antibody-mediated allograft injury, which is in part serum complement-dependent. Based on our previously delineated link between DAF and CD4 T cell responses, we evaluated the effects of donor Daf1 (the murine homolog of human DAF) deficiency on CD8 T cell-mediated cardiac allograft rejection. MHC-disparate Daf1(-/-) allografts were rejected with accelerated kinetics compared with wild-type grafts. The accelerated rejection predominantly tracked with DAF's absence on bone marrow-derived cells in the graft and required allograft production of C3. Transplantation of Daf1(-/-) hearts into wild-type allogeneic hosts augmented the strength of the anti-donor (direct pathway) T cell response, in part through complement-dependent proliferative and pro-survival effects on alloreactive CD8 T cells. The accelerated allograft rejection of Daf1(-/-) hearts occurred in recipients lacking anti-donor Abs. The results reveal that donor DAF expression, by controlling local complement activation on interacting T cell APC partners, regulates the strength of the direct alloreactive CD8(+) T cell response. The findings provide new insights into links between innate and adaptive immunity that could be exploited to limit T cell-mediated injury to an allograft following transplantation.     

Complement C5a receptor is essential for the optimal generation of antiviral CD8+ T cell responses

The complement system has been long regarded as an important effector of the innate immune response. Furthermore, complement contributes to various aspects of B and T cell immunity. Nevertheless, the role of complement in CD8(+) T cell antiviral responses has yet to be fully delineated. We examined the CD8(+) T cell response in influenza type A virus-infected mice treated with a peptide antagonist to C5aR to test the potential role of complement components in CD8(+) T cell responses. We show that both the frequency and absolute numbers of flu-specific CD8(+) T cells are greatly reduced in C5aR antagonist-treated mice compared with untreated mice. This reduction in flu-specific CD8(+) T cells is accompanied by attenuated antiviral cytolytic activity in the lungs. These results demonstrate that the binding of the C5a component of complement to the C5a receptor plays an important role in CD8(+) T cell responses.     

GM-CSF restores innate,but not adaptive,immune responses in glucocorticoid-immunosuppressed human blood in vitro

Infection remains the major complication of immunosuppressive therapy in organ transplantation. Therefore, reconstitution of the innate immunity against infections, without activation of the adaptive immune responses, to prevent graft rejection is a clinically desirable status in transplant recipients. We found that GM-CSF restored TNF mRNA and protein expression without inducing IL-2 production and T cell proliferation in glucocorticoid-immunosuppressed blood from either healthy donors or liver transplant patients. Gene array experiments indicated that GM-CSF selectively restored a variety of dexamethasone-suppressed, LPS-inducible genes relevant for innate immunity. A possible explanation for the lack of GM-CSF to restore T cell proliferation is its enhancement of the release of IL-1betaR antagonist, rather than of IL-1beta itself, since exogenously added IL-1beta induced an IL-2-independent Con A-stimulated proliferation of glucocorticoid-immunosuppressed lymphocytes. Finally, to test the in vivo relevance of our findings, we showed that GM-CSF restored the survival of dexamethasone- or cyclosporine A-immunosuppressed mice from an otherwise lethal infection with Salmonella typhimurium. In addition to this increased resistance to infection, GM-CSF did not induce graft rejection of a skin allotransplant in cyclosporine A-immunosuppressed mice. The selective restoration potential of GM-CSF suggests its therapeutic use in improving the resistance against infections upon organ transplantation.     

Innate and adaptive immune responses in asthma

The recognition that asthma is primarily an inflammatory disorder of the airways associated with T helper type 2 (T(H)2) cell-dependent promotion of IgE production and recruitment of mast cells and eosinophils has provided the rationale for disease control using inhaled corticosteroids and other anti-inflammatory drugs. As more has been discovered about the cytokine, chemokine and inflammatory pathways that are associated with T(H)2-driven adaptive immunity, attempts have been made to selectively inhibit these in the hope of discovering new therapeutics as predicted from animal models of allergic inflammation. The limited success of this approach, together with the recognition that asthma is more than allergic inflammation, has drawn attention to the innate immune response in this disease. Recent advances in our understanding of the sentinel role played by innate immunity provides new targets for disease prevention and treatment. These include pathways of innate stimulation by environmental or endogenous pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs) to influence the activation and trafficking of DCs, innate sources of cytokines, and the identification of new T cell subsets and lymphoid cells.     

The role of tumor-specific Lyt-1+2- T cells in eradicating tumor cells in vivo. I. Lyt-1+2- T cells do not necessarily require recruitment of host's cytotoxic T cell precursors for implementation of in vivo immunity

The present study determines the Ly phenotype of T cells mediating tumor cell rejection in vivo and investigates some of cellular mechanisms involved in the in vivo protective immunity. C3H/HeN mice were immunized to syngeneic X5563 plasmacytoma by intradermal (i.d.) inoculation of viable X5563 tumor cells, followed by the surgical resection of the tumor. Spleen cells from these immune mice were fractionated by treatment with anti-Lyt antibodies plus complement, and each Lyt subpopulation was tested for the reconstituting potential of in vivo protective immunity in syngeneic T cell-depleted mice (B cell mice). When C3H/HeN B cell mice were adoptively transferred with Lyt-1-2+ T cells from the above tumor-immunized mice, these B cell mice exhibited an appreciable cytotoxic T lymphocyte (CTL) response to the X5563 tumor, whereas they failed to resist the i.d. challenge of X5563 tumor cells. In contrast, the adoptive transfer of Lyt-1+2- anti-X5563 immune T cells into B cell mice produced complete protection against the subsequent tumor cell challenge. Although no CTL or antibody response against X5563 tumors was detected in the above tumor-resistant B cell mice, these mice were able to retain Lyt-1+2- T cell-mediated delayed-type hypersensitivity (DTH) responses to the X5563 tumor. These results indicate that Lyt-1+2- T cells depleted of the Lyt-2+ T cell subpopulation containing CTL or CTL precursors are effective in in vivo protective immunity, and that these Lyt-1+2- T cells implement their in vivo anti-tumor activity without inducing CTL or antibody responses. The mechanism(s) by which Lyt-1+2- T cells function in vivo for the implementation of tumor-specific immunity is discussed in the context of DTH responses to the tumor-associated antigens and its related Lyt-1+2- T cell-mediated lymphokine production.     

Lung effector memory and activated CD4+ T cells display enhanced proliferation in surfactant protein A-deficient mice during allergen-mediated inflammation

Although many studies have shown that pulmonary surfactant protein (SP)-A functions in innate immunity, fewer studies have addressed its role in adaptive immunity and allergic hypersensitivity. We hypothesized that SP-A modulates the phenotype and prevalence of dendritic cells (DCs) and CD4(+) T cells to inhibit Th2-associated inflammatory indices associated with allergen-induced inflammation. In an OVA model of allergic hypersensitivity, SP-A(-/-) mice had greater eosinophilia, Th2-associated cytokine levels, and IgE levels compared with wild-type counterparts. Although both OVA-exposed groups had similar proportions of CD86(+) DCs and Foxp3(+) T regulatory cells, the SP-A(-/-) mice had elevated proportions of CD4(+) activated and effector memory T cells in their lungs compared with wild-type mice. Ex vivo recall stimulation of CD4(+) T cell pools demonstrated that cells from the SP-A(-/-) OVA mice had the greatest proliferative and IL-4-producing capacity, and this capability was attenuated with exogenous SP-A treatment. Additionally, tracking proliferation in vivo demonstrated that CD4(+) activated and effector memory T cells expanded to the greatest extent in the lungs of SP-A(-/-) OVA mice. Taken together, our data suggested that SP-A influences the prevalence, types, and functions of CD4(+) T cells in the lungs during allergic inflammation and that SP deficiency modifies the severity of inflammation in allergic hypersensitivity conditions like asthma.     

The emerging role of platelets in adaptive immunity

Platelets' foremost role in survival is hemostasis. However, a significant quantity of research has demonstrated that platelets are an integral part of inflammation and can also be potent effector cells of the innate immune response. CD154, a molecule of vital importance to adaptive immune responses, is expressed by activated platelets and has been implicated in platelet-mediated modulation of innate immunity and inflammatory disease states. Recent studies in mice extend the role of platelet CD154 to the adaptive immune response demonstrating that platelets can enhance antigen presentation, improve CD8 T cell responses, and play a critical function in normal T-dependent humoral immunity. The latter studies suggest that the current paradigm for the B cell germinal center response should be modified to include a role for platelets.     

A protective role for innate immunity in systemic lupus erythematosus

Clinical and genetic studies in humans and animal models indicate a crucial protective role for the complement system in systemic lupus erythematosus (SLE). This presents a paradox because the complement system is considered to be an important mediator of the inflammation that is observed in patients with SLE. One current view is that complement provides protection by facilitating the rapid removal of apoptotic debris to circumvent an autoimmune response. In this Opinion article, I discuss an alternative model in which complement - together with other components of the innate immune system - participates in the 'presentation' of SLE-inducing self-antigens to developing B cells. In this way, the complement system and innate immunity protect against responses to SLE (self) antigens by enhancing the elimination of self-reactive lymphocytes.     

Immunotherapy with autologous tumor cells engineered to secrete Tag7/PGRP, an innate immunity recognition molecule

BACKGROUND: Recent studies indicate that the innate component of immune defense plays an important role in the establishment of antigen-specific immune response. We have previously isolated a novel mouse gene tag7/PGRP that was shown to be involved in the innate component of the immune system, and its insect homologue is an upstream mediator of Toll signaling in Drosophila. METHODS: Transiently or stably genetically modified mouse tumor cell lines expressing Tag7 were used. Tumor growth rate and animal survival were analyzed. Possible effector cells involved in tumor suppression were detected immunohistochemically. RESULTS: Transfection of mammary gland adenocarcinoma cells with the tag7 cDNA did not alter their growth rate in vitro but diminished their tumorogenicity in vivo in syngeneic and immunodeficient animals. Increased incidence of apoptosis was registered in the modified tumors. Transient expression of Tag7 by mouse melanoma M3 cells elicited protective immunity against parental tumor cells. Immunohistochemical analysis revealed that tumors after immunization with the genetically modified cells were infiltrated with Mac1(+) cells, B220(+) cells, and NK cells. Using nude mice we observed rejection of modified cells, but did not detect memory formation. CONCLUSIONS: We can conclude that secretion of the Tag7 protein by genetically modified cells can induce mobilization of antigen-presenting cells and innate effectors. Memory mechanisms are mediated by T cell response. For the first time our results demonstrate that local secretion of Tag7-the molecule involved in innate immunity-may play an important role in the induction of effective antitumor response in mice.     

IL-10-conditioned dendritic cells, decommissioned for recruitment of adaptive immunity, elicit innate inflammatory gene products in response to danger signals

Dendritic cells (DCs) are the professional APCs of the immune system, enabling T cells to perceive and respond appropriately to potentially dangerous microbes, while also being able to maintain T cell tolerance toward self. In part, such tolerance can be determined by IL-10 released from certain types of regulatory T cells. IL-10 has previously been shown to render DCs unable to activate T cells and it has been assumed that this process represents a general block in maturation. Using serial analysis of gene expression, we show that IL-10 pretreatment of murine bone marrow-derived DCs alone causes significant changes in gene expression. Furthermore, these cells retain the ability to respond to Toll-like receptor agonists, but in a manner skewed toward the selective induction of mediators known to enhance local inflammation and innate immunity, among which we highlight a novel CXCR2 ligand, DC inflammatory protein-1. These data suggest that, while the presence of a protolerogenic and purportedly anti-inflammatory agent such as IL-10 precludes DCs from acquiring their potential as initiators of adaptive immunity, their ability to act as initiators of innate immunity in response to Toll-like receptor signaling is enhanced.     

Micrometer-sized titanium particles can induce potent Th2-type responses through TLR4-independent pathways

Wear debris in joint replacements has been suggested as a cause of associated tissue-damaging inflammation. In this study, we examined whether solid titanium microparticles (mTi) of sufficient size to accumulate as wear debris could stimulate innate or adaptive immunity in vivo. mTi, administered in conjunction with OVA, promoted total and Ag-specific elevations in serum IgE and IgG1. Analysis of transferred transgenic OVA-specific naive T cells further showed that mTi acted as an adjuvant to drive Ag-specific Th2 cell differentiation in vivo. Assessment of the innate response indicated that mTi induced rapid recruitment and differentiation of alternatively activated macrophages in vivo, through IL-4- and TLR4-independent pathways. These studies suggest that solid microparticles alone can act as adjuvants to induce potent innate and adaptive Th2-type immune responses and further suggest that wear debris in joint replacements may have Th2-type inflammatory properties.     

Costimulation via CD55 on human CD4+ T cells mediated by CD97

Decay-accelerating factor (CD55) is a complement regulatory protein, which is expressed by most cells to protect them from complement-mediated attack. CD55 also binds CD97, an EGF-TM7 receptor constitutively expressed on granulocytes and monocytes and rapidly up-regulated on T and B cells upon activation. Early results suggested that CD55 could further enhance T cell proliferation induced by phorbol ester treatment. The present study demonstrates that coengagement of CD55, using either cross-linking mAbs or its natural ligand CD97, and CD3 results in enhanced proliferation of human peripheral blood CD4(+) T cells, expression of the activation markers CD69 and CD25, and secretion of IL-10 and GM-CSF. Recently, an increase in T cell responsiveness in CD55(-/-) mice was shown to be mediated by a lack of complement regulation. In this study, we show that direct stimulation of CD55 on CD4(+) T cells with CD97 can modulate T cell activation but does not interfere with CD55-mediated complement regulation. Our results support a multifaceted role for CD55 in human T cell activation, constituting a further link between innate and adaptive immunity.     

Pentraxins in innate immunity: lessons from PTX3

The innate immune system constitutes the first line of defence against microorganisms and plays a primordial role in the activation and regulation of adaptive immunity. The innate immune system is composed of a cellular arm and a humoral arm. Components of the humoral arm include members of the complement cascade and soluble pattern recognition molecules (PRMs). These fluid-phase PRMs represent the functional ancestors of antibodies and play a crucial role in the discrimination between self, non-self and modified-self. Moreover, evidence has been presented that these soluble PRMs participate in the regulation of inflammatory responses and interact with the cellular arm of the innate immune system. Pentraxins consist of a set of multimeric soluble proteins and represent the prototypic components of humoral innate immunity. Based on the primary structure of the protomer, pentraxins are divided into two groups: short pentraxins and long pentraxins. The short pentraxins C-reactive protein and serum amyloid P-component are produced by the liver and represent the main acute phase proteins in human and mouse, respectively. The long pentraxin PTX3 is produced by innate immunity cells (e.g. PMN, macrophages, dendritic cells), interacts with several ligands and plays an essential role in innate immunity, tuning inflammation and matrix deposition. PTX3 provides a paradigm for the mode of action of humoral innate immunity.     

Reovirus activates human dendritic cells to promote innate antitumor immunity

Oncolytic viruses can exert their antitumor activity via direct oncolysis or activation of antitumor immunity. Although reovirus is currently under clinical investigation for the treatment of localized or disseminated cancer, any potential immune contribution to its efficacy has not been addressed. This is the first study to investigate the ability of reovirus to activate human dendritic cells (DC), key regulators of both innate and adaptive immune responses. Reovirus induced DC maturation and stimulated the production of the proinflammatory cytokines IFN-alpha, TNF-alpha, IL-12p70, and IL-6. Activation of DC by reovirus was not dependent on viral replication, while cytokine production (but not phenotypic maturation) was inhibited by blockade of PKR and NF-kappaB signaling. Upon coculture with autologous NK cells, reovirus-activated DC up-regulated IFN-gamma production and increased NK cytolytic activity. Moreover, short-term coculture of reovirus-activated DC with autologous T cells also enhanced T cell cytokine secretion (IL-2 and IFN-gamma) and induced non-Ag restricted tumor cell killing. These data demonstrate for the first time that reovirus directly activates human DC and that reovirus-activated DC stimulate innate killing by not only NK cells, but also T cells, suggesting a novel potential role for T cells in oncolytic virus-induced local tumor cell death. Hence reovirus recognition by DC may trigger innate effector mechanisms to complement the virus's direct cytotoxicity, potentially enhancing the efficacy of reovirus as a therapeutic agent.     

The Cholinergic Anti-Inflammatory Pathway Delays TLR-Induced Skin Allograft Rejection in Mice: Cholinergic Pathway Modulates Alloreactivity

Activation of innate immunity through Toll-like receptors (TLR) can abrogate transplantation tolerance by revealing hidden T cell alloreactivity. Separately, the cholinergic anti-inflammatory pathway has the capacity to dampen macrophage activation and cytokine release during endotoxemia and ischemia reperfusion injury. However, the relevance of the α7 nicotinic acetylcholine receptor (α7nAChR)-dependent anti-inflammatory pathway in the process of allograft rejection or maintenance of tolerance remains unknown. The aim of our study is to investigate whether the cholinergic pathway could impact T cell alloreactivity and transplant outcome in mice. For this purpose, we performed minor-mismatched skin allografts using donor/recipient combinations genetically deficient for the α7nAChR. Minor-mismatched skin grafts were not rejected unless the mice were housed in an environment with endogenous pathogen exposure or the graft was treated with direct application of imiquimod (a TLR7 ligand). The α7nAChR-deficient recipient mice showed accelerated rejection compared to wild type recipient mice under these conditions of TLR activation. The accelerated rejection was associated with enhanced IL-17 and IFN-γ production by alloreactive T cells. An α7nAChR-deficiency in the donor tissue facilitated allograft rejection but not in recipient mice. In addition, adoptive T cell transfer experiments in skin-grafted lymphopenic animals revealed a direct regulatory role for the α7nAChR on T cells. Taken together, our data demonstrate that the cholinergic pathway regulates alloreactivity and transplantation tolerance at multiple levels. One implication suggested by our work is that, in an organ transplant setting, deliberate α7nAChR stimulation of brain dead donors might be a valuable approach for preventing donor tissue inflammation prior to transplant.     

Regulating the suppressors: apoptosis and inflammation govern the survival of tumor-induced myeloid-derived suppressor cells (MDSC)

Immune suppressive myeloid-derived suppressor cells (MDSC) are present in most cancer patients where they inhibit innate anti-tumor immunity and are a significant obstacle to cancer immunotherapy. Inflammation is a known inducer of Gr1(+)CD11b(+) MDSC; however, the factors/conditions that regulate MDSC survival and half-life have not been identified. We have used mass spectrometry (MS) and proteomic analysis to identify proteins and pathways that regulate MDSC survival. This analysis revealed high expression of caspase family proteins and the Fas-FasL, p38 MAPK, and TGFβ pathways, suggesting that Fas-FasL apoptosis regulates MDSC survival. Flow cytometry, confocal microscopy, and western blot analyses confirmed the MS findings and demonstrated that Fas(+) MDSC are susceptible to Fas-mediated killing in vitro. In vivo studies with FasL-deficient and Fas-deficient mice demonstrated that Fas-FasL interactions are essential for MDSC apoptosis and for rejection of established metastatic disease and survival and that FasL(+) T cells are the effector population mediating MDSC apoptosis. MS findings validated by biological experiments demonstrated that inflammation increases MDSC levels by protecting MDSC from Fas-mediated apoptosis, possibly by activating p38 MAPK. These results demonstrate that MDSC half-life in vivo is regulated by FasL(+) T cells and that inflammation increases MDSC levels by conferring resistance to Fas-mediated apoptosis and identifies T cells as the relevant effector cells causing MDSC apoptosis in vivo. This newly recognized mechanism for regulating MDSC levels identifies potential new targets for decreasing MDSC in cancer patients.     

Choosing between immunity and tolerance after transplantation

Transplantation of MHC disparate solid organ leads to activation of both innate and adaptive immune responses. These immune responses include both effector and regulatory limbs. Lymphodepletion and IL-2 blockade may control the acute alloimmune response in humans but may also limit the regulatory arm. Numerous cytokines involved in rejection appear to be critical for tolerance. TLRs, complement engagement can facilitate suppressive responses as well as effector responses. Facilitating antigen-specific tolerance while enabling robust immunity may require ex vivo manipulation of the immune system rather than relying on in vivo perturbations of the homeostatic process.     

The role of nitric oxide in inflammatory reactions

Nitric oxide (NO) was initially described as a physiological mediator of endothelial cell relaxation, an important role in hypotension. NO is an intercellular messenger that has been recognized as one of the most versatile players in the immune system. Cells of the innate immune system--macrophages, neutrophils and natural killer cells--use pattern recognition receptors to recognize the molecular patterns associated with pathogens. Activated macrophages then inhibit pathogen replication by releasing a variety of effector molecules, including NO. In addition to macrophages, a large number of other immune-system cells produce and respond to NO. Thus, NO is important as a toxic defense molecule against infectious organisms. It also regulates the functional activity, growth and death of many immune and inflammatory cell types including macrophages, T lymphocytes, antigen-presenting cells, mast cells, neutrophils and natural killer cells. However, the role of NO in nonspecific and specific immunity in vivo and in immunologically mediated diseases and inflammation is poorly understood. This Minireview will discuss the role of NO in immune response and inflammation, and its mechanisms of action in these processes.