Natural immunity最新文献

Under certain conditions, natural killer (NK) cells accumulate rapidly at extrahematic sites. In an effort to define the mechanisms underlying the recruitment of NK cells in tissues, we investigated their ability to adhere and transmigrate across endothelial cell (EC) monolayers. A considerable proportion of NK cells adhered to EC and about 30-40% of the adherent NK cells transmigrated across EC. NK cells were 2-3 times more efficient than resting T cells. Exposure of NK cells to IL-2 and IL-12 augmented their adhesive ability, while IL-4 had an inhibitory effect. mAb directed against CD18 and CD11a inhibited binding and migration of NK cells across resting or IL-1-activated EC, whereas anti-CD11b and Cd11c did not. Using IL-1-activated EC, it was found that anti-VLA-4 and anti-VCAM-1 mAb utilized in concert with anti-CD18 significantly reduced adhesion and transmigration. The CS-1 peptide of fibronectin (which recognizes VLA-4), when used in concert with anti-CD18 and anti-VCAM-1 (but not anti-VLA-4), caused a small, but significant increase in inhibition. Thus, LFA-1 and VLA-4 are crucial determinants of the adhesive and migratory interaction with the vascular endothelium.

Mixed suspensions of B16-F10 melanoma cells and murine interleukin 2 (IL2)-activated (adherent) natural killer (A-NK) cells cultured for 5 days were enclosed in gelled droplets of reconstituted basement membrane extracellular matrix (Matrigel). After incubation under cell culture conditions +/- IL2, samples were fixed for electron microscopy after 10 min and 2, 6, and 24 h. At the first time point cells were rounded and randomly distributed in the gel, at 2 h A-NK cells migrated vividly and formed contacts with target cells. At 6 h there were extensive effector:target conjugates and melanoma cell debris in the gel. Directed exocytosis of A-NK cell-specific granules could not be verified. At 24 h very few intact B16 cells remained in IL2-substituted specimens and there were large amounts of lytic melanoma cell remnants; in the absence of IL2 substantial numbers of surviving melanoma cells formed aggregates. At this time some A-NK cells had ingested melanoma cell components which probably fused with specific two-compartment granules to form large phagolysosomes. A-NK cells enlarged into a giant cell type with huge cytoplasmic accumulations of amorphous material described as mucoid masses by others.

Strains of mice differ greatly in resistance to infection in their lungs with virulent Mycoplasma pulmonis (MP) organisms even during the first 5 days, prior to detection of humoral or T cell mediated acquired immune responses. C57BL/6 mice are resistant, and BALB/c and C3H mice are susceptible, and one major gene, MP, not linked to the H2 major histocompatibility complex, regulates resistance. C57BL/6 x C3H (B x H) and BALB/c x C57BL/6 (C x B) recombinant inbred strain mice were infected intratracheally with the T2 strain of MP. Five days later, the recovery of organisms from tracheolung lavages and lung tissue was determined. The strain distribution pattern of resistance indicated that the MP gene maps to chromosome 4. B6.C-H18 (B6 mice congenic for the BALB/c H18 gene of chromosome 4) were much more susceptible than B6 mice, but were less susceptible than BALB/c mice, supporting the data obtained with the recombinant inbred strain mice, but suggesting that other genes may also influence resistance to infection with MP.

Studies were performed to determine whether resistance to Toxoplasma gondii infection in mice depends on a mechanism involving neutrophils. Immunocompetent C57BL/6 and C.B-17 mice infected with T. gondii by gavage had an increased percentage of neutrophils in their peripheral blood. C57BL/6 mice selectively depleted of neutrophils by injections of RB6-8C5 monoclonal antibody died during the acute phase of the disease. Depletion of neutrophils had no effect on interferon gamma production, but had a profound effect on the total numbers of peripheral blood CD4+ and CD8+ T cells. Neutrophil-depleted C.B-17 mice survived longer than neutrophil-depleted C57BL/6 mice when infected with T. gondii, however they became much sicker, and were less able to survive long-term than infected, control mAb-treated mice as indicated by severe sustained weight loss. This study shows that neutrophils play an important role in resistance to acute primary T. gondii infection and that depletion of neutrophils reduces the numbers of CD4+ and CD8+ lymphocytes recoverable from peripheral blood of infected but not uninfected mice. This effect on lymphocytes may contribute to the reduced long-term survival of neutrophil-depleted mice.

Bcl-2 is a major anti-apoptotic protein expressed in many normal and malignant cells. Recently, low to absent expression was reported in human natural killer (NK) cells cultured in serum-free media which could be induced with stem cell factor. We investigated the expression of bcl-2 protein of NK cells in normal blood donors and compared the bcl-2 expression in CD56+ NK cells with CD3+ T cells. To determine bcl-2 reactivity, a three-color flow-cytometric technique was used. CD56+ CD3- NK cells had an average bcl-2 expression of 83% compared with CD3+ T cells. CD56 and CD3 double positive T cells had an average content of 111% compared with all peripheral CD3+ T lymphocytes. When peripheral mononuclear cells were cultured with interleukin-2 (IL-2), bcl-2 could be upregulated by IL-2 in all cell populations studied. The induction of bcl-2 in these cell populations paralleled the induction in CD56- T lymphocytes cultured under identical conditions. The induction of bcl-2 by IL-2 was confirmed by Western blotting. The maximum induction of bcl-2 by IL-2 was observed at an IL-2 dose of 100-1,000 U/ml. Our data confirm the anti-apoptotic protein bcl-2 as an activation- or proliferation-associated marker of normal NK cells which can be induced by IL-2.

We investigated the deficiency of natural killer (NK) activity by contrasting healthy individuals with patients. Human NK activities of 125 individuals consisting of 68 healthy donors and 57 patients (36 autoimmune disease and 21 cancer patients) were measured by Eu-DTPA release assay in which the target cells were labeled by nonradioactive materials-Eu-DTPA, and they were phenotypically analyzed with three-color flow cytometry. Furthermore, a part of these donors was functionally studied on NK cells sorted out from PBL. 23.3% of healthy donors and approximately 70% of patients had low NK activity (LNK). In these healthy LNK and patient LNK, the population of CD3-CD16+CD56+ subset in PBL was significantly lower than that of the same subset in healthy individuals with high and medium NK activity (HMNK). The cytotoxicity of CD3-CD16+CD56+ cells sorted out from PBL in healthy LNK and patient LNK were approximately the same with or higher than that in healthy HMNK. No differences were found either in the expression of CD2 and LFA-1 antigens on the CD3-CD56+ NK cells or in the amount of granulous proteins such as perforin and granzyme A in these cells among healthy HMNK, healthy LNK and patient LNK. These results suggested that low NK activity of healthy LNK and patient LNK was more reflected by the diminution of the population of CD3-CD16+CD56+ subset in PBL rather than the functional defects of NK cells. A phenotypical and functional study on healthy LNK has not been reported extensively, and we found several differences between healthy LNK and patient LNK in this study. By stimulation with IL-2, the cytotoxicity of healthy LNK increased more rapidly than that of patient LNK, and at high effector:target cell ratio (> or = 40) it was significantly higher than that of patient LNK. The population of CD3+CD16+/CD56+ subset in PBL of healthy LNK was higher than that of patient LNK, but on the other hand it was about the same as that of healthy HMNK.

Murine CMV (MCMV) presents a model for the study of the role of the immune system in the pathogenesis of human CMV (HCMV) infection. MCMV causes T cell immune impairment in the infected mice, manifested by suppressed responses to T cell mitogens and a profound reduction of Con A induced IL-2 production. Thymic humoral factor (THF-gamma 2) is an octapeptide which was first isolated from calf thymus, characterized and chemically synthesized. This peptide has been shown to have immunoregulatory effects in various systems. Systemic treatment of MCMV-infected mice with THF-gamma 2 resulted in the enhancement of protective efficacy of MCMV immune spleen cells and the reconstitution of mitogenic responses and IL-2 secretion.

A widely accepted theory of lymphocyte-mediated cytotoxicity (CMC) proposes that upon effector cell (EC) and target cell (TC) interaction, release of perforin, serine proteases and other lytic moieties contained within cytoplasmic granules results in TC lysis. Complement activation and the activation of the various enzymatic activities associated with cytotoxic granules have strikingly similar modes of action and both lead to pore formation in their respective targets. We report here that by using antisera to early and late complement components we were able to inhibit CTL, NK and ADCC cytotoxicity up to 100%, even though binding of EC to TC was unaffected. Furthermore, we showed that addition of C1q or C1s (two serine proteases) antisera to C9 antisera, at titers too low to inhibit separately, resulted in synergistic inhibition of CMC. Anti-C1s together with anti-C1q (or anti-C8 with anti-C9) did not result in synergy. This finding supports a cascade model of activation for lytic molecules released from EC. In addition, we demonstrated that anti-C1q and anti-C1s bind to proteins in the 30-kD region and anti-C9 binds to proteins in the 70-kD region, coinciding with published molecular weights of granzymes and perforin, respectively. Finally, lytic ability of purified granules was also inhibited by complement antisera, further suggesting that activation occurs outside of TC. Taken as a whole, these data indicate that TC lysis may be the result of a cascade of events involving granzymes and perforin, analogous to that seen with the complement system.

In order to better understand the immunological functions of the HLA-G gene, expression of this gene has been studied with RT-PCR in human functional lymphocyte subpopulations. Only one population of cells has not shown any HLA-G mRNA expression, the BY55-mAb-defined natural killer cells in cord blood. This absence of transcription was not modulated by IL2, IFN-gamma or TNF-alpha. Several T clone lymphocytes isolated from human peripheral blood, bone marrow or thymus have shown a significant transcription of the HLA-G gene. Only one clone, with a natural killer phenotype, did not reveal full length or alternatively spliced transcripts of HLA-G. Intensity of HLA-G transcription was not affected by TNF-alpha, IL13 or IL4, but HLA-G transcripts appeared more abundant in the presence than in the absence of IL2.

To improve our understanding of the role natural-immunity cells play in regulating the immune response to Candida albicans (CA) we compared local versus systemic effects of intraperitoneal inoculations with inactivated CA cells in mice. Peritoneal exudate cells (PECs) and spleen cells (SCs) were recovered from CD2F1 mice after 5 intraperitoneal CA injections (2 x 10(7) cells/mouse on days -14, -10, -7, -3 and 0 (CA-5d) with respect to in vitro assays performed at 2 h, 24 h, 3 days and 5 days). Northern blot analysis revealed that 2 h after CA-5d, PECs expressed a high level of IL-2, IFN-gamma, IL-1 beta and a low level of IL-10 and TNF-alpha mRNAs, while IL-4 and IL-5 mRNAs were absent, suggesting the development of TH1 subset. At 24 h, while IL-2 mRNA remained high, IL-1 beta and IFN-gamma expression had decreased and IL-10 and TNF-alpha mRNAs were no longer detectable. Instead, in spleens of CA-treated mice, examined up to 5 days after CA-5d, only IL-2 and IL-1 beta mRNAs were detectable, but the expression level was similar to that of untreated control mice. CA-5d induced a high level of natural-killer (NK)/lymphokine-activated-killer (LAK) activity in the peritoneal cavity but did not affect spleen NK activity. After CA-5d, the proliferative response of PECs to mitogens and CA antigens was also different from that of SCs. Unfractionated PECs were unable to proliferate in response to concanavalin A (Con A), IL-2, CA cells and CA cell wall mannoprotein, but after removal of the nylon-wool-adherent fraction, the nonadherent peritoneal cells (Nad-PECs) showed a significant proliferative response to mitogens. After depletion of NK cells by anti-asialo-GM1 antibody plus complement, the proliferative response of Nad-PECs to Con A and CA increased further. Contrary to the PEC response, unfractionated SCs from the same animals responded very well to mitogens and CA antigens and the proliferative response was significantly higher compared to that of SC from control mice. In conclusion, these results cast some light on the mechanisms by which NK cells and macrophages regulated the development of the local specific response to CA: activated NK cells, by producing IFN-gamma, favor the development of TH1 subset, while suppressor macrophages keep proliferation of T lymphocytes under control because of the presence of highly activated NK cells.