Adoptive transfer of CD8+ T cells from immune animals does not transfer immunity to blood stage Plasmodium yoelii malaria

The malaria parasite, Plasmodium yoelii 17X, causes a self-limited, nonlethal infection characterized, in the blood stage, by preferential invasion of reticulocytes. Previous studies have suggested that immunity to the blood stage infection may be related to enhanced levels of class I MHC Ag on the parasitized reticulocyte surface and can be adoptively transferred to immunodeficient mice by immune CD8+ T cells in the absence of CD4+ T cells. To further examine the mechanisms of CD8+ T cell involvement in immunity to blood stage P. yoelii infection, we performed in vivo CD8 depletion and adoptive transfer experiments. Depletion of CD8+ T cells during primary blood stage infection in BALB/c mice did not diminish the ability of the mice to resolve their infections. Spleen cells from immune BALB/c and C57BL/10 mice were transferred to BALB/c-nu/nu and C57BL/10-nu/nu mice, respectively. The recipient mice were CD4 depleted in vivo to kill any transferred CD4+ T cells. The mice failed to control the infection. Populations of CD4-, CD8+ T cells were transferred from immune CBA/CaJ donors to in vivo CD4-depleted CBA/CaJ recipients. The mice were unable to control the infection. Although immune unfractionated spleen cells transferred rapid protection in all three mouse strains and immune CD4+ T cells transferred immunity in the two mouse strains studied, CD8+ T cells by themselves were neither protective nor did they enhance immunity.     

CD4+ T cells acting independently of antibody contribute to protective immunity to Plasmodium chabaudi infection after apical membrane antigen 1 immunization

Apical membrane Ag 1 (AMA1) is a leading malaria vaccine candidate. Homologues of AMA1 can induce protection in mice and monkeys, but the mechanism of immunity is not understood. Mice immunized with a refolded, recombinant, Plasmodium chabaudi AMA1 fragment (AMA1B) can withstand subsequent challenge with P. chabaudi adami. Here we show that CD4+ T cell depletion, but not gammadelta T cell depletion, can cause a significant drop in antiparasite immunity in either immunized normal or immunized B cell KO mice. In normal mice, this loss of immunity is not accompanied by a decline in Ab levels. These observations indicate a role for AMA1-specific Ab-independent T cell-mediated immunity. However, the loss of immunity in normal CD4+ T cell-depleted mice is temporary. Furthermore, immunized B cell KO mice cannot survive infection, demonstrating the absolute importance of B cells, and presumably Ab, in AMA1-induced immunity. CD4+ T cells specific for a cryptic conserved epitope on AMA1 can adoptively transfer protection to athymic (nu/nu) mice, the level of which is enhanced by cotransfer of rabbit anti-AMA1-specific antisera. Recipients of rabbit antisera alone do not survive. Some protected recipients of T cells plus antisera do not develop their own AMA 1-specific Ab response, suggesting that AMA 1-specific CMI alone can protect mice. These data are the first to demonstrate the specificity of any protective CMI response in malaria and have important implications for developing a malaria vaccine.     

IL-12 and NK cells are required for antigen-specific adaptive immunity against malaria initiated by CD8+ T cells in the Plasmodium yoelii model.

CD8+ T cells have been implicated as critical effector cells in protection against preerythrocytic stage malaria, including the potent protective immunity of mice and humans induced by immunization with radiation-attenuated Plasmodium spp. sporozoites. This immunity is directed against the Plasmodium spp. parasite developing within the host hepatocyte and for a number of years has been presumed to be mediated directly by CD8+ CTL or indirectly by IFN-gamma released from CD8+ T cells. In this paper, in BALB/c mice, we establish that after immunization with irradiated sporozoites or DNA vaccines parasite-specific CD8+ T cells trigger a novel mechanism of adaptive immunity that is dependent on T cell- and non-T cell-derived cytokines, in particular IFN-gamma and IL-12, and requires NK cells but not CD4+ T cells. The absolute requirement for CD8+ T cells to initiate such an effector mechanism, and the requirement for IL-12 and NK cells in such vaccine-induced protective immunity, are unique and underscore the complexity of the immune responses that protect against malaria and other intracellular pathogens.     

BALB/c小鼠感染不同疟原虫CD4＋ Th应答和凋亡特点的比较研究

探讨不同疟原虫感染BALB/c小鼠的免疫应答特点。BALB/c小鼠经腹腔注射致死型约氏疟原虫（P.y17XL）和夏氏疟原虫（P.cAS）感染的红细胞,计数红细胞感染率;ELISA动态检测感染小鼠脾细胞培养上清中IFN-γ和IL-4水平;流式细胞术检测脾中CD4＋T细胞凋亡数量。约氏疟原虫（P.y17XL）感染早期,小鼠原虫血症持续上升;IFN-γ和IL-4分泌水平仅在感染后第3天出现一过性有意义的升高,而且峰值较低;脾中CD4＋T细胞大量凋亡,小鼠全部死亡;而夏氏疟原虫（P.cAS）感染小鼠,原虫血症上升缓慢;IFN-γ分泌水平在感染后第5天达峰值;IL-4分泌水平在感染后第10天达峰值,且峰值较高维持时间较长;脾中CD4＋T细胞凋亡细胞于感染后8 d出现有意义升高,小鼠全部存活。抗疟保护性免疫有赖于Th1和Th2型免疫应答的有效建立和协调过渡,感染期间CD4＋T细胞凋亡的时相和数量可能是影响免疫应答的强度或引起宿主免疫抑制的原因,从而影响宿主疟原虫感染的结局。     

Requirement for NK cells in CD40 ligand-mediated rejection of Philadelphia chromosome-positive acute lymphoblastic leukemia cells.

We have previously developed a murine model of Philadelphia chromosome-positive acute lymphoblastic leukemia by i.v. injection of a pre-B ALL cell line (BM185) derived from Bcr-Abl-transformed BALB/c bone marrow. We are studying the potential to elicit autologous antileukemic immune responses by introducing genes encoding immunomodulators (CD40 ligand (CD40L), CD80, and GM-CSF) into leukemia cells. BM185 cells expressing CD40L or CD80 alone, when injected into BALB/c mice, were rejected in approximately 25% of mice, whereas cohorts receiving BM185 cells expressing two or more immunomodulator genes rejected challenge 50-76% of the time. The greatest protection was conferred in mice receiving BM185 cells expressing all three immunomodulators. Addition of murine rIL-12 treatments in conjunction with BM185/CD80/CD40L/GM-CSF vaccination allowed rejection of preestablished leukemia. BM185 cell lines expressing CD40L were rejected in BALB/c nu/nu (nude) mice, in contrast to cell lines expressing CD80 and/or GM-CSF. Nude mice depleted of NK cells were no longer protected when challenged with BM185/CD40L, demonstrating a requirement for NK cells. Similarly, NK cell depletion in immunocompetent BALB/c mice resulted in a loss of protection when challenged with BM185/CD40L, confirming the data seen in nude mice. The ability of CD40L to act in a T cell-independent manner may be important for clinical applications in patients with depressed cellular immunity following chemotherapy.     

Role of L3T4+ and LyT-2+ cells in experimental visceral leishmaniasis

In contrast to euthymic (nu/+) BALB/c mice, athymic nude (nu/nu) BALB/c mice fail to control the visceral intracellular replication of Leishmania donovani, do not generate the macrophage-activating lymphokine IFN-gamma, and show little or no granulomatous tissue response. To characterize the T cell requirement for successful defense against L. donovani, nude mice were first reconstituted with unfractionated nu/+ immune spleen cells, which readily conferred the capacity to control and eliminate visceral (hepatic) L. donovani. In reconstituted mice, acquired resistance was paralleled by the ability of spleen cells to generate high levels of leishmanial Ag-stimulated IFN-gamma and the development of well formed liver granulomas. In contrast, nude mice reconstituted with either L3T4+- or Lyt-2+-enriched immune spleen cells alone failed to control visceral parasite replication and did not develop effective granulomas despite the finding that transfer of L3T4+ cells largely and Lyt-2+ cells partially restored the capacity to secrete IFN-gamma. To determine whether both T cell subsets were also required in a normal host, nu/+ BALB/c mice were treated with cell-depleting anti-L3T4 and anti-Lyt-2 mAb. Depletion of either T cell subset inhibited the acquisition of resistance to L. donovani and impaired the tissue granulomatous response. Thus, successful T cell-dependent host defense towards intracellular L. donovani and the tissue expression (granulomas) of this mechanism appear to require both L3T4+ and Lyt-2+ cells. A primary role for the L3T4+ cell may be IFN-gamma production; the role of the Lyt-2+ cell and the precise interaction of the two T cell subsets remain to be identified.     

Central neuropathogenesis of vesicular stomatitis virus infection of immunodeficient mice.

To determine whether central neuropathogenesis associated with vesicular stomatitis virus (VSV) infection is regulated by T cells, we have examined the effects of intranasal infection of mice lacking T cells. The mice examined were of two kinds: (i) thymus-deficient BALB/c nu/nu nice and (ii) BALB/c mice experimentally depleted of T cells by systemic infusions of a monoclonal antibody to the CD4 or CD8 cell surface molecules. These mice were infected intranasally with a single dose of replication-competent VSV. Brain tissue homogenates were analyzed for the presence of infectious virus. For each population of mice, infection-related mortality was assessed. In histological sections of brain, the distribution of viral antigens (Ags) was examined by immunocytochemistry. We found that recovery of infectious virus from homogenates of tissues obtained from athymic nu/nu animals was more than 10 times greater than that from samples from their euthymic littermates. With a single exception in a BALB/c nu/nu mouse, virus was not isolated from the spleen when it was administered intranasally. In these experimental infections, athymic mice succumbed 1 to 2 days before their euthymic littermates. A dose of virus that resulted in half of the nu/+ survival rate was uniformly lethal to nu/nu mice. In experiments with BALB/c mice depleted of either CD4+ or CD8+ T cells by in vivo antibody treatment, histological analysis revealed an increase in viral Ag distribution in comparison with control (medium-infused) infected mice. Necrosis and inflammation paralleled the extent of viral Ag expression. Viral Ags were detected in discrete areas that usually remain uninfected in immunocompetent mice. These areas include the neocortex and caudate putamen nuclei, the piriform cortex, and the lateral olfactory tract. Neuronal loss and necrosis were consistently found in the olfactory bulb and the horizontal/vertical band of Broca. In some of the T-cell depleted mice, necrosis was also evident in the hippocampus, fimbria, mammillary bodies, and hypothalamic nuclei. In the brain stem, perivascular cuffing was evident, but with little necrosis. Collectively, these data suggest that CD4+ and CD8+ T cells make only a minor contribution to the development of histopathology but rather function together to limit viral replication and transsynaptic or ventricular spread of virus, thus promoting recovery. The primary effectors of histopathology appear to be related more to the cytopathologic nature of the virus infection and non-T-cell-mediated mechanisms.     

The early IL-4 response to Leishmania major and the resulting Th2 cell maturation steering progressive disease in BALB/c mice are subject to the control of regulatory CD4+CD25+ T cells

Susceptibility and development of Th2 cells in BALB/c mice infected with Leishmania major result from early IL-4 production by Vbeta4Valpha8 CD4+ T cells in response to the Leishmania homolog of mammalian RACK1 Ag. A role for CD4+CD25+ regulatory T cells in the control of this early IL-4 production was investigated by depleting in vivo this regulatory T cell population. Depletion induced an increase in the early burst of IL-4 mRNA in the draining lymph nodes of BALB/c mice, and exacerbated the course of disease with higher levels of IL-4 mRNA and protein in their lymph nodes. We further showed that transfer of 10(7) BALB/c spleen cells that were depleted of CD4+CD25+ regulatory T cells rendered SCID mice susceptible to infection and allowed Th2 differentiation while SCID mice reconstituted with 10(7) control BALB/c spleen cells were resistant to infection with L. major and developed a Th1 response. Treatment with a mAb against IL-4 upon infection with L. major in SCID mice reconstituted with CD25-depleted spleen cells prevented the development of Th2 polarization and rendered them resistant to infection. These results demonstrate that CD4+CD25+ regulatory T cells play a role in regulating the early IL-4 mRNA and the subsequent development of a Th2 response in this model of infection.     

Frequency of natural regulatory CD4+CD25+ T lymphocytes determines the outcome of tolerance across fully mismatched MHC barrier through linked recognition of self and allogeneic stimuli

We show in this study that long-term tolerance to allogeneic skin grafts can be established in the absence of immunosuppression by the combination of the following elements: 1) augmenting the frequency of regulatory CD4(+)CD25(+) T cells (Treg) and 2) presentation of the allogeneic stimuli through linked recognition of allo- and self-epitopes on semiallogeneic F(1) APCs. BALB/c spleen cells enriched for CD4(+)CD25(+) T lymphocytes were transferred either to BALB/c nu/nu mice or to BALB/c nu/nu previously injected with F(1)(BALB/c x B6.Ba) spleen cells, or else grafted with F(1)(BALB/c x B6.Ba) skin (chimeric BALB/c nu/nu-F(1)). Chimeric BALB/c nu/nu-F(1) reconstituted with syngeneic CD25(+)-enriched spleen cells were unable to reject the previously transferred F(1)(BALB/c x B6.Ba) spleen cells or F(1)(BALB/c x B6.Ba) skin grafts, and a specific tolerance to a secondary B6 graft was obtained, with rejection of third-party CBA grafts. BALB/c nu/nu mice reconstituted only with syngeneic CD25(+)-enriched spleen cells rejected both B6 and CBA skin grafts. In contrast, when chimeric BALB/c nu/nu-F(1) were reconstituted with spleen populations comprising normal frequencies of Treg cells, the linked recognition of allo and self resulted in breaking of self tolerance and rejection of syngeneic grafts, strongly suggesting that linked recognition works in both directions, either to establish tolerance to allo, or to break tolerance to self, the critical parameter being the relative number of Treg cells.     

Effector phenotypes and mechanisms of antitumor immune reactivity of tumor-immunized and tumor-bearing mice in two syngeneic tumors.

By using two different syngeneic tumors, Meth A sarcoma and RL male 1 lymphoma of BALB/c origin, the present study was designed to investigate the subset(s) of T cells mediating in vivo antitumor immune responses and some of the effector mechanisms of in vivo protective immunity in BALB/c mice immunized against tumor or bearing tumor. Spleen cells from the mice immunized against Meth A tumor or bearing Meth A tumor inhibited the growth of Meth A tumor in the Winn assay. In the Meth A-immunized mice, L3T4+ (CD4+) cells played a major role in mediating the inhibitory activity against Meth A tumor growth, whereas in the Meth A-bearing mice, the antitumor protective immunity was mediated by both L3T4+ and Lyt-2+ (CD8+) cells. Spleen cells from the Meth A-immunized or Meth A-bearing mice were not able to generate cytotoxic T lymphocytes (CTL) directed against Meth A tumor after the in vitro restimulation of spleen cells with mitomycin C (MMC)-treated Meth A cells, while fresh spleen cells from the Meth A-immunized or Meth A-bearing mice were able to induce the strong delayed-type hypersensitivity (DTH) responses to Meth A tumor. The DTH response to Meth A tumor was mediated by L3T4+ cells in the Meth A-immunized mice and by both L3T4+ and Lyt-2+ cells in the Meth A-bearing mice. In the similar experiments performed in the RL male 1 lymphoma, the antitumor activity in spleen cells from the RL male 1-immunized or RL male 1-bearing mice depended on Lyt-2+ but not L3T4+ cells in the Winn assay. When spleen cells from the RL male 1-immunized or RL male 1-bearing mice were cultured with MMC-treated RL male 1 cells for 5 days, an appreciable CTL response to RL male 1 tumor was induced. These results suggest that the nature of tumor and/or tumor antigens determines which T cell subset is required to exhibit the protective immunity against tumor and thus the different effector mechanisms could be induced in the different tumor models. Furthermore, these data support the conclusion that antitumor T cell responses are affected by the immune state of host to tumor.     

Changes in proliferation activity and relative distributions of lymphoid cell subpopulations in congenitally athymic nu/nu mice and lurcher mice with spontaneous olivopontocerebellar degeneration

Changes observed in mice with congenital damage of some part of the CNS-neuroendocrine-immune regulatory system are described. nu/nu mice with congenital absence of thymus and Lurcher mice with spontaneous olivopontocerebellar degeneration displayed changes in the histoarchitecture of adrenal gland, immune organs (thymus, spleen, axillar lymph nodes) and intestine. Changes were also observed in IgM+, IgG+, CD4+ and CD8+ lymphoid cell subpopulations in the main lymphoid organs--the spleen and axillar lymph nodes and in the proliferative ability of whole lymphoid cell populations. The extreme decrease of lymphoid T-cell subpopulations in athymic nu/nu mice is the consequence of the absence of thymus, the organ of their maturation. On the other hand, a relative increase of B-cell subpopulations was found in this mouse strain. A relative decrease of CD4+ lymphocytes and a different influence of immunization on B-cell subpopulations were found in the spleen in neurodeficient Lurcher mice. The high percentage of apoptotic cells, cells in the S-phase of cell cycle and increased proliferation index in nu/nu mice suggest that the turnover and renewal of lymphoid cells in the spleen in nu/nu mice is more rapid than in control immunocompetent BALB/c mice.     

Different subsets of T cells in the adult mouse bone marrow and spleen induce or suppress acute graft-versus-host disease.

Fractionation of normal adult mouse spleen and bone marrow cells (C57BL/Ka) was performed by discontinuous Percoll density gradients. The fractionated low density (1.050-1.060 g/ml) C57BL/Ka spleen cells completely suppressed acute lethal graft vs host disease (GVHD) when coinjected with unfractionated C57BL/Ka spleen cells into sublethally irradiated (400 rad) BALB/c mice. In dose response experiments, as few as 0.5 x 10(6) low density cells from the spleen fractions suppressed acute GVHD induced by 2.5 x 10(6) unfractionated allogeneic spleen cells. Although the low density spleen fractions inhibited acute GVHD, the high density (1.075-1.090 g/ml) spleen fractions induced acute GVHD in sublethally irradiated BALB/c recipients. Fractionation of C57BL/Ka bone marrow cells showed that none of the high or low density fractions or unfractionated cells induced lethal GVHD. When these fractions were tested for their capacity to suppress GVHD by coinjection with C57BL/Ka unfractionated spleen cells, all fractions protected the BALB/c recipients. Unfractionated bone marrow cells showed modest protection. Evaluation of the dose response characteristics of the suppressive activity of the low and middle density (1.060-1.068 g/ml) bone marrow cell fraction showed that reproducible protection could be achieved at a 5:1 ratio of inducing to suppressing cells. The low density fractions of both bone marrow and spleen cells had a marked depletion of typical TCR(+)-alpha beta CD4+ or CD8+ T cells, and a predominant population of TCR(+)-alpha beta CD4- CD8- T cells. Purified populations of the latter cells suppressed GVHD. Recipients given unfractionated C57BL/Ka spleen cells and protected with low-density bone marrow or spleen cells were chimeras.     

Recombinant viral-vectored vaccines expressing Plasmodium chabaudi AS apical membrane antigen 1: mechanisms of vaccine-induced blood-stage protection

Apical membrane Ag 1 (AMA1) is one of the leading candidate Ags for inclusion in a subunit vaccine against blood-stage malaria. However, the efficacy of Ab-inducing recombinant AMA1 protein vaccines in phase IIa/b clinical trials remains disappointing. In this article, we describe the development of recombinant human adenovirus serotype 5 and modified vaccinia virus Ankara vectors encoding AMA1 from the Plasmodium chabaudi chabaudi strain AS. These vectors, when used in a heterologous prime-boost regimen in BALB/c mice, are capable of inducing strong transgene-specific humoral and cellular immune responses. We show that this vaccination regimen is protective against a nonlethal P. chabaudi chabaudi strain AS blood-stage challenge, resulting in reduced peak parasitemias. The role of vaccine-induced, AMA1-specific Abs and T cells in mediating the antiparasite effect was investigated by in vivo depletion of CD4(+) T cells and adoptive-transfer studies into naive and immunodeficient mice. Depletion of CD4(+) T cells led to a loss of vaccine-induced protection. Adoptive-transfer studies confirmed that efficacy is mediated by both CD4(+) T cells and Abs functioning in the context of an intact immune system. Unlike previous studies, these results confirm that Ag-specific CD4(+) T cells, induced by a clinically relevant vaccine-delivery platform, can make a significant contribution to vaccine blood-stage efficacy in the P. chabaudi model. Given that cell-mediated immunity may also contribute to parasite control in human malaria, these data support the clinical development of viral-vectored vaccines that induce both T cell and Abs against Plasmodium falciparum blood-stage malaria Ags like AMA1.     

CTLA-4 and CD4+ CD25+ regulatory T cells inhibit protective immunity to filarial parasites in vivo

The T cell coinhibitory receptor CTLA-4 has been implicated in the down-regulation of T cell function that is a quintessential feature of chronic human filarial infections. In a laboratory model of filariasis, Litomosoides sigmodontis infection of susceptible BALB/c mice, we have previously shown that susceptibility is linked both to a CD4+ CD25+ regulatory T (Treg) cell response, and to the development of hyporesponsive CD4+ T cells at the infection site, the pleural cavity. We now provide evidence that L. sigmodontis infection drives the proliferation and activation of CD4+ Foxp3+ Treg cells in vivo, demonstrated by increased uptake of BrdU and increased expression of CTLA-4, Foxp3, GITR, and CD25 compared with naive controls. The greatest increases in CTLA-4 expression were, however, seen in the CD4+ Foxp3- effector T cell population which contained 78% of all CD4+ CTLA-4+ cells in the pleural cavity. Depletion of CD25+ cells from the pleural CD4+ T cell population did not increase their Ag-specific proliferative response in vitro, suggesting that their hyporesponsive phenotype is not directly mediated by CD4+ CD25+ Treg cells. Once infection had established, killing of adult parasites could be enhanced by neutralization of CTLA-4 in vivo, but only if performed in combination with the depletion of CD25+ Treg cells. This work suggests that during filarial infection CTLA-4 coinhibition and CD4+ CD25+ Treg cells form complementary components of immune regulation that inhibit protective immunity in vivo.     

High accumulation of T regulatory cells prevents the activation of immune responses in aged animals

In our previous in vivo study we demonstrated that young BALB/c mice effectively rejected the BM-185 tumor cells expressing enhanced GFP (EGFP) as a surrogate tumor Ag. In contrast, old BALB/c mice succumbed to the BM-185-EGFP tumors, indicating that there is a deficiency in old animals preventing the rejection of immunogenic tumors. There is cumulative evidence indicating that regulatory T (T(reg)) cells control the activation of primary and memory T cell responses. However, very little is known about whether there is a relation between T(regs) and the lack of immune responses in the aged. We evaluated young and aged animals, and our results demonstrated that there are significantly more CD4+CD25+FoxP3+ and CD8+CD25+FoxP3+ T(regs) in the spleen and lymph nodes of old animals when compared with the young. Depletion of CD25+ cells with anti-CD25 mAb induces the rejection of BM-185-EGFP cells, restores antitumor T cell cytotoxic activity, and results in the generation of a protective memory response against the BM-185 wild-type tumors in old mice. Furthermore, vaccination with CpG-oligodeoxynucleotide decreases the number of T(reg) cells in old animals to the same levels as young mice, restoring the primary and memory antitumor immune responses against BM-185-EGFP tumors. Taken together, these results indicate that there is a direct correlation between the expansion of T(reg) cells and immune deficiency in the old, and that depletion of these cells might be critical for restoring immune responses in aged animals.     

Evidence for the functional binding in vivo of tumor rejection antigens to antigen-presenting cells in tumor-bearing hosts

Spleen cells of BALB/c mice bearing a syngeneic CSA1M fibrosarcoma were treated with anti-Thy-1.2 antibody plus C, yielding a T cell-depleted, APC-containing fraction. The APC-containing fraction was first tested for its capacity to present exogenous modified-self or another tumor (Meth A) Ag after in vitro pulsing. The results showed comparable Ag-presenting capacities to those obtained by APC-containing fraction from normal spleen cells, indicating that APC function is not affected in tumor-bearing mice. We next examined whether APC from CSA1M-bearing mice bind endogenously generated CSA1M tumor Ag onto its surfaces to stimulate tumor-specific T cells. Five rounds of inoculation of APC-containing fraction from CSA1M-bearing mice without further in vitro pulsing resulted in the induction of potent anti-CSA1M immune resistance. The involvement of anti-CSA1M T cells in the induction of anti-CSA1M immunity was excluded by the fact that the in vivo immunity was excluded by the fact that the in vivo immunity was delivered by Thy-1+ cell-depleted, but not by Thy-1+ cell-enriched fractions of spleen cells from CSA1M-bearing mice. Moreover, the failure of Sephadex G10-passed spleen cells to deliver anti-CSA1M resistance demonstrated the absolute requirement of APC for inducing the in vivo immunity. Finally, this in vivo resistance was found to be tumor specific, because APC fractions from CSA1M-bearing and Meth A-bearing BALB/c mice induced immune resistance selective against the corresponding tumor cell challenge. These results indicate that APC from tumor-bearing hosts can not only exert unaffected APC function against exogenous Ag, but also function to present tumor Ag generated endogenously in the tumor-bearing state and to produce tumor-specific immunity in vivo.     

Susceptibility of nude mice carrying the Fv-4 gene to Friend murine leukemia virus infection.

Fv-4 is a mouse gene that dominantly confers resistance to infection with Friend murine leukemia virus (F-MuLV) (S. Suzuki, Jpn. J. Exp. Med. 45:473-478, 1975). Despite complete resistance to ecotropic MuLV infection in mice carrying the Fv-4 gene, it is known that cells carrying the resistance gene in tissue culture do not always show resistance as extensive as that in vivo (H. Yoshikura and T. Odaka, JNCI 61:461-463, 1978). To investigate the immunological effect on resistance in vivo, we introduced the Fv-4 gene into BALB/c nude mice (Fv-4-/- nude[nu/nu]) by mating them with Fv-4 congenic BALB/c mice (Fv-4r/r nude+/+) and examined the susceptibility of the F2 progeny to F-MuLV. All BALB/c nude mice without the Fv-4 gene (Fv-4-/- nude[nu/nu]) were permissive to F-MuLV and developed erythroleukemia within 2 weeks after virus inoculation. The BALB/c nude mice with the Fv-4 gene (Fv-4r/r nude[nu/nu]) did not develop leukemia, and no or little virus was detected in the spleen 7 weeks after virus inoculation. The resistance to F-MuLV was dominant in (Fv-4 congenic BALB/c x BALB/c nude) F1 mice with the Fv-4r/- nude(nu/+) genotype as strictly as in (Fv-4 congenic BALB/c x BALB/c) F1 mice with the Fv-4r/- nude+/+ genotype. However, almost all BALB/c nude mice with the Fv-4r/- nude(nu/nu) genotype developed the disease within 7 weeks, and the virus was detected in all of their spleens even in the mice without leukemia. These results show that the resistance caused by the Fv-4 gene is recessive in nude mice and dominant in BALB/c mice. Some immunological effects, perhaps cell-mediated immunity, may play important roles in the resistance to F-MuLV infection in vivo in addition to the dosage effect of the Fv-4 product.     

Murine CD8+ cytotoxic T lymphocytes lyse Toxoplasma gondii-infected cells

Studies were performed to determine whether CTL against Toxoplasma gondii-infected cells could be induced in a murine model of T. gondii infection in which CD8+ T lymphocytes have been shown to play a major role in resistance against this parasite. In 51Cr release assays, nylon wool nonadherent spleen cells from BALB/c (H-2d) mice immunized with the temperature-sensitive (ts-4) mutant strain of T. gondii were cytotoxic for T. gondii-infected P815 (H-2d) mastocytoma cells but not for uninfected cells. This cytotoxic activity was remarkably increased after in vitro stimulation with T. gondii-infected syngeneic spleen cells. The effector cells were shown to be CD8+ T lymphocytes, because the cytotoxicity was significantly inhibited by depletion of CD8+ T lymphocytes but not by depletion of CD4+ T lymphocytes. This cytotoxic activity was genetically restricted. Spleen cells from T. gondii-immune BALB/c mice were not cytotoxic for T. gondii-infected EL4 (H-2b) thymoma cells, whereas spleen cells from T. gondii-immune C57B1/6 (H-2b) mice were cytotoxic for T. gondii-infected EL4 cells but not for T. gondii-infected P815 cells. The cytolytic activity of CD8+ T lymphocytes against T. gondii-infected cells might be a mechanism whereby these cells confer resistance against T. gondii.     

Immunoregulatory function of CD3+, CD4-, and CD8- T cells. Gamma delta T cell receptor-positive T cells from nude mice abrogate oral tolerance

Previous work has shown that abrogation of oral tolerance is mediated by T cells which are found in the CD3+, L3T4- (CD4-), and Lyt-2- (CD8-) subset (termed double-negative; DN) in mice. Inasmuch as it is known that athymic, nude (nu/nu) mice possess Thy 1+, CD4-, and CD8- T cells which also exhibit a functionally rearranged TCR gamma-chain, we investigated whether this subset of nude T cells contained functional immunoregulatory cells. In this report, we examined the phenotype and distribution of CD3+ T cells in the spleen and in the mesenteric and peripheral lymph nodes of BALB/c nu/nu mice in comparison with normal mice (+/+). In the spleens of nude mice, the predominant CD3+ T cell subpopulation was DN. Further, in mesenteric and peripheral lymph nodes, approximately one-third and one-half of the CD3+ T cells were double negative, respectively. In contrast, CD3+, DN T cells represent a small subpopulation in normal (+/+) mice. We next showed that functional regulatory T cells which possess the ability to abrogate oral tolerance were induced in nu/nu mice by Ag priming. BALB/c nude mice were immunized with SRBC, and the splenic CD3+, Vicia villosa-adherent cells were obtained by panning. Adoptive transfer of CD3+, V. villosa-adherent T cells to orally tolerant BALB/c mice restored responsiveness to SRBC, whereas V. villosa nonadherent cells were without effect. In other experiments, CD3+ T cells from the spleens of SRBC-primed mice were further enriched for the CD5+, DN phenotype and adoptive transfer of this subset completely abrogated oral tolerance to SRBC. To characterize the nature of the TCR expressed on these CD3+, DN T cells, we developed a rabbit antibody to a synthetic peptide (residues 209-218: Tyr-Ala-Asn-Ser-Phe-Asn-Asn-Glu-Lys-Leu) which was synthesized from a deduced sequence of the murine delta-gene. Immunoprecipitation of a cell membrane fraction from CD3+, DN T cells with anti-delta TCR antibody isolated a 45-kDa band. Furthermore, immunoprecipitation of these cells with anti-CD3 (145-2C11) revealed bands at 45 and 35 kDa (corresponding to delta- and gamma-chains, respectively). Taken together, these results are the first to show that gamma delta-TCR bearing CD3+, CD4-, and CD8- T cells are functional and reverse oral tolerance when adoptively transferred.     

Genetic control of immunity to Plasmodium yoelii sporozoites

Using a rodent malaria system, we have shown that protective immunity to the preerythrocytic stages of malaria is genetically controlled by MHC and non-MHC genes. Ten congenic strains of mice were immunized with irradiated sporozoites of Plasmodium yoelii. When challenged with viable sporozoites, only two strains had a high proportion of animals that did not develop blood stage infections. Immunity did not correlate with antisporozoite antibody levels. Two protective mechanisms exist determined by non-H-2 genes, and each mechanism is further controlled by H-2-linked Ir genes. On the BALB background only H-2d mice are protected, and protection is abolished by depleting CD8+ T cells. In contrast, on the B10 background only H-2q mice are strongly protected, and protection is not affected by CD8+ T cell depletion. If similar complex genetic regulation of immunity occurs in the human malarias, it will be a major hurdle for vaccine development.