Adoptive transfer of immunity to Plasmodium berghei with spleen cells from drug cured mice

Plasmodium berghei infection gives rise to a fatal fulminating parasitaemia in mice. Resistance to the infection can be achieved if the mice are allowed to recover from the blood-induced infection by administration of chloroquine. In CBA/Caj, one cycle and in Balb/c mice, at least, two cycles of infection, alternating with drug-cure were necessary for the establishment of immunity. No parasitaemia was seen following further challenges after the third infection. Adoptive transfer of spleen cells from the immune mice showed that immunity could be transferred by 50 to 70 x 10(6) cells. On challenge the recipients develop high parasitaemia which was cleared in less than 5 weeks. Recipient mice given low number of spleen cells did not survive. The transfer of immune spleen cells depleted of either T or B lymphocytes abrogated the protective effect. Comparatively T lymphocytes were less effective in transferring protection than B lymphocytes. The study suggests that both humoral and cell-mediated effector mechanisms are needed for the maintenance of immunity in drug-cured immune mice.     

Both CD4+ and CD8+ T cells can mediate vaccine-induced protection against Coccidioides immitis infection in mice

To determine which lymphocytes are required for vaccine-induced immunity to coccidioidomycosis, we used a temperature-sensitive mutant of Coccidioides immitis to immunize mice lacking subsets of lymphocytes or specific cytokines and infected the mice 4 weeks later with virulent C. immitis. After 2 weeks, we determined the number of fungi in their lungs and spleens. Vaccine-induced immunity required alpha beta T lymphocytes. beta -2 microglobulin knockout (KO) mice were protected by immunization, and we transferred protection using CD4+ T cells from immunized mice. However, vaccination also protected CD4+ KO mice, which suggests that CD8+ T cells played a role in vaccine-induced immunity, even though they were not required. We adaptively transferred protection using spleen cells from immunized CD4+ KO mice to nonimmune B6 mice, but CD8+ -depleted spleen cells did not protect against infection. Recipients of spleen cells from immunized CD4+ KO mice had 6 times more tumor necrosis factor (TNF)- alpha mRNA in their lungs than did mice that received nonimmune spleen cells, and TNF receptor-1 KO mice were not fully protected by immunization. These results show that both CD4+ and CD8+ T cells can protect against coccidioidomycosis and that TNF- alpha is a necessary component of the acquired immune response.     

Significance of thymus-derived lymphocytes in immunity elicited by immunization with ribosomes or live yeast cells of Histoplasma capsulatum.

The lymphoid cells responsible for protective immunity to histoplasmosis were characterized. Adoptive transfer of spleen and peritoneal cells treated with antiserum to theta-antigen from mice immunized with ribosomes or live yeast cells of Histoplasma capsulatum abrogated the ability of these cells to protect the syngeneic recipients, whereas treatment of lymphoid cells with antiserum to IgG did not affect the immunity. Prior removal of glass-adhering cells from spleen and peritoneal cell suspensions did not alter their protective activity. Treatment with mitomycin C, an antimitotic agent, ablated the capacity of immune lymphocytes to protect the syngeneic recipients. These results indicate that the immune spleen and peritoneal cells that confer immunity to histoplasmosis are thymus-dependent (T) lymphocytes and that their active proliferation in the recipients is necessary for expression of the protective immunity. Furthermore, the immunity elicited by immunization with histoplasma ribosomes and live yeast cells is mediated by a similar mechanism.     

Immune spleen cell-mediated protection against fatal Hantaan virus infection in infant mice

To clarify the basis of the age-dependent susceptibility of infant mice to fatal meningoencephalitis caused by Hantaan virus, we investigated the ability of spleen cells from immune and nonimmune weanling BALB/c mice to confer protection to syngeneic infant mice. Intraperitoneal transfer of 10 X 10(7), 5 X 10(7), 2.5 X 10(7), and 1.2 X 10(7) immune spleen cells to infant mice 24 hr after intracerebral challenge with 100 50% lethal doses of Hantaan virus (strain 76-118) resulted in 100%, 70%, 64%, and 30% protection, respectively. Even as late as 48 hr after virus challenge, transfer of 10 X 10(7) immune spleen cells conferred complete protection. In contrast, nonimmune spleen cells offered no protection, even when cells were transferred 48 hr before virus challenge. The protective capacity of immune spleen cells was abolished following treatment with antibody to theta antigen and guinea pig complement, but was preserved after the depletion of B cells, a result suggesting that T cells play a crucial role in the resistance of mice to fatal Hantaan virus infection.     

T细胞亚群在感染约氏疟原虫小鼠中的保护性免疫作用

对约氏疟原虫有保护性免疫的BALB/c小鼠,体内注射单抗去除CD~(4 )或CD~(8 )T细胞后,对保护性免疫无明显影响。但将它们脾细胞中CD~(8 )T细胞注入裸鼠,可以转移部分免疫力,而注入CD~(4 )T细胞则不能。在约氏疟原虫感染小鼠模型中,由于在感染早期原虫侵入网织红细胞,CD~(8 )T细胞可被激活而有杀伤作用。在晚期,抗体在免疫中起主要作用。     

Anti-Thy-1 treated and irradiated spleen cells from (BALB/cXC57Bl/6) F1 mice infected with Plasmodium chabaudi chabaudi can transfer protection into irradiated hosts

The transfer of spleen cells from (BALB/cXC57Bl/6) F1 mice recovered from a Plasmodium chabaudi chabaudi AS infection into irradiated syngeneic recipients conferred protection. Neither elimination of Thy-1⁺ cells nor in vitro irradiation of immune cells before transfer affected protection while both anti-Thy-1 treatment and irradiation abolished the appearance of anti-P. c. chabaudi antibodies in the recipients. Superinfection of immune spleen cell donors did not improve their capability to transfer protection which was also unaffected by anti-Thy-1 treatment. The serum of mice after one infection was only marginally protective when transferred into irradiated recipients and a second infection improved the protective activity of serum which was not further improved by six infections. The cotransfer of immune serum and immune cells did not result in any synergistic effect. On the other hand, when P. c. chabaudi AS (BALB/cXC57B1/6)F1 infected mice were challenged with a high dose of Plasmodium yoelii 17XL at crisis, the mice were unable to control the heterologous parasite. When mice were challenged with P. yoelii 17XL several weeks after infection with P.c. chabaudi AS, a good degree of cross-protection was observed.     

特异性IgG抗体作为宿主抗约氏疟原虫再感染关键效应分子的研究

目的探讨再感染早期宿主保护性免疫应答的相关机制。方法采用致死型约氏疟原虫(Plasmodiumyoelii 17XL,Py17XL)感染抵抗型DBA/2小鼠,待全部小鼠自愈后用同种疟原虫株再次攻击。吉姆萨薄血膜染色法观察小鼠原虫血症水平;被动转移实验评估免疫血清的保护性作用;流式细胞仪检测脾中T、B细胞亚群动态变化。结果2/3小鼠能够完全抵御同种疟原虫再感染,仅1/3小鼠出现一过性低水平原虫血症,且于再感染后第8d自愈;免疫血清显著抑制初次感染小鼠原虫血症水平并延长其生存期;与初次感染相比,再感染早期脾CD4+T细胞、B220+和B220lowCD138+(浆细胞)亚群数目显著增加。结论特异性IgG抗体在宿主抗再感染免疫中发挥重要作用,记忆性T、B细胞可能为宿主保护性免疫应答的关键参与者。     

The nature of immunity against Trypanosoma cruzi in mice recovered from acute infection

Protective immunity against a lethal, Y strain, T. cruzi infection could be transferred to normal mice by either serum or spleen cells from mice which had recovered from the acute phase of infection. The ability of spleen cells to transfer immunity was abolished by B lymphocyte removal (anti-Ig column fractionation), but was relatively insensitive to T lymphocyte depletion (anti-Thy 1.2 plus complement) or macrophage removal (Sephadex G-10 fractionation). Immune spleen cells gave an anamnestic antibody response when injected together with T. cruzi antigen into lethally irradiated recipients and these antibodies conferred protection in a passive transfer system. T cell depletion reduced, but did not abolish, this antibody response. These data imply that the protective immunity of T. cruzi-convalescent mice is predominantly B cell-mediated with T cell involvement being restricted to a helper role.     

Inability of Plasmodium vinckei-immune spleen cells to transfer protection to recipient mice exposed to vaccine''vectors''or heterologous species of plasmodium

Mice can be immunized to Plasmodium vinckei by repeated infections followed by cure. Such immunity is dependent on CD4 T cells and an architecturally modified spleen, but has little requirement for antibody. Thus, athymic mice can be exposed to P. vinckei and cured, but do not develop immunity. They are resistant to challenge with parasites, however, if they are then given spleen cells from euthymic immunized animals. Such immune spleen cells, however, cannot transfer resistance to normal mice which have been exposed to BCG, Salmonella typhimurium, or vaccinia virus, and are only partially effective in transferring resistance to mice which have been previously immunized with heterologous plasmodia, P. yoelii, P. chabaudi and P. berghei. Mice exposed to varying numbers of irradiated P. vinckei-pRBC do not develop immunity and nor are such animals protected following adoptive transfer of immune spleen cells. Cellular immunity to malaria may not only be dependent on a population of immune CD4 T cells, but may require a specifically architecturally modified spleen which may not occur following either exposure to candidate vaccine vectors, heterologous plasmodia or non-viable homologous plasmodia.     

T cell-mediated immunity in murine malaria. II. Induction of protective immunity to P. chabaudi by antigen fed macrophages and antigen educated lymphocytes

We previously described assay systems for generating antigen specific proliferating T cells to P. chabaudi antigens. In the present study we examine whether the various sensitization approaches confer immunity against a cloned virulent strain IP-PCI of P. chabaudi. We present data indicating that effective specific protective immunity can be induced through P. chabaudi antigen fed macrophages and antigen educated spleen cells (initiator lymphocytes). The expression of this protective immunity is proposed to depend on (a) antigen presentation and/or accessory function of macrophages and (b) the subsequent activation of T cell functions related to protection. Indeed analysis of different macrophage populations revealed a correlation between the expression of Ia molecules and IL-1 secretion with their capacity to induce antigen specific T cells in vivo and subsequent protective immune mechanisms. Thus these results emphasize the critical functions of accessory cells in determining the outcome of malaria infections.     

T细胞在炭疽保护性抗原免疫中的作用——免疫T细胞继承转移试验

取炭疽保护性抗原(PA)免疫的小鼠脾脏,用尼龙棉柱过滤和EAC花环相结合的方法分离出免疫T细胞,继承转移给同系小鼠,可使受体鼠获得抗炭疽菌感染的保护力。     

Activated CD8+ T cells contribute to clearance of gastric Cryptosporidium muris infections

The role of CD4+ and CD8+ T lymphocytes in the development of a protective immune response against Cryptosporidium muris infection was studied by the reconstitution of severe combined immunodeficient (SCID) mice with well-defined populations of either naive or immune CD8+ or CD4+ T lymphocytes. Adoptive transfer of both naive and immune CD4+ T lymphocyte subpopulations protects SCID mice against cryptosporidiosis. Moreover, a significant biological impact of activated CD8+ T cells against gastric cryptosporidiosis was observed. The significant difference in the course and intensity of the infection in reconstituted SCID mice was found to be dependent on the protective function of both the CD4+ and CD8+ T-cell populations transferred. While SCID mice reconstituted with either immune or naive CD4+ or immune CD8+ T-cell subpopulations resolved the infection within 29, 37 and 51 days post-infection, respectively, those reconstituted with naive CD8+ T cells suffered from chronic infection similar to control SCID mice. Reconstitution with CD4+ T cells resulted in suppression of oocyst excretion and shortening of patent period in comparison with SCID mice reconstituted with CD8+ T cells. Thus, although CD4+ T cells are considered important in protective immunity, our results are the first to demonstrate the involvement of activated CD8+ T lymphocytes in the protection of mice against gastric cryptosporidiosis.     

Bacterial lipopolysaccharide activates suppressor B lymphocytes.

Lipopolysaccharide (LPS) extracted from the outer cell wall of Gram-negative bacteria modulates the immune response in vivo and in vitro. Depending on the experimental conditions, it may enhance or inhibit the production of humoral antibody. The pathway by which LPS suppresses antibody production is examined in this study. C57BL/6 spleen cells incubated with LPS (greater than 10 micrograms/ml) not only fail to produce antibody to sheep erythrocytes in vitro but also, when transferred 24 hr after stimulation with LPS, inhibit antibody production in spleen cells that were not treated with LPS. This observation suggested that LPS activates suppressor cells. We have identified a suppressor B cell as mediator of LPS-induced immune suppression and determined its cell surface antigen phenotype as Ig+, Ia+, CR+, Ly-B-2+,PC1-.LPS does not induce suppressor macrophages or suppressor T cells, nor are macrophages or T cells required for the generation of suppressor B cells by LPS.     

树突状细胞DNA多价疫苗抗日本血吸虫感染作用机制的研究

摘 要：目的 探讨树突状细胞（DC）DNA混合多价疫苗抗日本血吸虫感染保护性免疫作用机制。方法 BALB/c小鼠耳廓分别注射Sj26、Sj23和Sj14基因转染DC（A组）、Sj26基因转染DC（B组）、Sj23基因转染DC（C组）、Sj14基因转染DC（D组）、pcDNA3转染DC（E组）、未处理DC（F组）和RPMI-1640（G组）,共免疫3次,间隔2周,末次免疫后第2周,每鼠经皮肤感染40条日本血吸虫尾蚴。ELISA法检测血清特异性IgG抗体、干扰素-γ（IFN-γ）和白细胞介素-4（IL-4）水平,双夹心ELISA法检测脾淋巴细胞经ConA和可溶性虫卵抗原（SEA）刺激后培养上清中IFN-γ和IL-4水平,噻唑蓝（MTT）法检测脾淋巴细胞的增殖。结果 A组小鼠末次免疫后第2周血清特异性IgG抗体水平显著升高（与G组比较,P<0.001）。A组小鼠免疫后血清IFN-γ水平明显升高(P<0.01),而血清IL-4的水平,各组小鼠免疫前、后无明显变化。A组小鼠脾淋巴细胞经ConA和SEA刺激后诱生的IFN-γ水平显著增高,而IL-4水平显著降低(与G组比较,P<0.001)。A组小鼠脾淋巴细胞的刺激指数高于其他各组（与G组比较,P<0.001）。结论 体液免疫和细胞免疫共同参与了DC DNA混合多价疫苗诱导的保护性免疫作用,其中Th1型免疫应答在抗日本血吸虫感染的保护性免疫中起主要作用。 关键词：日本血吸虫;树突状细胞;DNA疫苗;保护性免疫     

Effective collaboration between marginal metallophilic macrophages and CD8+ dendritic cells in the generation of cytotoxic T cells

The spleen is the lymphoid organ that induces immune responses toward blood-borne pathogens. Specialized macrophages in the splenic marginal zone are strategically positioned to phagocytose pathogens and cell debris, but are not known to play a role in the activation of T-cell responses. Here we demonstrate that splenic marginal metallophilic macrophages (MMM) are essential for cross-presentation of blood-borne antigens by splenic dendritic cells (DCs). Our data demonstrate that antigens targeted to MMM as well as blood-borne adenoviruses are efficiently captured by MMM and exclusively transferred to splenic CD8⁺ DCs for cross-presentation and for the activation of cytotoxic T lymphocytes. Depletion of macrophages in the marginal zone prevents cytotoxic T-lymphocyte activation by CD8⁺ DCs after antibody targeting or adenovirus infection. Moreover, we show that tumor antigen targeting to MMM is very effective as antitumor immunotherapy. Our studies point to an important role for splenic MMM in the initial steps of CD8⁺ T-cell immunity by capturing and concentrating blood-borne antigens and the transfer to cross-presenting DCs which can be used to design vaccination strategies to induce antitumor cytotoxic T-cell immunity.     

树突状细胞多价核酸疫苗抗血吸虫感染作用及机制研究

目的 探讨基因转染对树突状细胞(DC)功能的影响及DC多价核酸疫苗抗血吸虫感染作用及机制。方法 利用脂质体介导的基因转染技术将Sj26、Sj23和Sj14基因分别转染DC,流式细胞术(FCM)检测DC摄取抗原能力,混合淋巴细胞反应(MLR)检测DC对同种异体T淋巴细胞的刺激作用。Sj26、Sj23和Sj14基因转染DC分别和联合免疫BALB/c小鼠3次,末次免疫2周后,每只小鼠感染日本血吸虫尾蚴40条,小鼠感染血吸虫6周后,计数成虫和虫卵。ELISA法检测血清特异性IgG、血清干扰素-γ(IFN-γ)和白细胞介素-4(IL-4)及脾淋巴细胞培养上清IFN-γ、IL-4水平,噻唑蓝(MTT)法检测脾淋巴细胞的增殖。结果 与真核表达载体pcDNA3转染DC和未处理DC相比较,基因转染DC摄取抗原的荧光强度明显降低(P<0.01),对同种异体T淋巴细胞的刺激指数明显升高(P<0.01)。各免疫组小鼠诱导的减虫率和减卵率均高于对照组(P<0.01),而基因转染DC联合免疫组小鼠抗血吸虫感染作用高于单一基因转染DC免疫组(P<0.001)。基因转染DC免疫组小鼠末次免疫2周后血清特异性IgG水平较免疫前显著升高(P<0.05),血清IFN-γ水平明显升高(P<0.01),而血清IL-4的水平无明显变化。与对照组比较,基因转染DC免疫组小鼠脾淋巴细胞经ConA和SEA刺激后培养上清IFN-γ水平显著增高,IL-4水平显著降低,刺激指数(SI)显著增高(P<0.001)。结论 基因转染能促进DC的成熟,增强DC的活性,DC多价核酸疫苗可增强抗血吸虫感染作用,其作用机制以Th1型免疫应答为主。     

Inflammation and immune response in atherosclerosis

Atherosclerosis is an inflammatory disease with a significant autoimmune component. Studies using transgenic murine models have clarified that recruitment of mononuclear leukocytes through vascular leukocyte-adhesion molecules and chemokines, differentiation of monocytes to macrophages, and endocytosis through scavenger receptors all are of decisive importance for atherosclerosis in hypercholesterolemic mice. T and B cells modulate disease progression and lesion development is reduced in mice lacking adaptive immunity. In particular, local immune responses eliciting Th1 effector mechanisms appear to be proatherogenic, whereas protective immune responses can be induced by immunization with oxidized low-density lipoprotein. Thus, innate immunity is necessary for atherosclerosis, whereas adaptive immunity is an important modulator of disease development.     

Prolonged Th1-like response generated by a Plasmodium yoeli-specific T cell clone allows complete clearance of infection in reconstituted mice

In the present study, we report the ability of in vitro cultured CD4⁺ T cells, generated following immunization with dead blood stage P. yoelii parasites, to mediate protection against homologous challenge infection in reconstituted nude mice. P. yoelii -specific T cell line cells produced IFN-γ after in vitro stimulation with specific antigen, and were protective when adoptively transferred into athymic nude mice. Following transfer of P. yoelii -specific T cell lines into nude and SCID mice, elevated levels of nitric oxide (NO) were detected during the first week of infection at a time when parasitaemias were suppressed. However, in vivo blocking of NO production through administration of L-NMMA, an inhibitor of NO synthase, increased mortality, but did not alter the course of primary parasitaemia in P. yoelii- specific T cell line-reconstituted nude mice. In addition, a P. yoelii -specific CD4⁺ T cell clone, which produced IFN-γ in vitro , afforded sterile protection via mechanisms other than NO. By ELISA, antibodies were undetectable on all but one day (day 79) post T cell clone transfer and parasite challenge, where very low levels of antibodies were detected, with some evidence of recognition of malaria proteins by Western blot. Collectively, our data suggest that T cell effector functions, independent of NO production and in the absence of high levels of parasite-specific antibodies, can contribute to sterile immunity to P. yoelii .     

Testosterone-unresponsiveness of existing immunity against Plasmodium chabaudi malaria

Testosterone (Te) is known to suppress immunity and to increase host susceptibility to many parasites. This study investigates the action of Te on immunity acquired against blood-stages of the malaria parasite Plasmodium chabaudi in female mice of the inbred strain C57BL/10. Our data show: (i) About 90% of mice infected with 10(6) P. chabaudi-infected erythrocytes are able to develop protective immune mechanisms which become evident in self-healing the infection. The capability of self-healing is lost when mice are pretreated with Te for 3 weeks. (ii) Mice which have self-healed infections acquire immunity to homologous rechallenge. Concomitantly, mice become Te-unresponsive in that their acquired immunity is not suppressible by Te-treatment. (iii) Flow cytometry reveals that Te-pretreatment entails an increase of CD8+ cells and a decrease of Ig+ cells by about 4% in spleens of non-immune mice. In immune mice, however, there is a Te-unresponsiveness of the percental distribution of splenic cell populations. (iv) Adoptive transfer experiments indicate that immunity is conferred by spleen cells, presumably non-T-cells. These cells are Te-unresponsive, since they exert their effect in Te-pretreated mice in the presence of Te. (v) Te-unresponsive immunity can be also transferred by serum, especially the IgG-fraction, obtained from immune mice. Our data demonstrate that Te prevents the development of protective immunity against P. chabaudi infections. However, when once established, protective immunity becomes unresponsive to Te. Our data suggest that the effector mechanisms of protective immunity involve Te-unresponsive B cells secreting protective IgG-antibodies.     

Antigen-specific production of colony-stimulating factors by Listeria monocytogenes-immune, L3T4-positive cells

We investigated production of colony-stimulating factors by Listeria monocytogenes-immune spleen cells. Levels of total colony-stimulating factors in supernatants from antigen-stimulated immune cells were increased two- to fourfold over those in supernatants from nonimmune cells. Immune supernatants primarily induced formation of granulocyte colonies, whereas nonimmune supernatants induced formation of macrophage colonies. Immune supernatants had two- to 10-fold higher levels of macrophage colony-stimulating factor, as determined by radioimmunoassay, and higher levels of interleukin-3 and possibly granulocyte-macrophage colony-stimulating factor, as determined by factor-dependent cell line growth, than did nonimmune supernatants. Using enrichment and depletion techniques we showed that L3T4-positive T lymphocytes were responsible for most of the colony-stimulating factor production in the immune reaction.