Suppression of experimentally induced autoimmune encephalomyelitis by cytolytic T-T cell interactions

Down-regulatory phenomena have been described in several experimental models of tissue-specific, T-cell-mediated autoimmunity. For example, resistance to active induction of experimental autoimmune encephalomyelitis (EAE) can be induced by pretreating animals with non-pathogenic inocula of autoantigen or effector cells. Moreover, animals that have recovered from one EAE episode are resistant to subsequent induction of EAE. In some models, resistance to EAE has been transferred with immune cells to naive recipients. These experiments, which were based on transfers of unseparated immune cell populations, are difficult to interpret. Immune suppression circuits are known to be complex and involve various distinct cellular subsets. To further complicate the issue, resistance to EAE can be transferred not only by suppressor cells, but also by encephalitogenic effector cells injected in 'subclinical' doses. We describe now the isolation of homogeneous T lymphocyte lines from the spleens of Lewis rats that had recovered from T-cell-mediated EAE (tEAE) caused by the MBP-specific T cell line S1. These spleen-derived T line cells express the CD8 phenotype and specifically respond to determinants on the inducing S1 line, but not to the autoantigen MBP. Furthermore, the anti-S1 cells selectively lyse the encephalitogenic S1 T line in vitro and efficiently neutralize their encephalitogenic capacity in vivo.     

Injection of adult neurospheres induces recovery in a chronic model of multiple sclerosis

Widespread demyelination and axonal loss are the pathological hallmarks of multiple sclerosis. The multifocal nature of this chronic inflammatory disease of the central nervous system complicates cellular therapy and puts emphasis on both the donor cell origin and the route of cell transplantation. We established syngenic adult neural stem cell cultures and injected them into an animal model of multiple sclerosis--experimental autoimmune encephalomyelitis (EAE) in the mouse--either intravenously or intracerebroventricularly. In both cases, significant numbers of donor cells entered into demyelinating areas of the central nervous system and differentiated into mature brain cells. Within these areas, oligodendrocyte progenitors markedly increased, with many of them being of donor origin and actively remyelinating axons. Furthermore, a significant reduction of astrogliosis and a marked decrease in the extent of demyelination and axonal loss were observed in transplanted animals. The functional impairment caused by EAE was almost abolished in transplanted mice, both clinically and neurophysiologically. Thus, adult neural precursor cells promote multifocal remyelination and functional recovery after intravenous or intrathecal injection in a chronic model of multiple sclerosis.     

T-cell epitope of the autoantigen myelin basic protein that induces encephalomyelitis

Chronic relapsing paralysis and demyelination within the central nervous system (CNS), features associated with the human disease multiple sclerosis (MS), develop in mice after injection of murine T-cell clones specific for the autoantigen myelin basic protein (MBP). We examined the fine specificity of three independently derived encephalitogenic T-cell clones using synthetic polypeptides derived from portions of the N-terminal sequence of MBP. These clones appear functionally identical; they all respond to an epitope in the N-terminal nine amino acid residues in association with the same class II (I-A) molecules of the major histocompatibility complex (MHC). Both the N-terminal acetyl moiety and the first residue (Ala) are necessary for recognition. Only N-terminal MBP peptides recognized by these clones were found to cause encephalomyelitis (EAE) in vivo. These results show that the N-terminal MBP-specific T lymphocytes that mediate autoimmune encephalomyelitis are a small population with a limited repertoire; they all recognise the same combination of MHC and target.     

Immunospecific inhibition of nerve conduction by T lymphocytes reactive to basic protein of myelin

Experimental autoimmune encephalomyelitis (EAE) induced by immunization to the basic protein of central nervous system myelin (BP) is a paralytic disease in which T lymphocytes attack the individual's own central nervous system. As the target is in white matter, EAE has been considered an experimental model of some aspects of human disease such as multiple sclerosis. To investigate whether autoimmune T lymphocytes could produce paralysis, we studied the effects on the electrophysiology of isolated nerves produced by T-lymphocyte lines reactive specifically to BP or other antigens. We now report that propagation of action potentials evoked by electrical stimulation was blocked by incubating optic nerves with specific anti-BP T cells. This blockade could be reversed for up to two hours by removing the anti-BP line cells from the optic nerve. The anti-BP line cells had no effect on conduction along allogeneic optic nerves or syngeneic peripheral nerves. This indicates that disruption of the function of myelin in neuroimmunological disease may result from an immunologically specific interaction between autoimmune T lymphocytes and myelin antigens.     

Increased levels of myelin basic protein transcripts in virus-induced demyelination

In multiple sclerosis, a demyelinating disease of young adults, there is a paucity of myelin repair in the central nervous system (CNS) which is necessary for the restoration of fast saltatory conduction in axons. Consequently, this relapsing disease often causes marked disability. In similar diseases of small rodents, however, remyelination can be quite extensive, as in the demyelinating disease caused by the A59 strain of mouse hepatitis virus (MHV-A59), a coronavirus of mice. To investigate when and where oligodendrocytes are first triggered to repair CNS myelin in such disease, we have used a complementary DNA probe specific for one major myelin protein gene, myelin basic protein (MBP), which hybridizes with the four forms of MBP messenger RNA in rodents. Using Northern blot and in situ hybridization techniques, we previously found that MBP mRNA is first detected at about 5 days after birth, peaks at 18 days and progressively decreases to 25% of the peak levels in the adult. We now report that in spinal cord sections of adult animals with active demyelination and inflammatory cells, in situ hybridization reveals a dramatic increase in probe binding to MBP-specific mRNA at 2-3 weeks after virus inoculation and before remyelination can be detected by morphological methods. This increase of MBP-specific mRNA is found at the edge of the demyelinating area and extends into surrounding areas of normal-appearing white matter. Thus, in situ hybridization with myelin-specific probes appears to be a useful method for detecting the timing, intensity and location of myelin protein gene reactivation preceding remyelination. This method could be used to elucidate whether such a reactivation occurs in multiple sclerosis brain tissue. Our results suggest that in mice, glial cells react to a demyelinating process with widespread MBP mRNA synthesis which may be triggered by a diffusible factor released in the demyelinated areas.     

Spreading of T-cell autoimmunity to cryptic determinants of an autoantigen.

Immunization with myelin basic protein (MBP) induces experimental allergic encephalomyelitis (EAE), a prototype of CD4+ T-cell mediated autoimmune disease. In rodents, MBP-reactive T-cell clones are specific for a single, dominant determinant on MBP and use a highly restricted number of T-cell receptor genes. Accordingly, EAE has been prevented by various receptor-specific treatments, suggesting similar strategies may be useful for therapy of human autoimmune disease. Here we report that in (SJL x B10.PL)F1 mice, immune dominance of a single determinant, MBP:Ac1-11, is confined to the inductive phase of EAE. In mice with chronic EAE, several additional determinants of MBP in peptides 35-47, 81-100 and 121-140 recall proliferative responses. Most importantly, reactivity to the latter determinants was also detected after induction of EAE with MBP peptide Ac1-11 alone; this demonstrates priming by endogenous MBP determinants. Thus, determinants of MBP that are cryptic after primary immunization can become immunogenic in the course of EAE. Diversification of the autoreactive T-cell repertoire due to 'determinant spreading' has major implications for the pathogenesis of, and the therapeutic approach to, T-cell driven autoimmune disease.     

A synthetic glycolipid prevents autoimmune encephalomyelitis by inducing TH2 bias of natural killer T cells

Experimental autoimmune encephalomyelitis (EAE) is a prototype autoimmune disease mediated by type 1 helper T (TH1) cells and under the control of regulatory cells. Here we report that a synthetic glycolipid ligand for CD1d-restricted natural killer T (NKT) cells expressing the semi-invariant T-cell receptor (Valpha14+) is preventive against EAE. The ligand is an analogue of alpha-galactosylceramide (alpha-GC), a prototype NKT cell ligand, with a truncated sphingosine chain. alpha-GC causes NKT cells to produce both interferon (IFN)-gamma and interleukin (IL)-4 (refs 4, 5). However, this new ligand can induce a predominant production of IL-4 by the NKT cells. A single injection of this glycolipid, but not of alpha-GC, consistently induced TH2 bias of autoimmune T cells by causing NKT cells to produce IL-4, leading to suppression of EAE. The lack of polymorphism of CD1d and cross-reactive response of mouse and human NKT cells to the same ligand indicates that targeting NKT cells with this ligand may be an attractive means for intervening in human autoimmune diseases such as multiple sclerosis.     

重建人类免疫系统的人源化小鼠模型及其在血液系统疾病研究中的应用

动物模型在人类疾病发病机制的研究及药理学实验研究治疗中发挥重要作用。小鼠是目前主要的实验动物,但其与人类背景存在差异,不能完全模拟人类免疫系统及疾病状态。研究人员通过将人类细胞或组织植入免疫缺陷小鼠体内,可构建免疫系统人源化小鼠模型及人源化小鼠疾病模型。人源化小鼠可通过优化人源造血干细胞来源等条件,可得到免疫系统人源化程度较高的小鼠模型。目前,GVHD人源化小鼠模型及白血病人源化小鼠模型已经应用在研究疾病发病机制及开发新型临床治疗药物等方面。     

A suppressor T-lymphocyte cell line for autoimmune encephalomyelitis

Experimental allergic encephalomyelitis (EAE) is a model for the in vitro and in vivo study of T-cell activation. It is an autoimmune disease mediated by T lymphocytes of the helper T-cell (Th) subset. After sensitization to guinea-pig myelin basic protein in complete Freund's adjuvant, Lewis rats develop an autoimmune response to central nervous system (CNS) myelin basic protein, manifested clinically as paralysis and histologically by a perivascular mononuclear cell infiltrate of the CNS parenchyma. Suppressor cell regulation of EAE has long been suspected because Lewis rats, which spontaneously recover from active disease, are resistant to reinduction of active EAE, even though effector T-cell lines can be rescued from these recovered rats. Using cyclosporin A, an immunosuppressive agent believed to inhibit Th cell function, suppressor T-cell (Ts) lines have now been generated from recovered Lewis rats. These Ts cells, when admixed with guinea pig myelin basic protein-specific Th cells, will prevent the adoptive transfer of EAE. The Ts cells appear to be CD4+, which explains previous observations that CD8+ lymphocytes are not important in the recovery of EAE in the rat. This is the first direct demonstration of Ts-cell regulation of EAE.     

氨基葡萄糖治疗实验性自身免疫性脑脊髓炎研究进展

氨基葡萄糖是葡萄糖的一种天然衍生物,是糖蛋白和蛋白聚糖的主要成分,可通过多种生理途径在细胞内合成.近年研究显示,氨基葡萄糖可作为免疫抑制剂和免疫调节剂在治疗多发性硬化（MS）及其动物模型——实验性自身免疫性脑脊髓炎（EAE）方面有一定的疗效.氨基葡萄糖可引起Th表型由Th1向Th2细胞转化,显著表现为TNF-α,IFN-γ,IL-17,IL-1分泌减少,IL-5和IL-10分泌增多.氨基葡萄糖通过不同给药方式,如口服、腹腔注射、静脉注射等对EAE都有一定治疗作用,具有低或无毒性、服用安全等优点.     

Sequence of simian immunodeficiency virus from macaque and its relationship to other human and simian retroviruses

Because of the growing incidence of AIDS (acquired immune deficiency syndrome), the need for studies on animal models is urgent. Infection of chimpanzees with the retroviral agent of human AIDS, the human immunodeficiency virus (HIV), will have only limited usefulness because chimpanzees are in short supply and do not develop the disease. Among non-human primates, both type D retroviruses and lentiviruses can be responsible for immune deficiencies. The D-type retroviruses, although important pathogens in macaque monkey colonies, are not satisfactory as a model because they differ in genetic structure and pathophysiological properties from the human AIDS viruses. The simian lentivirus, previously referred to as simian T-cell lymphotropic virus type III (STLV-III), now termed simian immunodeficiency virus (SIV) is related to HIV by the antigenicity of its proteins and in its main biological properties, such as cytopathic effect and tropism for CD4-bearing cells. Most importantly, SIV induces a disease with remarkable similarity to human AIDS in the common rhesus macaques, which therefore constitute the best animal model currently available. Natural or experimental infection of other monkeys such as African green monkeys or sooty mangabeys has not yet been associated with disease. Molecular approaches of the SIV system will be needed for biological studies and development of vaccines that could be tested in animals. We have cloned and sequenced the complete genome of SIV isolated from a naturally infected macaque that died of AIDS. This SIVMAC appears genetically close to the agent of AIDS in West Africa, HIV-2, but the divergence of the sequences of SIV and HIV-2 is greater than that previously observed between HIV-1 isolates.     

Adoptive transfer of EAE-like lesions from rats with coronavirus-induced demyelinating encephalomyelitis

Viruses have been found to induce inflammatory demyelinating lesions in central nervous system (CNS) tissue of both animal and man, either by natural infections or after vaccination. At least two different pathogenic mechanisms have been proposed for these changes, a cytopathic viral infection of oligodendroglia cells with subsequent cell death, and a host immune reaction against virus and brain antigens. We now report the occurrence of cell-mediated immune reactions against basic myelin proteins in the course of coronavirus infections in Lewis rats. Infection of rats with the murine coronavirus JHM leads to demyelinating encephalomyelitis developing several weeks to months postinfection. Lymphocytes from these diseased Lewis rats can be restimulated with basic myelin protein (BMP) and adoptive transfer of these cells leads to lesions resembling those of experimental allergic encephalomyelitis (EAE) in recipients, which can be accompanied by a mild clinical disease. This model demonstrates that a virus infection in CNS tissue is capable of initiating an autoimmune response which may be of pathogenic importance.     

Th1-specific cell surface protein Tim-3 regulates macrophage activation and severity of an autoimmune disease

Activation of naive CD4(+) T-helper cells results in the development of at least two distinct effector populations, Th1 and Th2 cells. Th1 cells produce cytokines (interferon (IFN)-gamma, interleukin (IL)-2, tumour-necrosis factor (TNF)-alpha and lymphotoxin) that are commonly associated with cell-mediated immune responses against intracellular pathogens, delayed-type hypersensitivity reactions, and induction of organ-specific autoimmune diseases. Th2 cells produce cytokines (IL-4, IL-10 and IL-13) that are crucial for control of extracellular helminthic infections and promote atopic and allergic diseases. Although much is known about the functions of these two subsets of T-helper cells, there are few known surface molecules that distinguish between them. We report here the identification and characterization of a transmembrane protein, Tim-3, which contains an immunoglobulin and a mucin-like domain and is expressed on differentiated Th1 cells. In vivo administration of antibody to Tim-3 enhances the clinical and pathological severity of experimental autoimmune encephalomyelitis (EAE), a Th1-dependent autoimmune disease, and increases the number and activation level of macrophages. Tim-3 may have an important role in the induction of autoimmune diseases by regulating macrophage activation and/or function.     

Transfer of diabetes in mice prevented by blockade of adhesion-promoting receptor on macrophages

Insulin-dependent diabetes mellitus (IDDM) is a disease with an autoimmune aetiology. The non-obese diabetic mouse is a good spontaneous animal model of the human disease, with IDDM developing in 50-80% of female mice by the age of 6 months. The disease can be transferred by splenic T cells from diabetic donors and is prevented by T-cell depletion. The mechanism(s) by which the beta cell is specifically destroyed is not known, but T cells and macrophages have both been implicated, based on the presence of macrophages in the infiltrated islet and the ability of chronic silica treatment to prevent disease. The monoclonal antibody 5C6 is specific for the myelomonocytic adhesion-promoting type-3 complement receptor (CR3 or CD11b/CD18) and does not bind to T cells. Here we show that blockade of macrophage CR3 in vivo prevents intra-islet infiltration by both macrophages and T cells and inhibits development of IDDM. We conclude that both T cells and macrophages have an essential role in the onset of IDDM.     

A microbial symbiosis factor prevents intestinal inflammatory disease

Humans are colonized by multitudes of commensal organisms representing members of five of the six kingdoms of life; however, our gastrointestinal tract provides residence to both beneficial and potentially pathogenic microorganisms. Imbalances in the composition of the bacterial microbiota, known as dysbiosis, are postulated to be a major factor in human disorders such as inflammatory bowel disease. We report here that the prominent human symbiont Bacteroides fragilis protects animals from experimental colitis induced by Helicobacter hepaticus, a commensal bacterium with pathogenic potential. This beneficial activity requires a single microbial molecule (polysaccharide A, PSA). In animals harbouring B. fragilis not expressing PSA, H. hepaticus colonization leads to disease and pro-inflammatory cytokine production in colonic tissues. Purified PSA administered to animals is required to suppress pro-inflammatory interleukin-17 production by intestinal immune cells and also inhibits in vitro reactions in cell cultures. Furthermore, PSA protects from inflammatory disease through a functional requirement for interleukin-10-producing CD4+ T cells. These results show that molecules of the bacterial microbiota can mediate the critical balance between health and disease. Harnessing the immunomodulatory capacity of symbiosis factors such as PSA might potentially provide therapeutics for human inflammatory disorders on the basis of entirely novel biological principles.     

C57BL/6小鼠实验性变态反应性脑脊髓炎模型的建立

目的 为更好地研究多发性硬化症(MS)的发病机理、开发具有良好效果的MS治疗药物,建立起能高度模拟人类多发性硬化症发生发展的小鼠模型则显得尤为必要且具有极高的应用价值。本文使用已广泛应用的标准化C57BL/6小鼠作为实验对象,成功地构建了实验性变态反应性脑脊髓炎模型(EAE)。此模型为研究人类多发性硬化症的发病机理及开发相关临床治疗药物提供了良好的小鼠模型。方法 C57BL/6雄性小鼠20只,分为2组。EAE组10只,皮下注射混匀的弗氏不完全佐剂与MOG53-58及结核分枝杆菌的悬浊液,同时分别在day0和day2腹腔注射百日咳毒素0.2μg/只;正常对照组10只,仅注射生理盐水。采用HE染色方法观察小鼠组织学变化;免疫组织化学方法进一步研究小鼠病理变化。结果 从抗原免疫后第10天开始EAE组小鼠陆续发生EAE临床症状且发病率达到100%,而正常对照组小鼠未发生任何EAE临床症状。组织病理学结果分析显示,EAE组小鼠脊髓及脑组织中出现大量炎性细胞浸润且脱髓鞘严重。结论 本实验成功构建的EAE小鼠模型病程稳定且发病率高,可用于MS机制研究及MS防治药物的筛选。     

Mouse skin carcinomas induced in vivo by chemical carcinogens have a transforming Harvey-ras oncogene

Several groups have shown that the malignant phenotype can be transferred to NIH/3T3 fibroblasts by incorporation of DNA isolated from tumour cell lines. These studies have demonstrated that the transforming activity of DNA isolated from human bladder, lung and colon carcinoma cell lines is related to an alteration of the cellular homologues of the ras genes of Harvey or Kirsten murine sarcoma viruses. It is, however, unclear what relevance these observations have to the multi-stage nature of tumorigenesis in vivo, in which several independent events are required in both humans and experimental animals. The activation of a cellular oncogene in a defined experimental system for the progressive induction of solid tumours has not yet been demonstrated. We report here that high molecular weight DNA from transplanted squamous cell carcinomas induced by sequential treatment of mouse skin with initiators and promoters of carcinogenesis causes morphological transformation of NIH/3T3 fibroblasts at high frequency. The transforming properties are due to the transfer of an activated cellular homologue of the Harvey-ras (rasH) oncogene.     

Ia-restricted encephalitogenic T lymphocytes mediating EAE lyse autoantigen-presenting astrocytes

T lymphocytes specific for myelin basic protein (MBP) are responsible for the cellular events leading to autoimmune disease within the central (CNS) and peripheral (PNS) nervous systems. Both in actively induced and T-cell transfer versions of experimental autoimmune encephalomyelitis (EAE) and neuritis (EAN), the autoaggressive T cells are activated outside the nervous system and reach their target tissue via the blood circulation. The target specificity of the autoaggressive T cells is impressive; T-cell lines specific for MBP predominantly home to and affect the white matter of the CNS whereas T cells specific for PNS myelin protein P2 exclusively infiltrate peripheral nerves. Having penetrated the tight blood tissue barriers, the lymphocytes seem to interact with local cells expressing the relevant autoantigen in an immunogenic form. Although the exact mechanism of target finding and destruction is unknown, studies from our laboratory have shown that astrocytes, a main component of the normal CNS glia, can actively present antigen to specific T cells. This observation suggests that astrocytes are involved in natural immune reactivity within the CNS, and that they may be involved in pathological aberrations, such as in the development of autoimmune lesions. Having studied astrocyte/T-cell interactions in more detail, we discovered that encephalitogenic T-cell lines recognizing MBP on astrocytes will subsequently proceed to kill the presenting cells. Here we report that astrocyte killing follows the rules governing 'classical' T-cell-mediated cytolysis; it is antigen-specific, restricted by antigens of the major histocompatibility complex (MHC) and apparently contact-dependent. Our data suggest that the nature of the recognized antigenic epitope determines whether or not antigen recognition is followed by killing; moreover, killing of antigen-presenting astrocytes seems to be correlated with the capacity to transfer encephalomyelitis to normal syngeneic rats.