氯化锂治疗实验性自身免疫性脑脊髓炎的实验研究
发布时间：2018-12-24 21:20
【摘要】：背景:多发性硬化(multiple sclerosis,MS)是一种中枢神经系统(central nerve system,CNS)的慢性自身免疫性疾病,以炎症复发和病情缓解交替发生为特征,好发于15~50岁的青壮年之中。目前临床中对MS患者大都采用综合性治疗的方式,但是即便治疗及时,这些疗法也只能起到缓解而非治愈MS的效果。因此,增强MS的治疗康复,已经成为临床及基础研究中的热点与重点。锂剂是临床治疗预防双相情感障碍躁狂的一线药物,具备对中枢神经系统的保护功能,同时能够对多种免疫细胞产生调节作用。目前的研究表明,使用锂剂能够在实验动物中对MS替代模型实验性自身免疫性脑脊髓炎(experimental autoimmune encephalomyelitis,EAE)产生显著的治疗作用,并且该治疗作用和IFN-γ干扰素γ(interferon-γ,IFN-γ)存在一定的联系。IFN-γ由Th1型细胞和单核细胞合成分泌,可以调节细胞因子之间的“级联作用模式”,增强免疫应答,在MS病人血清中表达升高。除了关注锂剂对于免疫系统的作用以外,我们还推测,锂剂治疗MS的过程,是一个抑制GSK3β、进而激活神经细胞内促细胞生存Wnt/β-catenin通路的过程。综上,探究锂剂对MS具体的治疗效果及相关机制,有助于提升MS病人的预后,同时发掘更多的治疗靶点。在本项研究中,我们重点关注锂剂在IFN-γ功能缺失时对CNS炎症的影响,同时还探究了β-catenin在锂剂治疗MS中扮演的具体角色。目的:探究锂剂在治疗MS过程中与IFN-γ和β-catenin的相互作用。方法:1.IFN-γR缺失小鼠PCR鉴定;2.使用MOG35-55和百日咳毒素联合注射的方法,诱导IFN-γR缺失小鼠发生EAE;3.对不同组中EAE小鼠脑部炎症非典型症状比例进行统计;4.对不同组中EAE小鼠脊髓炎症典型症状进行打分统计;5.收集小鼠脑和脊髓组织,进行HE染色分析;6.氯化锂和IFN-γ处理小鼠脑内皮细胞b END.3;7.免疫印迹检测b END.3细胞中β-catenin表达量的变化;结果:1.经PCR验证,实验用小鼠为IFN-γR-/-基因型,为IFN-γR缺失;2.MOG35-55和百日咳毒素联合注射的方法,成功在IFN-γR缺失小鼠中诱导了脑和脊髓炎症的发生;3.锂剂治疗降低了脑部炎症非典型症状的比例以及脊髓炎症典型症状的平均得分;4.锂剂治疗缓解了脑和脊髓组织中的炎症反应;5.锂剂对于CNS中炎症的治疗作用,可以不依赖于IFN-γ的功能;6.锂剂对于IFN-γR缺失小鼠脑和脊髓炎症的缓解作用,不具有长效性;7.锂剂和IFN-γ处理,显著提高了脑内皮细胞中β-catenin的表达水平;结论:锂剂可以激活脑内皮细胞中β-catenin信号,抑制脑和脊髓炎症的发生;锂剂对于CNS炎症的治疗,可以不依赖于IFN-γ的功能,但相应治疗作用不具有长效性。
[Abstract]:Background: multiple sclerosis (multiple sclerosis,MS) is a chronic autoimmune disease of central nervous system (central nerve system,CNS) characterized by recurrent inflammation and remission. At present, MS patients are mostly treated by comprehensive therapy, but even if the treatment is timely, these therapies can only alleviate rather than cure MS. Therefore, enhancing the treatment and rehabilitation of MS has become the focus of clinical and basic research. Lithium is a first-line drug for the prevention of bipolar affective disorder mania. It can protect the central nervous system and regulate many immune cells. Current studies have shown that the use of lithium can produce significant therapeutic effects on experimental autoimmune encephalomyelitis (experimental autoimmune encephalomyelitis,EAE) induced by MS in experimental animals, and the therapeutic effect is similar to that of IFN- interferon 纬 (interferon- 纬). IFN- 纬 was synthesized and secreted by Th1 type cells and monocytes, which could regulate the "cascade action mode" between cytokines, enhance the immune response, and increase the expression of IFN- 纬 in serum of MS patients. In addition to focusing on the role of lithium in the immune system, we also speculate that the treatment of MS with lithium is a process that inhibits GSK3 尾 and thus activates the Wnt/ 尾-catenin pathway that promotes cell survival in nerve cells. In conclusion, to explore the specific therapeutic effects and related mechanisms of lithium on MS is helpful to improve the prognosis of patients with MS, and to explore more therapeutic targets at the same time. In this study, we focused on the effects of lithium on CNS inflammation in the absence of IFN- 纬 function, and explored the role of 尾-catenin in the treatment of MS with lithium. Aim: to investigate the interaction of lithium with IFN- 纬 and 尾-catenin in the treatment of MS. Methods: 1. PCR identification of IFN- 纬 R deficient mice. Induction of EAE;3. in IFN- 纬 R deficient mice by combined injection of MOG35-55 and pertussis toxin The proportion of atypical symptoms of cerebral inflammation in EAE mice was analyzed in different groups. 4. The typical symptoms of spinal cord inflammation in EAE mice in different groups were evaluated. The brain and spinal cord of mice were collected and analyzed by HE staining. Treatment of b END.3;7. in mouse brain endothelial cells with lithium chloride and IFN- 纬 The expression of 尾-catenin in b END.3 cells was detected by Western blotting. Results: 1. PCR showed that the mice were IFN- 纬 R-r- genotype and IFN- 纬 R deletion, and the combination of 2.MOG35-55 and pertussis toxin induced the inflammation of brain and spinal cord in IFN- 纬 R deficient mice. 3. Lithium treatment decreased the proportion of atypical symptoms of cerebral inflammation and the average score of typical symptoms of spinal cord inflammation; 4. 5%. Lithium treatment alleviated inflammation in brain and spinal cord. The therapeutic effect of lithium on inflammation in CNS could not be dependent on the function of IFN- 纬. The effect of lithium on brain and spinal cord inflammation in mice with IFN- 纬 R deficiency was not long-lasting. Lithium and IFN- 纬 could significantly increase the expression of 尾-catenin in brain endothelial cells. Conclusion: lithium can activate 尾-catenin signal in brain endothelial cells and inhibit the occurrence of inflammation in brain and spinal cord. Lithium may not depend on the function of IFN- 纬 in the treatment of CNS inflammation, but the corresponding therapeutic effect is not long-lasting.
【学位授予单位】：新乡医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R744.51
本文编号：2390859
[Abstract]:Background: multiple sclerosis (multiple sclerosis,MS) is a chronic autoimmune disease of central nervous system (central nerve system,CNS) characterized by recurrent inflammation and remission. At present, MS patients are mostly treated by comprehensive therapy, but even if the treatment is timely, these therapies can only alleviate rather than cure MS. Therefore, enhancing the treatment and rehabilitation of MS has become the focus of clinical and basic research. Lithium is a first-line drug for the prevention of bipolar affective disorder mania. It can protect the central nervous system and regulate many immune cells. Current studies have shown that the use of lithium can produce significant therapeutic effects on experimental autoimmune encephalomyelitis (experimental autoimmune encephalomyelitis,EAE) induced by MS in experimental animals, and the therapeutic effect is similar to that of IFN- interferon 纬 (interferon- 纬). IFN- 纬 was synthesized and secreted by Th1 type cells and monocytes, which could regulate the "cascade action mode" between cytokines, enhance the immune response, and increase the expression of IFN- 纬 in serum of MS patients. In addition to focusing on the role of lithium in the immune system, we also speculate that the treatment of MS with lithium is a process that inhibits GSK3 尾 and thus activates the Wnt/ 尾-catenin pathway that promotes cell survival in nerve cells. In conclusion, to explore the specific therapeutic effects and related mechanisms of lithium on MS is helpful to improve the prognosis of patients with MS, and to explore more therapeutic targets at the same time. In this study, we focused on the effects of lithium on CNS inflammation in the absence of IFN- 纬 function, and explored the role of 尾-catenin in the treatment of MS with lithium. Aim: to investigate the interaction of lithium with IFN- 纬 and 尾-catenin in the treatment of MS. Methods: 1. PCR identification of IFN- 纬 R deficient mice. Induction of EAE;3. in IFN- 纬 R deficient mice by combined injection of MOG35-55 and pertussis toxin The proportion of atypical symptoms of cerebral inflammation in EAE mice was analyzed in different groups. 4. The typical symptoms of spinal cord inflammation in EAE mice in different groups were evaluated. The brain and spinal cord of mice were collected and analyzed by HE staining. Treatment of b END.3;7. in mouse brain endothelial cells with lithium chloride and IFN- 纬 The expression of 尾-catenin in b END.3 cells was detected by Western blotting. Results: 1. PCR showed that the mice were IFN- 纬 R-r- genotype and IFN- 纬 R deletion, and the combination of 2.MOG35-55 and pertussis toxin induced the inflammation of brain and spinal cord in IFN- 纬 R deficient mice. 3. Lithium treatment decreased the proportion of atypical symptoms of cerebral inflammation and the average score of typical symptoms of spinal cord inflammation; 4. 5%. Lithium treatment alleviated inflammation in brain and spinal cord. The therapeutic effect of lithium on inflammation in CNS could not be dependent on the function of IFN- 纬. The effect of lithium on brain and spinal cord inflammation in mice with IFN- 纬 R deficiency was not long-lasting. Lithium and IFN- 纬 could significantly increase the expression of 尾-catenin in brain endothelial cells. Conclusion: lithium can activate 尾-catenin signal in brain endothelial cells and inhibit the occurrence of inflammation in brain and spinal cord. Lithium may not depend on the function of IFN- 纬 in the treatment of CNS inflammation, but the corresponding therapeutic effect is not long-lasting.
【学位授予单位】：新乡医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R744.51
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