Basic Research for Regenerative Therapy of Multiple Sclerosis

目的和背景：多发性硬化症（MS）是一种影响中枢神经系统（CNS）白质的免疫介导疾病，受遗传和环境因素之间复杂的相互作用调节，以多灶性炎性脱髓鞘和轴突变性为特征，导致永久性神经功能缺损。MS表现出明显的临床病理异质性，根据病程分为复发缓解型MS（RRMS）、继发性进行性MS（SPMS）和原发性进行性MS(PPMS)，根据病变分布分为常规MS（CMS）和光脊髓型MS(OSMS)，从治疗反应分为干扰素- β (IFNB)应答和无应答，从病理角度分为淋巴细胞介导、抗体介导和少突胶质细胞凋亡介导的脱髓鞘。由于其可变性，由于缺乏可靠的诊断标志物，MS的准确临床诊断往往很困难。最近的研究表明，星形胶质细胞、少突胶质细胞上表达的髓鞘相关神经突生长抑制剂Nogo和激活的小胶质细胞释放的促炎性细胞因子在有效的髓鞘再生和轴突再生中起抑制作用。IFNB已被用作治疗MS急性复发最有效的药物之一，尽管其潜在机制尚不清楚，IFNB对诱导髓鞘再生和轴突再生无效。到目前为止，还没有针对多发性硬化症的再生治疗方法。本研究的目的是：(1)通过分析DNA微阵列上多发性硬化症的基因表达谱来鉴定一种新的多发性硬化症分子标志物，(2)建立一种在治疗前区分IFNB应答者和无应答者的方法，(3)阐明反应性胶质瘤的分子机制，(4)研究Nogo在多发性硬化症轴突再生失败中的病理作用。我们发现(1)通过DNA微阵列分析，一个参与细胞凋亡调控的基因家族在MS患者的外周血淋巴细胞中异常表达(2)通过分层聚类分析，一组基因在未治疗的MS患者和对照组之间的差异表达将MS患者分为四个不同的亚组，其中IFNβ应答者聚集在其中两个亚组中。(3)通过免疫组织化学和蛋白质组学分析，14-3-3 epsilon亚型在培养的人星形胶质细胞和MS病变中的反应性星形胶质细胞中与vimentin和GFAP结合。(4)免疫组化发现，在MS脱髓鞘病变中，Nogo- a在存活的少突胶质细胞中表达上调，而Nogo受体在反应性星形胶质细胞和小胶质细胞中的表达增强。这些结果提示了开发MS再生治疗的可能途径，例如靶向抑制反应性星形胶质细胞的14-3-3功能，可能有助于预防胶质瘤的发生。以及人源抗体阻断Nogo-A/NgR相互作用的应用，可能有助于支持ms Less的轴突再生

Objectives and Backgrounds : Multiple sclerosis(MS) is an immune-mediated disease affecting the central nervous system(CNS) white matter, regulated by a complex interplay between genetic and environmental factors, characterized by multifocal inflammatory demyelination and axonal degeneration that cause permanent neurological deficits. MS shows remarkable clinicopathological heterogeneity, categorized into relapsing-remitting MS(RRMS), secondary progressive MS(SPMS), and primary progressive MS(PPMS) based on the disease course, conventional MS(CMS) and opticospinal MS(OSMS) by lesion distribution, interferon-beta(IFNB) responder and nonresponder from the therapeutic response, and lymphocyte-mediated, antibody-mediated, and oligodendrocyte apoptosis-mediated demyelination from the pathological aspect. Because of its variability, the accurate clinical diagnosis of MS is often difficult due to lack of a reliable diagnostic marker. Recent studies showed that glial scar produced by reactive … More astrocytes, myelin-associated neurite outgrowth inhibitor Nogo expressed on oligodendrocytes, and proihflammatory cytokines released from activated microglia, play an inhibitory role in efficient remyelination and axonal regeneration. IFNB has been utilized as one of the most effective medications against acute relapse in MS, although the underlying mechanism remains unknown and IFNB is ineffective for induction of remyelination and axonal regeneration. Until present, no regenerative therapy is available for MS. The aim of this work is (1)to identify a novel molecular marker of MS by analyzing gene expression profile specific for MS on a DNA microarray, (2)to establish a method to distinguish IFNB responder and nonresponder before treatment, (3)to clarify the molecular mechanism of reactive gliosis, and (4)to investigate a pathological role of Nogo in the failure of axonal regeneration, in MS. Methods, Results, and Conclusions : We found that (1)by DNA microarray analysis, a family of genes involved in regulation of apoptosis are aberrantly expressed in peripheral blood lymphocytes in MS. (2)by hierarchial clustering analysis, a set of genes differentially expressed between untreated MS patients and control subjects separated four distinct subgroups of MS patients where IFNβ responders were clustered in two of these subgroups. (3)by immunohistochemical and proteome analysis, 14-3-3 epsilon isoform binds to vimentin and GFAP in cultured human astrocytes and reactive astrocytes in MS lesions. (4)by immunohistochemistry, Nogo-A is upregulated on surviving oligodendrocytes, while Nogo receptor expression is enhanced in reactive astrocytes and microglia in demyelinating lesions of MS. These results suggest a possible approach to develop regenerative therapy for MS, such as a targeted inhibition of 14-3-3 function in reactive astrocytes that might be useful for prevention of gliosis, and an application of humanized antibodies blocking Nogo-A/NgR interaction that might be helpful for supporting axonal regeneration in MS. Less