2018-08-012024-05-18https://scholars.lib.ntu.edu.tw/handle/123456789/713747摘要：脊髓肌肉萎縮症（Spinal Muscular Atrophy, 簡稱SMA）為一遺傳性運動神經元疾病，目前臨床上尚未有治療的方法。SMA的疾病遺傳模式為體染色體隱性遺傳，其發病率約為6,000至10,000分之一，而大多數SMA患者的兩股運動神經元存活基因 (Survival motor neuron gene, 簡稱SMN1)皆發生了基因突變。SMA模式小鼠已由小鼠Smn基因剔除合併人類SMN2基因轉殖的方法所建立，而我們已詳細的分析了SMA小鼠表現型的特色，因此這個SMA模式小鼠對後續SMA致病機制及治療性試驗的相關研究可帶來極大的幫助。骨形生成蛋白-4 (Bone morphogenetic proteins-4, 簡稱BMP-4)為一生長因子，其屬於TGFβ家族的一員，BMP-4在胚胎發育期間，對神經系統的發育有重要的地位；於成體期間，BMP-4亦能促進骨組織的生成。到目前為止，是否BMP-4於神經相關疾病具有調控或治療的角色仍然未知。我們最近以41組SMA病患的微陣列次分析(microarray meta-analysis)發現了包括BMP-4在內的7個具有潛力的SMA致病因子，將其以SMA小鼠驗證後發現，BMP-4於SMA小鼠的脊髓中表現較低，且該表現和SMA小鼠的疾病和發病時間呈現相關性。值得注意的是，過去曾有研究發現BMP之接受器的表現在SMA果蠅模式會下降，而提升BMP的訊息傳遞可減少SMA果蠅模式在神經肌肉交界的異常。此外，臨床上常用的降血脂用藥statin曾被發現能夠促進BMP的表現，而以atorvastatin(為一statin類藥物)治療某一運動神經元退化的小鼠模式時，可減少運動神經元的退化。因此，BMP-4的表現可能和SMA的病理機制相關，且可能可作為一具潛力的SMA治療標的。本研究計畫的第一個目的為：了解BMP-4於SMA-like的神經元中所扮演的角色。我們將嘗試驗證是否抑制細胞內SMN的表現會影響BMP-4系統的變化，而若進而增加BMP-4的表現(基因治療或atorvastatin治療)是否可改進神經元的異常細胞形態及細胞凋亡反應。第二個目的為：了解BMP-4於SMA模式小鼠中所扮演的角色。我們將製備傳統SMA小鼠和具BMP-4基因轉殖的SMA小鼠，以驗證小鼠體內在SMN表現不足時，不同組織中BMP-4系統的表現變化，並分析經BMP-4基因轉殖的SMA小鼠之表現型。第三個目的為：了解BMP-4基因治療及atorvastatin治療是否對SMA小鼠有所療效。我們將分別以腺相關病毒對SMA小鼠進行經腦室注射的BMP-4基因治療，及由經腹腔注射atorvastatin對SMA小鼠進行藥物治療。本研究不但有機會解答SMA的病態生理機制，亦有機會發現新的SMA治療方向。<br> Abstract: Spinal muscular atrophy (SMA) is a hereditary progressive motor neuron disease withoutavailable curative treatment currently. SMA exhibits an autosomal recessive pattern of inheritance withan incidence of 1 in 6,000-10,000 newborns. Most SMA patients have homozygous gene mutation onthe survival of motor neuron 1 gene (SMN1). The mice model of SMA has been generated by themouse Smn knockout-human SMN2 transgenic method. Following our characterization of thephenotype for SMA mice, this rodent model is suitable for use in investigating disease mechanisms andtesting therapies for SMA.Bone morphogenetic proteins (BMPs) are growth factors belonging to the transforming growthfactor β (TGFβ) superfamily. BMP-4 is one of BMPs and is an essential protein for embryonicdevelopment, including the neural system. For adult, BMP-4 is able to stimulate bone formation. Untilnow, the role of BMP-4 in neurological diseases outside from neural development is still unknown.We recently have performed a meta-analysis from 41 human SMA microarray datasets. Analysis ofgene networks from microarray databases revealed 7 target genes as potential SMA modifiers. Aftervalidation of these factors using SMA mice, we have demonstrated that the BMP-4 expression wasreduced in SMA mice and the BMP-4 down-regulation was parallel well to disease severity and stage ofSMA mice. Notably, according to previous studies, the expression of wishful thinking (wit), whichencodes a BMP receptor, was down-regulated in an SMA drosophila model. Further modulation of BMPsignaling was able to improve abnormal phenotypes at the neuromuscular junction of drosophila.Moreover, a group of lipid lowering agents, statin, can also enhance the BMP expression. In a mousemodel of motor neuron disease, atorvastatin (a kind of statin) can attenuate spinal motor neurondegeneration. Therefore, BMP-4 might be a potential disease modifier and a therapeutic target for SMA.The first aim of this project is to investigate the role of BMP-4 in SMA-like culture cells. We willtry to understand whether SMN knock-down will influence expression in BMP-4 system and whetherfurther up-regulation of BMP-4 using gene therapy or atorvastatin treatment will rescue abnormalphenotypes after SMN knock-down in SMA-like neurons. The second aim of this project is toinvestigate the role of BMP-4 in SMA using SMA model mice and BMP-4 transgenic mice. We willtry to analyze the expression of BMP-4 and BMP-4-related protein in various tissues of SMA mice. Wewill also investigate the effects of BMP-4 up-regulation in SMA mice by means of generating BMP-4transgenic SMA mice. The third aim of this project is to investigate the therapeutic responses ofBMP-4 gene therapy (AAV-BMP4) and atorvastatin treatment in SMA mice. We intend to test theeffects of intracerebroventricular delivery of AAV-BMP4 and intraperitoneal injection of atorvastatinin SMA mice using pathological and functional analyses.動物模式BMP4運動神經元疾病脊髓肌肉萎縮症animal modelBMP4motor neuron diseasespinal muscular atrophy