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研究生(外文):Ming-Wei Chen
論文名稱(外文):Comparison of Protein Expression Profiles Between Wild-type and SMA-like Motor neurons
指導教授(外文):Hung LiY.Henry Sun
外文關鍵詞:SMAmotor neuronmotoneuron
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人類脊髓肌肉萎縮症(SMA)是一種隱性染色體遺傳疾病,其病徵為脊髓前角α 運動神經元退化導致肌肉萎縮,是一種常見的新生兒致死性遺傳疾病,發生率約為一萬分之一。雖然在人類第五對體染色體上有兩個 SMN (survivalmotor neuron )基因,但是超過90%以上的病人在靠近染色體端粒的SMN1 基因發生基因刪除或突變的現象,所以稱SMN1 基因為SMA 決定基因。
SMN1 主要譯出全長38kDa 的SMN 蛋白,是個全身性表現的蛋白,特別在運動神經元有更高量的表現。之前的研究指出SMN 蛋白的功能與pre-mRNA progressing、抗細胞凋亡及調節基因表現有關,但脊髓中運動神經元死亡的機制仍屬未知的範疇。
為了瞭解在SMA 疾病中, 造成改變運動神經元特性的變異因子,因此在本實驗中首先依據前人建立脊髓肌肉萎縮症小鼠動物模式的原理,建立SMA-like 之運動神經細胞株。將人類SMN2 基因送入來自小鼠的類運動神經細胞株:NSC-34 細胞中,再以RNA interference 的技術,將細胞中相當於人類SMN1基因的小鼠Smn 基因knock down,藉此模仿SMA 病人運動神經元中SMN 蛋白的表現狀況。再藉由二維電泳分析觀察一般類運動神經細胞與SMA-like 運動神經細胞兩者在蛋白質表現上的差異性,尋找造成運動神經元死亡的可能因子。經過兩者蛋白質表現的比對,找到一個可能的candidate gene:profilin II。利用二維電泳所得之資料可幫助我們瞭解脊髓肌肉萎縮症的可能病理機制。
Spinal muscular atrophy (SMA) is an autosomal recessive disorder characterized by degeneration of spinal motor neurons. With an incidence of 1/10,000 live births,
SMA is one of the most common genetic lethal disorder to infants. Positional cloning has revealed that Survival Motor Neuron (SMN) is the determinant gene. Homozygous mutations or deletions of the telomeric copy of SMN gene (SMN1) are found in all SMA patients, but the centrameric copy of SMN gene (SMN2) is still present. The major detectable product of SMN1 gene, the SMN protein, is a
full-length 38kDa protein. SMN protein is ubiquitously expressed but is found at high levels in motor neurons. From previous studies, the biophysiological functions of
SMN protein is linked to the pre-mRNA processing, cell apoptosis and gene expression, but the reason account for the death involved in spinal cord motor neuron is still unknown, and the pathogenic mechanism of SMA is currently unclear.
We used a mouse motoneuron-like cell line (NSC-34) to establish the cell-base system mimicing SMA motor neurons. After transfecting human SMN2 genomic DNA, we knocked down mouse Smn gene expression of NSC-34 cells via RNA interference technique, and the SMA-like motor neurons were obtained. With proteomic analysis, the difference of protein expression between wild-type and SMA-like motor neurons could be identity. Profilin II, may be a candidate gene that expressed in different level between wild-type and SMA-like motor neuron. The results in this study may provide additional information for the pathogenic
mechanism of SMA.
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