脊髓小腦萎縮症第八型致病基因分析及建立脊髓小腦萎縮症第八型離體及活體模式

Analysis of spinocerebellar ataxia type 8 gene expression and establishment of spinocerebellar type 8 in vitro and in vivo models

陳韋倫(Wei Lun Chen)

指導教授 ： 謝秀梅

國立臺灣師範大學/理學院/生命科學研究所/碩士(2005年)

摘要

無資料

關鍵字

脊髓小腦萎縮症第八型

並列摘要

Spinocerebellar ataxia type 8 (SCA8) was reported caused by an unstable CTG repeat expansion in the 3-untranslated region of SCA8 gene on chromosome 13q21. How the trinucleotide expansion causes the disease is not clear now. Some studies indicate that SCA8 might not encode protein and play an anti-sense regulatory role on the sense strand gene, KLHL1. We first identified the expression patterns of KLHL1 and SCA8 in both human and mouse brain tissues with RT-PCR and in-situ hybridization. The results show that the expression of sca8 in the brain regions whose functions correlate SCA8 clinical symptoms. The similar expression patterns of sca8 and klhl1 in these regions further suggest an anti-sense regulation of KLHL1 by SCA8. To further investigate the molecular mechanism of SCA8, a transgenic mouse model of SCA8 was established. The human SCA8 genes with 23 or 157 CTG repeats was in frame fused with flag-EGFP and driven by NSE promoter, which is considered as a CNS specific promoter. The founder lines were generated by pronuclear microinjection. Expressional, behavioral and histological analyses were preceded to understand the impact of the CTG repeat expansion on these mice. We also transfected the same constructs into rat pheochromocytoma (PC12) cell line to generate the in vitro model. We found that cell line with expanded 157 CTG repeats was more vulnerable under oxidative compared to cells with normal CTG expansion or vector construct-transfected. In addition to the SCA8 model, an inducible purkinje cells albation transgenic mouse model was also generated to further mimic SCA’s clinical features. We found that NTR transgenic mouse with CB1954 administration shows impairment in balance in motor coordination. Immunohistochemistry shows purkinje cells loss and higher oxidative signal. With the model, we have obtain initial effects resulted from overexpression of SCA8 in vivo and in vitro and late onset damage of cerebellum, which should further provide more information for the therapeutic study of SCA, including SCA8.

並列關鍵字

spinocerebellar type 8 ； nitroreductase ； CB1954 ； in vitro ； in vivo