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学科主题: 临床医学
题名:
Inducing agent tamoxifen of CreER(T2) to reduce the side effects of gene therapy for Parkinson′s disease
作者: Li, Xiaogang1; Fan, Dongsheng1; Xiao, Weizhong1; Shen, Yang1; Muramatsu, Shin-ichi2; Ozawa, Keiya2; Nakano, Imaharu2
关键词: adeno-associated virus ; Cre recombinase ; tyrosine hydroxylase ; gene regulation ; Parkinson&prime ; s disease ; neural regeneration
刊名: NEURAL REGENERATION RESEARCH
发表日期: 2010-04-30
DOI: 10.3969/j.issn.1673-5374.2010.08.005
卷: 5, 期:8, 页:591-596
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology
类目[WOS]: Cell Biology ; Neurosciences
研究领域[WOS]: Cell Biology ; Neurosciences & Neurology
关键词[WOS]: ADENOASSOCIATED VIRUS VECTORS ; AMINO-ACID DECARBOXYLASE ; BEHAVIORAL RECOVERY ; TRIPLE TRANSDUCTION ; PRIMATE MODEL ; ANIMAL-MODEL ; RAT MODEL ; DELIVERY ; CELLS ; AAV
英文摘要:

BACKGROUND: Gene therapy for Parkinson′s disease is being explored as an effective strategy to restore and protect the function of neuronal cells in the substantia nigra. Regulation of gene expression is necessary for gene therapy to avoid adverse effects due to excessive synthesis of transgene products.

OBJECTIVE: Here we developed recombinant adeno-associated virus (AAV) as a viral vector-mediated gene regulation system based on Cre recombinase fused to the mutated ligand-binding domain of the estrogen receptor (CreER(T2)) + inducing agent tamoxifen. Inducible Cre recombinase was used to reduce tyrosine hydroxylase gene expression and to prevent the excessive increase in dopamine.

DESIGN, TIME AND SETTING: A genetic engineering in vitro comparative study and randomized controlled animal experiment. This study was conducted at the Gene Therapy Center, Jichi Medical School, Japan from June 2002 to June 2004.

METHODS: To construct a recombinant AAV vector carrying a dopamine synthase gene. The tyrosine hydroxylase gene was inserted using a loxP fragment that could be regulated by Cre recombinase. The recombinant AAV vector carrying the CreER(T2) gene was co-transduced with HEK293 cells and the corpus striatum in a rat model of Parkinson′s disease, with inducing agent tamoxifen to regulate gene expression.

MAIN OUTCOME MEASURES: The levels of dopamine and aromatic L-amino acid decarboxylase (AADC) activity were detected in HEK293 cell medium and in the corpus striatum in a rat model of Parkinson′s disease using high-performance liquid chromatography. Immunofluorescence double staining was used to observe tyrosine hydroxylase and Cre or AADC co-expression in HEK293 cell medium. Immunohistochemical staining was employed to observe tyrosine hydroxylase and AADC expression and behavioral changes were measured in Parkinson′s rats.

RESULTS: Transfected AAV-CreER(T2) and AAV expressing dopamine synthesis enzymes could increase the synthesis of dopamine in HEK293 medium and Parkinson′s rat striatum (P < 0.01) and improve the rotational behavior of Parkinson′s rats. While tamoxifen markedly reduced overproduction of dopamine caused by cotransfection of viral vectors (P < 0.01), but did not affect the expression and activity of AADC.

CONCLUSION: The application of AAV vector-encoded tyrosine hydroxylase gene under the gene regulation system of Cre-ERT2, after tamoxifen treatment, can effectively control the generation of genetically modified products to reduce the production of excessive dopamine in vivo and in vitro. Therefore, this method can increase the safety of gene therapy.

语种: 英语
项目资助者: Ministry of Education, Science, Sports and Culture ; Japanese Government ; Japan Ministry of Health, Labour and Welfare ; Cell Science Research Foundation
WOS记录号: WOS:000277425200005
Citation statistics:
内容类型: 期刊论文
URI标识: http://ir.bjmu.edu.cn/handle/400002259/52412
Appears in Collections:北京大学第三临床医学院_神经内科_期刊论文

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作者单位: 1.Jichi Med Sch, Dept Neurol, Jichi, Tochigi 3290498, Japan
2.Peking Univ, Hosp 3, Dept Neurol, Beijing 100083, Peoples R China

Recommended Citation:
Li, Xiaogang,Fan, Dongsheng,Xiao, Weizhong,et al. Inducing agent tamoxifen of CreER(T2) to reduce the side effects of gene therapy for Parkinson&prime;s disease[J]. NEURAL REGENERATION RESEARCH,2010,5(8):591-596.
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