关键词: 阿尔茨海默病, Complexin-1/2, 重组腺相关病毒载体, 基因过表达

Abstract:

Objective　To construct recombinant adeno-associated virus over-expressing vector of complexin-1/2 (CPLX-1/2) and investigate the effects of overexpression of  CPLX-1/2 on spatial memory impairment in Alzheimer’s Disease.　Methods 　The CPLX-1/2 gene sequences of mice reported in Genbank were analyzed by Oligo software, and the CPLX-1/2 gene was synthesized. The overexpressing vector was constructed and transfected into Escherichia coli TOP 10. Purified plasmids from the positive clones was confirmed by PCR and sequencing. The AAV293 cells were transfected by the recombinant adeno-associated virus vector to achieve the adeno-associated virus packaging production. The recombinant adeno-associated virus vector was injected into hippocampus of 3XTg-AD mice using a stereotaxic instrument guided by a computer and a motorized nanoinjector. The expression of CPLX-1/2 in hippocampus was detected by Western-blot and immunofluorescence; Morris water maze was performed to measure the change of spatial memory impairment in 3XTg-AD mice. 　Results　The PCR and sequencing confirmed that the recombinant adeno-associated virus vector was successfully constructed. Injection of the overexpessing vector led to significantly increased expression of CPLX-1/2 in hippocampus, and significantly improved the spatial memory impairment in 3XTg-AD mice.　Conclusion　The adeno-associated virus overexpressing vector of CPLX-1/2 gene was successfully constructed, and highly expressed in hippocampus of 3XTg-AD mice. The expression of CPLX-1/2 gene in hippocampus significantly improved spatial memory impairment in 3XTg-AD mice. This study provides a new approach to further study the function of CPLX-1/2 at in vivo levels. These findings suggest that CPLX-1/2 may serve as key therapeutic targets for AD.

Key words: Alzheimer’s disease (AD), Complexin-1/2 (CPLX-1/2), Adeno-associated virus vector, Gene over-expression