期刊
MOLECULAR BIOMEDICINE
卷 3, 期 1, 页码 -出版社
SPRINGERNATURE
DOI: 10.1186/s43556-021-00063-y
关键词
Recombinant adeno-associated virus; MicroRNA; Expression cassettes; MiR-22; Insulin
资金
- China's National Key RD programs [2020YFC2005600]
- National Natural Science Foundation of China (NSFC) [81571792, 81770199]
- National Clinical Research Center for Geriatrics, West China Hospital, Sichuan University [Z20191002]
This study successfully designed a series of expression cassettes for miR-22 using recombinant adeno-associated virus (rAAV) delivery. The optimal approach for efficient overexpression of miR-22 in vivo was determined by selecting appropriate promoters and longer primary miR-22. Overexpression of miR-22 in diabetic mice showed significant improvement in insulin sensitivity.
MicroRNA-22 (miR-22) was suggested to be important for type 2 diabetes but its functions for this disease remained unclear. Recombinant adeno-associated virus (rAAV)-mediated miR delivery is a powerful approach to study miR functions in vivo, however, the overexpression of miR-22 by rAAV remains challenging because it is one of the most abundant miRs in the liver. In this study, a series of expression cassettes were designed and compared. It was shown that different lengths of primary miR-22 were overexpressed in HEK293 and HeLa cells but the longer ones were more efficiently expressed. miR-22 may be placed in either introns or the 3 ' UTR of a transgene for efficient overexpression. RNA polymerase III or II promoters were successfully utilized for miR expression but the latter showed higher expression levels in cell lines. Specifically, miR-22 was expressed efficiently together with an EGFP gene. After screening, a liver-specific TTR promoter was chosen to overexpress miR-22 in diabetic mice fed a high-fat diet. It was shown that miR-22 was overexpressed 2-3 folds which improved the insulin sensitivity significantly. The approach utilized in this study to optimize miR overexpression is a powerful tool for the creation of efficient rAAV vectors for the other miRs.
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