关键词: Cyr61, 心肌, 骨骼肌, 条件性基因敲除小鼠

Abstract: Objective To construct a conditional knockout mice model of cysteine-rich angiogenesis inducer 61 (Cyr61, also known as Ccn1) gene in myocardium and skeletal muscle regulated by CRE recombinant enzyme. Methods C57BL/6 mice were used to create CKmm-Cre^+/- Cyr61^flox/flox mice by using CRISPR/Cas9 technology. The successful construction of conditional knockout mice was confirmed by identifying the Cyr61^flox/flox, 3'LoxP and Cre enzyme sequences in mice. The knockout efficiency of Cyr61 was confirmed by Western blot. The skeletal muscle function of Cyr61 knockout mice was evaluated and following related indicators in the myocardium and skeletal muscle were detected by Western blot: Aging (p53, p21),inflammatory response (TNF-α,IL-18,IL-1β),fibrosis (vimentin,TGF-β,α-SMA,COL1). Results The mice were successfully bred and identified. In comparison with the control group, Cyr61 protein level showed a significant decrease in both myocardium and skeletal muscle in the experimental group(P＜0.01). Compared with the control group, the experimental group mice showed increased skeletal muscle grip strength(P＜0.05), enhanced maximum single contraction force (P＜0.01), and increased average cross-sectional area of the anterior tibialis muscle(P＜0.05). The expression level of p21, TNF-α, IL-18, IL-1β, TGF-β, and COL1 proteins in the myocardium and skeletal muscle of the experimental group was all lower than those of the control group (P＜0.05). Conclusions Myocardial and skeletal muscle-specific Cyr61 gene conditional knockout mice were successfully constructed based on Cre-LoxP technology and heritable trait could be passed stably.

Key words: Cyr6, myocardium, skeletal muscle, gene conditional knockout mice