文献类型: 中文期刊
作者: 沙芳芳 1 ; 范沛 1 ; 杨培长 1 ; 张璐 1 ; 李建科 2 ;
作者机构: 1.河南工业大学生物工程学院
2.中国农业科学院蜜蜂研究所
关键词: CRISPR/Cas9;小鼠;基因敲入;蜂王浆主蛋白1;血压调节
期刊名称: 中国畜牧兽医
ISSN:
年卷期: 2023 年 011 期
页码: 4392-4402
收录情况: 北大核心 ; CSCD
摘要: 【目的】探究蜂王浆中含量最高的活性分子——蜂王浆主蛋白1(MRJP1)基因敲入血管平滑肌细胞后,在体内水平上是否具有调节小鼠血压的功能。【方法】利用CRISPR/Cas9技术,将可识别小鼠Hipp11位点的gRNA、Cas9和包含Sm22α启动子+Mrjp1 cDNA的载体共显微注射小鼠受精卵,制备在血管平滑肌细胞中特异性表达的Mrjp1基因定点敲入小鼠,并验证敲入位点的正确性。小鼠背部皮下包埋微渗透压缓释泵,持续将血管紧张素Ⅱ(AngⅡ)诱导野生型(WT)和纯合子(Mrjp1+/+)小鼠,比较不同时间收缩压和舒张压,以及胸主动脉中膜面积/管腔面积(Media/Lumen area)比值的差异。【结果】F0代嵌合体小鼠与野生型交配,产生杂合子(Mrjp1+/-)小鼠,对Mrjp1+/-小鼠敲入位点同源重组片段的PCR扩增、测序和Southern blotting检测结果显示,Mrjp1基因成功整合到小鼠染色体Hipp11位点。Mrjp1+/-小鼠之间交配产生WT、Mrjp1+/-和Mrjp1+/+小鼠,Mrjp1基因在胸主动脉中能够顺利转录和翻译。持续给予AngⅡ,Mrjp1+/+小鼠收缩压和舒张压升高趋势整体低于WT小鼠,收缩压在第3、6、9、12天差异显著(P<0.05),舒张压在第3(P<0.05)、6(P<0.01)天差异显著;经AngⅡ刺激后,Mrjp1+/+小鼠胸主动脉Media/Lumen area比值亦显著低于WT小鼠(P<0.05)。【结论】Mrjp1基因特异性敲入小鼠血管平滑肌细胞,可在体内水平上抑制AngⅡ诱导的高血压,为深入了解MRJP1蛋白功能及精准开发蜂王浆提供了重要的理论依据。
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