YBX1基因肠道特异性敲低小鼠模型的建立

The Establishment of a Mouse Model with Intestine-specific Knockdown of YBX1 Gene

Y-盒结合蛋白1(Y-box binding protein 1,YBX1)是冷休克蛋白超家族的成员,由YBX1基因编码。YBX1在机体多个组织中高度表达,参与多种生理功能的调节。然而目前YBX1在肠道中的作用尚不清楚。本研究先用CCK-8法在细胞水平上检测过表达/抑制YBX1基因对猪(Sus scrofa)肠上皮细胞IPEC-J2、大鼠(Rattus norvegicus)肠上皮细胞IEC-6增殖的影响。然后利用Cre-loxP基因敲除系统,构建YBX1基因肠道特异性敲低小鼠(Mus musculus)模型。通过YBX1flox/flox小鼠与PVillin-Cre^(+/-)小鼠杂交繁育获得子代小鼠。利用PCR方法鉴定小鼠基因型,采用实时定量PCR技术(real-time quantitative PCR,RT-qPCR)和Western blot法分别从mRNA水平和蛋白水平检测YBX1基因肠道特异性敲低小鼠和对照小鼠YBX1的表达情况,验证基因敲低的效率。结果表明,过表达/抑制YBX1基因不影响细胞增殖,模型小鼠YBX1的mRNA水平下降为对照鼠的62%,蛋白水平下降为对照鼠的63%,证明YBX1基因肠道特异性敲低小鼠构建成功,为深入研究YBX1在肠道中的功能和作用机制提供稳定可靠的动物模型。

Y-box binding protein 1(YBX1)is a member of the cold shock protein superfamily,encoded by the YBX1 gene.YBX1 is highly expressed in multiple tissues and participates in the regulation of various physiological functions.However,the role of YBX1 in the intestine remains unclear.This study first assessed the effects of overexpression/inhibition of the YBX1 gene on the proliferation of porcine(Sus scrofa)intestinal epithelial cells IPEC-J2 and rat(Rattus norvegicus)intestinal epithelial cells IEC-6 at the cellular level using the CCK-8 method.Subsequently,a mouse(Mus musculus)model of YBX1 gene intestine-specific knockdown was constructed utilizing the Cre-loxP gene knockout system.By crossing YBX1flox/flox mice with PVillin-Cre^(+/-)mice,we obtained offspring mice.To identify the mouse genotype using PCR method,real-time quantitative PCR(RT-qPCR)and Western blot were employed,YBX1 gene intestine-specific knockdown mice and control mice were subjected to mRNA and protein level detection to verify the efficiency of gene knockdown.The results showed that overexpression/suppression of the YBX1 gene did not affect cell proliferation.The mRNA level of YBX1 in the model mice decreased to 62%of that in the control mice,and the protein level decreased to 63%of that in the control mice,indicating successful construction of the mouse model with intestinal-specific knockdown of the YBX1 gene.This model provides a stable and reliable animal model for further investigation of the function and mechanism of action of YBX1 in the intestine.