动物造模 z-bio 2023-03-09 516

　　Objective To establish a mouse model of endothelial cell conditional knockout of EMCN gene, and to explore the differences in lung metastasis between C57BL/6 mice and endothelial cells conditional knockout of EMCN mouse model.

　　Methods The endothelial cell-specific Tek-CreERT2 mice were crossed with EMCNflox/flox mice, and the offspring male EMCNflox/wt/Tek-CreERT2+ mice were crossed with female EMCNflox/flox mice to obtain endothelial cell-specific knockout EMCN mice. (EMCNflox/flox/Tek-CreERT2+,EMCNecko). The accuracy and efficiency of gene knockout were verified at the DNA and protein levels. After extraction of mouse genomic DNA, PCR amplification was performed to detect Tek-CreERT2 and FLOX sites. After Tamoxifen induction, Western Blot was used to detect the expression of EMCN protein in tissues. The phenotypes of normal C57BL/6 mice and EMCNecko mice were observed and their organs were stained with HE. In order to confirm the effect of EMCN knockout on tumor metastasis, LLC cells were injected into the tail vein of C57BL/6 and EMCNecko mice, and the lungs were dissected 2 weeks later to detect lung metastasis, and the results of lung metastasis in the two groups of mice were statistically compared and analyzed. The LLC cells were subcutaneously injected into C57BL/6 and EMCNecko mice, and the subcutaneous tumors were surgically removed 2 weeks later. The lung metastasis was observed 2 and 3 weeks after the operation, and the lung tissues of the two groups were stained with HE. Statistical comparative analysis of lung metastases.

　　Results The successful construction of EMCNflox/flox/Tek-creERT2+ mice was confirmed from DNA and protein levels. Analysis of mouse tissue HE sections showed no abnormal organ development in knockout mice. Tumor-excised C57BL/6 and EMCNecko mice developed lung metastases after tail vein injection of LLC and subcutaneous injection of LLC. Compared with C57BL/6 mice, the absence of ECN in endothelial cells significantly promoted lung metastasis in mice.

　　Conclusion The endothelial cells EMCN conditional knockout mice were successfully obtained, and the ability of endothelial cells EMCN knockout mice to lung metastasis was significantly better than that of normal C57BL/6 mice. animal model.