| Summary: | Objective To construct NOD-SCID transgenic mice with Tet-on regulated liver specific expression of urokinase plasmin activator (uPA), repair subacute liver injury in mice through human liver cancer cell transplantation, and thereby establish humanized liver cell chimeric mice. Methods With CRISPR/Cas9 technology, under the guidance of gRNA, Cas9 endonuclease was cut and edited at Rosa26 locus, and then inserted into the target gene pTight-uPA-pA-Alb promoter rtTA-pA expression frame in order to enable the target gene to follow the stable inheritance of chromosomes and maintain stable biological functions. The newly cultivated transgenic mice were divided into induction group (n=17), transplantation group (n=16), non-induction group (n=5), and blank control group (n=5). The induction group was induced by doxycycline for 3 weeks, and then uPA expression in the liver caused subacute liver injury in mice, which was repaired through transplantation of human liver cancer cells. After 4 weeks of transplantation, the effect of humanized liver cell chimeric mice was evaluated through mouse serum biochemistry and histopathology. Results The serum levels of uPA and alanine aminotransferase (ALT) were significantly higher in the induction group than those the non-induction group and blank control group (P < 0.01), so was the human albumin content in the transplantation group than that of the 2 groups without transplantation (P < 0.01). HE staining showed that human hepatoma cells participated in the repair of mouse liver tissues. Laser confocal microscopy displayed that human hepatoma cells gathered in the mouse liver and expressed human albumin. Histochemical staining showed that human keratin (CK18) was positive in the mouse liver tissues. Conclusion The newly constructed transgenic mice can induce uPA expression in the mouse liver through doxycycline, causing subacute liver damage, and then construct humanized chimeric mice through transplantation and repair of human liver cancer cells.
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