2022, Vol. 26 ›› Issue (23): 3728-3732

CTLA4.FasL promotes the engraftment of xenogeneic hepatic oval cells in rat spleen

Wan Zhen1, Wang Xuzhen2, Zhang Xiaogang3   

Abstract: BACKGROUND: Cytotoxicn lymphocyte antigen 4.fasrymndrit ligand (CTLA4.FasL) can effectively inhibit alloreactive and autoreactive T lymphocytes. However, the inhibitory effect of CTLA4.FasL on xenogeneic T lymphocytes in hepatic oval cell transplantation via the spleen remains unclear.  
OBJECTIVE: To investigate the effect of CTLA4.FasL on the proliferation of xenogeneic lymphocytes, and to monitor the survival and engraftment of CTLA4.FasL-hepatic oval cells in rat spleen.
METHODS:   The recombinant lentiviral vectors carrying CTLA4.FasL fusion gene (Lv/CTLA4.FasL) and polybrene were added to the rat hepatic oval cell suspension. We used hepatic oval cells that were not infected with lentivirus and infected blank lentivirus-hepatic oval cells as controls, and then observed the viability of hepatic oval cells and the expression of red fluorescent protein lentiviral vector under a fluorescence microscope. Sprague-Dawley rats undergoing 2/3 hepatectomy were transplanted with CTLA4.FasL-hepatic oval cells, blank lentivirus-hepatic oval cells, hepatic oval cells, or phosphate buffered saline (without hepatic oval cells) through the spleen. At 1, 5, 14, and 21 days after hepatectomy, the concentration of CTLA4.FasL in rat serum was determined by enzyme linked immunosorbent assay, and the spleen of the recipient rats was taken for CK-19 immunohistochemical staining. C57BL/6 mouse lymphocytes were used as stimulator cells, and Sprague-Dawley rat lymphocytes as response cells. In the mixed lymphocyte reaction, CTLA4.FasL-hepatic oval cells, blank lentivirus-hepatic oval cells, and hepatic oval cells were inoculated, or CTLA4.FasL-hepatic oval cells (a mass concentration   of 10, 50, and 100 μg/L), blank lentivirus-hepatic oval cells, and hepatic oval cell supernatant. After 96 hours of culture, 5-bromodeoxyuridine method was used to detect the proliferation of lymphocytes in each group.  
RESULTS AND CONCLUSION: Both CTLA4.FasL-hepatic oval cells and the culture supernatant could effectively inhibit the proliferation of rat lymphocytes in the xenogeneic mixed lymphocyte culture system. Spleen lymphocytes from the recipient rats transplanted with CTLA4.FasL-hepatic oval cells had a low immune response to mouse lymphocytes, while the spleen cells derived from the recipient rats transplanted with blank lentivirus-hepatic oval cells and hepatic oval cells showed strong responses. CK-19 positive cells were detected in rat spleen parenchyma of the CTLA4.FasL-hepatic oval cell group but not in the phosphate buffered saline group, hepatic oval cell group, and blank lentivirus-hepatic oval cell group at 21 days after transplantation. The results show that CTLA4.FasL can promote the survival and engraftment of hepatic oval cells in the rat spleen parenchyma through inhibiting the proliferation of xenogeneic lymphocytes.
Key words: cytotoxicn lymphocyte antigen 4.fasrymndrit ligand, xenogeneic, hepatic oval cell, spleen, engraftment