Research On The Original Of The Primary Liver Cancer From Bone Marrow-Derived Cells And The Establishment Of The Animal Models Involved In This Study

BackgroundTumors are believed to originate from the transformation of normal stem cells. Current evidence indicates that bone marrow-derived stem cells (BMDCs) are frequently recruited to sites of tissue injury and inflammation, contribute to tissue regeneration and repair. For example, bone marrow-derived stem cells have an important role in liver regeneration and repair. These suggest that bone marrow-derived stem cells should also represent a potential source of malignancy. In 2004, Hougton et al. reported that mouse gastric cancer induced by Helicobacter felis infection originated from bone marrow-derived cells. The finding strongly supports that tumors derive from bone marrow-derived stem cells. In this study, our aim is to investigate Whether primary liver cancer also originates from bone marrow-derived stem cells.Part IThe establishment of syngeneic bone marrow transplantation in C57BL/6J mice Chapter 1Objective: To investigate the optimal irradiation dose for myeloablation in C57BL/6J mice. Method: C57BL/6J mice were irradiated with 8 Gy, 9 Gy, 10 Gy and 11 Gy from⁶⁰Co respectively , then transplanted or not transplanted with C57BL/6J marrow . The survival rates were obtained.Result: The non-transplanted mice were all death. But in transplanted mice, thesurvival rates were 100%, 95%, 55%, 5% with 8 Gy, 9 Gy, 10 Gy, 11 Gy respectively. The difference of the survival rate was significant between 9 Gy and 10 Gy (95% vs. 55%, p<0.01), and not between 8 Gy and 9 Gy (100% vs. 95%, p>0.05).Conclusion: 9.0 Gy is the optimal irradiation dose for myeloablation in C57BL/6J mouse.Chapter 2Objective: To observe and evaluate the reconstitution of bone marrow in transplanted mouse undergoing irradiation-induced myeloablation with 9.0 Gy. Method: Peripheral leukocytes were dynamically examined in transplanted mouse undergoing 9.0 Gy irradiation.Result: Peripheral leukocytes kept decreasing early, and begin to increase rapidly 6 days later, and were recovered completely 5 weeks after marrow transplantation. Conclusion: Bone marrow can be reconstituted completely after 5 weeks in transplanted mouse undergoing 9.0 Gy irradiation.Chapter 3Objective: To evaluate the overall level of engraftment in transplanted mouse with9.0 Gy irradiation.Method: Recipient C57BL/6J mice with 9.0 Gy irradiation were transplanted withmarrow from ROSA26. The overall level of engraftment was assessed by analysis of beta-galactosidase with immunocytochemistry in peripheral leukocytes. Result: The mean positive rate of beta-galactosidase was 95.1±3.0% (range from 91%-100%) in peripheral leukocytes.Conclusion: The overall level of engraftment is satisfactory in transplanted mouse with 9.0 Gy irradiation.Chapter 4Objective: To evaluate the feasibility of the retro-orbital plexus injection for marrowtransplantation in C57BL/6J mice.Method: The comparison between the retro-orbital plexus injection and tail veininjection was conducted in C57BL/6J mice with 9.0 Gy irradiation.Result: The marrow transplantation time was significant shorter in the retro-orbitalplexus injection compared to the tail vein injection (13 min vs. 5.5 min, p<0.01). But there was no difference between the two methods in the survival rate (100% vs. 100%) and the marrow reconstitution time (5 w vs. 5 w) .Conclusion: The retro-orbital plexus injection is feasibile, and is an optimal method for marrow transplantation in C57BL/6J mice.Part IIEstablishment of induced liver cancer model in C57BL/6J mice Chapter 1Objective: To establish chemical hepatocarcinogen-induced liver cancer model inC57BL/6J mice.Method: C57BL/6J mice undergoing low dose irradiation administrated AAF and DEN respectively. The hepatocarcinogenesis was examined by MRI.Result: The hepatocarcinogenesis rates were 77.8% in DEN and 68.8% in AAF. Andthe hepatocarcinogenesis times were 24-32 weeks in DEN and 40-52 weeks in AAF. The sensitivity of MRI for liver cancer diagnosis was 83.3% in C57BL/6J mice.Conclusion: Induced liver cancer model can be effectively established in C57BL/6Jmice by combination of irradiation and DEN or AAF. MRI is a valuable diagnostic instrument during the mouse hepatocarcinogenesis.Chapter 2Objective : To investigate the morphologic evolution during chemicalhepatocarcinogenesis in C57BL/6J mice.Method: A series histopathology research was employed to examine the hepaticlesions during chemical hepatocarcinogenesis.Result: Mouse hepatocarcinogenesis induced by chemical hepatocarcinogen hadsimilar procession and morphologic features to human.Conclusion: Chemical hepatocarcinogen-induced liver cancer model is analternative experimental model for study on human liver cancer. And further study on this model is required.Part IIIResearch on the original of the primary liver cancer from bonemarrow-derived cellsObjective: To investigate the role of BMDC in hepatocarcinogenesis.Method: Female recipient C57BL/6J mice irradiated with 9.0 Gy from ⁶⁰Co were transplanted with male donor C57BL/6J mice. And DEN was administrated after 5 weeks of recovery. All mice were sacrificed and the liver carcinomas were obtained at 30 weeks. Tumor tissues were analyzed by Y chromosome FISH and SRY gene PCR.Result: All the tumor cells were Y chromosome positive in the liver cancersanalyzed.Conclusion: Liver cancer can originate from bone marrow-derived cell. Furtherstudy on the mechanism is needed.