Establishment Of An Orthotopic Xenografted Model Of Human Wilms' Tumor In Nude Mice, And Investigation Of The Role Of All-trans-retinoic Acid In Inducing Differentiation Of Wilms' Tumor In Vivo

Retaining biologic properties of human origins, the subcutaneous implantation of human tumor in nude mice as a disease model has been widely applied to basic research for oncology and pre-clinical experiments. The orthotopic xenograft of human tumor in nude mice, with higher rate of successful grafting and tumor metastasis, was suggested to mimic more precisely human tumor in biologic behavior than subcutaneous model. In this study, we developed an orthotopic xenografted model of human Wilms' tumor in nude mice and investigated its biologic properties to provide an ideal platform for basic research on tumor genesis, metastasis and pre-clinical experimental interference of human Wilms' tumor.The utilization of retinoic acid as an agent of induction differentiation in clinical treatment for leukmia, particularlly in acut promyelocytic leukemia, obtained significant result. Wilms' tumor is one of the most common solid tumors in children, and the genesis of Wilms' tumor is believed to be associated with delayed differentiation of metanephric blastema in fetus and retaining proliferation in postnatal period. Retinoic acid has a key role in regulation of fetal tissue development. However, does it function as induction of differentiation in treatment for Wilms' tumor?A research group led by Gao, while attempting to establish a chemical induced model of Wilms' tumor in mice, found that rodent mice fed with low vitamin A are more likely to develop chemical induced tumor than those normally fed. Previous experiment conducted by Vencent TS also showed that proliferation of Wilms' tumor cell was inhibited in response to retinioc acid in vitro by way of dose-dependent. But, it has not been evidenced yet whether retinoic acid functions as interference with Wilms' tumor in experiment in vivo or not.Thus, the xenograft model of human Wilms' tumor was applied to investigate the effect of retinoic acid on the tumor in this study, to confirm the results from the previous research in vitro and to provide valuable data for clinical practice. It is established that retinoic acid has the ability to combine with type â…¡ of insulin-like growth factor receptor and regulates transcription of H19 gene. Because the IGF2 and H19 are two important factors involved in the process of fetal tissue differentiation and Wilms' tumor development, we hypothesize whether retinoic acid induces differentiation of Wilms' tumor by regulatiion of transcription of these two genes?Therefore, through utilization of reverse transcript-polymerase chains reaction to measure the level of IGF2mRNA and H19mRNA, we attempt to explore the mechanism by which retinoic acit induces Wilms' tumor differentiation.Section one. Establishment and biologic characterization of orthotopicxenografted model of human Wilms' tumor in nude miceMethod: Histological-intact patient's Wilm's tumor tissue fragments were inoculated into subrenal capsule of nude mice and subcutaneous site of the others as primary transplantation and then serial passage transplantations were carried out after formation and growth of tumor in primary tranplantation; the kidney innoculated tumor was anatomized for observation of tumor formation in the primary, the second, and the third passages of nude mice that were executed eight weeks after the inoculation was conducted. Then the transplanted tumors were removed and measured in weight in the 4th generation, of which three nude mice were executed weekly from the third to the eighth week after the inoculation, the purpose being to plot the time-tumor weight curve: The nude mice bearing tumor underwent an anatomic observation in its lung for metastasis. The distribution of cell cycle and apoptosis as well as of ploidies was monitored in tumor samples from both serial passages and original tissue with flow cytometry and the tumors were subjected to light microscopic observation for morphology and immunohistochemical staining of intermediate filaments, including CK18, vimentin, desmin; and PCR amplification for three microsatelli...