Incubation Of EGFP Nude Mice And Established Animal Models Of Human Glioma Using EGFP Nude Mice

Human glioma research is highly dependent on reliable animal models, which allow us to study different aspects of human glioma initiation and progression in vivo. In vitro studies will not allow us to elucidate the complex tumour-host interactions. When establishing a xenotransplantation human glioma model in a living animal, it is desirable that the animal is immunodeficient so that it will not reject foreign material. Immunodeficient mice, such as nude mice, and NOD/SCID mice, are therefore commonly used as in vivo human glioma models. In this study we used transgenic C57BL/6J mice expressing EGFP were crossed with nu/nu nude mice(genetic background was BALB/c) to obtain EGFP-expressing nude mouse, established animal models of human glioma using EGFP-expressing nude mouse, preliminary to explore tumor angiogenesis、the origin of blood vessels of tumor and to investigate the accuracy and feasibility of a clinical3.0T MR scanner in tumor animal models.The study involved two aspects of the xenotransplantation human glioma model in the EGFP nude mice:①To incubate the EGFP transgenic nude mice.②Established animal models of human glioma in the Enhanced Green Fluorescent Protein Nude Mouse.Part I:To incubate the EGFP nude miceObjectives To incubate the enhancement green fluoresent protein(EGFP) nude mice for the purpose of providing the experimental animals for establishment of animal models of glioma and clinical treatment of experimental research.Methods Transgenic C57BL/6J mice expressing EGFP were crossed with nu/nu nude mice(genetic background was BALB/c), then the hairless male nude mouse expressing EGFP was crossed with the hairy female pubescence mouse to obtain nude mouse bearing EGFP expression. The expression status of EGFP in the EGFP nude mice were evaluated under direct observation, by fluorescence microscope and PCR.Results Through the small animal in vivo imaging system2days after birth, can identify whether the expression of EGFP gene,4weeks after birth to the naked eye can identify whether the expression of nude gene propagated to18th generation; Gross anatomy of EGFP nude mice discovery athymismus, frozen sections of organs and tissues were seen green fluorescence under fluorescent microscope; EGFP gene PCR results showed that the brain tissue, subcutaneous muscle,tail and toes all expressed the EGFP gene.Conclusions The EGFP trangenic C57BL/6J mice crossing with nu/nu nude mice can obtain the EGFP nude mice. Tail EGFP genotype identification not to kill nude mice provides molecular evidence for EGFP gene expression,both accurate and reliable,but also reflects the humane care of experimental animals. The EGFP nude mice provides a new experimental animal for tumor model in nude mice. Part Ⅱ:Established animal models of human glioma in the Enhanced Green Fluorescent Protein Nude MouseObjective Establishment of the repeatability and stationary orthotopic and subcutaneous models of the human glioma cell line SHG-44in the EGFP nude mice, preliminary to explore tumor angiogenesis、the origin of blood vessels of tumor and to investigate the accuracy and feasibility of a clinical3.0T MR scanner in tumor animal models.Methods The human glioma cell line SHG-44were implanted into the brain of the EGFP-expressing nude mouse using surgical orthotopic implantation to produce orthotopic models of human glioma, The human glioma cell line SHG-44were injected into the forelimb subcutaneously with lml syringe of the EGFP-expressing nude mouse to produce subcutaneous models; The specimen of orthotopic models after HE and immunity histochemistry dyeing were observed with light microscope and fluorescence microscope,in order to study the transplant lump’s shape characteristic、the invasion and metastasis of neoplasm and the origin of blood vessels of tumor; The orthotopic and subcutaneous models after transplantation25days observation3.0T MRI scan characteristics and compared with pathological results.Result One mouse died before operation, and the remaining nine mice were sacrificed on25th day after tumor cell injection, the tumors were observed with HE staining, the tumor formation rate in orthotopic models was about100%; The cells of orthotopic model still have the basic characteristics of human glioma cells,the blood vessels of tumor were seen strong green fluorescence under fluorescent microscope; The orthotopic and subcutaneous models3.0T MRI scan were seen tumor growth, the scanned image is clear and consistent with the pathological results.Conclusion The EGFP-expressing nude mouse can be used for human tumors transplantation experiments and used the color to identify tumor angiogenesis,it have great significance of tumor angiogenesis and the origin of blood vessels of tumor; clinical3.0T MR examination is an effective and non-invasive way to detect the development of glioma in EGFP-expressing nude mouse.