The Expression Analysis Of EGFR And ABCG2 And Their Role In Angiogenesis In Human Glioma Tissues

Background and objective: There is high expression of EGFR in many human tumors, which has been taken as a treatment target and investigated extensively .But whether the EGFR can be considered as molecule etiological factor of tumors, especially for glioma still has few research now. Tumor angiopoiesis and anti-tumor vessels therapy have been the focus point in the field of phymatology research. Recently some research have indicated that tumor vessel is consisted by not only vascular endothelial cells but also tumor cells.For example ,the vasculogenic mimicry consisted by tumor cells and mosaic vessels consisted by endothelial cells and tumor cells.But it is still hard to confirm the two patterns mentioned above in most tumors including glioma,which can be found only in retina malignant melanoma. The objective of my experiment is to investigate the expression and meaning of EGFR in tissue array to aim directly at the formation and development of glioma, meanwhile to observe the angiogenesis patterns in xenografted human glioma in nude mouse by CD34-PAS dual staining and to investigate the expression of stem cell marker gene ABCG2 in transplanted tumors.Methods: We have made two different tissue arrays for the objective mentioned above.The first one contains 118 specimens which including different malignant grades of human glioma tissues,nude mouse-transplanted tumor specimens, glioma cell line multicellular spheroids, glioma stem cellular spheroids and all corresponding specimens for the use of control.Then we performed immunohistochemistry staining for EGFR by SABC to analyze its expression rate and amount, meanwhile we detected the expression amount of EGFR mRNA in the cell lines of tissue array by Fluorescence Real Time Quantitate PCR to check the protein level results further. The second one contains 30 models of subcutaneously transplanted human glioma ,20 models of intracranial transplanted tumor in nude mouse and 6 normal human brain tissue samples as control.Hematoxylin and eosin staining and immunohistochemistry techniques are applied to the tissue array.Detecting the angiogenesis patterns in xenografted human glioma and the positive expression rate and expression position of the ABCG2 with light microscope.Results: In the first tissue array,the staining of EGFR is chiefly located in the cytomembrane.Among 71 human glioma tissues, the positive expression rate of EGFR is 46.5% , 11.1% in Gradeâ… , 28.0% in Gradeâ…¡,60.9% in Gradeâ…¢and 78.6% in Gradeâ…£. The difference of EGFR expression rates between Gradeâ… -â…¡and Gradeâ…¢-â…£glioma is significant(X2= 13.813,P<0.001) and relate to the pathologic grade of glioma with linear positive correlation ( r=0.991,P=0.009).There is no expression of EGFR in normal human brain tissues, normal nude mouse bone marrow and neural stem cellular spheroids.But the expression of EGFR is positive in subcutaneously transplanted tumor in nude mouse, glioma cell line multicellular spheroids and glioma stem cellular spheroids.According to the Fluorescence Real Time Quantitate PCR test , EGFR expression amount is high in glioma cell line spheroids and glioma stem cellular spheroids, but low in neural stem cellular spheroids, which is consistent with the results of protein level determination. In the second tissue array ,we can observe most common endothelium-dependent vessels both in subcutaneously and intracranial transplanted human glioma in nude mouse .The mosaic vessels exist in 5/30 of subcutaneously transplanted human glioma and 6/20 of intracranial transplanted human glioma.The vasculogenic mimicry exist in 18/30 of subcutaneously transplanted human glioma and 13/20 of intracranial transplanted human glioma.In the normal brain tissue we can only find endothelium-dependent vessels,neither mosaic vessels nor vasculogenic mimicry are observed.Some tumor cells lost GFAP expression in the part areas of two kind transplanted tumors.ABCG2 expression exists in two kind transplanted tumors,the average positive cell percentage is 4%.The expression site of ABCG2 is consisted with blood vessel distribution of transplanted tumors.Conclusion: There is no expression of EGFR in normal tissues and neural stem cells.But high expression of EGFR is observated in glioma, specifically in brain tumor stem cells. The expression quantity of EGFR is related closely to malignant grades of glioma.So EGFR can be as molecule etiological factor of the formation and development of glioma for the further study.There are three different angiogenesis patterns in high malignant subcutaneously and intracranial transplanted human glioma in nude mouse.The expression of ABCG2 have associated closely with blood vessel distribution.