Biological Characteristics Of Glioma Stem/Progenitor Cells (GSPC) And The Relationship Between GSPCs And Angiogenesis

1, background and objectiveIt is well known that the angiogenesis is a key incident for the growth, infiltration ofglioma. However, the complete process of glioma angiogenesis remains to dispute, evenafter painful exploration for decades. Since the beginning of this century, with thegradually deep research on tumor stem cells and their Niche, some novel recognitionabout angiogenesis is being raised. At present, there mainly are three viewpoints:A, Neoformed tumor vessels of tumors were thought to arise by sprouting of pre-existingbrain capillaries induce by tumor cells.B, Tumor vessels were first arranged by tumor cells, and then host endothelial cellsjoined.C, From transdifferentiated tumor stem cells depending on their Niche (shown asfollowing). Figure1, Model of tumor angiogenesis: TC: tumor cells; EC: endothelial cells; GSCs: Gliomasstem cells; A：Zhang S Oncol Resp2006,15(1):15-20；B：Chang YS,Proc Natl Acad Sic USA2000,97(26):14608-14613；C：Ricc-Vitiani L,Natrue2010,133(pt4),973-982；Jun Dong, Stem CellRev and Rep2011,7(1):141-152.This paper aims to study the GSPCs and the relationship between GSPCs and gliomaangiogenesis, i.e. to verify the existence of the Model C in Figure.2, Methods1) To establish the growth model of GSPCs in vitro: fresh tumor specimens weresubjected to enzymatic dissociation into cell suspension, and cultivaed in DF containingEGF and bFGF. Then CD133~+cells, after being isolated by immunomagnetic beads,were seed into caudate nucleus of nude mice. The tumorigenesis incidence was recorded.2) To establish the RFP/GFP glioma model and detect blood vessels: GSCPs (25μl/1×106)transfected with Red Fluorescent Protein (RFP) were transplanted into NC nude miceexpressing Green Fluorescent Protein (GFP), intracranially or subcutaneously. Two to3weeks later, after tumorgenesis, tumor tissues were pressed into tabletting and tumorvessels were detected under under fluorescence microscope and confocal microscopy.3, results and discussion1) Characteristics of GSPCs: minority of cells in tumor spheres were CD133~+, but mostwere Nestin~+; in addition, some cells co-expressed both CD133and Nestin, accordingly,cells used in our research were called Glioma Stem/Progenitor Cells (GSPCs). The mostdifference between GSPCs and neural stem cells (NSCs) was that NSCs coulddifferentiate into mature end cells, but GSPCs could not, for the differentiated GSPCscould even differentiate backwards into progenitor cells. The tumorigenesis ratio were100%no matter GSPCs were transplanted intracranially or subcutaneously, and thetransplanted tumor grows at a highly-invasive pattern.2) Characteristics of RFP/GFP glioma models: the tumorigenesis ratio of GSPCs-RFP were100%, like GSPCs, no matter being transplanted intracranially or subcutaneously,with a highly-invasive growth pattern. The different fluorescence expressed by tumorcells (RFP) and host cells (GFP) make it very convenient to show their different rolesduring the tissue remodel process when observed under a fluorescence microscope orconfocal microscopy.3) Tumor vessels and host vessels: tumor vessels were made up of cells with red, greenor yellow fluorescence under a confocal microscopy, indicating the cellular originrespectively being tumor cells, host cells and their fusional cells. The thickness of tumortissue tabletting was about20μl, which was much thicker than common pathologicalsection, and it made the strip vessels much more visible.4, conclusion and Innovation1) CD133+glioma stem cells are only cells at quiescent periods, and once thedifferentiation program started, CD133+glioma stem cells will become Nestin+evenco-expressing CD133. We first demonstrated that GSPCs could not differentiate intomature end cells but have a potential to retro-differentiate.2) RFP/GFP bio-fluorescence labelled glioma models could clearly show the anatomyand histological relationships between tumor cells and host cells, which has an advantageat the research field of tumor tissue remodel and micro-environments. And theexperimental platform was reported first by us at home.3) With the RFP/GFP bio-fluorescence labelled glioma models, we first took advantageof tissue squash technique at home to show strip vessels, which could seldom be viewedby traditionally histological section. Our results showed that there were three types ofvessels cellular origins in tumor tissues: tumor cells, host cells and their fusional cells.These results had an important significance to the further study on tumor angiogenesisand transformation.