Preliminary Study on Relationship Between Temozolomide Chemotherapy-Resistant Cells and Stem Cells in Gliomas.

Abstract

To study the relationship between temozolomide (TMZ) chemotherapy-resistant cells and stem cells in gliomas. The U251 glioma cell line was exposed to TMZ to generate TMZ-resistant colonies (U251/TMZ cell line) using the pulse drug method. The TMZ sensitivity of U251/TMZ and parental cells was examined using an MTT assay. The cell growth curve was drawn to show the growth of the two kinds of cells. Glioma stem cells (GSCs) were cultured and differentiated in vitro. Immunofluorescence assays were used to identify the expression of CD133, Nestin, and ABCG2 in U251/TM and U251 cells. Western blot analysis was used to analyse protein expression levels. The U251/TMZ cell line was successfully cultured in vitro. The IC50 value of the U251/TMZ cell line is 8.1 times that of the parental U251 cell line (t=-63.28, p=0.00). The doubling time of U251/TMZ cells was long compared with the parental cells. GSC tumour spheres were successfully cultured in vitro, and they differentiated in medium containing serum. The expression of CD133, Nestin, and ABCG2 in U251/TMZ cells was significantly higher than that in the parental U251 cells (t=43.35, p=0.00; t=12.31, p=0.00; t=11.49, p=0.00). Immunofluorescence staining of CD133, Nestin, and ABCG2 was significantly higher in U251/TMZ than in the parental U251 cells (t=43.35, p=0.00; t=12.31, p=0.00; t=11.49, p=0.00). Moreover, Western blot results showed that CD133, Nestin, and ABCG2 expression was significantly higher in U251/TMZ cells than that in the parental U251 cells (t=17.76, p=0.00; t=18.78, p=0.00; t=6.19, p=0.00). The U251/TMZ cell line has the biological characteristics of GSCs. The relationship between GSCs and chemotherapy-resistant cells has been preliminary proven to be partially overlapping, which can provide a new perspective when using appropriate cell subpopulations as targets for glioma.