In vitro Tumorigenicity and Stemness Characterization of the U87MG Glioblastoma Cell Line based on the CD133 Cancer Stem Cell Marker

Abstract

The cancer stem cells (CSC) hypothesis is currently the most widely accepted theory regarding tumor formation and self-renewal ability. The need to find pharmacological compounds or biological agents capable of eliminating CSC makes the search for methods of recognition and isolation of these cells a matter of great urgency. The aim of the study was to separate CD133+ and CD133- cell subpopulations from the U87MG glioblastoma cell line by magnetic activated cell sorting (MACS), and to analyze the tumorigenic and stem cell differences among four sets of cells: the cell line grown in serum (monolayer group), the neurospheres of the same cell line, and the CD133+ and CD133- sorted cell fractions from the neurospheres. Our results showed that the neurospheres and the CD133+ cells overexpressed stem cell marker genes such as SOX2, Nestin, Musashi1 and CD133, and formed more colonies in soft agar than the cell groups with less CD133 expression (monolayer and CD133- group). On the other hand the CD133- and the monolayer groups had similar expression levels of CD133, while the CD133- cells expressed higher levels of Nestin, SOX2 and Musashi1 than the monolayer cells did. In addition, cell migration assessed by the scratching test showed that the CD133- and the monolayer cells migrated more than the neurospheres and the CD133+ cells did. Moreover, the CD133- cells had a higher proliferation rate than the CD133+ cells and the neurospheres, with all of these three groups being cultured in the medium for neurospheres. In conclusion, we might point out the presence of CSC in the CD133- group, as suggested by others. Therefore, CD133+ cells are not necessarily equivalent to CSC. Perhaps CD133+ cells present a higher probability of including CSC than CD133- do. Further functional and genetic analyses must be performed in order to reach optimal isolation of CSC. Keywords: