Immunohistochemical expression of stem cell, endothelial cell, and chemosensitivity markers in primary glioma spheroids cultured in serum-containing and serum-free medium.
OBJECTIVE
To investigate the influence of serum-free medium (SFM) supplemented with epidermal growth factor and basic fibroblast growth factor compared with conventional serum-containing medium (SCM) on the phenotype of organotypic primary spheroids from seven gliomas.


METHODS
Paraffin sections of the original surgical specimens, primary glioma spheroids, and U87 derived spheroids were stained immunohistochemically with the stem cell markers CD133, podoplanin, Sox2, Bmi-1, and nestin; the endothelial cell markers CD31, CD34, and Von Willebrand Factor (VWF); the chemosensitivity markers P-glycoprotein and tissue inhibitor of metalloproteinases-1 (TIMP-1); and glial fibrillary acidic protein, neural cell adhesion molecule CD56, and the proliferation marker Ki67.


RESULTS
Scoring of the immunohistochemical stainings showed that the expression of CD133 and all other markers included was preserved in primary spheroids, confirming the in vivo-like nature of these spheroids. Spheroids in SFM better mimicked the in vivo phenotype with significantly more CD133, CD34, VWF, P-glycoprotein, TIMP-1, and Ki67 compared with SCM.


CONCLUSIONS
In this first study of the influence of SFM on primary glioma spheroids, the conditions favored an in vivo-like phenotype with increased expression of CD133. More vascular structures were found in SFM, suggesting that the close relationship between blood vessels and tumor stem-like cells was better preserved in this medium.
Christensen
Karina
K
Department of Pathology, Odense University Hospital, 5000 Odense, Denmark.
Aaberg-Jessen
Charlotte
C
Andersen
Claus
C
Goplen
Dorota
D
Bjerkvig
Rolf
R
Kristensen
Bjarne Winther
BW
D000328:Adult; D000368:Aged; D014408:Biomarkers, Tumor / Q000032:analysis*; D001932:Brain Neoplasms / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods*; D003470:Culture Media; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005910:Glioma / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D010641:Phenotype; D018874:Spheroids, Cellular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D014407:Tumor Cells, Cultured
D014408:Biomarkers, Tumor; D003470:Culture Media; D016895:Culture Media, Serum-Free; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor