Induced differentiation inhibits sphere formation in neuroblastoma.
Neuroblastoma arises from the neural crest, the precursor cells of the sympathoadrenal axis, and differentiation status is a key prognostic factor used for clinical risk group stratification and treatment strategies. Neuroblastoma tumor-initiating cells have been successfully isolated from patient tumor samples and bone marrow using sphere culture, which is well established to promote growth of neural crest stem cells. However, accurate quantification of sphere-forming frequency of commonly used neuroblastoma cell lines has not been reported. Here, we show that MYCN-amplified neuroblastoma cell lines form spheres more frequently than non-MYCN-amplified cell lines. We also show that sphere formation is directly sensitive to cellular differentiation status. 13-cis-retinoic acid is a clinically used differentiating agent that induces a neuronal phenotype in neuroblastoma cells. Induced differentiation nearly completely blocked sphere formation. Furthermore, sphere formation was specifically FGF-responsive and did not respond to increasing doses of EGF. Taken together, these data suggest that sphere formation is an accurate method of quantifying the stemness phenotype in neuroblastoma.
Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN, USA.
Brian T
BT
Craig
Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN, USA.
Eric J
EJ
Rellinger
Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN, USA.
Alexandra L
AL
Alvarez
Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN, USA.
Haley L
HL
Dusek
Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN, USA.
Jingbo
J
Qiao
Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN, USA; Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, TN, USA. Electronic address: dai.chung@vanderbilt.edu.
Dai H
DH
Chung
D061249:Batch Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D065150:Cellular Reprogramming; D006801:Humans; D014411:Neoplastic Stem Cells / Q000473:pathology*; D009447:Neuroblastoma / Q000473:pathology*; D018874:Spheroids, Cellular / Q000473:pathology*
Cancer stem cell; Limiting dilution analysis; Neuroblastoma; Retinoic acid; Sphere assay