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Intermittent hypoxia regulates stem-like characteristics and differentiation of neuroblastoma cells.

ABSTRACT: Neuroblastomas are the most common extracranial solid tumors in children. Neuroblastomas are derived from immature cells of the sympathetic nervous system and are characterized by clinical and biological heterogeneity. Hypoxia has been linked to tumor progression and increased malignancy. Intermittent hypoxia or repeated episodes of hypoxia followed by re-oxygenation is a common phenomenon in solid tumors including neuroblastoma and it has a significant influence on the outcome of therapies. The present study focuses on how intermittent hypoxia modulates the stem-like properties and differentiation in neuroblastoma cells.Cell survival was assessed by clonogenic assay and cell differentiation was determined by morphological characterization. Hypoxia-inducible genes were analyzed by real-time PCR and Western blotting. Immunofluorescence, real-time PCR and Western blotting were utilized to study stem cell markers. Analysis of neural crest/sympathetic nervous system (SNS) markers and neuronal differentiation markers were done by real-time PCR and Western blotting, respectively. Intermittent hypoxia stimulated the levels of HIF-1? and HIF-2 ? proteins and enhanced stem-like properties of neuroblastoma cells. In intermittent hypoxia-conditioned cells, downregulation of SNS marker genes and upregulation of genes expressed in the neural crest were observed. Intermittent hypoxia suppressed the retinoic acid-induced differentiation of neuroblastoma cells.Our results suggest that intermittent hypoxia enhances stem-like characteristics and suppresses differentiation propensities in neuroblastoma cells.

SUBMITTER: Bhaskara VK

PROVIDER: S-EPMC3281893 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Intermittent hypoxia regulates stem-like characteristics and differentiation of neuroblastoma cells.

Bhaskara Vasantha Kumar VK Mohanam Indra I Rao Jasti S JS Mohanam Sanjeeva S

PloS one 20120217 2

<h4>Background</h4>Neuroblastomas are the most common extracranial solid tumors in children. Neuroblastomas are derived from immature cells of the sympathetic nervous system and are characterized by clinical and biological heterogeneity. Hypoxia has been linked to tumor progression and increased malignancy. Intermittent hypoxia or repeated episodes of hypoxia followed by re-oxygenation is a common phenomenon in solid tumors including neuroblastoma and it has a significant influence on the outcom ...[more]

PMID: 22363512

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Project description:BackgroundNeuroblastoma is the most common extracranial pediatric solid tumor. Intermittent hypoxia, which is characterized by cyclic periods of hypoxia and reoxygenation, has been shown to positively modulate tumor development and thereby induce tumor growth, angiogenic processes, and metastasis. Bone is one of the target organs of metastasis in advanced neuroblastoma Neuroblastoma cells produce osteoclast-activating factors that increase bone resorption by the osteoclasts. The present study focuses on how intermittent hypoxia preconditioned SH-SY5Y neuroblastoma cells modulate osteoclastogenesis in RAW 264.7 cells compared with neuroblastoma cells grown at normoxic conditions.MethodsWe inhibited HIF-1α and HIF-2α in neuroblastoma SH-SY5Y cells by siRNA/shRNA approaches. Protein expression of HIF-1α, HIF-2α and MAPKs were investigated by western blotting. Expression of osteoclastogenic factors were determined by real-time RT-PCR. The influence of intermittent hypoxia and HIF-1α siRNA on migration of neuroblastoma cells and in vitro differentiation of RAW 264.7 cells were assessed. Intratibial injection was performed with SH-SY5Y stable luciferase-expressing cells and in vivo bioluminescence imaging was used in the analysis of tumor growth in bone.ResultsUpregulation of mRNAs of osteoclastogenic factors VEGF and RANKL was observed in intermittent hypoxia-exposed neuroblastoma cells. Conditioned medium from the intermittent hypoxia-exposed neuroblastoma cells was found to enhance osteoclastogenesis, up-regulate the mRNAs of osteoclast marker genes including TRAP, CaSR and cathepsin K and induce the activation of ERK, JNK, and p38 in RAW 264.7 cells. Intermittent hypoxia-exposed neuroblastoma cells showed an increased migratory pattern compared with the parental cells. A significant increase of tumor volume was found in animals that received the intermittent hypoxia-exposed cells intratibially compared with parental cells.ConclusionsIntermittent hypoxic exposure enhanced capabilities of neuroblastoma cells in induction of osteoclast differentiation in RAW 264.7 cells. Increased migration and intratibial tumor growth was observed in intermittent hypoxia-exposed neuroblastoma cells compared with parental cells.

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Intermittent hypoxia regulates stem-like characteristics and differentiation of neuroblastoma cells.

Publications

Intermittent hypoxia regulates stem-like characteristics and differentiation of neuroblastoma cells.

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