Three-dimensional neuroblastoma cell culture: Proteomic analysis between monolayer and multicellular tumor spheroids

Hari R. Kumar, Xiaoling Zhong, Derek J. Hoelz, Frederick Rescorla, Robert J. Hickey, Linda H. Malkas, John A. Sandoval

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

Introduction: Solid tumors, such as neuroblastoma (NB), are associated with a heterogeneous cell environment. Multicellular tumor spheroid (MCTS) cultures have been shown to better mimic growth characteristics of in vivo solid tumors. Because tumor spheroid growth patterns may be quite different from standard two-dimensional culture systems, we sought to compare the protein expression profiles of two- and three-dimensional in vitro NB cultures, i.e., monolayers and MCTS. Materials and methods: Human NB cells were grown as both monolayers and spheres. Nuclear and cytosolic proteins were analyzed for differentially secreted proteins by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and selected polypeptides were identified by mass spectrometry (LC-MS/MS). Results: Several metabolic (transketolase, triosephosphate isomerase, pyruvate kinase M1/M2, alpha enolase, and phosphoglycerate mutase-1), cell stress response (heat shock proteins (HSP) 90, 70, and 60; antioxidant, thioredoxin), cell structure (septin 2, adenyl cyclase-associated protein-1), tubulin β-2 chain, actin, translationally controlled tumor protein and cofilin), signal transduction (peptidyl prolyl cis/trans isomerase A), biosynthetic (phosphoserine aminotransferase) and transport (cellular retinoic acid binding protein 1) polypeptides were overexpressed in spheroids. Several protein groups were differentially expressed between NB monolayers and spheroids. Conclusion: The altered proteins among NB spheroids may represent an important link between monolayer cell cultures and in vivo experiments and thus a more ideal in vitro culture system for determining the precise threedimensional microenvironment of NB.

Original languageEnglish
Pages (from-to)1229-1234
Number of pages6
JournalPediatric Surgery International
Volume24
Issue number11
DOIs
StatePublished - Nov 2008

Fingerprint

Cellular Spheroids
Neuroblastoma
Proteomics
Cell Culture Techniques
Neoplasms
phosphoserine aminotransferase
Proteins
Phosphoglycerate Mutase
Septins
Cyclophilin A
Actin Depolymerizing Factors
Transketolase
Triose-Phosphate Isomerase
Chaperonin 60
HSP90 Heat-Shock Proteins
Peptides
Retinoic Acid Receptors
Thioredoxins
HSP70 Heat-Shock Proteins
Pyruvate Kinase

Keywords

  • 3-D Culture
  • Monolayers
  • Multicellular spheroid
  • Neuroblastoma
  • Proteomics
  • Tumor microenvironment

ASJC Scopus subject areas

  • Pediatrics, Perinatology, and Child Health
  • Surgery

Cite this

Three-dimensional neuroblastoma cell culture : Proteomic analysis between monolayer and multicellular tumor spheroids. / Kumar, Hari R.; Zhong, Xiaoling; Hoelz, Derek J.; Rescorla, Frederick; Hickey, Robert J.; Malkas, Linda H.; Sandoval, John A.

In: Pediatric Surgery International, Vol. 24, No. 11, 11.2008, p. 1229-1234.

Research output: Contribution to journalArticle

Kumar, Hari R. ; Zhong, Xiaoling ; Hoelz, Derek J. ; Rescorla, Frederick ; Hickey, Robert J. ; Malkas, Linda H. ; Sandoval, John A. / Three-dimensional neuroblastoma cell culture : Proteomic analysis between monolayer and multicellular tumor spheroids. In: Pediatric Surgery International. 2008 ; Vol. 24, No. 11. pp. 1229-1234.
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T1 - Three-dimensional neuroblastoma cell culture

T2 - Proteomic analysis between monolayer and multicellular tumor spheroids

AU - Kumar, Hari R.

AU - Zhong, Xiaoling

AU - Hoelz, Derek J.

AU - Rescorla, Frederick

AU - Hickey, Robert J.

AU - Malkas, Linda H.

AU - Sandoval, John A.

PY - 2008/11

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N2 - Introduction: Solid tumors, such as neuroblastoma (NB), are associated with a heterogeneous cell environment. Multicellular tumor spheroid (MCTS) cultures have been shown to better mimic growth characteristics of in vivo solid tumors. Because tumor spheroid growth patterns may be quite different from standard two-dimensional culture systems, we sought to compare the protein expression profiles of two- and three-dimensional in vitro NB cultures, i.e., monolayers and MCTS. Materials and methods: Human NB cells were grown as both monolayers and spheres. Nuclear and cytosolic proteins were analyzed for differentially secreted proteins by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and selected polypeptides were identified by mass spectrometry (LC-MS/MS). Results: Several metabolic (transketolase, triosephosphate isomerase, pyruvate kinase M1/M2, alpha enolase, and phosphoglycerate mutase-1), cell stress response (heat shock proteins (HSP) 90, 70, and 60; antioxidant, thioredoxin), cell structure (septin 2, adenyl cyclase-associated protein-1), tubulin β-2 chain, actin, translationally controlled tumor protein and cofilin), signal transduction (peptidyl prolyl cis/trans isomerase A), biosynthetic (phosphoserine aminotransferase) and transport (cellular retinoic acid binding protein 1) polypeptides were overexpressed in spheroids. Several protein groups were differentially expressed between NB monolayers and spheroids. Conclusion: The altered proteins among NB spheroids may represent an important link between monolayer cell cultures and in vivo experiments and thus a more ideal in vitro culture system for determining the precise threedimensional microenvironment of NB.

AB - Introduction: Solid tumors, such as neuroblastoma (NB), are associated with a heterogeneous cell environment. Multicellular tumor spheroid (MCTS) cultures have been shown to better mimic growth characteristics of in vivo solid tumors. Because tumor spheroid growth patterns may be quite different from standard two-dimensional culture systems, we sought to compare the protein expression profiles of two- and three-dimensional in vitro NB cultures, i.e., monolayers and MCTS. Materials and methods: Human NB cells were grown as both monolayers and spheres. Nuclear and cytosolic proteins were analyzed for differentially secreted proteins by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and selected polypeptides were identified by mass spectrometry (LC-MS/MS). Results: Several metabolic (transketolase, triosephosphate isomerase, pyruvate kinase M1/M2, alpha enolase, and phosphoglycerate mutase-1), cell stress response (heat shock proteins (HSP) 90, 70, and 60; antioxidant, thioredoxin), cell structure (septin 2, adenyl cyclase-associated protein-1), tubulin β-2 chain, actin, translationally controlled tumor protein and cofilin), signal transduction (peptidyl prolyl cis/trans isomerase A), biosynthetic (phosphoserine aminotransferase) and transport (cellular retinoic acid binding protein 1) polypeptides were overexpressed in spheroids. Several protein groups were differentially expressed between NB monolayers and spheroids. Conclusion: The altered proteins among NB spheroids may represent an important link between monolayer cell cultures and in vivo experiments and thus a more ideal in vitro culture system for determining the precise threedimensional microenvironment of NB.

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KW - Multicellular spheroid

KW - Neuroblastoma

KW - Proteomics

KW - Tumor microenvironment

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