An in vitro three-dimensional co-culture system for ameloblastoma modelling

Soo Leng Lee, Zainal Ariff Abdul Rahman, Hidetsugu Tsujigiwa, Mei Hamada, Kiyofumi Takabatake, Keisuke Nakano, Hitoshi Nagatsuka, Chong Huat Siar

Research output: Contribution to journalArticle

Abstract

Ameloblastoma, the most clinically significant odontogenic epithelial tumor, is a locally-invasive and destructive lesion in the jawbones. However, the nature of this infiltrativeness and destructive behavior remains ill-understood. To address this, we established an in vitro three-dimensional (3D) co-culture system to simulate an amelobastoma disease model aimed at investigating the interactions between tumor cells and osteoblasts. Osteoblastic cell lines (KUSA/A1 and MC3T3-E1) and one stromal cell line (ST2) were separately co-seeded with ameloblastoma-derived cell line (AM-1) in a collagen scaffold (representing the extracellular bone matrix) and incubated with mineralization medium. Immunohistochemistry, double immunofluorescence and mineralization assay were performed. Only AM-1/KUSA-A1 co-culture showed a significant increase in AM-1 cell count, suggesting that heterotypic cell-cell interaction promotes tumoral cell growth, while formation of visible AM-1 epithelial nest-like structures resembling ameloblastoma cells in their native state, suggest morphodifferentiation. A RANK-high, RANKL-low and osteoprotegerin-low immunoprofile in co-culture AM-1 cells implies deregulated osteoclastogenesis. Mineralization assays showed diminished calcification in AM-1/KUSA-A1 co-culture extracellular matrix suggesting an altered local bone metabolism. In contrast, KUSA/A1 monocultures showed abundant extracellular matrix calcification. Taken together, these results suggest that a 3D co-culture system as an amelobastoma disease model provides insights that bidirectional ameloblastoma-osteoblastic interactions might play a role in modulating tumor growth and osteoclastogenesis.

Original languageEnglish
Pages (from-to)1697-1706
Number of pages10
JournalSains Malaysiana
Volume48
Issue number8
DOIs
Publication statusPublished - Jan 1 2019

Fingerprint

Ameloblastoma
Coculture Techniques
Extracellular Matrix
Osteogenesis
Cell Line
Odontogenic Tumors
Osteoprotegerin
Bone Matrix
Stromal Cells
Growth
Osteoblasts
Cell Communication
Fluorescent Antibody Technique
Neoplasms
Collagen
Cell Count
Immunohistochemistry
In Vitro Techniques
Bone and Bones

Keywords

  • Ameloblast
  • Ameloblastoma modelling
  • Co-culture system
  • Pre-osteoblast

ASJC Scopus subject areas

  • General

Cite this

An in vitro three-dimensional co-culture system for ameloblastoma modelling. / Lee, Soo Leng; Rahman, Zainal Ariff Abdul; Tsujigiwa, Hidetsugu; Hamada, Mei; Takabatake, Kiyofumi; Nakano, Keisuke; Nagatsuka, Hitoshi; Siar, Chong Huat.

In: Sains Malaysiana, Vol. 48, No. 8, 01.01.2019, p. 1697-1706.

Research output: Contribution to journalArticle

Lee, Soo Leng ; Rahman, Zainal Ariff Abdul ; Tsujigiwa, Hidetsugu ; Hamada, Mei ; Takabatake, Kiyofumi ; Nakano, Keisuke ; Nagatsuka, Hitoshi ; Siar, Chong Huat. / An in vitro three-dimensional co-culture system for ameloblastoma modelling. In: Sains Malaysiana. 2019 ; Vol. 48, No. 8. pp. 1697-1706.
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AB - Ameloblastoma, the most clinically significant odontogenic epithelial tumor, is a locally-invasive and destructive lesion in the jawbones. However, the nature of this infiltrativeness and destructive behavior remains ill-understood. To address this, we established an in vitro three-dimensional (3D) co-culture system to simulate an amelobastoma disease model aimed at investigating the interactions between tumor cells and osteoblasts. Osteoblastic cell lines (KUSA/A1 and MC3T3-E1) and one stromal cell line (ST2) were separately co-seeded with ameloblastoma-derived cell line (AM-1) in a collagen scaffold (representing the extracellular bone matrix) and incubated with mineralization medium. Immunohistochemistry, double immunofluorescence and mineralization assay were performed. Only AM-1/KUSA-A1 co-culture showed a significant increase in AM-1 cell count, suggesting that heterotypic cell-cell interaction promotes tumoral cell growth, while formation of visible AM-1 epithelial nest-like structures resembling ameloblastoma cells in their native state, suggest morphodifferentiation. A RANK-high, RANKL-low and osteoprotegerin-low immunoprofile in co-culture AM-1 cells implies deregulated osteoclastogenesis. Mineralization assays showed diminished calcification in AM-1/KUSA-A1 co-culture extracellular matrix suggesting an altered local bone metabolism. In contrast, KUSA/A1 monocultures showed abundant extracellular matrix calcification. Taken together, these results suggest that a 3D co-culture system as an amelobastoma disease model provides insights that bidirectional ameloblastoma-osteoblastic interactions might play a role in modulating tumor growth and osteoclastogenesis.

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