Analysis of Extracellular Matrix Formed by Hertwig's Epithelial Root Sheath Cells in vitro

2004 Fiscal Year Final Research Report Summary

• Project/Area Number: 15592191
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Periodontal dentistry
• Research Institution: The University of Tokushima
• Principal Investigator: OISHI Keiji Tokushima Univ., Hospital, Assistant Professor, 医学部・歯学部附属病院, 講師 (00253211)
• Co-Investigator(Kenkyū-buntansha): KIDO Jun-ichi The University of Tokushima, Institute of Health Biosciences, Dept of Periodontology and Endodontology, Associate Professor, 大学院・ヘルスバイオサイエンス研究部, 助教授 (10195315)
• Project Period (FY): 2003 – 2004
• Keywords: Hertwig's Epithelial Root Sheath / Extracellular Matrix Proteins / basement membrane

Research Abstract

Hertwig's epithelial root sheath (HERS) cells were thought to induce cementogenesis on the tooth root through direct or indirect mechanisms. To elucidate the mechanism, we investigated extracellular matrix formed by HERS cells in vitro.
In this study, we used clonal HERS cells we have established and primary culture of isolated HERS cells. The mRNA expression of some extracellular matrix proteins in the cells were analyzed by RT-PCR. Both cells expressed mRNAs of collagen type IV and laminin which were major components of basement membrane. In addition, mRNA of fibronectin, also one of components of basement membrane, was detected in isolated HERS cells. To check the differentiation stage of the cells, we examined mRNAs of TrkA and calbindin D28k, which were known to express in cells of epithelial rests of Malassez. Both mRNAs were not detected in the isolated HERS cells, suggesting that the cells had not differentiated into cells of epithelial rests in vitro.
HERS cells were cultured on culture plates coated with some extracellular matrix proteins. However, the mRNA expressions of laminin and collagen type IV were not affected by the coating.
Next, extracellular matrix formed by the cultured HERS cells were analyzed at the protein levels. Isolated HERS cells were cultured and their conditioned medium was collected for Western blot analysis. Laminin and collagen type IV were not detected in the culture medium. Then HERS cells were cultured on collagen membranes for histochemical analysis of extracellular matrix. However, the cells did not attach and grow actively on the membrane. The extracellular matrix formed by the cells could not be analyzed on this culture system.
These results suggest that cultured HERS cells maintain their phenotype in vitro and possibly form extracellular matrix. However, at the protein levels, the matrix formation was not confirmed in our culture system.