Expression of growth-factor Receptors in Rat Socket coagulum-derived Cells

Abstract

The purpose of this study was to determine if periodontal ligament (PDL) fibroblasts are capable of expressing growth factor receptors during their differentiation. PDL cells, obtained from 2-day-old coagulum in the tooth socket of Sprague-Dawley rats after tooth extraction, were cultured in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum and antibiotics. Confluent cells were grown in the presence of 50 μg/ml ascorbic acid (AA), 10mM β-glycerophosphate (βGP), and 5 µM dexamethasone (Dex) for up to 3 weeks. Morphological changes in PDL cells during the 3-week cultures in the presence of AA, βGP, and Dex were investigated under a phase-contrast microscope. The messenger ribonucleic acid (mRNA) expression pattern of growth factors, including platelet-derived growth factor receptor-alpha and beta (PDGF-Rα and PDGFRβ), insulin-like growth factor type 1 and 2 receptors (IGF-1R and IGF-2R), and fibroblast growth factor receptor-4 (FGF-R4), were investigated in PDL cells, using Northern blot analysis. Based on the appearance of PDL cell cultures under the phase-contrast microscope, 4 morphologically distinct stages were identified: the confluent stage, the multilayer stage, the nodule stage and the mineralization stage. In the confluent stage (stage Ⅰ), PDL cells had a fibroblast-like or polygonal shape and formed a monolayer. On the 7(superscript th) day after treatment with AA, βGP, and Dex (stage Ⅱ), cell proliferation occurred in focal areas generating multilayered cells with a fibroblastic shape. After the 14(superscript th) day (stage Ⅲ) the multilayered cells formed round nodules by depositing a matrix between the cell layers. Heavy mineralization of the matrix was first seen on the 21st day after mineralized medium treatment and was characterized by the presence of refringent material covering the nodules, referred to as the mineralization stage (stage Ⅳ). By Northern blot analysis, it was determined that PDGF-Rα was weakly expressed in the confluent stage and then increased with the progression of PDL cell differentiation, while the level of PDGF-Rβ transcripts remained high throughout the 4 stages. IGF-1R gene activities were also high throughout the four stages. In contrast, expressions of IGF-2R and FGF-R4 mRNA were undetectable throughout differentiation. These results indicate that growth factor receptors are differentially expressed by PDL cells, which may be involved in the regulation of growth and differentiation of PDL cells.