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

老鼠齒糟窩衍生細胞生長因子感受器的表現

摘要


本實驗的目的在探討老鼠的牙周韌帶細胞在進行分化期間,對生長因子感受器表現的情形。牙周韌帶細胞取自老鼠拔牙後第二天傷口內的凝塊,將細胞培養在含有10%牛胎兒血清及抗生素的DMEM培養液中,等細胞融合後給予維生素C、β甘油磷酸鹽和皮質脂酮治療三週。我們使用像位差顯微鏡來觀察牙周韌帶細胞在進行分化期間形態學上的變化,和使用Northern blot分析來檢測PDGF、IGF及FGF等生長因子感受器信使RNA(mRNA)的表現。分化期間的牙周韌帶細胞在形態學上有四個階段的變化,第一期為細胞融合期,呈現多角形的單層細胞;第二期為多層細胞期,細胞在局部區域增生並形成具有成纖維細胞形狀的多層細胞;第三期為結節期,基質沉積使多層細胞形成圓形結節;第四期為礦質化期,具有屈光的物質覆蓋在結節上並形成礦質化。PDGF感受器α型的表現剛開始時非常弱,但隨著牙周韌帶細胞分化的進行而增強;然而PDGF感受器β型與IGF-1感受器的表現則在四個階段都維持在差不多的強度,至於IGF-2與FGF-4感受器在這四個階段都不表現。生長因子感受器在牙周韌帶細胞進行分化時的表現都不一致,這些感受器的表現可能與調節牙周韌帶細胞的生長與分化有關。

並列摘要


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.

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