Isolation, Characterization and Osteogenic differentiation potential of periodontal ligament stem cells (in vitro study)

Document Type : Original Article

Authors

1 Lecturer of Oral biology , Faculty of Oral & Dental Medicine, Cairo University

2 Lecturer of Oral biology, Faculty of Dentistry, Ain Shams University

3 Professor of Medical Biochemistry and Molecular Biology, Department of Medical biochemistry and Molecular Biology, Faculty of medicine, Cairo University.

Abstract

Background: Stem cells are unspecialized and immature cells that have the potential to develop into many different cell lineages via differentiation in a suitable culture media. Periodontal ligament stem cells (PDLSCs), which reside within the perivascular space periodontium, possess characteristics of mesenchymal stem cells and are a promising tool for periodontium regeneration.
Purpose: This study aimed to isolate and characterize periodontal ligament stem cells (PDLSCs) from albino rat, Identification of surface markers expression utilizing flow cytometric analysis (FACS) and assessment of the osteogenic differentiation capability of these cells.
Material and methods: PDLSCs were collected from the root surfaces of mandibular 1st molars from 6 adult male albino rats. Collected samples were enzymatically treated and centrifuged to allow cells release. Cells were cultured for fourteen days. After reaching confluence, the isolated cells were characterized by flow cytometry using CD73, CD133 and C-Kit. The cells were induced for osteogenic differentiation by placing them in osteogenic culture media. Osteogenic differentiation was evaluated by Alizarin Red stain and by Molecular assessment for Bone sialoprotein (BSP) and alkaline phosphatase (ALP) genes expression by real time PCR (q RT-PCR).
Results: The results showed successful isolation of stem cells from periodontal ligament. PLSCs were identified as mesenchymal stem cells by expression of mesenchymal stem cell markers through FACS. PDLSCs showed positive expression to CD73 marker and negative expression to hematopoietic markers CD133 and C-kit. Our results showed successful differentiation of PDLSCs into osteogenic lineage. The formed mineralized matrix stained by alizarin red and was more advanced at 21 days after osteoblastic differentiation. Real Time PCR (q RT-PCR) for quantitative expression of BSP and ALP showed significant increase in the expression of BSP and ALP from day 14 to day 21 in differentiated PDLSCs.
Conclusion: According to the obtained results, we conclude that PDLSCs could be successively isolated from periodontal ligament tissues. Isolated PDLSCs possess an osteogenic potential and show differentiation into osteogenic lineage when cultured in suitable osteogenic medium.

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