Effect of human periodontal ligament-derived mesenchymal stem cells with high and low osteogenic potential on bone repair of defects created in rat calvaria

Author(s)

Letícia Faustino Adolpho

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Odontologia de Ribeirão Preto (PCARP/BC)
• Defense date: 2021-07-20

Examining board members

Márcio Mateus Beloti; Denise Carleto Andia; Paula Gabriela Faciola Pessôa de Oliveira

• Advisor: Márcio Mateus Beloti

Abstract

Therapy with mesenchymal stem cells (MSCs) is one of the strategies used in regenerative medicine that can be applied to regenerate bone tissue. MSCs can be isolated from several sites of the body, such as bone marrow, adipose tissue, umbilical cord, muscle, and periodontal ligament. Our research group has evaluated strategies to promote bone regeneration in a model of defects created in rat calvaria and, in recent studies, we have shown that locally injected MSCs are more effective compared to the use of biomaterial itself or in combination with cells. The periodontal ligament harvest from extracted teeth is a simple procedure and this tissue is a source of periodontal ligament stem cells (PDLSCs). In vitro and in vivo studies have shown that these cells are capable of osteoblastic differentiation and to form mineralized matrix. In addition, PDLSCs may have different osteogenic potentials. In this context, the aim of this study was to evaluate the capacity of PDLSCs with high (AP-PDLSCs) and low osteogenic potential (BP-PDLSCs), as well as both mixed cell populations (AP-PDLSCs+BP-PDLSCs), to repair the bone tissue after locally injected in defects created in rat calvaria. The cells were obtained from the periodontal ligament of extracted third molars, in vitro characterized in terms of their capacity for osteoblastic proliferation and differentiation, and sorted as AP-PDLSCs or BP-PDLSCs. Then, unilateral defects, 5 mm in diameter, were created in calvaria of Sprague-Dawley rats and 2 weeks after the creation of the defects, 5x106 APPDLSCs, BP-PDLSCs, or AP-PDLSCs + BP-PDLSCs (1:1) were directly injected into bone defects. Defects treated with vehicle (PBS) were used as control. Four weeks post-injection, bone formation was evaluated by computed microtomography, histological analysis, and gene expression of Runx2, Sp7, Alp, Bglap, Ibsp, Opg, Rankl, Rank, Cd31, Vegfa, and Vegfr2 by real-time PCR. The results demonstrated that PDLSCs can adhere to plastic and a typical MSC phenotype, evidenced by the expression of mesenchymal markers and the absence of hematopoietic markers on cell surface. Furthermore, AP-PDLSCs and BP-PDLSCs exhibited distinct in vitro potential to differentiate into osteoblasts, demonstrated by the higher in situ ALP activity of the AP-PDLSCs. The formation of a mineralized matrix was, however, similar in both cell populations. In vivo, the ability to induce repair of bone defects was similar for AP-PDLSCs, BP-PDLSCs, and AP-PDLSCs + BP-PDLSCs and in all cases the bone formation was greater compared to defects treated with PBS. Neoformed tissue in defects treated with cells also exhibited the same histological features. Gene expression in newly formed tissues indicated the occurrence of bone remodeling in defects treated with cells, especially in that ones treated with AP-PDLSCs. Taken together, the data demonstrated that, at the evaluated period, regardless of the in vitro osteogenic potential, PDLSCs induced the similar bone formation in defects created in rat calvaria and therefore they can be considered to the development of cell therapies to treat bone defects. (AU)

• FAPESP's process: 19/10076-0 - Effect of mesenchymal stem cells from periodontal ligament with high and low osteogenic potential on bone repair
• Grantee: Letícia Faustino Adolpho
• Support Opportunities: Scholarships in Brazil - Master