The application of human periodontal ligament stem cells and biomimetic silk scaffold for in situ tendon regeneration

Abstract

Background With the development of tissue engineering, enhanced tendon regeneration could be achieved by exploiting suitable cell types and biomaterials. The accessibility, robust cell amplification ability, superior tendon differentiation potential, and immunomodulatory effects of human periodontal ligament stem cells (hPDLSCs) indicate their potential as ideal seed cells for tendon tissue engineering. Nevertheless, there are currently no reports of using PDLSCs as seed cells. Previous studies have confirmed the potential of silk scaffold for tendon tissue engineering. However, the biomimetic silk scaffold with tendon extracellular matrix (ECM)-like structure has not been systematically studied for in situ tendon regeneration. Therefore, this study aims to evaluate the effects of hPDLSCs and biomimetic silk scaffold on in situ tendon regeneration. Methods Human PDLSCs were isolated from extracted wisdom teeth. The differentiation potential of hPDLSCs towards osteo-, chondro-, and adipo-lineage was examined by cultured in different inducing media. Aligned and random silk scaffolds were fabricated by the controlled directional freezing technique. Scaffolds were characterized including surface structure, water contact angle, swelling ratio, degradation speed and mechanical properties. The biocompatibility of silk scaffolds was evaluated by live/dead staining, SEM observation, cell proliferation determination and immunofluorescent staining of deposited collagen type I. Subsequently, hPDLSCs were seeded on the aligned silk scaffold and transplanted into the ruptured rat Achilles tendon. Scaffolds without cells served as control groups. After 4 weeks, histology evaluation was carried out and macrophage polarization was examined to check the repair effects and immunomodulatory effects. Results Human PDLSCs were successfully isolated, and their multi-differentiation potential was confirmed. Compared with random scaffold, aligned silk scaffold had more elongated and aligned pores and promoted the proliferation and ordered arrangement of hPDLSCs. After implantation into rat Achilles tendon defect, hPDLSCs seeded aligned silk scaffold enhanced tendon repair with more tendon-like tissue formation after 4 weeks, as compared to the scaffold-only groups. Higher expression of CD206 and lower expression of iNOS, IL-1β and TNF-α were found in the hPDLSCs seeded aligned silk scaffold group, which revealed its modulation effect of macrophage polarization from M1 to M2 phenotype. Conclusions In summary, this study demonstrates the efficacy of hPDLSCs as seed cells and aligned silk scaffold as a tendon-mimetic scaffold for enhanced tendon tissue engineering, which may have broad implications for future tendon tissue engineering and regenerative medicine researches.

Advanced Nanofiber-Based Scaffolds for Achilles Tendon Regenerative Engineering

“…The MSCs are pluripotent non-hematopoietic adult stem cells generated from the germinal layer of mesoderm ( Pillai et al, 2017 ). The MSCs can be found in bone marrow, adipose, periosteum, skin, blood vessel walls, tendons, muscles, peripheral circulation, umbilical cord blood, periodontal ligament, and tooth tissue ( Wang et al, 2013 ; Chen J. et al, 2021 ; El Khatib et al, 2021 ). A range of active molecules (e.g., BMP-12/7 and growth factor) can promote further expansion and tendinogenic differentiation ( Tang and Lane, 2012 ; Zhou et al, 2019a ; Perucca Orfei et al, 2019 ).…”

“…Bioactive molecules are molecules or compounds (including growth factors, angiogenic factors, cytokines, siRNA, DNA, hormones, and others) that are synthesized and secreted by cells, as controllers of cell proliferation, differentiation, and matrix deposition and play an important role in the development and repair of AT tissue ( Raghavan et al, 2012 ; Ruiz-Alonso et al, 2021 ; Chen et al, 2022a ). Bioactive compounds, particularly recombinant growth factors, are effective, readily available, dose-controlled, and stable.…”

“…Different types, concentrations, or release gradients of the bioactive molecule are appropriate for the proliferation and differentiation of different types of cells ( Teh et al, 2015 ; Shiroud Heidari et al, 2021 ). The significant growth factors for tendon tissue engineering are IGF-1 ( Olesen et al, 2021 ), PDGF-BB ( Evrova et al, 2020 ), BMP-7 ( Zhang P. et al, 2020 ), TGF-β1 ( Rajpar and Barrett, 2019 ), VEGF ( Zhou Y. L. et al, 2019 ; Yuan et al, 2022 ), BMP-12 ( Rinoldi et al, 2019 ), SDF-1α ( Chen C. et al, 2020 ), bFGF ( Liu et al, 2013 ; Zhao et al, 2019 ), and TGF-β3 ( Chen et al, 2022a ). Raghavan et al (2012) discovered that IGF-1, bFGF, and PDGF-BB increased the proliferation of all cell types (hADSCs, fibroblasts, and tenocytes) in in vitro scaffold experiments.…”

“…AT reconstruction ( Sankar et al, 2017 ) employs natural polymers such as silk fibroin protein ( Yi et al, 2018 ), collagen ( Sandri et al, 2016 ), chitosan ( Chen et al, 2018 ), sodium alginate, and gelatin, as well as degradable synthetic polymers such as PCL ( Fotticchia et al, 2018 ), PLA ( Wu et al, 2020b ), PLGA ( Hasmad et al, 2018 ), PET ( Cai et al, 2018a ), and PEO ( Table1 ). Different materials used as scaffolds for tendon tissue engineering have their advantages and disadvantages ( Chen et al, 2021a ). The advantage of synthetic polymer scaffold polymers is that their sizes may be easily modified to meet requirements and they degrade slowly.…”

“…The advantage of synthetic polymer scaffold polymers is that their sizes may be easily modified to meet requirements and they degrade slowly. In general, the use of synthetic polymer scaffolds constrained by the adverse effects of their degradation products in vivo (such as inflammatory reactions) and their relatively poor integration with host tissues ( Chen et al, 2021a ). PGA, PLA, and PLGA are considered biocompatible and cause only mild or minor foreign body reactions because their hydrolysis degradation product (glycolate lactate) is typically present in the metabolic pathway of the human body ( Bianchi et al, 2021 ).…”