School of Medicine, Southeast University, Nanjing, 210009, China.
Jiangsu Key Laboratory for Biomaterials and Devices, Southeast University, Nanjing, 210096, China.
Stem Cell Res Ther. 2021 Dec 4;12(1):596. doi: 10.1186/s13287-021-02661-7.
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.
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.
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.
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.
随着组织工程学的发展，通过利用合适的细胞类型和生物材料，可以实现增强的肌腱再生。人牙周膜干细胞（hPDLSCs）的易得性、强大的细胞扩增能力、优异的肌腱分化潜力和免疫调节作用，表明其具有成为理想的肌腱组织工程种子细胞的潜力。然而，目前尚无使用 PDLSCs 作为种子细胞的报道。先前的研究已经证实了丝素支架在肌腱组织工程中的潜力。然而，具有肌腱细胞外基质（ECM）样结构的仿生丝素支架尚未被系统地研究用于原位肌腱再生。因此，本研究旨在评估 hPDLSCs 和仿生丝素支架对原位肌腱再生的影响。
从拔出的智齿中分离出人牙周膜干细胞。通过在不同诱导培养基中培养来检测 hPDLSCs 向成骨、软骨和成脂谱系的分化潜能。通过受控定向冷冻技术制备了定向和随机丝素支架。对支架的表面结构、水接触角、溶胀比、降解速度和机械性能进行了表征。通过活/死染色、SEM 观察、细胞增殖测定和沉积的 I 型胶原的免疫荧光染色评估丝素支架的生物相容性。随后，将 hPDLSCs 接种到定向丝素支架上，并移植到大鼠跟腱断裂处。无细胞的支架作为对照组。4 周后，进行组织学评估，并检查巨噬细胞极化，以检查修复效果和免疫调节作用。
成功分离出人牙周膜干细胞，并证实其多向分化潜能。与随机支架相比，定向丝素支架具有更细长和定向的孔，并促进 hPDLSCs 的增殖和有序排列。植入大鼠跟腱缺损后，与支架仅组相比，接种定向丝素支架的 hPDLSCs 增强了肌腱修复，形成更多类似肌腱的组织。在 hPDLSCs 接种定向丝素支架组中发现 CD206 表达较高，而 iNOS、IL-1β 和 TNF-α 表达较低，表明其对巨噬细胞极化从 M1 表型向 M2 表型的调节作用。
总之，本研究证明了 hPDLSCs 作为种子细胞和定向丝素支架作为肌腱模拟支架用于增强肌腱组织工程的功效，这可能对未来的肌腱组织工程和再生医学研究具有广泛的意义。
