Liang Qingqing, Liang Cheng, Liu Xiaojing, Xing Xiaotao, Ma Shixing, Huang Haisen, Liang Chao, Liu Lei, Liao Li, Tian Weidong
State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, No. 14, 3rd Section, Renmin South Road, Chengdu, Sichuan 610041, China.
National Engineering Laboratory for Oral Regenerative Medicine & Engineering Research Center of Oral Translational Medicine, Ministry of Education, West China Hospital of Stomatology, Sichuan University, Chengdu, Sichuan 610041, China.
J Mater Chem B. 2022 Dec 14;10(48):10097-10111. doi: 10.1039/d2tb01825j.
Regeneration of dental pulp the transplantation of dental pulp stem cells (DPSCs) has emerged as a novel therapy for dental pulp necrosis after inflammation and injury. However, providing sufficient oxygen and nutrients to support stem cell survival, self-renewal, and differentiation in the narrow root canal remains a great challenge. In this study, we explored a novel strategy based on cell-laden microfibers for dental pulp regeneration. Firstly, we fabricated suitable GelMA hydrogels that facilitate the survival and proliferation of DPSCs and human umbilical vein endothelial cells (HUVECs) and possess satisfactory biomechanical properties to generate microfibers. Two kinds of GelMA microfibers were fabricated with DPSCs and HUVECs a silicone-tube-based coagulant bath-free method. Live/dead and Ki-67 immunofluorescence staining assays identified that these two cell lines maintained high survival rate and proliferation ability in GelMA microfibers. Immunofluorescence staining confirmed that DPSCs fully spread in the microfibers and highly expressed CD90 and laminin. HUVECs positively express CD31 and VE-cad in microfibers and could migrate well in the GelMA hydrogel. permeation experiments confirmed the superiority of microfiber aggregates (MAs) in liquid permeation compared to GelMA hydrogel blocks. We further adopted an ectopic pulp regeneration assay in nude mice to validate the regeneration of the aggregates of mixed DPSC-microfibers and HUVEC-microfibers . Compared to a conventional mixture of DPSCs and HUVECs in GelMA hydrogel blocks, the aggregates of cell-laden microfibers generated more pulp-like tissue, blood vessels, and odontoblast-like cells that positively express DMP-1 and DSPP. To our knowledge, this is the first attempt to apply cell-laden MAs for pulp regeneration. Our study proposes a new solution to the challenge of pulp regeneration, which might promote the clinical translation and application of stem cell-based therapy.
牙髓再生——牙髓干细胞（DPSCs）移植已成为治疗炎症和损伤后牙髓坏死的一种新疗法。然而，在狭窄的根管中提供足够的氧气和营养以支持干细胞的存活、自我更新和分化仍然是一个巨大的挑战。在本研究中，我们探索了一种基于载细胞微纤维的牙髓再生新策略。首先，我们制备了合适的甲基丙烯酰化明胶（GelMA）水凝胶，其有助于DPSCs和人脐静脉内皮细胞（HUVECs）的存活和增殖，并具有令人满意的生物力学性能以生成微纤维。采用基于硅胶管的无凝固浴方法制备了两种载有DPSCs和HUVECs的GelMA微纤维。活/死和Ki-67免疫荧光染色分析表明，这两种细胞系在GelMA微纤维中保持高存活率和增殖能力。免疫荧光染色证实，DPSCs在微纤维中充分铺展并高表达CD90和层粘连蛋白。HUVECs在微纤维中阳性表达CD31和血管内皮钙黏蛋白（VE-cad），并且可以在GelMA水凝胶中良好迁移。渗透实验证实了微纤维聚集体（MAs）在液体渗透方面优于GelMA水凝胶块。我们进一步在裸鼠中采用异位牙髓再生试验来验证混合DPSC-微纤维和HUVEC-微纤维聚集体的再生情况。与传统的DPSCs和HUVECs在GelMA水凝胶块中的混合物相比，载细胞微纤维聚集体产生了更多的牙髓样组织、血管和成牙本质细胞样细胞，这些细胞阳性表达牙本质基质蛋白-1（DMP-1）和牙本质涎磷蛋白（DSPP）。据我们所知，这是首次尝试将载细胞MAs应用于牙髓再生。我们的研究为牙髓再生的挑战提出了一种新的解决方案，这可能会促进基于干细胞治疗的临床转化和应用。
