Zeng Jingjie, Deng Huidan, Li Quanjie, Kang Jingyi, Wu Yu
Guangxi Medical University, Nanning, China; Guangxi Key Laboratory of Oral and Maxillofacial Surgery Disease Treatment, Nanning, China; Guangxi Clinical Research Center for Craniofacial Deformity, Nanning, China.
Department of Pediatric Dentistry, College ＆ Hospital of Stomatology, Guangxi Medical University, No. 10 Shuangyong Road, Nanning 530021, China.
Tissue Cell. 2024 Dec;91:102549. doi: 10.1016/j.tice.2024.102549. Epub 2024 Aug 30.
The formation of dentin-pulp complex determines the success of vital pulp therapy. Human umbilical cord mesenchymal stem cell-derived small extracellular vesicles (hUCMSC-sEVs) appeared to have stronger effect in anti-inflammatory and promoting the proliferation and migration of human dental pulp stem cells (hDPSCs). Moreover, Lipopolysaccharides (LPS) pretreatment can enhance the rapeutic potency of extracellular vesicles. LPS pretreatment hUCMSC-sEVs have the potential to regenerate the dentin-pulp complex by recruiting hDPSCs. This paper aims to develop collagen sponge/self-assembling peptide nanofiber scaffold (CS/SAPNS) composite scaffold loaded with LPS pretreatment hUCMSC-sEVs (CS/SAPNS-sEVs), and assess the release characteristics of hUCMSC-sEVs and the effect of this composite scaffold on osteo/odontogenic differentiation and angiogenic potential in hDPSCs.
LPS pretreatment hUCMSC-sEVs (LPS-hUCMSC-sEVs) were mixed with self-assembling peptide hydrogel and loaded onto collagen sponge to obtain the CS/SAPNS-sEVs. BCA assay, nanoparticle analysis, transmission electron microscopy and laser confocal microscopy were used to investigate the characteristics of LPS-hUCMSC-sEVs loaded on CS/SAPNS. Osteo/odontogenic differentiation ability of hDPSCs were analyzed by ALP stainning, alizarin red staining. RT-PCR and Western blot analysis were performed to confirm the levels of osteo/odontogenic factors and angiogenic factors, and the involvement of NF-κB pathway was verified by immunocytochemical staining and Western blot analysis.
CS/SAPNS could control LPS-hUCMSC-sEVs release for 7 days and keep their structural integrity. CS/SAPNS-sEVs promoted deposition of calcified nodules and expression of osteogenic/odontogenic and angiogenic factors in hDPSCs. On the contrary, inhibition of the NF-κB pathway down-regulated the expression of CS/SAPNS-sEVs-regulated osteo/odontogenic and angiogenic factors.
CS/SNAPS could be used as scaffold for LPS-hUCMSC-sEVs, and CS/SAPNS-sEVs may promote osteo/odontogenic differentiation and enhance the angiogenic potential of hDPSCs through activating the NF-κB pathway.
牙本质-牙髓复合体的形成决定了牙髓活力治疗的成功与否。人脐带间充质干细胞来源的小细胞外囊泡（hUCMSC-sEVs）在抗炎以及促进人牙髓干细胞（hDPSCs）增殖和迁移方面似乎具有更强的作用。此外，脂多糖（LPS）预处理可增强细胞外囊泡的治疗效力。经LPS预处理的hUCMSC-sEVs有通过募集hDPSCs使牙本质-牙髓复合体再生的潜力。本文旨在研发负载经LPS预处理的hUCMSC-sEVs（CS/SAPNS-sEVs）的胶原海绵/自组装肽纳米纤维支架（CS/SAPNS）复合支架，并评估hUCMSC-sEVs的释放特性以及该复合支架对hDPSCs成骨/成牙分化及血管生成潜力的影响。
将经LPS预处理的hUCMSC-sEVs（LPS-hUCMSC-sEVs）与自组装肽水凝胶混合，负载于胶原海绵上，从而获得CS/SAPNS-sEVs。采用BCA法、纳米颗粒分析、透射电子显微镜和激光共聚焦显微镜研究负载于CS/SAPNS上的LPS-hUCMSC-sEVs的特性。通过碱性磷酸酶染色、茜素红染色分析hDPSCs的成骨/成牙分化能力。进行逆转录-聚合酶链反应（RT-PCR）和蛋白质免疫印迹分析以确认成骨/成牙因子和血管生成因子的水平，并通过免疫细胞化学染色和蛋白质免疫印迹分析验证核因子-κB（NF-κB）通路的参与情况。
CS/SAPNS能够控制LPS-hUCMSC-sEVs释放7天，并保持其结构完整性。CS/SAPNS-sEVs促进了hDPSCs中钙化结节的沉积以及成骨/成牙和血管生成因子的表达。相反，抑制NF-κB通路会下调CS/SAPNS-sEVs调节的成骨/成牙和血管生成因子的表达。
CS/SNAPS可用作LPS-hUCMSC-sEVs的支架，且CS/SAPNS-sEVs可能通过激活NF-κB通路促进hDPSCs的成骨/成牙分化并增强其血管生成潜力。
