Fujian Key Laboratory of Oral Diseases, Fujian Medical University, Fujian, 350000, China.
School and Hospital of Stomatology, Fujian Medical University, Fujian, 350000, China.
BMC Oral Health. 2024 Mar 27;24(1):394. doi: 10.1186/s12903-024-04130-y.
The objective of this study was to assess the characterization of human acellular amniotic membrane (HAAM) using various decellularization methods and their impact on the proliferation and differentiation of human dental pulp stem cells (DPSCs). The goal was to identify scaffold materials that are better suited for pulp regeneration.
Six different decellularization methods were used to generate the amniotic membranes. The characteristics of these scaffolds were examined through hematoxylin and eosin (H&E) staining, scanning electron microscopy (SEM), and immunohistofluorescence staining (IHF). The DPSCs were isolated, cultured, and their capacity for multidirectional differentiation was verified. The third generation (P3) DPSCs, were then combined with HAAM to form the decellularized amniotic scaffold-dental pulp stem cell complex (HAAM-DPSCs complex). Subsequently, the osteogenic capacity of the HAAM-DPSCs complex was evaluated using CCK8 assay, live-dead cell staining, alizarin red and alkaline phosphatase staining, and real-time quantitative PCR (RT-PCR).
Out of the assessed decellularization methods, the freeze-thaw + DNase method and the use of ionic detergent (CHAPS) showed minimal changes in structure after decellularization, making it the most effective method. The HAAM-DPSCs complexes produced using this method demonstrated enhanced biological properties, as indicated by CCK8, alizarin red, alkaline phosphatase staining, and RT-PCR.
The HAAM prepared using the freeze-thaw + DNase method and CHAPS methods exhibited improved surface characteristics and significantly enhanced the proliferation and differentiation capacity of DPSCs when applied to them. The findings, therefore demonstrate the capacity for enhanced pulp regeneration therapy.
本研究旨在评估使用各种脱细胞方法对人羊膜进行表征及其对人牙髓干细胞（DPSCs）增殖和分化的影响。目的是确定更适合牙髓再生的支架材料。
使用六种不同的脱细胞方法生成羊膜。通过苏木精和伊红（H&E）染色、扫描电子显微镜（SEM）和免疫荧光染色（IHF）来检查这些支架的特性。分离、培养 DPSCs，并验证其多向分化能力。然后将第三代（P3）DPSCs 与 HAAM 结合形成脱细胞羊膜-牙髓干细胞复合物（HAAM-DPSCs 复合物）。随后，通过 CCK8 测定、死活细胞染色、茜素红和碱性磷酸酶染色以及实时定量 PCR（RT-PCR）评估 HAAM-DPSCs 复合物的成骨能力。
在所评估的脱细胞方法中，冻融+DNase 法和离子去污剂（CHAPS）法在脱细胞后结构变化最小，是最有效的方法。使用该方法制备的 HAAM-DPSCs 复合物表现出增强的生物学特性，CCK8、茜素红、碱性磷酸酶染色和 RT-PCR 均表明了这一点。
使用冻融+DNase 法和 CHAPS 方法制备的 HAAM 表现出改善的表面特性，并显著增强了 DPSCs 的增殖和分化能力，因此证明了增强牙髓再生治疗的能力。
