Human deciduous pulp stem cells (hDPSCs) have remarkable stem cell potency associated with cell proliferation, mesenchymal multipotency, and immunosuppressive function and have shown beneficial effects in a variety of animal disease models. Recent studies demonstrated that hDPSCs exhibited in vivo anti-fibrotic and anti-inflammatory action and in vivo hepatogenic-associated liver regeneration, suggesting that hDPSCs may offer a promising source with great clinical demand for treating liver diseases. However, how to manufacture ex vivo large-scale clinical-grade hDPSCs with the appropriate quality, safety, and preclinical efficacy assurances remains unclear. We isolated hDPSCs from human deciduous dental pulp tissues formed by the colony-forming unit-fibroblast (CFU-F) method and expanded them under a xenogeneic-free and serum-free (XF/SF) condition; hDPSC products were subsequently stored by two-step banking including a master cell bank (MCB) and a working cell bank (WCB). The final products were directly thawed hDPSCs from the WCB. We tested the safety and quality check, stem cell properties, and preclinical potentials of final hDPSC products and hDPSC products in the MCB and WCB. We optimized manufacturing procedures to isolate and expand hDPSC products under a XF/SF culture condition and established the MCB and the WCB. The final hDPSC products and hDPSC products in the MCB and WCB were validated the safety and quality including population doubling ability, chromosome stability, microorganism safety, and stem cell properties including morphology, cell surface marker expression, and multipotency. We also evaluated the in vivo immunogenicity and tumorigenicity and validated in vivo therapeutic efficacy for liver regeneration in a CCl4-induced chronic liver fibrosis mouse model in the final hDPSC products and hDPSC products in the WCB. The manufacture and quality control results indicated that the present procedure could produce sufficient numbers of clinical-grade hDPSC products from a tiny deciduous dental pulp tissue to enhance clinical application of hDPSC products in chronic liver fibrosis.

中文翻译：

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用于慢性肝纤维化治疗的临床级乳牙牙髓干细胞的制造和验证的模型研究。

人乳髓果干细胞（hDPSC）具有与细胞增殖，间质多能性和免疫抑制功能相关的显着干细胞潜能，并已在多种动物疾病模型中显示出有益的作用。最近的研究表明，hDPSCs表现出体内抗纤维化和抗炎作用以及体内与肝相关的肝再生，这表明hDPSCs可能为治疗肝脏疾病提供巨大的临床需求。但是，如何制造具有适当质量，安全性和临床前疗效保证的离体大规模临床级hDPSCs尚不清楚。我们从通过集落形成单位成纤维细胞（CFU-F）方法形成的人乳牙牙髓组织中分离了hDPSC，并在无异种和无血清（XF / SF）的条件下扩增了hDPSC；hDPSC产品随后通过两步存储（包括主单元存储库（MCB）和工作单元存储库（WCB））进行存储。最终产品是直接从WCB解冻的hDPSC。我们测试了MCB和WCB中最终hDPSC产品和hDPSC产品的安全性和质量检查，干细胞特性以及临床前潜力。我们优化了制造程序，以在XF / SF培养条件下分离和扩展hDPSC产品，并建立了MCB和WCB。验证了MCB和WCB中最终的hDPSC产品和hDPSC产品的安全性和质量，包括种群倍增能力，染色体稳定性，微生物安全性以及干细胞特性，包括形态，细胞表面标志物表达和多能性。我们还评估了在最终hDPSC产品和WCB中hDPSC产品中CCl4诱导的慢性肝纤维化小鼠模型中肝脏再生的体内免疫原性和致瘤性，并验证了体内治疗功效。制造和质量控制结果表明，本程序可从微小的乳牙牙髓组织中产生足够数量的临床级hDPSC产品，以增强hDPSC产品在慢性肝纤维化中的临床应用。我们还评估了在最终hDPSC产品和WCB中hDPSC产品中CCl4诱导的慢性肝纤维化小鼠模型中肝脏再生的体内免疫原性和致瘤性，并验证了体内治疗功效。制造和质量控制结果表明，本程序可从微小的乳牙牙髓组织中产生足够数量的临床级hDPSC产品，以增强hDPSC产品在慢性肝纤维化中的临床应用。我们还评估了在最终hDPSC产品和WCB中hDPSC产品中CCl4诱导的慢性肝纤维化小鼠模型中肝脏再生的体内免疫原性和致瘤性，并验证了体内治疗功效。制造和质量控制结果表明，本程序可从微小的乳牙牙髓组织中产生足够数量的临床级hDPSC产品，以增强hDPSC产品在慢性肝纤维化中的临床应用。