Bárcia Rita N, Santos Jorge M, Teixeira Mariana, Filipe Mariana, Pereira Ana Rita S, Ministro Augusto, Água-Doce Ana, Carvalheiro Manuela, Gaspar Maria Manuela, Miranda Joana P, Graça Luis, Simões Sandra, Santos Susana Constantino Rosa, Cruz Pedro, Cruz Helder
ECBio, Investigação e Desenvolvimento em Biotecnologia, S.A., Amadora, Portugal.
Centro Cardiovascular da Universidade de Lisboa, Lisboa, Portugal.
Cytotherapy. 2017 Mar;19(3):360-370. doi: 10.1016/j.jcyt.2016.11.008. Epub 2016 Dec 28.
The effect of cryopreservation on mesenchymal stromal cell (MSC) therapeutic properties has become highly controversial. However, data thus far have indiscriminately involved the assessment of different types of MSCs with distinct production processes. This study assumed that MSC-based products are affected differently depending on the tissue source and manufacturing process and analyzed the effect of cryopreservation on a specific population of umbilical cord tissue-derived MSCs (UC-MSCs), UCX.
Cell phenotype was assessed by flow cytometry through the evaluation of the expression of relevant surface markers such as CD14, CD19, CD31, CD34, CD44, CD45, CD90, CD105, CD146, CD200, CD273, CD274 and HLA-DR. Immunomodulatory activity was analyzed in vitro through the ability to inhibit activated T cells and in vivo by the ability to reverse the signs of inflammation in an adjuvant-induced arthritis (AIA) model. Angiogenic potential was evaluated in vitro using a human umbilical vein endothelial cell-based angiogenesis assay, and in vivo using a mouse model for hindlimb ischemia.
Phenotype and immunomodulatory and angiogenic potencies of this specific UC-MSC population were not impaired by cryopreservation and subsequent thawing, both in vitro and in vivo.
This study suggests that potency impairment related to cryopreservation in a given tissue source can be avoided by the production process. The results have positive implications for the development of advanced-therapy medicinal products.
冷冻保存对间充质基质细胞(MSC)治疗特性的影响极具争议。然而,迄今为止的数据不加区分地涉及了对不同生产过程产生的不同类型MSC的评估。本研究假定基于MSC的产品会因组织来源和制造工艺的不同而受到不同影响,并分析了冷冻保存对特定脐带组织来源的MSC群体(UC-MSC),即UCX的影响。
通过流式细胞术评估细胞表型,评估相关表面标志物如CD14、CD19、CD31、CD34、CD44、CD45、CD90、CD105、CD146、CD200、CD273、CD274和HLA-DR的表达。通过抑制活化T细胞的能力在体外分析免疫调节活性,并在佐剂诱导的关节炎(AIA)模型中通过逆转炎症体征的能力在体内分析免疫调节活性。使用基于人脐静脉内皮细胞的血管生成试验在体外评估血管生成潜力,并在小鼠后肢缺血模型中在体内评估血管生成潜力。
无论是在体外还是体内,这种特定的脐带间充质干细胞群体的表型、免疫调节和血管生成能力都不会因冷冻保存和随后的解冻而受损。
本研究表明,通过生产过程可以避免特定组织来源中与冷冻保存相关的效力受损。这些结果对先进治疗药物产品的开发具有积极意义。