Department of Dentistry, Charles University, Faculty of Medicine in Hradec Kralove and University Hospital Hradec Kralove, Sokolska 581, 50005 Hradec Kralove, Czech Republic.
Department of Histology and Embryology, Faculty of Medicine in Hradec Kralove, Charles University, Simkova 870, 50003 Hradec Kralove, Czech Republic.
Biomolecules. 2022 Apr 20;12(5):610. doi: 10.3390/biom12050610.
The physical stresses during cryopreservation affect stem cell survival and further proliferation. To minimize or prevent cryoinjury, cryoprotective agents (CPAs) are indispensable. Despite the widespread use of 10% dimethyl sulfoxide (DMSO), there are concerns about its potential adverse effects. To bypass those effects, combinations of CPAs have been investigated. This study aimed to verify whether high-molecular-hyaluronic acid (HMW-HA) serves as a cryoprotectant when preserving human mesenchymal stem cells (hMSCs) to reduce the DMSO concentration in the cryopreservation medium. We studied how 0.1% or 0.2% HMW-HA combined with reduced DMSO concentrations (from 10% to 5%, and 3%) affected total cell count, viability, immunophenotype, and differentiation potential post-cryopreservation. Immediately after cell revival, the highest total cell count was observed in 10% DMSO-stored hMSC. However, two weeks after cell cultivation an increased cell count was seen in the HMW-HA-stored groups along with a continued increase in hMSCs stored using 3% DMSO and 0.1% HMW-HA. The increased total cell count corresponded to elevated expression of stemness marker CD49f. The HA-supplemented cryomedium did not affect the differential potential of hMSC. Our results will participate in producing a ready-to-use product for cryopreservation of mesenchymal stem cells.
冷冻保存过程中的物理应激会影响干细胞的存活和进一步增殖。为了最小化或预防冷冻损伤，需要使用冷冻保护剂（CPAs）。尽管 10%二甲基亚砜（DMSO）被广泛应用，但人们对其潜在的不良反应仍存在担忧。为了避免这些影响，人们研究了 CPAs 的组合。本研究旨在验证高分子量透明质酸（HMW-HA）是否可以作为人骨髓间充质干细胞（hMSC）冷冻保存的保护剂，以降低冷冻保存培养基中 DMSO 的浓度。我们研究了 0.1%或 0.2%的 HMW-HA 与降低的 DMSO 浓度（从 10%降至 5%和 3%）结合使用对总细胞计数、活力、免疫表型和冷冻保存后分化潜能的影响。细胞复苏后立即，在储存于 10%DMSO 的 hMSC 中观察到最高的总细胞计数。然而，细胞培养两周后，在 HMW-HA 储存组中观察到细胞计数增加，同时使用 3%DMSO 和 0.1%HMW-HA 储存的 hMSCs 持续增加。总细胞计数的增加与干细胞标志物 CD49f 的表达增加相对应。添加 HA 的冷冻培养基不影响 hMSC 的分化潜能。我们的研究结果将有助于开发用于冷冻保存间充质干细胞的即用型产品。
