Zhou Hongfeng, Song Mengyao, Zhang Xiaohu, Ke Tao, Shi Guosheng, Wu Yaojiong, Geng Hongya
Institute of Biopharmaceutical and Health Engineering, Key Laboratory of Active Proteins and Peptides Green Biomanufacturing of Guangdong Higher Education Institutes, Shenzhen International Graduate School, Tsinghua University, Shenzhen, Guangdong 518055, China.
Shanghai Applied Radiation Institute, State Key Lab. Advanced Special Steel, Shanghai University, Shanghai 200444, China.
Langmuir. 2025 Apr 15;41(14):9532-9541. doi: 10.1021/acs.langmuir.5c00569. Epub 2025 Mar 31.
Cryopreservation is fundamental to cell-based therapeutics but is severely limited by ice formation and growth, which causes irreversible membrane rupture, osmotic imbalance, and chilling injury. Antifreeze proteins and conventional cryoprotectants like gold standard dimethyl sulfoxide (DMSO) struggle to offer a cost-effective and biocompatible strategy fitting various cellular storage. Herein, we design and fabricate a class of oligopeptides exhibiting potent ice recrystallization inhibition (IRI) activity, achieving a 55.5% reduction in the mean largest grain size at a concentration of less than 0.1 wt %. The side-chain functional groups (e.g., hydroxyl and amine groups) and length (<10 amino acids) are meticulously optimized to avoid the thermal hysteresis (TH) activity that causes sudden burst of fatal needle-like ice crystals. Simulation and experimental results illustrate that ice inhibition mechanism of oligopeptides involves binding to the ice crystal surface and disrupting the ordering of water molecules, thereby preventing the formation of well-structured ice crystals. Notably, the employment of oligopeptides as cryoprotectants maintains cell proliferation and differentiation capabilities while having a high cell viability of 90-95%, comparable to 10% DMSO.
冷冻保存是基于细胞的治疗方法的基础，但受到冰的形成和生长的严重限制，冰的形成和生长会导致不可逆的膜破裂、渗透失衡和冷损伤。抗冻蛋白和传统的冷冻保护剂，如金标准二甲基亚砜（DMSO），难以提供一种适用于各种细胞储存的经济高效且生物相容的策略。在此，我们设计并制造了一类具有强大的抑制冰重结晶（IRI）活性的寡肽，在浓度低于0.1 wt%时，平均最大晶粒尺寸降低了55.5%。对侧链官能团（如羟基和胺基）和长度（<10个氨基酸）进行了精心优化，以避免导致致命针状冰晶突然形成的热滞（TH）活性。模拟和实验结果表明，寡肽的抑冰机制包括与冰晶表面结合并破坏水分子的有序排列，从而防止形成结构良好的冰晶。值得注意的是，使用寡肽作为冷冻保护剂可维持细胞增殖和分化能力，同时具有90 - 95%的高细胞活力，与10% DMSO相当。
