Tan Jia, Li Jiahui, Lin Chunyan, Zhang Yuqi, Zhang Hui, Ye Na, Liu Chenxi, Han Sha, Li Zheng, Zhou Xinli
Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai, 200093, China; Key Laboratory for Tissue Engineering of Jiangxi Province, School of Medical Information Engineering, Gannan Medical University, Ganzhou 341000, China.
Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai, 200093, China.
Reprod Biomed Online. 2025 Jul;51(1):104479. doi: 10.1016/j.rbmo.2024.104479. Epub 2024 Oct 10.
Can a novel hydrogel-encapsulated cryopreservation system reduce cryoinjury while maintaining the viability and functionality of testicular tissue, and support subsequent in-vitro culture for fertility preservation?
Various pre-treatment methods that combined cryoprotectants (CPA) with gelatin methacryloyl (GelMA) hydrogels to enhance the rapid freezing of encapsulated testicular tissue were investigated. Key parameters, including CPA concentrations and compositions, were evaluated alongside different cryocarriers to establish an effective freezing system. Fresh and frozen-warmed testicular tissues were cultured in vitro for 1week, comparing the effects of two hydrogel systems: agarose and alginate methacryloyl (AlgMA), based on organ culture method.
Encapsulating testicular tissue with GelMA hydrogel, followed by the application of a 30% penetrating CPA, was identified as the optimal pre-treatment strategy. The combination of 15% dimethyl sulfoxide and 15% ethylene glycol significantly enhanced mitochondrial activity compared with 30% concentrations of either CPA alone (both P < 0.05). Among the cryocarriers tested, cryovials demonstrated superior preservation of cellular viability, mitochondrial function and antioxidant capacity. After in-vitro culture, AlgMA hydrogels further preserved basement membrane morphology and germ cell survival in frozen-warmed testicular tissues at levels comparable to fresh tissue, outperforming the traditional agarose hydrogels system in minimizing nuclear condensation, preventing cell loss and preserving tubular structure.
GelMA hydrogel functions as an osmotic barrier, regulating CPA diffusion rates to prevent apoptosis, oxidative stress and other cryoinjuries typically caused by direct exposure to high CPA concentrations. These findings advance the field of fertility preservation, showcasing the potential of novel materials and technologies to improve male fertility restoration outcomes.
一种新型水凝胶包裹的冷冻保存系统能否在保持睾丸组织活力和功能的同时减少冷冻损伤，并支持后续用于生育力保存的体外培养？
研究了多种将冷冻保护剂（CPA）与甲基丙烯酰化明胶（GelMA）水凝胶相结合以增强包裹睾丸组织快速冷冻的预处理方法。评估了包括CPA浓度和组成在内的关键参数，以及不同的冷冻载体，以建立有效的冷冻系统。根据器官培养方法，将新鲜和冻融后的睾丸组织进行体外培养1周，比较两种水凝胶系统（琼脂糖和甲基丙烯酰化海藻酸盐（AlgMA））的效果。
用GelMA水凝胶包裹睾丸组织，然后应用30%的渗透性CPA，被确定为最佳预处理策略。与单独使用30%浓度的任何一种CPA相比，15%二甲基亚砜和15%乙二醇的组合显著增强了线粒体活性（两者P < 0.05）。在测试的冷冻载体中，冻存管在细胞活力、线粒体功能和抗氧化能力的保存方面表现优异。体外培养后，AlgMA水凝胶在冻融后的睾丸组织中进一步保持了基底膜形态和生殖细胞存活，水平与新鲜组织相当，在最小化核浓缩、防止细胞丢失和保持管状结构方面优于传统的琼脂糖水凝胶系统。
GelMA水凝胶作为一种渗透屏障，调节CPA扩散速率以防止细胞凋亡、氧化应激和其他通常由直接暴露于高浓度CPA引起的冷冻损伤。这些发现推动了生育力保存领域的发展，展示了新型材料和技术在改善男性生育力恢复结果方面的潜力。
