Laboratory of Carnivores Reproduction, State University of Ceará, Fortaleza, Brazil.
Laboratory of Animal Germplasm Conservation, Federal Rural University of Semi-Arid, Mossoró, Brazil.
Biopreserv Biobank. 2020 Oct;18(5):415-424. doi: 10.1089/bio.2020.0048. Epub 2020 Aug 11.
Anhydrous preservation is a promising approach for storage of living biomaterials at nonfreezing temperatures. Using the domestic cat model, the objectives of this study were to characterize changes in histology, DNA integrity, and viability of testicular tissues from adult versus prepubertal individuals during microwave-assisted drying. Testes from each age group were cut into small pieces before reversible membrane permeabilization, exposure to trehalose, and microwave-assisted drying during different time periods. In Experiment 1, water content was monitored for up to 40 minutes of drying. Tissues from adult or prepubertal cats experienced similar decreases of water content during the first 10 minutes. Desiccation progressed slowly between 10 and 20 minutes and then remained stable. In Experiment 2, structural properties were explored at 5, 10, and 20 minutes of desiccation. Percentages of normal seminiferous tubules were lower after 20 minutes drying in adult (43%) than in prepubertal tissues (61%). At the same time point, the proportion of cell degeneration was higher in adult (53%) than prepubertal tissues (28%). Percentages of intact DNA in tissues remained above 85% regardless of the microwave time in both age groups. Lastly, adult and prepubertal tissues only lost 33% of viability in both age groups. Collective results demonstrated for the first time that normal morphology, incidence of degeneration, DNA integrity, and viability of testicular tissues remained at acceptable levels during microwave-assisted drying for 20 minutes. Overall, prepubertal testicular tissues appeared to be more resilient to microwave-assisted desiccations than adult tissues. Importantly, water loss in the presence of trehalose after 20 minutes of desiccation already is compatible with long-term storage of testicular tissues at temperatures above -20°C, which is one step closer to future storage at supra-zero temperatures.
无水保存是一种有前途的方法，可以在非冻结温度下储存活体生物材料。本研究使用家猫模型，旨在研究在微波辅助干燥过程中，成年和未成熟个体的睾丸组织在组织学、DNA 完整性和活力方面的变化特征。每个年龄组的睾丸在可逆膜通透性、暴露于海藻糖和微波辅助干燥之前，先切成小块，然后在不同时间进行干燥。在实验 1 中，监测了长达 40 分钟的干燥过程中的水分含量。成年或未成熟猫的组织在最初的 10 分钟内经历了相似的水分含量下降。在 10 到 20 分钟之间，干燥过程进展缓慢，然后保持稳定。在实验 2 中，在干燥 5、10 和 20 分钟时探索了结构特性。在成年（43%）组织中，干燥 20 分钟后正常生精小管的比例低于未成熟组织（61%）。在同一时间点，成年组织（53%）的细胞变性比例高于未成熟组织（28%）。无论微波时间如何，两个年龄组的组织中完整 DNA 的百分比均保持在 85%以上。最后，成年和未成熟组织在两个年龄组中的活力仅损失了 33%。总体而言，在微波辅助干燥 20 分钟内，正常形态、变性发生率、DNA 完整性和睾丸组织活力仍保持在可接受的水平。未成熟睾丸组织在微波辅助干燥过程中似乎比成年组织更能耐受干燥。重要的是，在干燥 20 分钟后，海藻糖存在下的水分损失已经与睾丸组织在-20°C 以上温度下的长期储存兼容，这是朝着未来在超零温度下储存更近了一步。
