Laboratory of Carnivore Reproduction, School of Veterinary Medicine, State University of Ceará, Fortaleza, Brazil.
Laboratory of Goat and Sheep Reproduction, School of Veterinary Medicine, State University of Ceará, Fortaleza, Brazil.
Reprod Domest Anim. 2021 Oct;56(10):1342-1348. doi: 10.1111/rda.13997. Epub 2021 Aug 12.
Testicular vitrification is an alternative to preserve the genetic material of pre-pubertal animals. However, there are few studies on post-vitrification warming. Hence, the aim was to compare the influence of different warming temperatures on vitrified testicular fragments from pre-pubertal cats. The testicles were fragmented and divided into a control group (non-vitrified) and vitrified, using an association between dimethylsulphoxide and glycerol. The vitrified fragments were warmed at 50, 55 and 60°C/5 s. Morphological and morphometric evaluations were carried out using classical histology. Afterwards, the mitochondrial activity was evaluated using Rhodamine 123. The data were expressed in mean and standard error. The differences were considered significant when p < .05. In the histomorphological analysis, the testicular fragment presented seminiferous tubules with poorly developed germinal epithelium, compatible with pre-pubertal animals. The group warmed at 50°C presented similar to the control regarding the maintenance of the integrity of the tubules and cells, without stromal rupture and lamina propria alteration, as well as regarding the maintenance of the junctions between the cells. The group warmed at 55°C showed reduction of the cell junctions, and the one warmed at 60°C had increased detachment of the basement membrane (p < .05). The warming caused a reduction in the tubular diameter inversely proportional and progressive to the increase in temperature, with the highest diameter in the control group and the lowest in the 60°C group. The control group showed a lower incidence of Rhodamine 123, followed in ascending order of the warmings at 55 and 60°C. The higher mitochondrial activity was obtained with 50°C, showing an increase of the metabolic cell function at this temperature. It was concluded that the testicular fragment of pre-pubertal cats presents a better preserved morphology, morphometry and viability when warmed at 50°C.
睾丸玻璃化是保存未成年动物遗传物质的一种替代方法。然而，关于玻璃化后解冻的研究较少。因此，本研究旨在比较不同解冻温度对未成年猫睾丸玻璃化碎片的影响。将睾丸组织碎片化并分为对照组（非玻璃化）和玻璃化组，使用二甲基亚砜和甘油联合处理。玻璃化的碎片在 50、55 和 60°C/5s 下进行解冻。使用经典组织学进行形态学和形态计量学评估。然后，使用 Rhodamine 123 评估线粒体活性。数据以平均值和标准误差表示。当 p<0.05 时，认为差异具有统计学意义。在组织形态学分析中，睾丸组织碎片显示出具有发育不良的生殖上皮的生精小管，与未成年动物相符。在 50°C 下解冻的组在保持小管和细胞完整性方面与对照组相似，没有基质破裂和固有层改变，以及细胞之间的连接保持不变。在 55°C 下解冻的组显示细胞连接减少，而在 60°C 下解冻的组显示基膜脱落增加（p<0.05）。解冻导致管腔直径减小，与温度升高呈反比且呈渐进性，对照组的管腔直径最大，60°C 组的管腔直径最小。对照组的 Rhodamine 123 发生率较低，随后依次为 55°C 和 60°C 的升温组。在 50°C 下获得了更高的线粒体活性，表明在该温度下细胞代谢功能增加。综上所述，50°C 下解冻未成年猫的睾丸组织碎片形态、形态计量和活力保存更好。
