Fernandes Denilsa Pires, Praxedes Érika Almeida, da Silva Viana João Vitor, de Aquino Leonardo Vitorino Costa, Vieira Rodrigues Luanna Lorenna, Moura Yasmin Beatriz França, de Oliveira Moacir Franco, Alves Freitas Carlos Iberê, Fernandes Pereira Alexsandra
Laboratory of Animal Biotechnology, Federal Rural University of Semi-Arid (UFERSA), Mossoro, RN, Brazil.
Laboratory of Applied Animal Morphophysiology, UFERSA, Mossoro, RN, Brazil.
Cryobiology. 2023 Oct 28:104788. doi: 10.1016/j.cryobiol.2023.104788.
Considering the importance of efficiently obtaining somatic resource banks as an ex-situ conservation strategy for wild mammals, we evaluated two techniques (slow freezing - SF and solid-surface vitrification - SSV) for the cryopreservation of ear cartilage and skin from six-banded armadillos. Additionally, we analyzed the effects of two combinations of intracellular cryoprotectants (3.0 M or 6.0 M ethylene glycol - EG and dimethyl sulfoxide - MeSO) on SSV. Tissues not subjected to cryopreservation were used as controls. All samples were evaluated for morphological analysis and cell ability during culture. The thickness of the basal layer was similar to the control only for tissues derived from SF, while SSV ensured the preservation of the cartilage thickness. Moreover, fragments derived from SF and SSV, especially in the 3.0 M EG-MeSO group, resulted in dermis thickness and total skin similar to the control. All cryopreserved techniques maintained normal patterns of the fibroblasts, epidermal cells, and melanocytes. While only SF-derived fragments maintained the number of degenerated chondrocytes similar to the control, no difference was observed between groups for normal chondrocytes and lacunae. Moreover, SSV maintained the collagen fibers percentage of the tissues even after warming. After culture, SF and SSV techniques were efficient for the recovery of the somatic cells in all parameters evaluated, except the day of subconfluence, which was greater for the SSV group with 6.0 M EG-MeSO. In summary, SSV, especially with 3 M EG-MeSO, was as efficient as SF in preserving ear skin and cartilage derived from six-banded armadillos.
考虑到高效获取体细胞资源库作为野生哺乳动物迁地保护策略的重要性，我们评估了两种用于冷冻保存六带犰狳耳软骨和皮肤的技术（慢速冷冻 - SF和固体表面玻璃化 - SSV）。此外，我们分析了两种细胞内冷冻保护剂组合（3.0 M或6.0 M乙二醇 - EG和二甲基亚砜 - MeSO）对SSV的影响。未进行冷冻保存的组织用作对照。对所有样本进行了培养期间的形态学分析和细胞能力评估。仅SF来源的组织基底层厚度与对照相似，而SSV确保了软骨厚度的保存。此外，SF和SSV来源的碎片，尤其是在3.0 M EG-MeSO组中，真皮厚度和总皮肤面积与对照相似。所有冷冻保存技术均维持了成纤维细胞、表皮细胞和黑素细胞的正常模式。虽然只有SF来源的碎片中退化软骨细胞数量与对照相似，但各组间正常软骨细胞和腔隙数量无差异。此外，即使在复温后，SSV仍维持了组织的胶原纤维百分比。培养后，除汇合期外，SF和SSV技术在所有评估参数中对体细胞的恢复均有效，汇合期在6.0 M EG-MeSO的SSV组中更长。总之，SSV，尤其是与3 M EG-MeSO组合时，在保存六带犰狳的耳皮肤和软骨方面与SF一样有效。
