Clinical Research Center for Reproduction and Genetics in Hunan Province, Reproductive and Genetic Hospital of CITIC-Xiangya, Changsha, Hunan, China; Institute of Reproductive and Stem Cell Engineering, Basic Medicine College, Central South University, Changsha, Hunan, China.
Clinical Research Center for Reproduction and Genetics in Hunan Province, Reproductive and Genetic Hospital of CITIC-Xiangya, Changsha, Hunan, China.
Biomol Biomed. 2024 Jan 3;24(1):51-60. doi: 10.17305/bb.2023.9421.
The global practice of cryopreservation of human semen is commonplace in Assisted Reproductive Technology (ART) labs and sperm banks. However, information on the effects of long-term cryopreservation on semen is limited to clinical data summaries and descriptions. For this study, we prepared 4 semen specimens of fresh semen, 4 specimens cryostored for at least 1 year, 3 specimens cryostored for at least 5 years, 4 specimens cryostored for at least 10 years, and 3 specimens cryostored for at least 15 years. Total RNA was extracted from each sample, amplified, labeled, and mapped to the known primary microRNA (miRNA) in the miRBase database, enabling the prediction of novel miRNAs. We found that cryopreservation can lead to changes in miRNA expression, and with the increase in storage time, these changes became more pronounced. Meanwhile, the expression of let-7d-3p, let-7c-5p and let-7i-3p miRNAs changed dynamically over cryostorage time in frozen-thawed human sperm. Furthermore, we analyzed the time-dependent dynamics of cryostorage-expressed miRNAs and their target mRNAs and found that half of the target genes were expressed in oocytes. These intersection genes were mainly enriched in cancer and cytoskeletal signaling pathways. Our findings showed that the miRNA expression profile of cryopreserved human semen is modified by long-term storage. Furthermore, as the storage time increases, the impact on human sperm becomes more pronounced in terms of miRNAs, which may have an effect on subsequent fertilization and embryonic development.
全球范围内，在辅助生殖技术（ART）实验室和精子库中，人类精液的冷冻保存实践已非常普遍。然而，关于长期冷冻保存对精液的影响的信息仅限于临床数据总结和描述。在这项研究中，我们准备了 4 份新鲜精液标本、4 份至少冷冻保存 1 年的标本、3 份至少冷冻保存 5 年的标本、4 份至少冷冻保存 10 年的标本和 3 份至少冷冻保存 15 年的标本。从每个样本中提取总 RNA，进行扩增、标记，并映射到 miRBase 数据库中已知的主要 microRNA（miRNA），从而预测新的 miRNA。我们发现，冷冻保存会导致 miRNA 表达发生变化，并且随着储存时间的增加，这些变化变得更加明显。同时，在冷冻解冻的人类精子中，let-7d-3p、let-7c-5p 和 let-7i-3p 等 miRNA 的表达在冷冻储存时间上呈现动态变化。此外，我们分析了冷冻储存表达 miRNA 及其靶 mRNA 的时间依赖性动态变化，发现一半的靶基因在卵母细胞中表达。这些交集基因主要富集在癌症和细胞骨架信号通路中。我们的研究结果表明，长期储存会改变冷冻保存的人类精液的 miRNA 表达谱。此外，随着储存时间的增加，miRNA 对人类精子的影响变得更加明显，这可能会对随后的受精和胚胎发育产生影响。
