Melatonin application during cryopreservation improves the development and clinical outcomes of human vitrified-warmed oocytes.

In this clinical study, we investigated the potential of melatonin (MT) supplementation in the freeze-thaw medium used for cryopreserved human oocytes. In total, 152 patients who underwent in vitro fertilization between January 2020 and December 2022 were included and categorized into different groups as follows: the donor group, comprising 108 patients who donated their oocytes, with 34 patients using a vitrification and warming medium supplemented with MT (D-MT subgroup) and 74 patients using conventional medium without MT (D-0 subgroup); and the autologous group, comprising 38 patients who used their own oocytes, with 19 patients using medium supplemented with MT (A-MT subgroup) and 19 patients using medium without MT (A-0 subgroup). After thawing, the surviving oocytes in the D-MT and A-MT subgroups and D-0 and A-0 subgroups were cultured in a fertilization media with and without 10-9 MMT for 2.5 h, respectively, followed by intracytoplasmic sperm injection insemination, embryo culture, and transfer. The survival, cleavage, high-quality embryo, clinical pregnancy, ongoing pregnancy, and implantation rates were significantly higher in the D-MT subgroup than in the D-0 subgroup (all P < 0.05). Similarly, the survival, fertilization, high-quality embryo, and high-quality blastocyst rates were significantly higher in the A-MT subgroup than in the A-0 subgroup (all P < 0.05). These findings indicate that MT addition during cryopreservation can enhance the development of vitrified-warmed human oocytes and improve clinical outcomes.

Melatonin enhances the developmental potential of immature oocytes from older reproductive-aged women by improving mitochondrial function.

Aims: To evaluate whether melatonin (MT) supplementation during in vitro maturation (IVM) of human oocytes can reverse the age-related decline in oocyte quality. Main methods: We enrolled 172 patients aged ≥35 years (older reproductive-aged women) and 83 patients aged <35 years (young women) who underwent in vitro fertilization between 2019 and 2022. We conducted IVM with and without 10 µM MT in immature oocytes of different ages. Oocyte fertilization and embryo development were observed using a stereomicroscope. We assessed the immunofluorescence intensity of mitochondrial function, measured the copy number of mitochondrial DNA (mtDNA), and examined the spindle and chromosome composition in in vitro mature stage II (IVM-MII) oocytes using immunofluorescence and second-generation sequencing. Key findings: MT supplementation significantly improved the redox level in the IVM medium and IVM-MII oocytes in older reproductive-aged women. It also significantly increased the proportion of circular mtDNA and the adenosine triphosphate content in IVM-MII oocytes. In addition, the IVM-MII oocytes obtained with MT supplementation showed a significant improvement in the normal composition of the spindle and chromosomes. Thus, the aged immature oocytes also showed significantly improved maturation and blastocyst formation rates owing to the role of MT. Significance: Supplementation with 10 µM MT in the IVM medium reverses the age-related decline in oocyte quality. Our findings provide a viable solution for enhancing fertility in older reproductive-aged women.

Blastocyst Transplantation Strategies in Women of Different Ages.

(1) Background: Single blastocyst transfers (SBT) and double blastocyst transfers (DBT) are widely used in clinical practice. The objective of this study was to investigate the application of these two strategies in women of different ages. (2) Methods: Analysis was carried out on 5477 frozen embryo transfer cycles of women in different ages. All the cycles were divided into three groups according to the age of the included women: <35, 35-39, and >39. The live birth rate (LBR) and multiple birth rate (MBR) between the SBT and DBT among these groups, respectively, were compared. (3) Results: In the women < 35 group, the LBR was similar in SBT and DBT, but the MBR was higher in DBT than SBT. In women 35-39, with >10 oocytes retrieved, the MBR in DBT was significantly higher than SBT, but there was no significant difference in LBR between the two groups; with ≤10 oocytes retrieved, the LBR in DBT were significantly higher than SBT, but the MBT was not significantly different between the two groups. In women > 39, the LBR and MBR were lower in the SBT than DBT, but the differences were not statistically significant. (4) Conclusions: Selective SET is appropriate for most young women, whereas older women are advised to make individualized choices based on the number of oocytes retrieved and blastocyst quality.

Comparative transcriptomics analysis reveals a calcineurin B-like gene to positively regulate constitutive and acclimated freezing tolerance in potato.

Freezing stress is a major limiting factor in crop production. To increase frost-hardiness of crops via breeding, deciphering the genes conferring freezing-tolerance is vital. Potato cultivars (Solanum tuberosum) are generally freezing-sensitive, but some potato wild species are freezing-tolerant, including Solanum commersonii, Solanum malmeanum and Solanum acaule. However, the underlying molecular mechanisms conferring the freezing-tolerance to the wild species remain to be deciphered. In this study, five representative genotypes of the above-mentioned species with distinct freezing-tolerance were investigated. Comparative transcriptomics analysis showed that SaCBL1-like (calcineurin B-like protein) was upregulated substantially in all of the freezing-tolerant genotypes. Transgenic overexpression and known-down lines of SaCBL1-like were examined. SaCBL1-like was shown to confer freezing-tolerance without significantly impacting main agricultural traits. A functional mechanism analysis showed that SaCBL1-like increases the expression of the C-repeat binding factor-regulon as well as causes a prolonged higher expression of CBF1 after exposure to cold conditions. Furthermore, SaCBL1-like was found to only interact with SaCIPK3-1 (CBL-interacting protein kinase) among all apparent cold-responsive SaCIPKs. Our study identifies SaCBL1-like to play a vital role in conferring freezing tolerance in potato, which may provide a basis for a targeted potato breeding for frost-hardiness.

Salicylic Acid Improves the Constitutive Freezing Tolerance of Potato as Revealed by Transcriptomics and Metabolomics Analyses.

Freezing severely impacts potato production. Deciphering the pathways and metabolites that regulate the freezing tolerance of potato is useful in cultivation and breeding for hardiness. In the present study, Solanum acaule was identified to be more freezing tolerant than S. tuberosum. Furthermore, the two genotypes before/after exposure to 4 °C for 7 d with additional -1 °C for 12 h were analysed by RNA-seq and metabolomics, and the results were compared with the previous -1 °C for 12 h. The results showed that S. acaule activated numerous genes that differed from those of S. tuberosum. Among the genes, five pathways, such as the hormone signalling pathway, which includes salicylic acid, were enriched. Further metabolomics analysis showed that the content of salicylic acid was improved in S. acaule in response to -1 °C for 12 h. Moreover, exogenous application of 0.1 mM salicylic acid to potato was shown to improve constitutive freezing tolerance and increase the expression of HSFC1. Following transcriptome and metabolome analyses, it was documented that the content of SA that increased in freezing-tolerant S. acaule after exposure to cold condition, associated with the SA signalling pathway, enhanced potato freezing tolerance, probably through HSFC1.