Combined positive effect of oocyte extracts and brilliant cresyl blue stained recipient cytoplasts on epigenetic reprogramming and gene expression in buffalo nuclear transfer embryos

Abstract

This study examined the effects of buffalo oocyte extracts (BOE) on donor cells reprogramming and molecular characterisation of oocytes screened via brilliant cresyl blue (BCB) staining and comparison of gene expression profiles of developmentally important genes in blastocysts from IVF and cloned derived from BOE treated donor cells with BCB selected recipient cytoplasts. Relative abundance (RA) of OCT4 and NANOG was increased (P < 0.05) and HDAC-1, DNMT-1, and DNMT-3A decreased (P < 0.05) in extract treated cells (ETCs). This ETCs dedifferentiated into neuron-like lineage under appropriate induction condition. The RA of NASP, EEF1A1, DNMT1, ODC1 and RPS27A was increased (P < 0.05) in BCB+ oocytes, whereas ATP5A1 and S100A10 increased (P < 0.05) in BCB- oocytes. Total cell number and RA of OCT4, NANOG, SOX2, DNMT1, IGF2, IGF2R, MNSOD, GLUT1, BAX and BCL2 in cloned blastocysts derived from BCB+ oocytes with ETC more closely followed that of IVF counterparts compared to BCB+ oocytes with extract untreated cell and BCB- oocytes with ETC derived blastocysts. In conclusion, BOE influenced epigenetic reprogramming of buffalo fibroblasts making them suitable donors for nuclear transfer (NT). BCB staining can be effectively used for selection of developmentally competent oocytes for NT. The combined effects of epigenetic reprogramming of donor nuclei by BOE and higher nuclear reprogramming capacity of BCB+ oocytes improve developmentally important gene expression in cloned blastocysts. Whether these improvements have long-term effects on buffalo calves born following embryo transfer remains unknown.

Transcriptome profile and association study revealedSTAT3gene as a potential quality marker of bovine gametes

“…Localization of ODC1 in rat oocytes suggested its involvement in maintenance of oocyte physiology (Fernandes et al, 2017). Additionally, ODC1 was among overexpressed transcripts in developmentally competent bovine (Torner et al, 2008) and equine (Mohammadi-Sangcheshmeh et al, 2011) and buffalo (Sadeesh et al, 2017) oocytes of BCB model. However, it was not stably overexpressed in competent oocytes at both days of estrous cycle, but it rather overexpressed at the luteal phase.…”

Strategies to Improve the Efficiency of Somatic Cell Nuclear Transfer

“…Therefore, the fully grown oocytes have blue cytoplasm (BCB+) [52]. BCB staining has been widely used for selecting good quality oocytes in various species, including cattle [53,54], goats [55], horses [56], pigs [57,58], sheep [59,60], buffaloes [61,62], rabbits [63], mice [64], rats [65], dogs [66], cats [67,68], camels [69], and humans [70]. BCB+ oocytes had a higher maturation rate [56,62,63,[69][70][71], mitochondrial DNA (mtDNA) copy number [57,58], blastocyst rate [53,54,61,63], calving rate [54], as well as total cell number [54,63] and inner cell mass (ICM) to trophectoderm (TE) ratio [54,61,63] when compared with those of BCB− oocytes.…”

“…BCB staining has been widely used for selecting good quality oocytes in various species, including cattle [53,54], goats [55], horses [56], pigs [57,58], sheep [59,60], buffaloes [61,62], rabbits [63], mice [64], rats [65], dogs [66], cats [67,68], camels [69], and humans [70]. BCB+ oocytes had a higher maturation rate [56,62,63,[69][70][71], mitochondrial DNA (mtDNA) copy number [57,58], blastocyst rate [53,54,61,63], calving rate [54], as well as total cell number [54,63] and inner cell mass (ICM) to trophectoderm (TE) ratio [54,61,63] when compared with those of BCB− oocytes. Furthermore, cloned blastocysts derived from BCB+ oocytes had higher levels of histone H3 lysine 18 acetylation (H3K18ac) [54,61], OCT4, SOX2, and CDX2 expression [54,61], and also had lower apoptosis cells [54,63].…”

Current status of assisted reproductive technologies in buffaloes

“…Treatment of buffalo donor cells with cell-free extracts from buffalo matured oocyte (BOE) decreased expression levels of HDAC1, and increased H3K9ac level as well as OCT4 and NANOG pluripotency-related gene expression levels in the donor cells. Moreover, HMC river buffalo embryos produced from BOE-treated donor cells had similar OCT4, NANOG, and SOX2 expression levels to those in IVEP blastocysts (Sadeesh et al, 2017). microRNAs (miRNAs) are single-stranded noncoding RNA molecule (about 22 nucleotides) that are involved in oocyte maturation and fertilization (Gilchrist et al, 2016), embryo development (Hossain et al, 2012), maternal-to-zygotic transition (Mondou et al, 2012), and epigenetic reprogramming and pluripotency (Onder & Daley, 2011).…”