Impact of bisphenol-A on the spliceosome and meiosis of sperm in the testis of adolescent mice.

BACKGROUND: Bisphenol-A (BPA) has estrogenic activity and adversely affects humans and animals' reproductive systems and functions. There has been a disagreement with the safety of BPA exposure at Tolerable daily intake (TDI) (0.05 mg/kg/d) value and non-observed adverse effect level (5 mg/kg/d). The current study investigated the effects of BPA exposure at various doses starting from Tolerable daily intake (0.05 mg/kg/d) to the lowest observed adverse effect level (50 mg/kg/d) on the testis development in male mice offspring. The BPA exposure lasted for 63 days from pregnancy day 0 of the dams to post-natal day (PND) 45 of the offspring. RESULTS: The results showed that BPA exposure significantly increased testis (BPA ≥ 20 mg/kg/d) and serum (BPA ≥ 10 mg/kg/d) BPA contents of PND 45 mice. The spermatogenic cells became loose, and the lumen of seminiferous tubules enlarged when BPA exposure at 0.05 mg/kg/d TDI. BPA exposure at a low dose (0.05 mg/kg/d) significantly reduced the expression of Scp3 proteins and elevated sperm abnormality. The significant decrease in Scp3 suggested that BPA inhibits the transformation of spermatogonia into spermatozoa in the testis. The RNA-seq proved that the spliceosome was significantly inhibited in the testes of mice exposed to BPA. According to the RT-qPCR, BPA exposure significantly reduced the expression of Snrpc (BPA ≥ 20 mg/kg/d) and Hnrnpu (BPA ≥ 0.5 mg/kg/d). CONCLUSIONS: This study indicated that long-term BPA exposure at Tolerable daily intake (0.05 mg/kg/d) is not safe because low-dose long-term exposure to BPA inhibits spermatogonial meiosis in mice testis impairs reproductive function in male offspring.

α-Ketoglutarate Improves Meiotic Maturation of Porcine Oocytes and Promotes the Development of PA Embryos, Potentially by Reducing Oxidative Stress through the Nrf2 Pathway.

α-Ketoglutarate (α-KG) is a metabolite in the tricarboxylic acid cycle. It has a strong antioxidant function and can effectively prevent oxidative damage. Previous studies have shown that α-KG exists in porcine follicles, and its content gradually increases as the follicles grow and mature. However, the potential mechanism of supplementation of α-KG on porcine oocytes during in vitro maturation (IVM) has not yet been reported. The purpose of this study was to explore the effect of α-KG on the early embryonic development of pigs and the mechanisms underlying these effects. We found that α-KG can enhance the development of early pig embryos. Adding 20 µM α-KG to the in vitro culture medium significantly increased the rate of blastocyst formation and the total cell number. Compared with to that of the control group, apoptosis in blastocysts of the supplement group was significantly reduced. α-KG reduced the production of reactive oxygen species and glutathione levels in cells. α-KG not only improved the activity of mitochondria but also inhibited the occurrence of apoptosis. After supplementation with α-KG, pig embryo pluripotency-related genes (OCT4, NANOG, and SOX2) and antiapoptotic genes (Bcl2) were upregulated. In terms of mechanism, α-KG activates the Nrf2/ARE signaling pathway to regulate the expression of antioxidant-related targets, thus combating oxidative stress during the in vitro culture of oocytes. Activated Nrf2 promotes the transcription of Bcl2 genes and inhibits cell apoptosis. These results indicate that α-KG supplements have a beneficial effect on IVM by regulating oxidative stress during the IVM of porcine oocytes and can be used as a potential antioxidant for IVM of porcine oocytes.

Nivalenol affects spindle formation and organelle functions during mouse oocyte maturation.

Oocyte maturation is essential for fertilization and early embryo development, and proper organelle functions guarantee this process to maintain high-quality oocytes. The type B trichothecene nivalenol (NIV) is a mycotoxin produced by Fusarium oxysporum and is commonly found in contaminated food. NIV intake affect growth, the immune system, and the female reproductive system. Here, we investigated NIV toxicity on mouse oocyte quality. Transcriptome analysis results showed that NIV exposure altered the expression of multiple genes involved in spindle formation and organelle function in mouse oocytes, indicating its toxicity on mouse oocyte maturation. Further analysis indicated that NIV exposure disrupted spindle structure and chromosome alignment, possibly through tubulin acetylation. NIV exposure induced aberrant mitochondria distribution and reduced mitochondria number, mitochondria membrane potential (MMP), and ATP levels. In addition, NIV caused the abnormal distribution of the Golgi apparatus and altered the expression of the vesicle trafficking protein Rab11. ER distribution was also disturbed under NIV exposure, indicating the effects of NIV on protein modification and transport in oocytes. Thus, our results demonstrated that NIV exposure affected spindle structure and organelles function in mouse oocytes.

Toxic effects of patulin on mouse oocytes and its possible mechanisms.

Patulin is a secondary metabolite mainly secreted by fungi and is the most common mycotoxin found in apples and apple-based products. For the past few years, numerous studies suggested the wide distribution and toxicity of patulin. In this study, we investigated the toxic effect of patulin on mouse oocytes and its possible mechanisms. The results showed that patulin treatment did not affect meiotic resumption, but inhibited oocyte maturation as indicated by failure of first polar body extrusion. Further mechanistic study showed that patulin treatment disturbed normal spindle assembly, chromosome alignment and morphology. We also found increased oxidative stress by testing the level of ROS and decreased mitochondrial membrane potential, indicating mitochondria dysfunction. In summary, our results suggest that patulin treatment causes oocyte meiotic arrest by disturbing normal spindle assembly and chromosome alignment, which may be caused by dysfunctions of mitochondria.

Gene expression profiles of two testicular somatic cell lines respond differently to 4-nitrophenol mediating vary reproductive toxicity.

4-Nitrophenol (PNP) has been extensively used in manufacturing for several decades. Its toxic effects on the male reproductive system have been reported, but the underlying mechanisms remain unclear. In this study, we utilized two testicular somatic cell lines (TM3 and TM4 cells) to explore the possible toxic effects of PNP on the male reproductive system. The activity of the cells after exposure to different doses of PNP (0.01, 0.1, 1, 10 and 100 µM) was evaluated. PNP treatment at 10 µM significantly inhibited cell viability, and 10 µM PNP was thus selected for subsequent experiments. Although PNP (10 µM) inhibited cell proliferation, promoted cell apoptosis, and changed the cell cycle distribution and ultrastructure in both types of cells, these effects were more significant in the TM4 cells. In addition, an Agilent mouse mRNA array was used to identify the gene expression differences between the control and PNP (10 µM) exposed TM3 and TM4 cells. The microarray analysis identified 67 and 1372 differentially expressed genes mainly concentrated in endothelial cell morphogenesis and anatomical structure development in TM3 cells and associated with cardiovascular system development and circulatory system development in TM4 cells. Moreover, a pathway analysis revealed that PNP not only predominately affected meiotic recombination and meiosis in TM3 cells, but also influenced axon guidance and developmental biology in TM4 cells. These results suggest that TM3 and TM4 cells exhibit different responses to PNP, which might mediate different toxic mechanisms.

The Role of MAPK3/1 and AKT in the Acquisition of High Meiotic and Developmental Competence of Porcine Oocytes Cultured In Vitro in FLI Medium.

The developmental potential of porcine oocytes cultured in vitro was remarkably enhanced in a medium containing FGF2, LIF and IGF1 (FLI) when compared to a medium supplemented with gonadotropins and EGF (control). We analyzed the molecular background of the enhanced oocyte quality by comparing the time course of MAPK3/1 and AKT activation, and the expression of genes controlled by these kinases in cumulus-oocyte complexes (COCs) cultured in FLI and the control medium. The pattern of MAPK3/1 activation in COCs was very similar in both media, except for a robust increase in MAPK3/1 phosphorylation during the first hour of culture in the FLI medium. The COCs cultured in the FLI medium exhibited significantly higher activity of AKT than in the control medium from the beginning up to 16 h of culture; afterwards a deregulation of AKT activity occurred in the FLI medium, which was not observed in the control medium. The expression of cumulus cell genes controlled by both kinases was also modulated in the FLI medium, and in particular the genes related to cumulus-expansion, signaling, apoptosis, antioxidants, cell-to-cell communication, proliferation, and translation were significantly overexpressed. Collectively, these data indicate that both MAPK3/1 and AKT are implicated in the enhanced quality of oocytes cultured in FLI medium.

NAMPT reduction-induced NAD+ insufficiency contributes to the compromised oocyte quality from obese mice.

Maternal obesity is associated with multiple adverse reproductive outcomes, whereas the underlying molecular mechanisms are still not fully understood. Here, we found the reduced nicotinamide phosphoribosyl transferase (NAMPT) expression and lowered nicotinamide adenine dinucleotide (NAD+ ) content in oocytes from obese mice. Next, by performing morpholino knockdown assay and pharmacological inhibition, we revealed that NAMPT deficiency not only severely disrupts maturational progression and meiotic apparatus, but also induces the metabolic dysfunction in oocytes. Furthermore, overexpression analysis demonstrated that NAMPT insufficiency induced NAD+ loss contributes to the compromised developmental potential of oocytes and early embryos from obese mice. Importantly, in vitro supplement and in vivo administration of nicotinic acid (NA) was able to ameliorate the obesity-associated meiotic defects and oxidative stress in oocytes. Our results indicate a role of NAMPT in modulating oocyte meiosis and metabolism, and uncover the beneficial effects of NA treatment on oocyte quality from obese mice.

The role of retinoic acid in the commitment to meiosis.

Male meiosis is a complex process whereby spermatocytes undergo cell division to form haploid cells. This review focuses on the role of retinoic acid (RA) in meiosis, as well as several processes regulated by RA before cell entry into meiosis that are critical for proper meiotic entry and completion. Here, we discuss RA metabolism in the testis as well as the roles of stimulated by retinoic acid gene 8 (STRA8) and MEIOSIN, which are responsive to RA and are critical for meiosis. We assert that transcriptional regulation in the spermatogonia is critical for successful meiosis.

Insulin/IGF-1 signaling and heat stress differentially regulate HSF1 activities in germline development.

Germline development is sensitive to nutrient availability and environmental perturbation. Heat shock transcription factor 1 (HSF1), a key transcription factor driving the cellular heat shock response (HSR), is also involved in gametogenesis. The precise function of HSF1 (HSF-1 in C. elegans) and its regulation in germline development are poorly understood. Using the auxin-inducible degron system in C. elegans, we uncovered a role of HSF-1 in progenitor cell proliferation and early meiosis and identified a compact but important transcriptional program of HSF-1 in germline development. Interestingly, heat stress only induces the canonical HSR in a subset of germ cells but impairs HSF-1 binding at its developmental targets. Conversely, insulin/insulin growth factor 1 (IGF-1) signaling dictates the requirement for HSF-1 in germline development and functions through repressing FOXO/DAF-16 in the soma to activate HSF-1 in germ cells. We propose that this non-cell-autonomous mechanism couples nutrient-sensing insulin/IGF-1 signaling to HSF-1 activation to support homeostasis in rapid germline growth.

Perfluorodecanoic acid induces meiotic defects and deterioration of mice oocytes in vitro.

Perfluorodecanoic acid (PFDA) is a member of the perfluoroalkyl substances, which are toxic to organic functions. Recently, it has been found in follicular fluid, seriously interfering with reproduction. Follicular fluid provides the oocyte with necessary resources during the process of oocytes maturation. However, the effects of PFDA on the oocyte need investigation. Our study evaluated the impacts of PFDA on the meiosis and development potential of mouse oocytes by exposing oocytes to PFDA in vitro at 350, 400, and 450 µM concentrations. The results showed that exposure to PFDA resulted in the first meiotic prophase arrest by obstructing the function of the maturation-promoting factor. It also induced the dysfunction of the spindle assembly checkpoint, expedited the progression of the first meiotic process, and increased the risk of aneuploidy. The oocytes treated with PFDA had a broken cytoskeleton which also contributed to meiotic maturation failure. Besides, PFDA exposure caused mitochondria defections, increased the reactive oxygen species level in oocytes, and consequently induced oocyte apoptosis. Moreover, PFDA produced epigenetic modifications in oocytes and increased the frequency of mature oocytes with declined development potential. In summary, our data indicated that PFDA disturbs the meiotic process and induces oocyte quality deterioration.

HDAC8 drives spindle organization during meiotic maturation of porcine oocytes.

OBJECTIVES: Histone deacetylase 8 (HDAC8) is one of the class I HDAC family proteins, which participates in the neuronal disorders, parasitic/viral infections, tumorigenesis and many other biological processes. However, its potential function during female germ cell development has not yet been fully understood. MATERIALS AND METHODS: HDAC8-targeting siRNA was microinjected into GV oocytes to deplete HDAC8. PCI-34051 was used to inhibit the enzyme activity of HDAC8. Immunostaining, immunoblotting and fluorescence intensity quantification were applied to assess the effects of HDAC8 depletion or inhibition on the oocyte meiotic maturation, spindle/chromosome structure, γ-tubulin dynamics and acetylation level of α-tubulin. RESULTS: We observed that HDAC8 was localized in the nucleus at GV stage and then translocated to the spindle apparatus from GVBD to M II stages in porcine oocytes. Depletion of HDAC8 led to the oocyte meiotic failure by showing the reduced polar body extrusion rate. In addition, depletion of HDAC8 resulted in aberrant spindle morphologies and misaligned chromosomes due to the defective recruitment of γ-tubulin to the spindle poles. Notably, these meiotic defects were photocopied by inhibition of HDAC8 activity using its specific inhibitor PCI-34051. However, inhibition of HDAC8 did not affect microtubule stability as assessed by the acetylation level of α-tubulin. CONCLUSIONS: Collectively, our findings demonstrate that HDAC8 acts as a regulator of spindle assembly during porcine oocyte meiotic maturation.

Leukaemia inhibitory factor modulates the differentiation of granulosa cells during sheep in vitro preantral to antral follicle development and improves oocyte meiotic competence.

In vitro follicle development from cryopreserved ovarian tissue could become an invaluable assisted reproduction technology for women with early ovarian failure. The challenge lies in producing, from small follicles present in the ovarian cortex, high-quality mature oocytes able to sustain embryo development. In vivo, an optimal combination of hormones and other factors coordinates the development of follicles and their enclosed oocyte. We have investigated the effect of the leukaemia inhibitory factor (LIF) cytokine, alone or in combination with FSH, on sheep in vitro follicle development from the preantral stage onwards. LIF did not alter follicle growth or antrum formation, but it modulated the differentiation of granulosa cells, as revealed by decreased production of anti-Müllerian hormone and abolished FSH-induced stimulation of oestradiol secretion. This modulatory role was also reflected in the abundance of mRNA from 35 genes, analysed by reverse-transcription coupled to microfluidic quantitative PCR. LIF stimulated or at least maintained the expression of genes involved in the dialogue between the oocyte and granulosa cells, through gap junctions (GJA4 encoding connexin 37) or paracrine signalling (Bone morphogenetic protein 15, KIT ligand and their receptors). Finally, the presence of both LIF and FSH during follicle growth strongly improved oocyte meiotic competence: most oocytes (56%) underwent subsequent nuclear maturation, a significant increase compared with their counterparts from follicles of similar size (550-900 µm) cultured with FSH only (28%) or developed in vivo (9%). Their ability to sustain embryo development remains to be evaluated. Combined supplementation with FSH and LIF certainly merits investigation with human follicles.

Effects of oocyte-derived growth factors on the growth of porcine oocytes and oocyte-cumulus cell complexes in vitro.

During oocyte growth and follicle development, oocytes closely communicate with cumulus cells. We examined the effects of oocyte-derived growth factors, growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), on the growth and acquisition of meiotic competence of porcine oocytes collected from early antral follicles (1.2-1.5 mm). First, we confirmed that GDF9 and BMP15 mRNAs were expressed almost exclusively in the oocytes. Oocyte-cumulus cell complexes (OCCs) collected from early antral follicles were cultured in growth medium supplemented with 0-100 ng/ml of GDF9 or BMP15 for 5 days. GDF9 dose-dependently increased the OCC diameter, while BMP15 did not. GDF9 and BMP15 had no significant effects on oocyte growth (P > 0.05). When OCCs that had been cultured with 50 and 100 ng/ml BMP15 were subjected to a subsequent maturation culture, they expanded fully by gonadotropic stimulation and 49% and 61% of oocytes matured to metaphase II (MII), respectively. In contrast, GDF9 did not promote cumulus expansion, and < 10% of oocytes matured to MII. Based on the difference in cumulus expansion, we compared the expression of luteinizing hormone/choriogonadotropin receptor (LHCGR) and follicle stimulating hormone receptor (FSHR) mRNAs in cumulus cells. The level of LHCGR mRNA was increased in cumulus cells of the BMP15 group, although there were no significant differences in FSHR mRNA levels among the groups. These results suggest that GDF9 promotes the growth of OCCs and that BMP15 promotes LHCGR mRNA expression in cumulus cells during oocyte growth culture, which may contribute to cumulus expansion and oocyte maturation.

Effect of combined exposure to silica nanoparticles and cadmium chloride on female zebrafish ovaries.

Silica nanoparticles (SiNPs) and cadmium chloride (CdCl2) are two important environmental pollutants. In previous research, found that SiNPs in zebrafish larvae can amplify the cardiovascular damage caused by cadmium. Whether SiNPs in the ovaries can amplify the adverse effects of cadmium on the zebrafish ovaries is worth studying problem. In this study, sexually mature female zebrafish were used as model organisms and exposed to 1 µmol/L CdCl2 and/or 25 µg/mL SiNPs for 30 days. The results showed that the structure and function of ovaries in the sole and combined exposure groups changed significantly, resulting in reduced ovarian quality, decreased number of mature oocytes, and the development of malformed offspring. A deep-sequencing analysis showed that organisms' lipid metabolism and transportation, estrogen metabolism, and response to the maturation, meiosis, and vitellogenin synthesis of oocytes were significantly affected by single exposure or combined exposure. These findings provide further insights into the harm of cooperation of CdCl2 and/or SiNPs to the aquatic ecosystems.

Interaction between thyroid hormones and gonadotropin inhibitory hormone in ex vivo culture of zebrafish testis: An approach to study multifactorial control of spermatogenesis.

Reproduction is under multifactorial control of neurohormones, pituitary gonadotropins, as well as of local gonadal signaling systems including sex steroids, growth factors and non-coding RNAs. Among the factors, gonadotropin-inhibitory hormone (Gnih) is a novel RFamide neuropeptide which directly modulates gonadotropin synthesis and release from pituitary, and in the gonads, Gnih mediated inhibitory actions on gonadotropin response of zebrafish spermatogenesis. Thyroid hormones are peripheral hormones which are also known to interact with reproductive axis, in particular, regulating testicular development and function. This study investigated the interaction between Gnih and thyroid hormones in zebrafish spermatogenesis using in vivo and ex vivo approaches. Three experimental groups were established: "control" (non-treated fish), "methimazole" and "methimazole + T4". Fish were exposed to goitrogen methimazole for 3 weeks; T4 (100 µg/L) was added in the water from the second week only in the "reversal treatment" group. After exposure, testes were dissected out and immediately incubated in Leibovitz's L-15 culture medium containing hCG, Gnih or hCG + Gnih for 7 days. Germ cell cysts and haploid cell population were evaluated by histomorphometry and flow cytometry, respectively. Our results showed that hypothyroidism affected germ cell development in basal and gonadotropin-induced spermatogenesis, in particular, meiosis and spermiogenesis. Hypothyroid testes showed lower amount of spermatozoa, and decreased potency of hCG. We also showed that goitrogen treatment nullified the inhibitory actions of Gnih on the gonadotropin-induced spermatogenesis. This study provided evidences that thyroid hormones are important regulatory factors for hCG- and Gnih-mediated functions in zebrafish spermatogenesis.

Dysfunction in Sertoli cells participates in glucocorticoid-induced impairment of spermatogenesis.

The effect of stress on male fertility is a widespread public health issue, but less is known about the related signaling pathway. To investigate this, we established a hypercortisolism mouse model by supplementing the drinking water with corticosterone for four weeks. In the hypercortisolism mice, the serum corticosterone was much higher than in the control, and serum testosterone was significantly decreased. Moreover, corticosterone treatment induced decrease of sperm counts and increase of teratozoospermia. Increased numbers of multinucleated giant cells and apoptotic germ cells as well as downregulated meiotic markers suggested that corticosterone induced impaired spermatogenesis. Further, upregulation of macrophage-specific marker antigen F4/80 as well as inflammation-related genes suggested that corticosterone induced inflammation in the testis. Lactate content was found to be decreased in the testis and Sertoli cells after corticosterone treatment, and lactate metabolism-related genes were downregulated. In vitro phagocytosis assays showed that the phagocytic activity in corticosterone-treated Sertoli cells was downregulated and accompanied by decreased mitochondrial membrane potential, while pyruvate dehydrogenase kinase-4 inhibitor supplementation restored this process. Taken together, our results demonstrated that dysfunctional phagocytosis capacity and lactate metabolism in Sertoli cells participates in corticosterone-induced impairment of spermatogenesis.

Anticentromere antibody induced by immunization with centromere protein a and Freund's complete adjuvant may interfere with mouse oocyte meiosis.

BACKGROUND: Anticentromere antibody (ACA) is a member of the antinuclear antibody (ANA) family, and recent studies have found that ACA may be associated with oocyte maturation disorders; however, the possible mechanism behind this phenomenon remains unknown. We conducted this study to investigate whether ACA could penetrate into the living oocytes and interfere with oocyte meiosis in a mouse model. METHODS: We divided mice into three groups: human recombinant centromere protein-A (human CENP-A, HA) and complete Freund's adjuvant (CFA) were used to immunize mice for the study group (HA + CFA), and mice injected with CFA (CFA group) or saline (Saline group), respectively, served as controls. After immunization, serum anti-CENP-A antibody was detected by indirect immunofluorescence assay (IIFT) and enzyme-linked immunosorbent assay (ELISA). Chromosome alignment and intracellular IgG localization in MI- and MII-stage oocytes were investigated by immunofluorescence analysis. RESULTS: Positive ACAs were successfully induced by immunization with CENP-A and CFA, and results showed that the serum level of anti-CENP-A antibody was significantly higher in the HA + CFA group compared with the control groups. There was marked increase of chromosome misalignments in MI and MII oocytes in the HA + CFA group compared to the control groups. However, no oocytes from any of the three groups showed intracellular antibody immunofluorescence. CONCLUSIONS: The development and maturation of oocytes were impaired in peripheral ACA positive mice, which exhibited severe chromosomal misalignments in metaphase meiosis; however, no evidence of ACAs entering the oocytes was observed, thus the underlying mechanism needs further exploration.

Nutrient restriction synergizes with retinoic acid to induce mammalian meiotic initiation in vitro.

The molecular machinery and chromosome structures carrying out meiosis are frequently conserved from yeast to mammals. However, signals initiating meiosis appear divergent: while nutrient restriction induces meiosis in the yeast system, retinoic acid (RA) and its target Stra8 have been shown to be necessary but not sufficient to induce meiotic initiation in mammalian germ cells. Here, we use primary culture of mouse undifferentiated spermatogonia without the support of gonadal somatic cells to show that nutrient restriction in combination with RA is sufficient to induce Stra8- and Spo11-dependent meiotic gene and chromosome programs that recapitulate the transcriptomic and cytologic features of in vivo meiosis. We demonstrate that neither nutrient restriction nor RA alone exerts these effects. Moreover, we identify a distinctive network of 11 nutrient restriction-upregulated transcription factor genes, which are associated with early meiosis in vivo and whose expression does not require RA. Our study proposes a conserved model, in which nutrient restriction induces meiotic initiation by upregulating key transcription factor genes for the meiotic gene program and provides an in vitro platform for meiotic induction that could facilitate research and haploid gamete production.

Histone deacetylase inhibitor during in vitro maturation decreases developmental capacity of bovine oocytes.

This study aimed to evaluate the effect of scriptaid during pre-maturation (PIVM) and/or maturation (IVM) on developmental competence of bovine oocytes. Cumulus-oocyte complexes (COCs) were submitted to PIVM for 6 h in the presence or absence of scriptaid. COCs were distributed into five groups: T1-IVM for 22 h, T2-PIVM for 6 h and IVM for 22 h, T3-PIVM with scriptaid for 6 h and IVM for 22 h, T4-PIVM for 6 h and IVM with scriptaid for 22 h, and T5-PIVM with scriptaid for 6 h and IVM with scriptaid for 22 h. Nuclear maturation, gene expression, cumulus cells (CCs) expansion, and embryo development and quality were evaluated. At the end of maturation, all groups presented the majority of oocytes in MII (P>0.05). Only HAT1 gene was differentially expressed (P<0.01) in oocytes with different treatments. Regarding embryo development at D7, T4 (23%) and T5 (18%) had lower blastocyst rate (P<0.05) than the other treatments (T1 = 35%, T2 = 37% and T3 = 32%). No effect was observed when scriptaid in PIVM was used in less competent oocytes (P>0.05). In conclusion, presence of scriptaid in PIVM and/or IVM did not improve developmental competence or embryo quality.

HDAC11 inhibition disrupts porcine oocyte meiosis via regulating α-tubulin acetylation and histone modifications.

HDAC11, the sole member of HDAC class IV family, plays vital roles in activating mitosis and apoptosis of tumor cells, but its functions in meiosis are rarely investigated. In the present study, the effect of HDAC11 on meiosis during porcine oocytes maturation was fully studied. The results showed that HDAC11 inhibition by its specific inhibitor JB-3-22 dramatically decreased the porcine oocyte maturation rate by disturbing spindle organization and chromosomes alignment without affecting the cytoplasmic maturation. Further study indicated that HDAC11 inhibition significantly elevated the acetylation levels of α-tubulin and H4K16, which are crucial for spindle organization and chromosomes alignment. Moreover, immunofluorescence staining results showed that HDAC11 inhibition also disturbed other meiosis-related histone modifications, such as increased H3S10pho, H4K5ac and H4K12ac levels and reduced H3T3pho level. Furthermore, RNA-seq analysis results indicated that HDAC11 inhibition disturbed porcine oocytes transcriptome (157 up-regulation, 106 down-regulation). In addition, HDAC11 inhibition compromised oocytes quality and subsequent development after parthenogenetic activation, which may be caused by the aberrant nuclear maturation and transcriptome expression profile during oocytes maturation. Therefore, our results elucidate the function of HDAC11 in porcine oocytes maturation and embryos development through regulating α-tubulin acetylation, meiosis-related histone modifications and transcriptome.