Estrogen influences the transzonal projection assembly of cumulus-oocyte complexes through G protein-coupled estrogen receptor during goat follicle development.

Estrogen is an important hormone that plays a role in regulating follicle development and oocyte maturation. Transzonal projections (TZPs) act as communication bridges between follicle somatic cells and oocytes, and their dynamic changes are critical for oocyte development and maturation. However, the roles and mechanisms of estrogen in regulating TZPs during follicular development are not yet understood. We found that the proportion of oocytes spontaneously resuming meiosis increases as the follicle grows, which is accompanied by rising estrogen levels in follicles and decreasing TZPs in cumulus-oocyte complex. To further explore the effect of elevated estrogen levels on TZP assembly, additional estrogen was added to the culture system. The increased estrogen level significantly decreased the mRNA and protein expression levels of TZP assembly-related genes. Subsequent research revealed that TZP regulation by estrogen was mediated by the membrane receptor GPER and downstream ERK1/2 signaling pathway. In summary, our study suggests that estrogen may regulate goat oocyte meiosis arrest by decreasing TZP numbers via estrogen-mediated GPER activation during follicle development.

E2/ER signaling mediates the meiotic arrest of goat intrafollicular oocytes induced by follicle-stimulating hormone.

The increased production of high-quality oocytes lies at the heart of the search to accelerate the reproduction of high-quality breeding livestock using assisted reproductive technology. Follicle-stimulating hormone (FSH) maintains the arrest of oocyte meiosis during early follicular development in vivo and promotes the synchronous maturation of nucleus and cytoplasm to improve oocyte quality. However, the mechanism by which FSH maintains meiotic arrest in oocytes is still not fully understood. Oocytes spontaneously resume meiosis once released from the arrested state. In this study, we isolated goat antral follicles with a diameter of 2.0-4.0 mm, cultured them in vitro either with or without added FSH, and finally collected the oocytes to observe their meiotic state. The results showed that FSH effectively inhibited the meiotic recovery of oocytes in follicles [4 h: control (n = 84) vs. with FSH (n = 86), P = .0115; 6 h: control (n = 86) vs. FSH (n = 85), P = 0.0308; and 8 h: control (n = 95) vs. FSH (n = 101), P = 0.0039]. FSH significantly inhibited the downregulation of natriuretic peptide receptor 2 (NPR2) expression and cyclic guanosine monophosphate (cGMP) synthesis during follicular culture in vitro (P < 0.05). Further exploration found that FSH promoted the synthesis of 17ß-estradiol (E2) (P = .0249 at 4 h and P = .0039 at 8 h) and maintained the expression of the estrogen nuclear receptor ERß, but not the estrogen nuclear receptor ERα during follicle culture in vitro (P = .0190 at 2 h, and P = .0100 at 4 h). In addition, E2/ER (estrogen nuclear receptors ERα and ERß) mediated the inhibitory effect of FSH on the downregulation of NPR2 expression and cGMP synthesis, ultimately preventing the meiotic recovery of oocytes (P < .05). In summary, our study showed that FSH-induced estrogen production in goat follicles, and the E2/ER signaling pathway, both mediated meiotic arrest in FSH-induced goat oocytes.

Obtaining a greater number of high-quality oocytes to accelerate the reproduction of high-quality breeding livestock using artificial-assisted reproductive technology remains a pressing problem in animal husbandry and requires further research into the mechanism of oocyte maturation. We investigated the regulatory action of follicle-stimulating hormone (FSH) on the meiosis of oocytes during goat follicle culture in vitro. We found that FSH promoted 17ß-estradiol (E2) synthesis and that E2/ER (estrogen nuclear receptors ERα and ERß)-mediated FSH regulation of the CNP/NPR2 (C-type natriuretic peptide/natriuretic peptide receptor 2) signaling pathway and oocyte meiosis in goat follicles. This study provided an improved theoretical foundation for the increased production of high-quality oocytes using in vitro culture methods.

FASCIN regulates actin assembly for spindle movement and polar body extrusion in mouse oocyte meiosis.

During mouse oocyte meiotic maturation, actin filaments play multiple roles in meiosis such as spindle migration and cytokinesis. FASCIN is shown to be an actin-binding and bundling protein, making actin filaments tightly packed and parallel-aligned, and FASCIN is involved in several cellular processes like adhesion and migration. FASCIN is also a potential prognostic biomarker and therapeutic target for the treatment of metastatic disease. However, little is known about the functions of FASCIN in oocyte meiosis. In the present study, we knocked down the expression of FASCIN, and our results showed that FASCIN was essential for oocyte maturation. FASCIN was all expressed in the different stages of oocyte meiosis, and it mainly localized at the cortex of oocytes from the GV stage to the MII stage and showed a similar localization pattern with actin and DAAM1. Depletion of FASCIN affected the extrusion of the first polar body, and we also observed that some oocytes extruded from the large polar bodies. This might have resulted from the defects of actin assembly, which further affected the meiotic spindle positioning. In addition, we showed that inhibition of PKC activity decreased FASCIN expression, indicating that FASCIN might be regulated by PKC. Taken together, our results provided evidence for the important role of FASCIN on actin filaments for spindle migration and polar body extrusion in mouse oocyte meiosis.

Nonylphenol exposure affects mouse oocyte quality by inducing spindle defects and mitochondria dysfunction.

Nonylphenol (NP) is a chemical raw material and intermediate which is mainly used in the production of surfactants, lubricating oil additives and pesticide emulsifiers. NP is reported to be toxic on the immune system, nervous system and reproductive system due to its binding to estrogen receptors. However, the toxicity of NP on mammalian oocyte quality remains unclear. In present study, we explored the effects of NP exposure on mouse oocyte maturation. Our results showed that 4 weeks of NP exposure increased the number of atresia follicles and decreased oocyte developmental competence. Transcriptomic analysis indicated that NP exposure altered the expression of more than 800 genes in oocytes, including multiple biological pathways. Subcellular structure examination indicated that NP exposure disrupted meiotic spindle organization and caused chromosome misalignment. Moreover, aberrant mitochondrial distribution and decreased membrane potential were also observed, indicating that NP exposure caused mitochondria dysfunction. Further analysis showed that NP exposure resulted in the accumulation of reactive oxygen species (ROS), which causes oxidative stress; and the NP-exposed oocytes showed positive Annexin-V signal, indicating the occurrence of early apoptosis. In summary, our results indicated that NP exposure reduced oocyte quality by affecting cytoskeletal dynamics and mitochondrial function, which further induced oxidative stress and apoptosis in mice.

Vesicular transport protein Arf6 modulates cytoskeleton dynamics for polar body extrusion in mouse oocyte meiosis.

Arf6 (ADP-ribosylation factor 6) is known to play important roles in membrane dynamics through the regulation of actin filament reorganization for multiple cellular processes such as cytokinesis, phagocytosis, cell migration and tumor cell invasion. However, the functions of Arf6 in mammalian oocyte meiosis have not been clarified. In present study we showed that Arf6 expressed in mouse oocytes and was mainly distributed around the spindle during meiosis. Depletion of Arf6 by morpholino microinjection caused oocytes failing to extrude first polar body. Further analysis indicated that Arf6 knock down caused the aberrant actin distribution, which further induced the failure of meiotic spindle movement. And the loss of oocyte polarity also confirmed this. The regulation of Arf6 on actin filaments in mouse oocytes might be due to its effects on the phosphorylation level of cofilin and the expression of Arp2/3 complex. Moreover, we found that the decrease of Arf6 caused the disruption of spindle formation, indicating the multiple roles of Arf6 on cytoskeleton dynamics in meiosis. In summary, our results indicated that Arf6 was involved in mouse oocyte meiosis through its functional roles in actin-mediated spindle movement and spindle organization.