In mammalian ovaries, oocytes are physically coupled to somatic granulosa cells, and this coupling is crucial for the growth and development of competent oocytes as it mediates the transfer of metabolic support molecules. However, aging-mediated dysregulation in communication between the oocytes and granulosa cells affects the oocyte quality. In the present study, we examined the defected germline-soma communication and reduced mRNA levels encoding key structural components of transzonal projections (TZPs) in maternally aged oocytes. Oral administration of melatonin to aged mice substantially increased TZPs and maintained the cumulus cells-oocyte communication, which played a central role in the production of adequate oocyte ATP levels and reducing the accumulation of reactive oxygen species (ROS), apoptosis, DNA damage, endoplasmic reticulum (ER) stress and spindle/chromosomal defects. This beneficial effect of melatonin was inhibited by carbenoxolone (CBX), a gap junctional uncoupler, which disrupts bidirectional communications between oocyte and somatic cells. Simultaneously, melatonin significantly increased the mRNA and protein levels corresponding to genes associated with TZPs and prevented TZP retraction in in vitro-cultured cumulus-oocyte complex (COCs). Furthermore, we infused melatonin and CBX into the COCs in vitro culture system and monitored the levels of nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione (GSH) in cumulus cells and oocytes. Notably, COCs treated with melatonin demonstrated improved NADPH and GSH levels. Of note, CBX was capable of reducing NADPH and GSH levels, aggravated the ROS accumulation and ER stress. Collectively, our data demonstrate the role of melatonin in preventing age-associated germline-soma communication defects, aiding the relay of antioxidant metabolic molecules for the maintenance of oocyte quality from cumulus cells, which have important potential for improving deficient phenotypes of maternally aged oocytes and the treatment of woman infertility.

Background: Mastitis not only deteriorates the composition or quality of milk, but also damages the health and productivity of dairy goats. Sulforaphane (SFN) is a phytochemical isothiocyanate compound with various pharmacological effects such as anti-oxidant and anti-inflammatory. However, the effect of SFN on mastitis has yet to be elucidated. This study aimed to explore the anti-oxidant and anti-inflammatory effects and potential molecular mechanisms of SFN in lipopolysaccharide (LPS)-induced primary goat mammary epithelial cells (GMECs) and a mouse model of mastitis. Results: In vitro, SFN downregulated the mRNA expression of inflammatory factors (tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6), inhibited the protein expression of inflammatory mediators (cyclooxygenase-2 (COX2), and inducible nitric oxide synthase (iNOS)) while suppressing nuclear factor kappa-B (NF-κB) activation in LPS-induced GMECs. Additionally, SFN exhibited an antioxidant effect by increasing Nrf2 expression and nuclear translocation, up-regulating antioxidant enzymes expression, and decreasing LPS-induced reactive oxygen species (ROS) production in GMECs. Furthermore, SFN pretreatment promoted the autophagy pathway, which was dependent on the increased Nrf2 level, and contributed significantly to the improved LPS-induced oxidative stress and inflammatory response. In vivo, SFN effectively alleviated histopathological lesions, suppressed the expression of inflammatory factors, enhanced immunohistochemistry staining of Nrf2, and amplified of LC3 puncta LPS-induced mastitis in mice. Mechanically, the in vitro and in vivo study showed that the anti-inflammatory and anti-oxidative stress effects of SFN were mediated by the Nrf2-mediated autophagy pathway in GMECs and a mouse model of mastitis. Conclusions: These results indicate that the natural compound SFN has a preventive effect on LPS-induced inflammation through by regulating the Nrf2-mediated autophagy pathway in primary goat mammary epithelial cells and a mouse model of mastitis, which may improve prevention strategies for mastitis in dairy goats.

During mammalian oocyte meiosis, spindle migration and asymmetric cytokinesis are unique steps for the successful polar body extrusion. The asymmetry defects of oocytes will lead to the failure of fertilization and embryo implantation. In present study, we reported that an actin nucleating factor Formin-like 2 (FMNL2) played critical roles in the regulation of spindle migration and organelle distribution in mouse and porcine oocytes. Our results showed that FMNL2 mainly localized at the oocyte cortex and periphery of spindle. Depletion of FMNL2 led to the failure of polar body extrusion and large polar bodies in oocytes. Live-cell imaging revealed that the spindle failed to migrate to the oocyte cortex, which caused polar body formation defects, and this might be due to the decreased polymerization of cytoplasmic actin by FMNL2 depletion in the oocytes of both mice and pigs. Furthermore, mass spectrometry analysis indicated that FMNL2 was associated with mitochondria and endoplasmic reticulum (ER)-related proteins, and FMNL2 depletion disrupted the function and distribution of mitochondria and ER, showing with decreased mitochondrial membrane potential and the occurrence of ER stress. Microinjecting Fmnl2-EGFP mRNA into FMNL2-depleted oocytes significantly rescued these defects. Thus, our results indicate that FMNL2 is essential for the actin assembly, which further involves into meiotic spindle migration and ER/mitochondria functions in mammalian oocytes.

Oxidative stress in high-yielding dairy goats adversely affects lactation length, milk quality, and the economics of dairy products. During the lactation period, goat mammary epithelial cells (GMECs) are often in a state of disordered metabolic homeostasis primarily caused by the overproduction of reactive oxygen species (ROS). Sulforaphane (SFN), an electrophilic compound that is enriched in broccoli, is a promising antioxidant agent for future potential clinical applications. The objective of the present study was to investigate the function of SFN on hydrogen peroxide (H₂O₂)-induced oxidative damage in primary GMECs and the underlying molecular mechanisms. Isolated GMECs in triplicate were pretreated with SFN (1.25, 2.5, and 5 μM) for 24 h in the absence or presence of H₂O₂ (400 μM) for 24 h. The results showed that SFN effectively enhanced superoxide dismutase (SOD) activity, elevated the ratio of glutathione (GSH)/glutathione oxidized (GSSG), and reduced H₂O₂-induced ROS and malondialdehyde (MDA) production and cell apoptosis. Mechanically, SFN-induced nuclear factor erythroid 2-related factor 2 (NRF2/NFE2L2) translocation to the nucleus through the activation of the adenosine monophosphate-activated protein kinase (AMPK) signaling pathway coupled with inhibition of the caspase apoptotic pathway. In addition, GMECs were transfected with NFE2L2 small interfering RNA (NFE2L2 siRNA) for 48 h and/or treated with SFN (5 μM) for 24 h before being exposed to H₂O₂ (400 μM) for 24 h. We found that knockdown of NFE2L2 by siRNA abrogated the preventive effect of SFN on H₂O₂-induced ROS overproduction and apoptosis. Taken together, sulforaphane suppressed H₂O₂-induced oxidative stress and apoptosis via the activation of the AMPK/NFE2L2 signaling pathway in primary GMECs.

The gut microbiota play a vital role in regulating endocrine-mediated metabolism, immunity, and energy metabolism. However, little is known about the gut microbiota and metabolite composition and development throughout pregnancy and lactation. Here, we used amplicon sequencing to analyze the gut microbiota composition of 35 female mice in five stages of pregnancy and lactation, namely, non-receptive (NR) stages, sexually-receptive (SR) stages, at day 15 of pregnancy (Pre_D15), at the day of birth (Del), and at day 10 of lactation (Lac_D10). The results revealed that the α diversity of gut microbiota was significantly increased during pregnancy and lactation. In addition, the principal coordinate analysis (PCoA) conducted on the amplicon sequence variants’ (ASVs’) distribution of the 16S rRNA amplicons indicated that the microbiota composition was significantly different among the five groups. Based on a random forest analysis, Oscillospira, Dehalobacterium, and Alistipes were the most important microbiota. The abundance of Allobaculum, Oscillospira, and Ruminococcus were negatively correlated with the serum progesterone concentration, while the abundance of Oscillospira was positively correlated with the propionate and valerate concentration in the caecal contents. Moreover, the concentration of acetate and propionate in the Del and Lac_D10 stages was significantly lower than in the SR and Pre_D15 stages. Our findings indicate that some gut microbes and metabolites vary considerably at the different stages of pregnancy and during lactation stages, which can potentially be used as microbial biomarkers. These results provide information on the potential use of the identified microbes as probiotics to maintain a healthy pregnancy and lactation.