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.

In vitro maturation (IVM) of oocytes is an important assisted reproductive technology for infertility treatment and livestock breeding programs. Because of asynchronous nuclear and cytoplasmic maturation, the developmental competence of oocytes matured in vitro is compromised. C-Type natriuretic peptide (CNP), which has been proved to be an inhibitor of oocyte maturation, provides a new alternative to optimise synchronisation of nuclear and cytoplasmic maturation and improve developmental capacity of immature oocytes in vitro. To investigate the effect of temporary meiotic arrest mediated by CNP on maturation and subsequent development of immature oocytes, immature mouse oocytes from small antral follicles were temporarily arrested in meiosis by CNP (0, 5, 10 and 50 nM) for 24 h and then matured for 16 h. CNP treatment significantly increased the oocyte maturation rate from less than half to above 80%. After IVF, temporary meiotic arrest mediated by 10 and 50 nM CNP significantly improved fertilisation and blastocyst rate of oocytes matured in vitro up to approximately 55% and 30% respectively. Moreover, this positive effect of CNP was attributed, in part, to an increase in the number of mature oocytes with aligned chromosomes and a normal spindle. The present findings indicate the potential to use CNP to improve the efficiency of oocyte IVM.

The overall oocyte quality declines with aging, and this effect is strongly associated with a higher reactive oxygen species (ROS) level and the resultant oxidative damage. C-type natriuretic peptide (CNP) is a well-characterized physiological meiotic inhibitor that has been successfully used to improve immature oocyte quality during in vitro maturation. However, the underlying roles of CNP in maternally aged oocytes have not been reported. Here, we found that the age-related reduction in the serum CNP concentration was highly correlated with decreased oocyte quality. Treatment with exogenous CNP promoted follicle growth and ovulation in aged mice and enhanced meiotic competency and fertilization ability. Interestingly, the cytoplasmic maturation of aged oocytes was thoroughly improved by CNP treatment, as assessed by spindle/chromosome morphology and redistribution of organelles (mitochondria, the endoplasmic reticulum, cortical granules, and the Golgi apparatus). CNP treatment also ameliorated DNA damage and apoptosis caused by ROS accumulation in aged oocytes. Importantly, oocyte RNA-seq revealed that the beneficial effect of CNP on aged oocytes was mediated by restoration of mitochondrial oxidative phosphorylation, eliminating excessive mitophagy. CNP reversed the defective phenotypes in aged oocytes by alleviating oxidative damage and suppressing excessive PINK1/Parkin-mediated mitophagy. Mechanistically, CNP functioned as a cAMP/PKA pathway modulator to decrease PINK1 stability and inhibit Parkin recruitment. In summary, our results demonstrated that CNP supplementation constitutes an alternative therapeutic approach for advanced maternal age-related oocyte deterioration and may improve the overall success rates of clinically assisted reproduction in older women.

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.

Time-restricted eating (TRE) and low-carbohydrate diet (LCD) can improve multiple cardiometabolic parameters in patients with metabolic syndrome (MetS), but their effects on psychosocial health and satiety are unclear. In this study, we aimed to evaluate the effects of TRE, LCD, and their combination (TRE + LCD) on quality of life (QoL), sleep, mood, appetite, and metabolic hormones in patients with MetS. This is a secondary analysis of a single-center, 3-month, open-label, randomized clinical trial investigating the effects of TRE, LCD, and TRE + LCD on weight and cardiometabolic parameters in individuals with MetS. This secondary analysis examined QoL, sleep, mood, and appetite using the Rand 36-Item Short Form (SF-36); Pittsburgh Sleep Quality Index (PSQI); Depression, Anxiety, and Stress Scale; and Eating Behavior Rating Scale, respectively, as well as measured levels of metabolic hormones including leptin, amylin, glucose-dependent insulinotropic polypeptide, glucagon-like peptide-1 (GLP-1), pancreatic polypeptide (PP), and peptide YY. Between-group comparisons were conducted via one-way ANOVAs and post hoc LSD tests for normally distributed variables or Kruskal‒Wallis H tests and the Nemenyi test for abnormally distributed variables. P < 0.017 was considered significant in multiple comparisons following Bonferroni adjustment. A total of 162 participants (mean [SD] age, 41.2 [9.9] years; mean [SD] body mass index, 29.3 [3.4] kg/m²; 102 [63%] men) who started the intervention were analyzed. After 3 months, only the TRE group decreased GLP-1 levels (-0.9 [IQR, -1.9 to -0.3] pg/mL; P = 0.002), increased PP levels (8.9 [IQR, -7.6 to 71.8] pg/mL; P = 0.011), physical functioning in the SF-36 (5.2 [95% CI, 1.9 to 8.5]; P = 0.001), social functioning in the SF-36 (9.1 [95% CI, 2.5 to 15.6]; P = 0.005), role-physical in the SF-36 (24.1 [95% CI, 11.8 to 36.4]; P < 0.001), role-emotional in the SF-36 (22.4 [95% CI, 12.6 to 32.2]; P < 0.001), and sleep efficiency in the PSQI (0.29 [95% CI, 0.03 to 0.55]; P = 0.021). Compared with changes in LCD, TRE further increased general health in the SF-36 (9.7 [95% CI, 3.3 to 16.0]; P = 0.006). Relative to the changes of TRE + LCD, TRE significantly increased role-emotional in the SF-36 (19.9 [95% CI 4.9 to 34.8]; P = 0.006). Changes in sleep quality, mood status, appetite, and metabolic hormones did not differ among three groups. Greater weight loss was associated with decreased leptin levels (r = 0.538), decreased amylin levels (r = 0.294), reduced total appetite scores (r = 0.220), and improved general health (r = -0.253) (all P ≤ 0.01). TRE, LCD, and TRE + LCD all could improve psychosocial health and reduce appetite. Notably, TRE yielded greater benefits in QoL compared with LCD or TRE + LCD in individuals with MetS. ClinicalTrials.gov Identifier: NCT04475822.