Multiomics analyses of Jining Grey goat and Boer goat reveal genomic regions associated with fatty acid and amino acid metabolism and muscle development.

Objective: Jining Grey Goat is a local Chinese goat breed that is well known for its high fertility and excellent meat quality but shows low meat production performance. Numerous studies have focused on revealing the genetic mechanism of its high fertility, but its highlighting meat quality and muscle growth mechanism still need to be studied. Methods: In this research, an integrative analysis of the genomics and transcriptomics of Jining Grey goats compared with Boer goats was performed to identify candidate genes and pathways related to the mechanisms of meat quality and muscle development. Results: Our results overlap among five genes (ABHD2, FN1, PGM2L1, PRKAG3, RAVER2) and detected a set of candidate genes associated with fatty acid metabolism (PRKAG3, HADHB, FASN, ACADM), amino acid metabolism (KMT2C, PLOD3, NSD2, SETDB1, STT3B, MAN1A2, BCKDHB, NAT8L, P4HA3) and muscle development (MSTN, PPARGC1A, ANKRD2). Several pathways have also been detected, such as the FoxO signaling pathway and Apelin signaling pathway that play roles in lipid metabolism, lysine degradation, N-glycan biosynthesis, valine, leucine and isoleucine degradation that involving with amino acid metabolism. Conclusion: The comparative genomic and transcriptomic analysis of Jining Grey Goat and Boer Goat revealed the mechanisms underlying the meat quality and meat productive performance of goats. These results provide valuable information for future breeding in goats.

Whole genome sequencing of Luxi Black Head sheep for screening selection signatures associated with important traits.

OBJECTIVE: Luxi Black Head sheep (LBH) is the first crossbreed specialized for meat production and was developed by crossbreeding Black Head Dorper sheep (DP) and Small Tailed Han sheep (STH) in the farming areas of northern China. Research on the genomic variations and selection signatures of LBH caused by continuous artificial selection is of great significance for identifying the genetic mechanisms of important traits of sheep and for the continuous breeding of LBH. METHODS: We explored the genetic relationships of LBH, DP, and several Mongolian sheep breeds by constructing phylogenetic tree, principal component analysis and linkage disequilibrium analysis. In addition, we analysed 29 whole genomes of sheep. The genomewide selection signatures have been scanned with four methods heterozygosity (HP), fixation index (FST), cross-population extended haplotype homozygosity (XP-EHH) and the nucleotide diversity (θπ) ratio. RESULTS: The genetic relationships analysis showed that LBH appeared to be an independent cluster closer to DP. The candidate signatures of positive selection in sheep genome revealed candidate genes for developmental process (HoxA gene cluster, BCL2L11, TSHR), immunity (CXCL6, CXCL1, SKAP2, PTK6, MST1R), growth (PDGFD, FGF18, SRF, SOCS2), and reproduction (BCAS3, TRIM24, ASTL, FNDC3A). Moreover, two signalling pathways closely related to reproduction, the thyroid hormone signalling pathway and the oxytocin signalling pathway, were detected. CONCLUSION: The selective sweep analysis of LBH genome revealed candidate genes and signalling pathways associated with developmental process, immunity, growth, and reproduction. Our findings provide a valuable resource for sheep breeding and insight into the mechanisms of artificial selection.

Gallic acid as a key substance to inhibit proliferation and adipogenesis in bovine subcutaneous adipocyte.

Gallic acid (GA) is a widespread naturally occurring phenolic acid and one of the main active monomers that forms polyphenols such as tannins. In recent years, GA has been found as a potential regulator in lipid metabolism. However, effects and possible mechanisms of GA on cell growth and lipid metabolism of bovine subcutaneous adipocytes remain unknown. In this study, we investigated whether GA could affect proliferation and adipogenesis of subcutaneous adipocyte in beef cattle. We found that GA possesses inhibitive effects on proliferation and adipogenesis of bovine subcutaneous adipocyte via activating the metabolic master factor AMP-activated protein kinase alpha (AMPKα) to promote programmed cell death and lipolysis. The findings prove GA is a key substance to inhibit proliferation and adipogenesis of bovine subcutaneous adipocyte in vitro. Further in vivo study needs conducted to verify the reductive effects of GA on subcutaneous fat in beef cattle.

Protective effect of resveratrol against hydrogen peroxide-induced oxidative stress in bovine skeletal muscle cells.

The objective of this study was to investigate the protective effects and the underlying mechanisms of resveratrol (RES) against hydrogen peroxide (H2O2)-induced oxidative stress in bovine skeletal muscle cells (BMCs). Pretreatment of BMCs with RES prior to H2O2 exposure increased cell viability, attenuated reactive oxygen species, and stabilized the redox state. H2O2 exposure activated sirtuin type 1 (SIRT1) and nuclear factor E2-related factor 2 (NRF2)-mediated signaling pathways. Pretreatment with RES did not alter SIRT1-regulated genes but inhibited the upregulation of NRF2, whereas enhanced heme oxygenase 1 (HO-1) expression. Pretreatment with RES prior to H2O2 exposure failed to suppress NRF2 expression when NRF2 was knocked down by RNA interference. However, HO-1 expression still could be induced by RES. These results suggest that RES has benifical effects against oxidative stress. NRF2-mediated pathway play an important role, and HO-1 upregulation is the key process in RES regulation. RES may be used as a therapeutic agent for meat quality improvement in beef cattle.

Caffeic acid modulates methane production and rumen fermentation in an opposite way with high-forage or high-concentrate substrate in vitro.

BACKGROUND: Plant secondary metabolites, including tannins, saponins and phenolic acids, possess potential methane (CH4 ) inhibition bioactivity. Caffeic acid (CA), as one of the typical phenolic acids, serves as a promising rumen CH4 inhibitor, but the underlying mechanisms and investigations with typical formulated rations are still not well documented. Therefore, a batch culture study was conducted to investigate the effects of CA on methanogenesis, rumen fermentation and growth of ruminal microorganisms when high-forage or high-concentrate substrates are fermented. RESULTS: After 48 h incubations, adding CA up to 40 g kg-1 dry matter linearly reduced (P < 0.05) the disappearance of dry matter, neutral detergent fiber (NDFD), total gas, methanogenesis, total volatile fatty acid and 16S rDNA copy numbers of Ruminococcus albus and Butyrivibrio fibrisolvens, and increased 16S rDNA copy numbers of methanogens for the high-forage treatment. For the high-concentrate treatment, CA exerted opposite effects (P < 0.05) on the above variables, except that CA did not affect (P > 0.05)16S rDNA copy numbers of methanogens or R. albus. CONCLUSION: Caffeic acid inhibited in vitro methanogenesis and rumen fermentation with high-forage substrate incubation. Contrarily, CA benefited in vitro fermentation and enhanced methanogenesis with high-concentrate substrate incubation. It suggests that CA modulates methanogenesis and rumen fermentation mainly by affecting the growth of cellulolytic bacteria in vitro. © 2020 Society of Chemical Industry.

HMGR overexpression and interference affect the expression of steroidogenic genes and cholesterol content in bovine intramuscular adipocytes.

Previously, we found that mevalonic acid stimulates 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGR) expression in bovine intramuscular adipocytes to influence adipocyte differentiation. However, any direct links among HMGR, steroidogenic genes, and cholesterol content remain unclear. RNA-Seq was conducted to determine the differences between the gene expression profiles of bovine adipocytes containing different HMGR expression constructs. In total, 10,234 differentially expressed genes (DEGs) were found. Of these, 35 and 6 DEGs between the control and the overexpression groups were functionally related to lipid and energy metabolism, respectively. In addition, 43 and 8 DEGs between the control and the HMGR inhibition groups were related to lipid and energy metabolism, respectively. Several DEGs related to lipid and energy metabolism were also identified between the HMGR overexpression group and the HMGR interference group, and many DEGs were correlated positively or negatively with the overexpression or inhibition of HMGR. We also found that, following the activation or inhibition of the HMGR gene, AMP-activated protein kinase (AMPK) and sirtuin type 1 (SIRT1) had opposite expression patterns in bovine intramuscular adipocytes. Interestingly, the HMGR gene was downregulated when HMGR was overexpressed, and upregulated when HMGR was inhibited. Our findings establish a theoretical understanding of signaling pathways involved in cholesterol synthesis by elucidating the relationships between key genes.

In vitro degradability of corn silage and Leymus chinensis silage and evaluation of their mixed ratios on performance, digestion and serum parameters in beef cattle.

This study investigated the degradability of corn silage (CS) and Leymus chinensis silage (LS) in vitro, and evaluated the effect of various ratios on growth performance, digestion and serum parameters in beef cattle. A 72-hr bath culture trial was performed to evaluate degradability and rumen fermentation characteristics of CS, LS and their combinations [67:33, 33:67, dry matter (DM) basis]. Forty Simmental steers, averaging 441.46 ± 4.45 kg of body weight (BW), were randomly allocated into four dietary treatments for 120-d period. Diets were given as total mixed rations with a forage-to-concentrate ratio of 60:40 and CS:LS ratios of 100:0, 67:33, 33:67 and 0:100 (DM basis). The in vitro trial showed that DM and neutral detergent fibre (NDF) degradability decreased linearly as LS proportion increased, whereas CP degradability increased linearly. Additionally, increased acid detergent fibre (ADF) degradability was detected at 48 hr of incubation. Increasing the proportion of LS increased rumen liquor pH and decreased volatile fatty acid linearly including acetate, propionate and butyrate, whereas the ammonia-N increased linearly at 12 and 72 hr of incubation. With increasing LS ratio, final BW, average daily gain and feed conversion ratio of steers decreased linearly, whereas DMI was not affected. Additionally, apparent digestibility of DM, organic matter, NDF and ADF linearly and quadratically decreased while ether extract apparent digestibility decreased linearly, and CP apparent digestibility was not affected. Serum glucose and urea nitrogen linearly and quadratically decreased while glutamic-pyruvic transaminase activity linearly decreased as the proportion of LS increased. Other serum parameters including total triglycerides, total cholesterol, total protein, albumin and glutamic-oxalacetic transaminease were not affected. Overall, enhancing ratio of LS caused inferior DM and NDF degradability but improved CP degradability in the combinations of LS and CS. A CS:LS ratio of 67:33 resulted in the best growth performance and nutrient utilization in steers.

Protective effects of astaxanthin on lipopolysaccharide-induced inflammation in bovine endometrial epithelial cells†.

Astaxanthin (AST), a natural antioxidant carotenoid, has been shown to exert anti-inflammatory effects. However, to our knowledge, no study has specifically addressed the potential protective effects of AST against bovine endometritis. The purpose of this study was to examine whether treatment with AST could protect endometrial epithelial cells against lipopolysaccharide (LPS)-induced inflammatory injury. Treatment of bovine endometrial (BEND) epithelial cell line with AST reduced LPS-induced production of interleukin-6 and tumor necrosis factor-alpha, increased the cellular activity of superoxide dismutase and catalase, decreased the proportion of apoptotic cells, and promoted the production of insulin-like growth factor and epithelial growth factor. The effects of AST were mediated through the downregulation of B-cell lymphoma 2 (Bcl-2) associated X, apoptosis regulator (Bax), and cleaved caspase-3 and through the upregulation of Bcl-2. Moreover, AST significantly increased the expression of the tight junction proteins (TJP) claudin, cadherin-1, and TJP1, which play an essential role in the maintenance of host endometrial defense barrier against pathogen infection. Collectively, these results demonstrated that treatment with AST protected against oxidative stress, prevented cell apoptosis, promoted BEND cells viability, and increased the production of growth factors, in addition to activating the endometrial defense barrier. Therefore, AST is a promising therapeutic agent for the prevention and treatment of endometritis. This finding is of utmost importance in the present times when the excessive use of antibiotics has resulted in the development of antibiotic-resistant bacteria.

Effect of simvastatin on bovine intramuscular and subcutaneous adipocytes proliferation and gene expression in vitro.

Simvastatin (SIM) is a widely used anticholesterolemic drug that blocks the biosynthesis of cholesterol. However, SIM also has pleiotropic effects on 3-hydroxy-3-methyglutary-CoA reductase (HMGR), cholesteryl ester transfer protein (CETP), and lipoprotein lipase (LPL), which are important genes in the cholesterol biosynthesis and transport processes. We investigated the effects of different concentrations of SIM on the mRNA expression of these genes in bovine intramuscular and subcutaneous adipocytes from the longissimus dorsi muscle and subcutaneous fat tissues of Luxi Yellow cattle. The results showed that SIM treatment showed dose-dependent toxicity on normal adipose cells, but no effect on cell proliferation. SIM decreased HMGR expression in a dose-dependent manner but showed no significant effect on CETP and LPL expression. Thus, SIM may lower the cholesterol content by decreasing the HMGR expression level, but CETP and LPL may be regulated through other mechanisms, which require further investigation.

Comparison of apoptosis between bovine subcutaneous and intramuscular adipocytes by resveratrol via SIRT1.

A better understanding of the differential mechanisms regulating the deposition and release of fat between intramuscular and external adipose tissues is very important to the quality of beef. Resveratrol is a natural activator of sirtuin type 1 (SIRT1), a NAD-dependent deacetylase involved in regulating the cell cycle, energy homeostasis and apoptosis in adipose tissue. To compare the molecular mechanisms underlying differential apoptosis in bovine intramuscular and subcutaneous adipocytes, we evaluated the effect of resveratrol on differentiated adipocytes. We found that resveratrol-induced apoptosis in bovine adipocytes by regulating SIRT1 activity. In addition, we report that bovine intramuscular and subcutaneous adipocytes exhibited differential responses to resveratrol. In particular, gene and protein expression of Bcl-2 was higher, whereas that of SIRT1, AMPKα, FOXO1, Bax and caspase-3 were lower in bovine subcutaneous adipocytes than in intramuscular adipocytes. After resveratrol-treatment, the extent of up- or down-regulation was higher in subcutaneous adipocytes than in intramuscular adipocytes. These data indicate that bovine subcutaneous adipocytes are more sensitive to apoptosis than intramuscular adipocytes following treatment with resveratrol by regulating SIRT1 activity.

Astragaloside IV Protects Against Oxidative Stress in Calf Small Intestine Epithelial Cells via NFE2L2-Antioxidant Response Element Signaling.

Oxidative stress can damage intestinal epithelial cell integrity and function, causing gastrointestinal disorders. Astragaloside IV (ASIV) exhibits a variety of biological and pharmacological properties, including anti-inflammatory and antioxidant effects. The purpose of this research was to investigate the cytoprotective action of ASIV and its mechanisms in calf small intestine epithelial cells with hydrogen peroxide (H2O2)-induced oxidative stress. ASIV pretreatment not only increased cell survival, but it also decreased reactive oxygen species generation and apoptosis, enhanced superoxide dismutase, catalase, and glutathione peroxidase levels, and it reduced malondialdehyde formation. Furthermore, pretreatment with ASIV elevated the mRNA and protein levels of nuclear factor erythroid 2-related factor 2 (NFE2L2), heme oxygenase-1 (HMOX1), and NAD(P)H quinone dehydrogenase 1 (NQO1). The NFE2L2 inhibitor ML385 inhibited NFE2L2 expression and then blocked HMOX1 and NQO1 expression. These results demonstrate that ASIV treatment effectively protects against H2O2-induced oxidative damage in calf small intestine epithelial cells through the activation of the NFE2L2-antioxidant response element signaling pathway.

Alanyl-glutamine ameliorates lipopolysaccharide-induced inflammation and barrier function injury in bovine jejunum epithelial cells.

The aim of this study was to investigate the effects of alanyl-glutamine (Ala-Gln) on the regulation of lipopolysaccharide (LPS)-induced inflammation and barrier function in bovine jejunum epithelial cells (BJECs). BJECs were exposed (or not) to 1 µg/mL LPS for 24 h to generate a pro-inflammatory model. The cells were then treated with different concentrations of Ala-Gln (0.25, 0.5, 1.0, 2.0, or 4.0 mmol/L) to detect any regulatory effects on the inflammation and barrier function of BJECs. LPS decreased cell viability and enhanced the production of the pro-inflammatory cytokines interleukin (IL)-6 and IL-8. LPS induced inflammation and damaged the barrier function of BJECs, as evidenced by up-regulated mRNA and protein expression of inflammatory factors and down-regulated expression of tight junction proteins. Conversely, Ala-Gln rescued the decrease in cell viability and prevented the accumulation of ILs after LPS exposure by reducing the mRNA and protein expression levels of inflammatory factors. In addition, Ala-Gln induced the mRNA and protein expression of multiple tight junction proteins, and thus reconstituted the barrier function of BJECs. In conclusion, Ala-Gln attenuates injury from inflammation and repairs damaged intestinal barrier induced with LPS, suggesting its potential as a therapeutic agent against intestinal inflammation in mammals.

Resveratrol induces apoptosis and inhibits adipogenesis by stimulating the SIRT1-AMPKα-FOXO1 signalling pathway in bovine intramuscular adipocytes.

Sirtuin type 1 (SIRTl) and AMP-activated protein kinase (AMPK) play important roles in regulating energy metabolism, cell proliferation and differentiation, ageing, apoptosis, and metabolism. The effect of 100, 200, and 400 µm Resveratrol (RES), an activator of SIRT1, on apoptosis of bovine intramuscular adipocytes was investigated by nuclear staining, flow cytometry, quantitative real-time polymerase chain reaction, and western blotting. Results show that RES inhibited adipogenesis, decreased cell viability, and increased apoptotic rates in a dose-dependent way. RES up-regulated SIRT1, AMPKα, forkhead box O1 (FOXO1), hormone-sensitive lipase (HSL), lipoprotein lipase (LPL), caspase-3, and Bax; and down-regulated peroxisome proliferator-activated receptor-gamma (PPARγ), fatty acid synthase (FAS), and Bcl-2, at both mRNA and protein level. The effect of RES was abolished by addition of sirtinol (an inhibitor of SIRT1). This is the first study demonstrating a role for AMPK-SIRT1-FOXO1 signalling pathway in regulating apoptosis in bovine intramuscular adipocytes. Our findings provide important information on the mechanism by which RES controls deposition of cattle intramuscular fat via adipocyte apoptosis.

Corticotrophin-releasing hormone and corticosterone impair development of preimplantation embryos by inducing oviductal cell apoptosis via activating the Fas system: an in vitro study.

STUDY QUESTION: What are the mechanisms by which corticotrophin-releasing hormone (CRH) and corticosterone impair the development of preimplantation embryos in the oviduct. SUMMARY ANSWER: CRH and corticosterone do not affect preimplantation embryos directly, but impair their development indirectly by triggering apoptosis of oviductal epithelial cells (OECs) through activation of the Fas system. WHAT IS KNOWN ALREADY: Studies report that stress impairs embryo development with facilitated secretion of CRH and glucocorticoids. Although an in vivo study demonstrated that preimplantation stress impaired embryo development in conjunction with oviductal apoptosis and activation of the Fas system, whether CRH or glucocorticoids damage embryos directly or indirectly by way of oviductal cells remains to be clarified. STUDY DESIGN, SIZE, DURATION: Mice of Kunming strain, the generalized lymphoproliferative disorder (gld) mice with a germline mutation F273L in Fas ligand in a C57BL/6J genomic background and the wild-type C57BL/6J mice were used. Female mice were used 8-10 weeks after birth. PARTICIPANTS/MATERIALS, SETTING, METHODS: While some female mice were killed 48 h after being injected with equine CG to collect oviducts and prepare OECs, others were killed to recover zygotes after mating with males following superovulation with eCG and hCG. The zygotes obtained were cultured with or without CRH or corticosterone (CRH/Cort) either in Chatot-Ziomek-Bavister (CZB) medium with or without OECs or in conditioned medium (CM) conditioned with OECs pretreated or not with CRH/Cort. Preimplantation development, levels of redox potential and apoptosis, and expression of CRH receptor 1 (CRHR1), glucocorticoid receptor (GR), Fas and 11ß-hydroxysteroid dehydrogenase (HSD) were observed in embryos recovered at different times of in vitro culture. After culture of OECs with or without CRH/Cort, levels of redox potential and apoptosis, mRNA and protein expression of growth factors, and protein expression of CRHR1, GR and Fas were examined in OECs and the level of FasL was measured in CM. The gld mice were used to confirm a role for the Fas system in triggering apoptosis of embryos and oviducts. MAIN RESULTS AND THE ROLE OF CHANCE: This study showed that blastocyst development was unaffected when mouse zygotes were cultured in CZB medium containing various concentrations of CRH/Cort but was impaired when embryos were cultured with CRH/Cort plus OECs or in CM conditioned with OECs pretreated with CRH/Cort (treatment CM). Culture in treatment-CM induced oxidative stress and apoptosis in embryos. Preimplantation embryos expressed GR and Fas at all stages and CRHR1 at the blastocyst stage only. Mouse 4-cell embryos and blastocysts expressed HSD2 but not HSD1. Culture of OECs with CRH/Cort increased their oxidative stress, apoptosis, CRHR1, Fas and FasL while decreasing their GR and growth factors. Blastocyst development in treatment-CM conditioned with OECs from gld mice harboring FasL mutations was superior to treatment-CM conditioned with wild-type mouse OECs. The results suggest that CRH/Cort impairs embryo development indirectly by inducing oviductal apoptosis via activating the Fas system. The insensitivity of preimplantation embryos to CRH and corticosterone is due to, respectively, a lack of CRHR and the exclusive expression of HSD2 that inactivate corticosterone. LARGE SCALE DATA: Not applicable. LIMITATIONS, REASONS FOR CAUTION: Although significant, the conclusions were drawn from limited results obtained using mice and thus they need further verification in other species. For example, bovine embryos express both HSD1 and HSD2 at all the preimplantation stages whereas mouse preimplantation embryos express HSD2 exclusively without HSD1. WIDER IMPLICATIONS OF THE FINDINGS: The data are important for our understanding of the mechanisms by which stress affects female reproduction in both human and animals, as early stages of pregnancy are considered more vulnerable to stress than the late stages. STUDY FUNDING AND COMPETING INTEREST(S): This study was supported by grants from the National Basic Research Program of China (Nos. 2014CB138503 and 2012CB944403), the China National Natural Science Foundation (Nos. 31272444 and 30972096) and the Animal breeding improvement program of Shandong Province. All authors declare that their participation in the study did not involve factual or potential conflicts of interests.

Preimplantation maternal stress impairs embryo development by inducing oviductal apoptosis with activation of the Fas system.

STUDY QUESTION: What are the mechanisms by which the preimplantation restraint stress (PIRS) impairs embryo development and pregnancy outcome? SUMMARY ANSWER: PIRS impairs embryo development by triggering apoptosis in mouse oviducts and embryos,and this involves activation of the Fas system. WHAT IS KNOWN ALREADY: Although it is known that the early stages of pregnancy are more vulnerable than later stages to prenatalstress, studies on the effect of preimplantation stress on embryo developmentare limited. Furthermore, the mechanisms by which psychological stress impairs embryo development are largely unknown. These issues are worth exploring using the mouse PIRS models because restraint of mice is an efficient experimental procedure developed for studies of psychogenic stress. STUDY DESIGN, SIZE AND DURATION: Mice of Kunming strain, the generalized lymphoproliferative disorder (gld) mice with a germline mutation F273L in FasL in a C57BL/6J genomic background and the wild-type C57BL/6J mice were used. Female and male mice were used 8-10 weeks and 10-12 weeks after birth, respectively. Female mice showing vaginal plugs were paired by weight and randomly assigned to restraint treatments or as controls. For restraint treatment, an individual mouse was put in a micro-cage with food and water available. Control mice remained in their cages with food and water during the time treated females were stressed. PARTICIPANTS/MATERIALS, SETTING, METHODS: Female mice were exposed to PIRS for 48 h starting from 16:00 on the day of vaginal plug detection. At the end of PIRS, levels of glucorticoids (GC), corticotropin-releasing hormone (CRH)and redox potential were measured in serum, while levels of GC, GC receptor (GR), CRH, CRH receptor (CRHR), Fas and Fas ligand (FasL) protein, mRNAs for brain derived neurotrophic factor (BDNF) and insulin-like growth factor-1 (IGF-1), oxidative stress (OS) and apoptosis were examined in oviducts. Preimplantation development and levels of GR, Fas, redox potential and apoptosis were observed in embryos recovered at different times after the initiation of PIRS. The gld mice were used to confirm a role for the Fas system in triggering apoptosis of embryos and oviducts. MAIN RESULTS AND THE ROLE OF CHANCE: Compared to those in control mice, while the number of blastocysts/mouse (5.0 ± 0.7 versus 11.1 ± 0.5), cell number/blastocyst (49.1 ± 1.3 versus 61.5 ± 0.9), percentages of term pregnancy (37.5% versus 90.9%) and litter size (3.7 ± 0.1versus 9.6 ± 0.6) decreased, blood CRH (560 ± 23 versus 455 ± 37 pg/ml), cortisol (27.3 ± 3.4 versus 5 ± 0.5 ng/ml) and OS index (OSI: 2.8 versus 1.7) increased significantly (all P < 0.05) following PIRS. In the oviduct, while levels of CRH (1175 ± 85 versus 881 ± 33 pg/100 mg), cortisol (28.9 ± 1.7 versus14 ± 4 ng/g), CRHR (2.3 ± 0.3 versus 1.0 ± 0.0), FasL (1.31 ± 0.06 versus 1.08 ± 0.05 ng/g), Fas (1.42 ± 0.13 versus 1.0 ± 0.0) and apoptotic cells (19.1 ± 0.5% versus 8.4 ± 0.4%) increased, levels of GR proteins (0.67 ± 0.14 versus 1.0 ± 0.0) and Igf-1 (0.6 ± 0.09 versus 1.0 ± 0.0) and Bdnf (0.73 ± 0.03 versus 1.0 ± 0.0) mRNAs decreased significantly (all P < 0.05 versus control) after PIRS. Mouse embryos expressed GR and Fas at all stages of preimplantation development and embryo OS (GSH/GSSG ratio: 0.88 ± 0.03 versus 1.19 ± 0.13) and annexin-positive cells (blastocysts: 31.4 ± 3.8% versus 10.96 ± 3.4%) increased significantly (P < 0.05) following PIRS. Furthermore, the detrimental effects of PIRS on embryo development and oviductal apoptosis were much reduced in gld mice. Thus, PIRS triggered apoptosis in oviductal cells with activation of the Fas/FasL system. The apoptotic oviductal cells promoted embryo apoptosis with reduced production of IGF-1 and BDNF and increased production of FasL. LIMITATIONS, REASONS FOR CAUTION: Although important, the conclusions were drawn from limited results obtained using a single model in one species and thus they need further verification using other models and/or in other species. Furthermore, as differences in stressed samples were modest and sometimes not significant between gld and wild-type mice whereas differences between control and stressed samples were always present within gld mice, it is deduced that signaling pathways other than the Fas/FasL system might be involved as well in the PIRS-triggered apoptosis of oviducts and embryos. WIDER IMPLICATIONS OF THE FINDINGS: The data are important for studies on the mechanisms by which psychological stress affects female reproduction, as FasL expression has been observed in human oviduct epithelium. LARGE SCALE DATA: Not applicable. STUDY FUNDING AND COMPETING INTERESTS: This study was supported by grants from the National Basic Research Program of China (Nos. 2014CB138503 and 2012CB944403), the China National Natural Science Foundation (Nos. 31272444 and 30972096) and the Animal breeding improvement program of Shandong Province. All authors declare that their participation in the study did not involve factual or potential conflicts of interests.

The signaling pathways by which the Fas/FasL system accelerates oocyte aging.

In spite of great efforts, the mechanisms for postovulatory oocyte aging are not fully understood. Although our previous work showed that the FasL/Fas signaling facilitated oocyte aging, the intra-oocyte signaling pathways are unknown. Furthermore, the mechanisms by which oxidative stress facilitates oocyte aging and the causal relationship between Ca2+ rises and caspase-3 activation and between the cell cycle and apoptosis during oocyte aging need detailed investigations. Our aim was to address these issues by studying the intra-oocyte signaling pathways for Fas/FasL to accelerate oocyte aging. The results indicated that sFasL released by cumulus cells activated Fas on the oocyte by increasing reactive oxygen species via activating NADPH oxidase. The activated Fas triggered Ca2+ release from the endoplasmic reticulum by activating phospholipase C-γ pathway and cytochrome c pathway. The cytoplasmic Ca2+ rises activated calcium/calmodulin-dependent protein kinase II (CaMKII) and caspase-3. While activated CaMKII increased oocyte susceptibility to activation by inactivating maturation-promoting factor (MPF) through cyclin B degradation, the activated caspase-3 facilitated further Ca2+releasing that activates more caspase-3 leading to oocyte fragmentation. Furthermore, caspase-3 activation and fragmentation were prevented in oocytes with a high MPF activity, suggesting that an oocyte must be in interphase to undergo apoptosis.

Effects of feeding ß-carotene on levels of ß-carotene and vitamin A in blood and tissues of beef cattle and the effects on beef quality.

The effects of feeding ß-carotene (ßC) on levels of ßC and vitamin A (retinol) in blood and tissues, and on beef quality, were evaluated in 120 steers. Each steer received supplementary ßC (at concentrations of 0, 600, 1200, or 1800 mg/day) for 90 days and then received no supplementary ßC for 60 days. ßC significantly increased in blood serum, liver, and subcutaneous and omental fat; linearly increased in the intestine and muscle; and remained unchanged in perirenal fat during supplementation. Differences between treatment groups were eliminated in subcutaneous and omental fat and in the liver by days 120 and 150, respectively, but remained significant at day 150 in blood. Retinol increased significantly in the liver and intestine during supplementation. Intramuscular fat content, meat color, and retinol in blood, muscle, or adipose tissues were not affected. Backfat thickness decreased slightly with increasing ßC supplementation and significantly differed between groups during depletion.

Role of cytoskeleton in regulating fusion of nucleoli: a study using the activated mouse oocyte model.

Although fusion of nucleoli was observed during pronuclear development of zygotes and the behavior of nucleoli in pronuclei has been suggested as an indicator of embryonic developmental potential, the mechanism for nucleolar fusion is unclear. Although both cytoskeleton and the nucleolus are important cellular entities, there are no special reports on the relationship between the two. Role of cytoskeleton in regulating fusion of nucleoli was studied using the activated mouse oocyte model. Mouse oocytes were cultured for 6 h in activating medium (Ca²âº-free CZB medium containing 10 mM SrCl2) supplemented with or without inhibitors for cytoskeleton or protein synthesis before pronuclear formation, nucleolar fusion, and the activity of maturation-promoting factor (MPF) were examined. Whereas treatment with microfilament inhibitor cytochalasin D or B or intermediate filament inhibitor acrylamide suppressed nucleolar fusion efficiently, treatment with microtubule inhibitor demecolcine or nocodazole or protein synthesis inhibitor cycloheximide had no effect. The cytochalasin D- or acrylamide-sensitive temporal window coincided well with the reported temporal window for nucleolar fusion in activated oocytes. Whereas a continuous incubation with demecolcine prevented pronuclear formation, pronuclei formed normally when demecolcine was excluded during the first hour of activation treatment when the MPF activity dropped dramatically. The results suggest that 1) microfilaments and intermediate filaments but not microtubules support nucleolar fusion, 2) proteins required for nucleolar fusion including microfilaments and intermediate filaments are not de novo synthesized, and 3) microtubule disruption prevents pronuclear formation by activating MPF.

Gene expression profiling of SIRT1, FoxO1, and PPARγ in backfat tissues and subcutaneous adipocytes of Lilu bulls.

The temporal pattern of gene expression of sirtuin 1 (SIRT1), forkhead box O1 (FoxO1), and peroxisome proliferator-activated receptor-γ (PPARγ) in differentiating bovine preadipocytes and in backfat tissue from Lilu bulls 12, 18, 24, and 30 months old was investigated using real-time quantitative PCR; Carcass characteristics and adipocyte diameters were also measured. The upregulation of PPARγ and the downregulation of SIRT1 and FoxO1 were observed in the backfat tissue of Lilu cattle with increasing age. Moreover, the results showed that fat accumulation in Lilu cattle may primarily be related to an increase in mature fat cell numbers after 18 months of age. The present study indicates SIRT1 may play an important role in the development of bovine adipose tissue in vivo. Although SIRT1, FoxO1, and PPARγ expression appeared to be nonlinear during the stages of preadipocyte differentiation, these genes play an important role during bovine adipocyte development in Lilu cattle.

Successful vitrification of mouse ovaries using less-concentrated cryoprotectants with Supercool X-1000 supplementation.

The purpose of our study was to investigate the feasibility of using less-concentrated cryoprotectants supplemented with ice blocker Supercool X-1000 to vitrify ovarian tissues. Mouse ovaries were cryopreserved in different concentrations of vitrification solution alone or with Supercool X-1000, and fresh non-frozen ovaries were used as control. The proportions of morphological normality of follicles, normal GCs in follicular fluids and developing to blastocysts were higher in 12.5% ethylene glycol (EG) + 12.5% dimethylsulfoxide (DMSO) with Supercool X-1000 than those of treated in 10% EG + 10% DMSO or 15% EG + 15% DMSO alone or with Supercool X-1000. In conclusion, the inclusion of Supercool X-1000 in less-concentrated vitrification solution was effective to improve the efficiency and efficacy of cryopreservation of ovarian tissues.