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The meat of local livestock breeds often has unique qualities and flavors. In this study, three Shanghai native pig breeds (MSZ, SWT, and SHB) exhibited better meat quality traits than globalized commercial pig breeds (DLY). Subsequently, metabolomic and lipidomic differences in the longissimus dorsi (L) and gluteus (T) muscles of the Shanghai native pig breeds and DLY pig breed were compared using liquid chromatography-mass spectrometry (LC-MS). The results demonstrated that the metabolites mainly consisted of (28.16%) lipids and lipid-like molecules, and (25.87%) organic acids and their derivatives were the two most dominant groups. Hundreds of differential expression metabolites were identified in every compared group, respectively. One-way ANOVA was applied to test the significance between multiple groups. Among the 20 most abundant differential metabolites, L-carnitine was significantly different in the muscles of the four pig breeds (p-value = 7.322 × 10-11). It was significantly higher in the L and T muscles of the two indigenous black pig breeds (MSZ and SWT) than in the DLY pigs (p-value < 0.001). Similarly, lipidomic analysis revealed the PA (18:0/18:2) was significantly more abundant in the muscle of these two black breeds than that in the DLY breed (p-value < 0.001). These specific metabolites and lipids might influence the meat quality and taste properties and lead to customer preferences. Therefore, this study provided insights into the characterization of meat metabolites and lipids in Shanghai native pig breeds.
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This study was conducted in nutrient-restricted pregnant Hu ewes to determine whether rumen-protected arginine (RP-Arg) or N-carbamylglutamate (NCG) supplementation affects fetal liver growth and development. From 35 d to 110 d of gestation, 32 Hu ewes were randomly divided into four groups: a control group (100% of the National Research Council (NRC) requirements), a nutrient-restricted group (50% of the NRC requirements), and two treatment groups (ARG and NCG, 50% of the NRC requirements, supplemented with 20 g/day RP-Arg or 5 g/day NCG, respectively). Fetal body weights, fetal liver growth performance, the capability of antioxidation, and the expression of the mRNA and proteins of apoptosis-related genes in the fetal liver were determined and analyzed at 110 d of gestation. The dry matter, water, fat, protein, and ash components of the fetal livers in the RG group were found to be lower than in the CG group, and these components were significantly higher in the NCG group than in the RG group (p < 0.05). A decrease in DNA, RNA, and protein concentrations and contents, as well as in protein/DNA ratios, was observed in the RG group in comparison to the CG group (p < 0.05). Compared with the RG group, the NCG group had higher concentrations of DNA, RNA, and protein, as well as higher protein/DNA ratios (p < 0.05). The RG group had lower concentrations of cholinesterase, nitric oxide, nitric oxide synthase, superoxide dismutase, alanine aminotransferase, and total protein than the CG group (p < 0.05). The RG group had higher levels of glutathione peroxidase, maleic dialdehyde, and aspartate aminotransferase than the CG group (p < 0.05). In the RG group, the mRNA and protein expression of p53 and Bax was significantly increased (p < 0.05) compared with the CG group, and the gene expression of FasL and Bcl-2, the ratio of Bcl-2 to Bax, and the protein expression of Bcl-2 in the RG group were lower (p < 0.05) than in the CG group. It appears that RP-Arg and NCG supplementation during pregnancy could influence fetal liver growth and development. A nutrition-based therapeutic intervention to alleviate reduced fetal growth can be developed based on this study, which has demonstrated that maternal undernutrition during pregnancy induces the maldevelopment of the fetal liver.
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BACKGROUND: The transcriptome and metabolome dissection of the skeletal muscle of high- and low- growing individuals from a crossbred population of the indigenous Chongming white goat and the Boer goat were performed to discover the potential functional differentially expressed genes (DEGs) and differential expression metabolites (DEMs). RESULTS: A total of 2812 DEGs were detected in 6 groups at three time stages (3,6,12 Month) in skeletal muscle using the RNA-seq method. A DEGs set containing seven muscle function related genes (TNNT1, TNNC1, TNNI1, MYBPC2, MYL2, MHY7, and CSRP3) was discovered, and their expression tended to increase as goat muscle development progressed. Seven DEGs (TNNT1, FABP3, TPM3, DES, PPP1R27, RCAN1, LMOD2) in the skeletal muscle of goats in the fast-growing and slow-growing groups was verified their expression difference by reverse transcription-quantitative polymerase chain reaction. Further, through the Liquid chromatography-mass spectrometry (LC-MS) approach, a total of 183 DEMs in various groups of the muscle samples and these DEMs such as Queuine and Keto-PGF1α, which demonstrated different abundance between the goat fast-growing group and slow-growing group. Through weighted correlation network analysis (WGCNA), the study correlated the DEGs with the DEMs and identified 4 DEGs modules associated with 18 metabolites. CONCLUSION: This study benefits to dissection candidate genes and regulatory networks related to goat meat production performance, and the joint analysis of transcriptomic and metabolomic data provided insights into the study of goat muscle development.
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Cabras , Carne , Músculo Esquelético , Transcriptoma , Animales , Cabras/genética , Cabras/metabolismo , Músculo Esquelético/metabolismo , Carne/análisis , Metabolómica , Perfilación de la Expresión Génica , MetabolomaRESUMEN
The aim of this study was to identify effective genetic markers for the Antigen Processing Associated Transporter 1 (TAP1), α (1,2) Fucosyltransferase 1 (FUT1), Natural Resistance Associated Macrophage Protein 1 (NRAMP1), Mucin 4 (MUC4) and Mucin 13 (MUC13) diarrhea-resistance genes in the local pig breeds, namely Shanghai white pigs, Fengjing pigs, Shawutou pigs, Meishan pigs and Pudong white pigs, to provide a reference for the characterization of local pig breed resources in Shanghai. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLR) and sequence sequencing were applied to analyze the polymorphisms of the above genes and to explore the effects on the immunity of Shanghai local pig breeds in conjunction with some immunity factors. The results showed that both TAP1 and MUC4 genes had antidiarrheal genotype GG in the five pig breeds, AG and GG genotypes of the FUT1 gene were detected in Pudong white pigs, AA antidiarrheal genes of the NRAMP1 gene were detected in Meishan pigs, the AB type of the NRAMP1 gene was detected in Pudong white pigs, and antidiarrheal genotype GG of the MUC13 gene was only detected in Shanghai white pigs. The MUC13 antidiarrhea genotype GG was only detected in Shanghai white pigs. The TAP1 gene was moderately polymorphic in Shanghai white pigs, Fengjing pigs, Shawutou pigs, Meishan pigs and Pudong white pigs, among which TAP1 in Shanghai white pigs and Shawutou pigs did not satisfy the Hardy-Weinberg equilibrium. The FUT1 gene of Pudong white pigs was in a state of low polymorphism. NRAMP1 of Meishan pigs and Pudong white pigs was in a state of moderate polymorphism, which did not satisfy the Hardy-Weinberg equilibrium. The MUC4 genes of Shanghai white pigs and Pudong white pigs were in a state of low polymorphism, and the MUC4 genes of Fengjing pigs and Shawutou pigs were in a state of moderate polymorphism, and the MUC4 genes of Fengjing pigs and Pudong white pigs did not satisfy the Hardy-Weinberg equilibrium. The MUC13 gene of Shanghai white pigs and Pudong white pigs was in a state of moderate polymorphism. Meishan pigs had higher levels of IL-2, IL-10, IgG and TNF-α, and Pudong white pigs had higher levels of IL-12 than the other pigs. The level of interleukin 12 (IL-12) was significantly higher in the AA genotype of the MUC13 gene of Shanghai white pigs than in the AG genotype. The indicator of tumor necrosis factor alpha (TNF-α) in the AA genotype of the TAP1 gene of Fengjing pigs was significantly higher than that of the GG and AG genotypes. The indicator of IL-12 in the AG genotype of the Shawutou pig TAP1 gene was significantly higher than that of the GG genotype. The level of TNF-α in the AA genotype of the NRAMP1 gene of Meishan pigs was markedly higher than that of the AB genotype. The IL-2 level of the AG type of the FUT1 gene was obviously higher than that of the GG type of Pudong white pigs, the IL-2 level of the AA type of the MUC4 gene was dramatically higher than that of the AG type, and the IgG level of the GG type of the MUC13 gene was apparently higher than that of the AG type. The results of this study are of great significance in guiding the antidiarrhea breeding and molecular selection of Shanghai white pigs, Fengjing pigs, Shawutou pigs, Meishan pigs and Pudong white pigs and laying the foundation for future antidiarrhea breeding of various local pig breeds in Shanghai.
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Diarrea , Animales , Porcinos/genética , China , Diarrea/genética , Diarrea/veterinaria , Fucosiltransferasas/genética , Proteínas de Transporte de Catión/genética , Cruzamiento , Galactósido 2-alfa-L-Fucosiltransferasa , Mucina 4/genética , GenotipoRESUMEN
To address the safety problems posed by the transportation of boar semen using LN, this study was conducted on the short-term storage of frozen boar semen in dry ice (-79 °C). Boar semen frozen in LN was transferred to dry ice, kept for 1 day, 3 days, 5 days, 7 days, or 8 days, and then moved back to LN. The quality of frozen semen stored in LN or dry ice was determined to evaluate the feasibility of short-distance transportation with dry ice. The results showed that 60 °C for 8 s was the best condition for thawing frozen semen stored in dry ice. No significant differences in spermatozoa motility, plasma membrane integrity, or acrosome integrity were observed in semen after short-term storage in dry ice compared to LN (p > 0.05). There were no significant changes in antioxidant properties between storage groups either (p > 0.05). In conclusion, dry ice could be used as a cold source for the short-term transportation of frozen boar semen for at least 7 days, without affecting sperm motility, morphological integrity, or antioxidant indices.
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Reproductive performance is one of the most important economic traits in the goat industry. Increasing the number of goats is an effective measure to improve production efficiency and reduce production costs. Ovaries are important reproductive organs in female mammals that directly affect the estrous cycle and reproductive abilities. Understanding the complex transcription network of non-coding RNAs (lncRNAs, circRNAs, and miRNAs) and messenger RNA (mRNA) could lead to significant insights into the ovarian regulation of the reproductive processes of animals. However, the whole-transcriptome analysis of the non-coding RNAs and mRNA of the ovaries in Chongming white goats between high-fecundity (HP) and low-fecundity (LP) groups is limited. In this study, a whole-transcriptome sequencing approach was used to identify lncRNA, circRNA, miRNA, and mRNA expression in the ovaries of Chongming white goats during the estrus phase using RNA-Seq technology. More than 20,000 messenger RNAs (mRNAs), 10,000 long non-coding RNAs (lncRNAs), 3500 circular RNAs (circRNAs), and 1000 micro RNAs (miRNAs) were identified. A total of 1024 differential transcripts (724 mRNAs, 112 lncRNAs, 178 circRNAs, and 10 miRNAs) existing between the HP and the LP groups were revealed through a bioinformatics analysis. They were enriched in the prolactin signaling pathway, the Jak-STAT signaling pathway, and the GnRH signaling pathway, as well as various metabolic pathways. Differentially expressed mRNAs (such as LYPD6, VEGFA, NOS3, TNXB, and EPHA2) and miRNAs (such as miR-10a-5p) play key roles in the regulation of goat ovaries during the estrus phase. The enrichment of pathways related to reproduction, such as the Hippo, Hedgehog, PI3K-AKT, and MAPK signaling pathways, suggests that they might be involved in the prolificacy of goat ovaries. Overall, we identified several gene modules associated with goat fecundity and provided a basis for a molecular mechanism in the ovaries of Chongming white goats.
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China has rich genetic resources of local pig breeds. In this study, whole-genome resequencing was performed on five Shanghai local pig breeds, aiming to analyze their population genetic structure and unique genomic characteristics. Tens of millions of single nucleotide variants were obtained through the resequencing of a total of 150 individual pigs from five local pig breeds (Meishan, Fengjing, Shawutou, Pudong White, and Shanghai White) after mapping them with the pig reference genome of Sus scrofa 11.1. The results of admixture structure analysis also clearly demonstrated the genetic differences between the Shanghai local pig breeds and the three commercial pig breeds (Duroc, Landrace, and Yorkshire). The genetic infiltration of Landrace and Yorkshire pig breeds in the SHW breed was detected, which is consistent with the early history of crossbreeding in this breed. Selective sweep analysis between four indigenous Shanghai pig breed populations and three commercial pig breed populations identified 270 and 224 genes with selective signatures in the commercial and indigenous Shanghai pig populations, respectively. Six genes (TGS1, PLAG1, CHCHD7, LCORL, TMEM68, and TMEM8B) were found to be associated with animal growth in the commercial pig population through gene enrichment and protein-protein interaction analysis. In contrast, the MSRB3 gene in the indigenous Shanghai pig population was significantly under selection, which correlated with the long pendulous ear phenotype of the indigenous Shanghai pig population. In conclusion, this study is the first genomic profiling of five representative local pig breeds in Shanghai, which provides molecular genetic data and foundations for better conservation and utilization of local pig breed resources in Shanghai, China.
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A substantial body of evidence points to the heritability of dietary preferences. While vegetarianism has been practiced for millennia in various societies, its practitioners remain a small minority of people worldwide, and the role of genetics in choosing a vegetarian diet is not well understood. Dietary choices involve an interplay between the physiologic effects of dietary items, their metabolism, and taste perception, all of which are strongly influenced by genetics. In this study, we used a genome-wide association study (GWAS) to identify loci associated with strict vegetarianism in UK Biobank participants. Comparing 5,324 strict vegetarians to 329,455 controls, we identified one SNP on chromosome 18 that is associated with vegetarianism at the genome-wide significant level (rs72884519, ß = -0.11, P = 4.997 x 10-8), and an additional 201 suggestively significant variants. Four genes are associated with rs72884519: TMEM241, RIOK3, NPC1, and RMC1. Using the Functional Mapping and Annotation (FUMA) platform and the Multi-marker Analysis of GenoMic Annotation (MAGMA) tool, we identified 34 genes with a possible role in vegetarianism, 3 of which are GWAS-significant based on gene-level analysis: RIOK3, RMC1, and NPC1. Several of the genes associated with vegetarianism, including TMEM241, NPC1, and RMC1, have important functions in lipid metabolism and brain function, raising the possibility that differences in lipid metabolism and their effects on the brain may underlie the ability to subsist on a vegetarian diet. These results support a role for genetics in choosing a vegetarian diet and open the door to future studies aimed at further elucidating the physiologic pathways involved in vegetarianism.
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Dieta Vegetariana , Estudio de Asociación del Genoma Completo , Humanos , Dieta , Dieta Vegana , EncéfaloRESUMEN
In this study, we explore the effects of Poria cocos mushroom polysaccharides (PCPs) on the quality and DNA methylation of the cryopreserved spermatozoa of Shanghai white pigs. A total of 24 ejaculates (three ejaculate samples per boar) from eight Shanghai white pigs were manually collected. The pooled semen was diluted with a based extender supplemented with different concentrations of PCPs (0, 300, 600, 900, 1200, and 1500 µg/mL). Once thawed, the quality of the spermatozoa and their antioxidant function were assessed. In the meantime, the effect of spermatozoa DNA methylation was also analyzed. The results show that compared with the control group, 600 µg/mL of PCPs significantly improves the spermatozoa viability (p < 0.05). The motility and plasma membrane integrity of the frozen-thawed spermatozoa are significantly higher after treatment with 600, 900, and 1200 µg/mL of PCPs compared with the control group (p < 0.05). In comparison with the control group, the percentages of acrosome integrity and mitochondrial activity are significantly enhanced after the application of 600 and 900 µg/mL PCPs (p < 0.05). The reactive oxygen species (ROS), the malondialdehyde (MDA) levels, and the glutathione peroxidase (GSH-Px) activity, in comparison with the control group, are significantly decreased in all groups with PCPs (all p < 0.05). The enzymatic activity of superoxide dismutase (SOD) in spermatozoa is significantly higher in the treatment with 600 µg/mL of PCPs than in the other groups (p < 0.05). As compared with the control group, a significant increase in the catalase (CAT) level is found in the groups with PCPs at 300, 600, 900, and 1200 µg/mL (all p < 0.05). In comparison with the control group, the 5-methylcytosine (5-mC) levels are significantly decreased in all groups with PCPs (all p < 0.05). As a result of these findings, a certain amount of PCPs (600-900 µg/mL) added to the cryodiluent can significantly improve the quality of Shanghai white pig spermatozoa and can also reduce the methylation of spermatozoa DNA caused by cryopreservation. This treatment strategy may establish a foundation for the cryopreservation of semen from pigs.
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Agaricales , Preservación de Semen , Wolfiporia , Masculino , Animales , Porcinos , Wolfiporia/metabolismo , Agaricales/metabolismo , Metilación de ADN , Preservación de Semen/métodos , China , Espermatozoides/metabolismo , Criopreservación/métodos , Antioxidantes/farmacología , Antioxidantes/metabolismo , Polisacáridos/farmacología , Polisacáridos/metabolismoRESUMEN
Chinese indigenous pig breeds have unique genetic characteristics and a rich diversity; however, effective breed identification methods have not yet been well established. In this study, a genotype file of 62,822 single-nucleotide polymorphisms (SNPs), which were obtained from 1059 individuals of 18 Chinese indigenous pig breeds and 5 cosmopolitan breeds, were used to screen the discriminating SNPs for pig breed identification. After linkage disequilibrium (LD) pruning filtering, this study excluded 396 SNPs on non-constant chromosomes and retained 20.92~-27.84% of SNPs for each of the 18 autosomes, leaving a total of 14,823 SNPs. The principal component analysis (PCA) showed the largest differences between cosmopolitan and Chinese pig breeds (PC1 = 10.452%), while relatively small differences were found among the 18 indigenous pig breeds from the Yangtze River Delta region of China. Next, a random forest (RF) algorithm was used to filter these SNPs and obtain the optimal number of decision trees (ntree = 1000) using corresponding out-of-bag (OOB) error rates. By comparing two different SNP ranking methods in the RF analysis, the mean decreasing accuracy (MDA) and mean decreasing Gini index (MDG), the effects of panels with different numbers of SNPs on the assignment accuracy, and the statistics of SNP distribution on each chromosome in the panels, a panel of 1000 of the most breed-discriminative tagged SNPs were finally selected based on the MDA screening method. A high accuracy (>99.3%) was obtained by the breed prediction of 318 samples in the RF test set; thus, a machine learning classification method was established for the multi-breed identification of Chinese indigenous pigs based on a low-density panel of SNPs.
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Polimorfismo de Nucleótido Simple , Bosques Aleatorios , Animales , Genotipo , Desequilibrio de Ligamiento , Polimorfismo de Nucleótido Simple/genética , Porcinos/genéticaRESUMEN
In this study, we aimed to determine the benefit of mitoquinone (MitoQ) in rooster semen extenders on sperm quality, motility parameters, antioxidant capacities, and apoptotic changes in post-thawed rooster semen. A total of 85 ejaculates from 18 roosters were collected and then divided into five equal aliquots and cryopreserved in extenders with 1.0% soy lecithin nanoparticles that contained various concentrations of MitoQ (0 nM (M0), 50 nM (M50), 100 nM (M100), 150 nM (M150), and 200 nM (M200)). By using a computer-assisted semen analyzer, sperm motility parameters were assessed after freeze thawing. The M150 group had significantly higher percentages of total motility, progressive motility, viability, acrosome membrane integrity, and mitochondrial activity than the other groups (p < 0.05). Compared to other groups, M100 and M150 groups produced a higher percentage of plasma membrane integrity and ATP contents (p < 0.05). Additionally, the lowest levels of ROS and MDA in spermatozoa were observed in M150 group (p < 0.05), whereas the highest levels of ROS and MDA were observed in sperm in the controls or the M200 group (p < 0.05). Significantly higher values of SOD, GPx, and Cas-3 were found in the M150 group compared to other groups (p < 0.05). Overall, these results demonstrate that MitoQ at 150 nM not only ameliorates post-thawed sperm quality and motility parameters by restoring ATP levels and preventing membrane damage, but also improves redox balance and antiapoptotic activities.
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Mitochondria hold redox homeostasis and energy metabolism as a crucial factor during oocyte maturation, while the exposure of estrogenic mycotoxin zearalenone causes developmental incapacity in porcine oocyte. This study aimed to reveal a potential resistance of phytoalexin resveratrol against zearalenone during porcine oocyte maturation and whether its mechanism was related with PTEN-induced kinase 1 (PINK1)/Parkin-mediated mitophagy. Porcine oocytes were exposed to 20 µM zearalenone with or without 2 µM resveratrol during in vitro maturation. As for the results, zearalenone impaired ultrastructure of mitochondria, causing mitochondrial depolarization, oxidative stress, apoptosis and embryonic developmental incapacity, in which mitophagy was induced in response to mitochondrial dysfunction. Phytoalexin resveratrol enhanced mitophagy through PINK1/Parkin in zearalenone-exposed oocytes, manifesting as enhanced mitophagy flux, upregulated PINK1, Parkin, microtubule-associated protein light-chain 3 beta-II (LC3B-II) and downregulated substrates mitofusin 2 (MFN2), voltage-dependent anion channels 1 (VDAC1) and p62 expressions. Resveratrol redressed zearalenone-induced mitochondrial depolarization, oxidative stress and apoptosis, and accelerated mitochondrial DNA copy during maturation, which improved embryonic development. This study offered an antitoxin solution during porcine oocyte maturation and revealed the involvement of PINK1/Parkin-mediated mitophagy, in which resveratrol mitigated zearalenone-induced embryonic developmental incapacity.
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Antitoxinas , Zearalenona , Animales , Antitoxinas/metabolismo , ADN Mitocondrial , Proteínas Asociadas a Microtúbulos/metabolismo , Mitocondrias/metabolismo , Mitofagia/genética , Oocitos/metabolismo , Proteínas Quinasas , Resveratrol/farmacología , Sesquiterpenos , Porcinos , Ubiquitina-Proteína Ligasas/genética , Zearalenona/metabolismo , Zearalenona/toxicidad , FitoalexinasRESUMEN
Mycotoxins such as zearalenone (ZEN), deoxynivalenol (DON) and T-2 toxin (T-2) are the most poisonous biological toxins in food pollution. Mycotoxin contaminations are a global health issue. The aim of the current study was to use porcine Leydig cells as a model to explore the toxic effects and underlying mechanisms of ZEN, DON and T-2. The 50% inhibitory concentration (IC50) of ZEN was 49.71 µM, and the IC50 values of DON and T-2 were 2.49 µM and 97.18 nM, respectively. Based on the values of IC50, ZEN, DON and T-2 exposure resulted in increased cell apoptosis, as well as disrupted mitochondria membrane potential and cell cycle distribution. The results also showed that ZEN and DON significantly reduced testosterone and progesterone secretion in Leydig cells, but T-2 only reduced testosterone secretion. Furthermore, the expression of steroidogenic acute regulatory (StAR) protein and 3ß-hydroxysteroid dehydrogenase (3ß-HSD) were significantly decreased by ZEN, DON and T-2; whereas the protein expression of cholesterol side-chain cleavage enzyme (CYP11A1) was only significantly decreased by ZEN. Altogether, these data suggest that the ZEN, DON and T-2 toxins resulted in reproductive toxicity involving the inhibition of steroidogenesis and cell proliferation, which contributes to the cellular apoptosis induced by mitochondrial injury in porcine Leydig cells.
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Células Intersticiales del Testículo/efectos de los fármacos , Toxina T-2/toxicidad , Tricotecenos/toxicidad , Zearalenona/toxicidad , 3-Hidroxiesteroide Deshidrogenasas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Enzima de Desdoblamiento de la Cadena Lateral del Colesterol/metabolismo , Células Intersticiales del Testículo/metabolismo , Masculino , Fosfoproteínas/metabolismo , Progesterona/metabolismo , Porcinos , Testosterona/metabolismoRESUMEN
Vitrification is an effective technique for fertility preservation, but is known to lead to mitochondrial dysfunction in porcine oocytes. Mitophagy is induced to rebalance mitochondrial function, a process in which reactive oxygen species (ROS) plays a role. In this study, vitrified-warmed porcine oocytes were incubated for 4 h with the oxidant AAPH or antioxidant α-tocopherol to alter ROS levels. A series of tests suggested that vitrification damaged mitochondrial structure and caused dysfunction, including blurred mitochondrial cristae, decreased mitochondrial membrane potential, decreased mtDNA copy number and increased ROS generation. This dysfunction resulted in mitophagy and the loss of embryonic developmental potential. Incubation with AAPH or α-tocopherol altered mitochondrial function and mitophagy flux status in vitrified oocytes. The PINK1/Parkin pathway was involved in oxidative stress regulation in vitrified oocytes. Under AAPH-induced oxidative stress, increased fluorescence intensity of Parkin, increased expression of PINK1, Parkin, and LC3B-II, and decreased expression of MFN2 and p62 were observed, whereas the opposite effects were induced under α-tocopherol treatment. The inhibition of ROS by α-tocopherol benefitted mitochondrial homeostasis and alleviated PINK1/Parkin-mediated mitophagy, resulting in the recovery of embryonic developmental potential in vitrified porcine oocytes. Therefore, this study provides a new mechanism for the application of antioxidants to aid the cryopreservation of porcine oocytes.
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Mitocondrias , Mitofagia , Animales , Potencial de la Membrana Mitocondrial , Mitocondrias/metabolismo , Oocitos/metabolismo , Estrés Oxidativo , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Porcinos , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
The present study aimed to investigate the impact of different concentrations (0%, 0.5%, 1.0%, 1.5%, and 2.0%) of nano-soybean lecithin (SL) in the extender on sperm quality, sperm motion characteristics, and fertility outcomes of post-thawed rooster semen. Adult Ross broiler breeder roosters (n = 20) were subjected to semen collections twice a week for three weeks. At each collection, semen samples were pooled and allocated into five treatments corresponding to different nano-SL concentrations (control, SL0.5, SL1.0, SL1.5, and SL2.0). Sperm parameters, including motility (collected using a computer-assisted sperm analysis system), plasma membrane and acrosome integrities, and mitochondrial activity were assessed. Sperm malondialdehyde (MDA) and antioxidant activities (total antioxidant capacity (TAC); superoxide dismutase (SOD); glutathione peroxidase (GPx)) were evaluated. The fertility and hatchability obtained with frozen-thawed rooster semen supplemented with the optimum nano-SL concentration were assessed after artificial insemination. The results showed that the addition of 1% nano-SL into the extender led to a higher semen motility in roosters, improved plasma membrane and acrosome integrities, and higher mitochondrial activity of post-thawed rooster semen in comparison to controls (p < 0.05). The MDA levels in the SL0.5 and SL1.0 groups were lower than the other groups (p < 0.05). TAC activities in SL0.5, SL1.0, and SL1.5 groups were significantly higher than those in the other groups (p < 0.05). It was observed that the concentration of SOD was higher in the SL1.0 group than in the other groups (p < 0.05). The activity of GPx was not influenced in any of the cases (p > 0.05). Moreover, the percentages of fertility and hatchability in the SL1.0 group were higher (56.36% and 58.06%) than those in the control group (42.72% and 40.43%). In summary, the addition of nano-SL to the extenders enhanced the post-thawed semen quality and fertility of roosters by reducing the level of oxidative stress. The optimum nano-SL concentration was 1.0%. These results may be beneficial for improving the efficacy of semen cryopreservation procedures in poultry breeding.
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Mitochondrial dysfunction is considered a crucial factor aggravating oocyte viability after vitrification-warming. To clarify the role of mitophagy in mitochondrial extinction of vitrified porcine oocytes, mitochondrial function, ultrastructural characteristics, mitochondria-lysosomes colocalization, and mitophagic proteins were detected with or without chloroquine (CQ) treatment. The results showed that vitrification caused mitochondrial dysfunction, including increasing reactive oxygen species production, decreasing mitochondrial membrane potential, and mitochondrial DNA copy number. Damaged mitochondrial cristae and mitophagosomes were observed in vitrified oocytes. A highly fused fluorescence distribution of mitochondria and lysosomes was also observed. In the detection of mitophagic flux, mitophagy was demonstrated as increasing fluorescence aggregation of microtubule-associated protein light chain 3B (LC3B), enhanced colocalization between LC3B, and voltage-dependent anion channels 1 (VDAC1), and upregulated LC3B-II/I protein expression ratio. CQ inhibited the degradation of mitophagosomes in vitrified oocytes, manifested as decreased mitochondria-lysosomes colocalization, increased fluorescence fraction of VDAC1 overlapping LC3B, increased LC3B-II/I protein expression ratio, and p62 accumulation. The inhibition of mitophagosomes degradation by CQ aggravated mitochondrial dysfunction, including increased oxidative damage, reduced mitochondrial function, and further led to loss of oocyte viability and developmental potentiality. In conclusion, mitophagy is involved in the regulation of mitochondrial function during porcine oocyte vitrification.
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Mitofagia , Oocitos/fisiología , Vitrificación , Animales , Cloroquina/farmacología , Cloroquina/toxicidad , Criopreservación/métodos , Desarrollo Embrionario/efectos de los fármacos , Femenino , Lisosomas/efectos de los fármacos , Lisosomas/ultraestructura , Microscopía Confocal , Microscopía Electrónica de Transmisión , Proteínas Asociadas a Microtúbulos/análisis , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/ultraestructura , Mitofagia/efectos de los fármacos , Oocitos/efectos de los fármacos , Oocitos/ultraestructura , Fagosomas/efectos de los fármacos , Fagosomas/ultraestructura , Preservación Biológica/métodos , Especies Reactivas de Oxígeno/metabolismo , Porcinos , Canal Aniónico 1 Dependiente del Voltaje/análisisRESUMEN
Hu sheep is a valuable sheep breed in China, and semen cryopreservation of Hu sheep is important for sustainable development of the agri-food industry. This study aimed to evaluate the effect of thawing rate and antioxidants (procyanidins [PC] and mitoquinone [MitoQ]) on the quality and antioxidant enzyme activity of post-thaw sperm in Hu sheep. Our results showed that the highest sperm quality was obtained from the group thawed at 70°C for 5 seconds. Furthermore, addition of 150 nM MitoQ in the extender could enhance motility, integrity of the membrane and acrosome, and mitochondrial activity, whereas only sperm motility and membrane integrity were increased with 10 µg/mL of PC supplementation, compared with the control group. Meanwhile, both PC (10 µg/mL) and MitoQ (150 nM) supplementation increased the levels of superoxide dismutase and glutathione peroxidase and decreased the levels of reactive oxygen species and malondialdehyde. In conclusion, the optimal thawing protocol of semen cryopreservation in Hu sheep was 70°C for 5 seconds. MitoQ supplementation (150 mM) in the extender could improve sperm quality and reduce the level of oxidative stress in Hu sheep semen after cryopreservation. Further studies are needed to evaluate fertility of the post-thaw semen using MitoQ.
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Preservación de Semen , Animales , Antioxidantes , China , Crioprotectores , Masculino , Semen , Análisis de Semen , Ovinos , Motilidad Espermática , EspermatozoidesRESUMEN
Oxidative stress can impede maturation of the nucleus and cytoplasm of oocytes during in vitro maturation (IVM). Rhodiola sachalinensis, an herb commonly used in traditional Chinese medicine, conveys antioxidative effects to cryopreserved bovine sperm. Therefore, the aims of this study were to evaluate the effects of different concentrations of R. sachalinensis aqueous extract (RSAE) on IVM and subsequent in vitro embryonic development after parthenogenetic activation (PA), in vitro fertilization (IVF), and somatic cell nuclear transfer (SCNT). The results showed that RSAE supplementation (6 and 60 mg/L) significantly increased intracellular glutathione levels, but had no effect on maturation rates or reactive oxygen species. After in vitro culture, greater blastocyst formation was observed in PA embryos (6 mg/L RSAE), as well as in IVF and SCNT embryos (60 mg/L) matured in RSAE-supplemented IVM media. In conclusion, although there was no significant improvement in the maturation rate, RSAE supplementation conveyed an antioxidative effect during IVM, and improved subsequent embryonic development in vitro. Further studies are needed to explore gene expression pattern in oocytes and embryos treated with RSAE.
Asunto(s)
Antioxidantes/farmacología , Embrión de Mamíferos/citología , Desarrollo Embrionario/efectos de los fármacos , Técnicas de Maduración In Vitro de los Oocitos/métodos , Oocitos/citología , Extractos Vegetales/farmacología , Rhodiola/química , Animales , Embrión de Mamíferos/efectos de los fármacos , Femenino , Fertilización In Vitro , Glutatión/metabolismo , Técnicas In Vitro , Oocitos/efectos de los fármacos , Partenogénesis , Especies Reactivas de Oxígeno/metabolismo , PorcinosRESUMEN
The aim of this study was to evaluate the interaction of different concentrations of butylated hydroxytoluene (BHT) in a tris-based extender on semen quality parameters in post-thawed dog semen. Twenty-four ejaculates were collected from eight male Beagle dogs using an artificial vagina. Pooled semen was diluted with a tris-based extender supplemented with 0 (control), 0.5, 1.0, 1.5, and 2.0 mM BHT, at a final concentration of 200 × 106 spermatozoa/mL. After thawing, sperm samples were assessed for motility parameters (CASA), membrane integrity (SYBR-14/PI), acrosome integrity (FITC-PNA), mitochondrial activity (JC-1/PI), malondialdehyde (MDA) concentration, and glutathione peroxidase (GPx) activity. The total motility, progressive motility, and average path velocity of the frozen-thawed sperm were significantly higher in the BHT1.5 group than in the control and the other sample groups (P < 0.05). Higher values of straight-line velocity, curvilinear velocity, amplitude of the lateral head displacement, and linearity were observed in the BHT1.0, BHT1.5, and BHT2.0 groups than in the control (P < 0.05). The BHT1.0 and BHT1.5 groups had higher percentages of straightness and acrosome integrity than the other groups (P < 0.05). Beat cross frequency, plasma membrane integrity, and GPx activity of the BHT1.5 and BHT2.0 groups were higher than those of the control (P < 0.05). A lower concentration of MDA was observed in the BHT1.0, BHT1.5, and BHT2.0 groups than in the control (BHT0) (P < 0.05). Our results indicate that 1.5 mM BHT is the optimal concentration for improving the post-thaw quality of canine spermatozoa.
Asunto(s)
Hidroxitolueno Butilado , Preservación de Semen , Acrosoma , Animales , Hidroxitolueno Butilado/farmacología , Criopreservación/métodos , Crioprotectores/farmacología , Perros , Masculino , Análisis de Semen , Preservación de Semen/veterinaria , Motilidad Espermática , EspermatozoidesRESUMEN
T-2 toxin, one of the most toxic mycotoxins, is commonly presented along with its metabolites, HT-2 toxin, neosolaniol (NEO), T-2 triol, and T-2 tetraol in foodstuff and feed. The aim of this study was to evaluate the cytotoxic effects of T-2 toxin alone and in combination with its metabolites on porcine Leydig cells. Based on the determination of cell viability with CCK-8, toxicological interactions were investigated using Combination Index method. The cytotoxic potency of five tested mycotoxins individual and their mixtures all showed with a dose-dependent manner. In view of IC50 values, the decreasing cytotoxicity of mycotoxins was ranking: T-2 toxin > HT-2 toxin > T-2 triol > NEO > T-2 tetraol. Combinations of T-2+HT-2, T-2+NEO, and HT-2+NEO displayed synergism at low doses but antagonism at high doses, while the ternary combination of T-2+HT-2+NEO revealed adverse situation from antagonism to synergism. All binary and ternary combinations of T-2 toxin, T-2 triol, and T-2 tetraol exhibited antagonistic interactions. Our results suggest that the co-occurrence of T-2 toxin and its metabolites might pose a slight threat to reproductive health due to antagonistic interactions. However, the synergy observed should be not ignored especially at low doses of mycotoxins co-occurrence in the diet.