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The objective of this study was to uncover the genetic background of wool quality, a production trait, by estimating genomic heritability and implementing GWAS in Akkaraman sheep. The wool characteristics measured included fibre diameter (FD) and staple length (SL) at the age of 8 months and yearling fibre diameter (YFD), yearling staple length (YSL) and yearling greasy fleece weight (YGFW) at 18 months of age. Animals were genotyped using the Axiom 50 K Ovine Genotyping Array. Maximum likelihood estimations of a linear mixed model (LMM) were used to estimate genomic heritability, where GWAS was conducted following a score test of each trait. Genomic heritability estimates for the traits ranged between 0.22 and 0.63, indicating that phenotypes have a moderate range of heritability. One genome- and six chromosome-wide significant SNPs were associated with the wool traits in Akkaraman lambs. Accordingly, TRIM2, MND1, TLR2, RNF175, CEP290, TMTC3, RERE, SLC45A1, SOX2, MORN1, SKI, FAAP20, PRKCZ, GABRD, CFAP74, CALML6 and TMEM52 genes as well as nine uncharacterized regions (LOC101118971, LOC105609137, LOC105603067, LOC101122892, LOC106991694, LOC106991467, LOC106991455, LOC105616534 and LOC105609719) were defined as plausible candidates. The findings of this study shed light on the genetics of wool quality and yield for the Akkaraman breed and suggests targets for breeders during systematic breeding programmes.

© 2022 by the authors.In the current study, the genetic architecture of growth and linear type traits were investigated in Akkaraman sheep. Estimations of genomic heritability, genetic correlations, and phenotypic correlations were implemented for 17 growth and linear type traits of 473 Akkaraman lambs by the univariate and multivariate analysis of animal mixed models. Correspondingly, moderate heritability estimates, as well as high and positive genetic/phenotypic correlations were found between growth and type traits. On the other hand, 2 genome-wide and 19 chromosome-wide significant single nucleotide polymorphisms were found to be associated with the traits as a result of animal mixed model-based genome-wide association analyses. Accordingly, we propose several genes located on different chromosomes (e.g., PRDM2, PTGDR, PTPRG, KCND2, ZNF260, CPE, GRID2, SCD5, SPIDR, ZNF407, HCN3, TMEM50A, FKBP1A, TLE4, SP1, SLC44A1, and MYOM3) as putative quantitative trait loci for the 22 growth and linear type traits studied. In our study, specific genes (e.g., TLE4, PTGDR, and SCD5) were found common between the traits studied, suggesting an interplay between the genetic backgrounds of these traits. The fact that four of the proposed genes (TLE4, MYOM3, SLC44A1, and TMEM50A) are located on sheep chromosome 2 confirms the importance of these genomic regions for growth and morphological structure in sheep. The results of our study are therefore of great importance for the development of efficient selection indices and marker-assisted selection programs, as well as for the understanding of the genetic architecture of growth and linear traits in sheep.

The aim of this study was to estimate genomic heritability and the impact that genetic backgrounds have on blood parameters in Akkaraman sheep by conducting genome-wide association studies and regional heritability mapping analysis. Genomic heritability estimates for blood parameters ranged from 0.00 to 0.55, indicating that measured phenotypes have a low to moderate heritability. A total of 7 genome- and 13 chromosome-wide significant SNPs were associated with phenotypic changes in 15 blood parameters tested. Accordingly, SCN7A, SCN9A, MYADM-like, CCDC67, ITGA9, MGAT5, SLC19A1, AMPH, NTRK2, MSRA, SLC35F3, SIRT6, CREB3L3, and NAV3 genes as well as three undefined regions (LOC101117887, LOC106991526 and LOC105608461) were suggested as candidates. Most of the identified genes were involved in basic biological processes that are essential to immune system function and cellular growth; specific functions include cellular transport, histone deacetylation, cell differentiation, erythropoiesis, and endocytosis. The top significant SNP for HCT, MCH, and MCHC was found within a genomic region mainly populated by the MYADM-like gene family. This region was previously suggested to be under historical selection pressure in many sheep breeds from various parts of the world. These results have implications on animal breeding program studies due to the effect that the genetic background has on blood parameters, which underlying many productive and wellness related traits.

© 2020 Taylor & Francis Group, LLC.This study was conducted to ivnestigate the associations of GH-AluI, STAT5A-AvaI and MYF5-TaqI gene polymorphisms with milk somatic cell count (SCC), electrical conductivity (EC) and pH levels in Holstein dairy cows. For this purpose, 167 blood and 1670 milk samples of 167 Holstein cows in their 2nd lactation were used. There were significant relationships between GH-AluI genotypes and milk EC (p < 0.001) and between STAT5A-AvaI genotypes and milk EC (p = 0.007), but there were not any significant relationships between MYF5 gene polymorphism and the investigated traits (p > 0.05). The greatest EC values were observed in GH-AluI-LV and STAT5A-AvaI-TT-genotyped individuals. Just because of association of EC with mastitis, it was concluded that present GH-AluI and STAT5A-AvaI polymorphisms could be used in further studies to be conducted to improve mastitis resistance and milk quality traits of Holstein dairy cows.

Use of genomic information in ruminant production systems can help alleviating concerns related to food security and sustainability of production. Nutritional genomics (i.e., nutrigenomics) is the field of research that is interested in all types of reciprocal interactions between nutrients and the genomes of organisms, i.e., variable patterns of gene expression and effect of genetic variations on the nutritional environment. Devising a revolutionary analytical approach to the traditional ruminant nutrition research, the relatively novel area of ruminant nutrigenomics has several studies concerning different aspects of animal production systems. This paper aims to review the current nutrigenomics research in the frame of how nutrition of ruminants can be modified accounting for individual genetic backgrounds and gene/diet relationships behind productivity, quality, efficiency, disease resistance, fertility, and greenhouse gas emissions. Furthermore, current challenges facing ruminant nutrigenomics are evaluated and future directions for the novel area are strongly argued by this review.

Simple Summary Epigenetic modifications of the genes regulate the inflammation process that includes the DNA methylation and histone acetylation. Sulforaphane is well known for its immunomodulatory properties. Notably, the mechanism of its anti-inflammatory functions involving epigenetic modifications is unclear. This study highlighted the regulatory mechanism of sulforaphane in the innate immunity responses in an acute inflammatory state employ in vivo cell culture model. Porcine monocyte-derived dendritic cells were exposed to LPS with or without sulforaphane pre-treatment for these purposes. Epigenetics modulations of the important genes and regulatory factors were studies as well as the immune responses of the cells were vigorously studied over the period of time. This study deciphers the mechanism of SFN in restricting the excessive inflammatory reactions, thereby, exerting its protective and anti-inflammatory function though epigenetic mechanism. Abstract Inflammation is regulated by epigenetic modifications, including DNA methylation and histone acetylation. Sulforaphane (SFN), a histone deacetylase (HDAC) inhibitor, is also a potent immunomodulatory agent, but its anti-inflammatory functions through epigenetic modifications remain unclear. Therefore, this study aimed to investigate the epigenetic effects of SFN in maintaining the immunomodulatory homeostasis of innate immunity during acute inflammation. For this purpose, SFN-induced epigenetic changes and expression levels of immune-related genes in response to lipopolysaccharide (LPS) stimulation of monocyte-derived dendritic cells (moDCs) were analyzed. These results demonstrated that SFN inhibited HDAC activity and caused histone H3 and H4 acetylation. SFN treatment also induced DNA demethylation in the promoter region of the MHC-SLA1 gene, resulting in the upregulation of Toll-like receptor 4 (TLR4), MHC-SLA1, and inflammatory cytokines’ expression at 6 h of LPS stimulation. Moreover, the protein levels of cytokines in the cell culture supernatants were significantly inhibited by SFN pre-treatment followed by LPS stimulation in a time-dependent manner, suggesting that inhibition of HDAC activity and DNA methylation by SFN may restrict the excessive inflammatory cytokine availability in the extracellular environment. We postulate that SFN may exert a protective and anti-inflammatory function by epigenetically influencing signaling pathways in experimental conditions employing porcine moDCs.

The objective of this study was to determine fattening performance data for Charolais, Limousin and Blonde d’Aquitaine beef cattle and associate these data with NPR2 gene 8:g.59961937 T>C (rs208158047) mutation. Experiments were conducted with 176 beef cattle (77 Charolais, 66 Limousin and 33 Blonde d’Aquitaine) at nine months of age. Experiments lasted for 9 months and animals were slaughtered at the age of 18 months. Cattle body weights were determined at four different periods: beginning of fattening (d0), 60th day of fattening (d60), 120th day of fattening (d120) and at the end of fattening (sw). In terms of rs208158047 mutation of Charolais, Limousin and Blonde d’Aquitaine breeds, TT and CT genotypes were identified, and CC genotype was not encountered. The association of average daily gain (ADG) in d0-d60, d0-d120 and d0-sw periods with the genotypes of rs208158047 mutation was found to be significant (Prs208158047-CT genotypes compared to rs208158047-TT genotypes. These results indicate that the selection of bovine NPR2 gene could be used to ensure the breeding direction for growth related traits of the beef cattle.

Signature of selection studies have identified many genomic regions with known functional importance and some without verified functional roles. Multiple studies have identified Transmembrane protein 8B (TMEM8B)rs426272889 as having been recently under extreme selection pressure in domesticated sheep, but no study has provided sheep phenotypic data clarifying a reason for extreme selection. We tested rs426272889 for production trait association in 770 U.S. Rambouillet, Targhee, Polypay, and Suffolk sheep. TMEM8Brs426272889 was associated with mature weight at 3 and 4 years (p &lt, 0.05). This suggested selection for sheep growth and body size might explain the historical extreme selection pressure in this genomic region. We also tested Sperm-associated antigen 8 (SPAG8) rs160159557 encoding a G493C substitution. While this variant was associated with mature weights at ages 3 and 4, it was not as strongly associated as TMEM8Brs426272889. Transmembrane protein 8B has little functional information except as an inhibitor of cancer cell proliferation. To our knowledge, this is the first study linking TMEM8B to whole organism growth and body size under standard conditions. Additional work will be necessary to identify the underlying functional variant(s). Once identified, such variants could be used to improve sheep production through selective breeding.

Maternal nutrition during pregnancy is one of the major intrauterine environmental factors that influence fetal development by significantly altering the expression of genes that might have a consequence on the physiological, morphological, and metabolic performance of the offspring in the postnatal period. The impact of maternal dietary protein on the expression of genes in sheep fetal skeletal muscle development is not well understood. The current study aims to investigate the impact of high and low maternal dietary protein on the holistic mRNA expression in the sheep fetal skeletal muscle. Dams were exposed to an isoenergetic high-protein diet (HP, 160-270 g/day), low-protein diet (LP, 73-112 g/day), and standard protein (SP, 119-198 g/day) diets during pregnancy. Fetal skeletal muscles were obtained at the 105th day of pregnancy and mRNA expression profiles were evaluated using Affymetrix GeneChip (TM) Ovine Gene 1.0 ST Array. The transcriptional analysis revealed a total of 323, 354, and 14 genes were differentially regulated (fold change > 2 and false discovery rate

This study was conducted to determine LEP-Sau3AI, SCD-Fnu4HI, NR1H3-HpyCH4IV and FABP4-HinII gene polymorphisms and to investigate the association between these SNPs and somatic cell count (SCC), electrical conductivity (EC) and pH in Holstein cow milk. LEP-Sau3AI polymorphism found associated with SCC (p < 0.01), EC (p < 0.01) and pH (p < 0.05). LEP-Sau3AI-BB genotype resulted with higher SCC, EC and pH compared to other genotypes. SCD-Fnu4HI polymorphism showed differences in genotypes for EC (p < 0.05) and pH (p

Paratuberculosis (pTB), also known as Johne's disease (JD), is a contagious, chronic, and granulomatous inflammatory disease of the intestines of ruminants which is caused by Mycobacterium avium subsp. paratuberculosis (MAP) infection, resulting in billions of dollars in economic losses worldwide. Since, currently, no effective cure is available for MAP infection, it is important to explore the genetic variants that affect the host MAP susceptibility. The aim of this study was to analyze a potential association between EDN2 synonymous gene mutations (rs110287192, rs109651404 and rs136707411), that modifies susceptibility to pTB. EDN2 rs110287192, rs109651404 and rs136707411 mutations were genotyped in 68 infected and 753 healthy animals from East Anatolian Red crossbred, Anatolian Black crossbred and Holstein breed cattle by using Custom TaqMan SNP Genotyping Assays. For pTB status, serum antibody levels S/P ≥ 1.0 were assessed in carriers of the different EDN2 genotypes. EDN2 rs110287192 mutation showed a significant association with bovine pTB (adj. p < 0.05). For rs110287192 locus, the odd ratios for GG and TG genotypes versus TT genotypes were 1.73; (95% CI = 0.34–8.59) and 0.53 (95% CI = 0.12–2.37) respectively, which indicated that proportion of TG heterozygotes were significantly higher in control animals as compared to pTB animals. On the other hand, while rs136707411 mutation showed a suggestive association with pTB status in the examined cattle population (nominal p < 0.05); no association was detected between rs109651404 genotypes and pTB status. Selecting animals against rs110287192-GG genotype may decrease the risk of pTB in cattle of the Bos taurus taurus subspecies.

Sheep are economically important worldwide, and growth-associated genetic variants, such as a TNXB charged amino acid substitution E2004G and a silent DGAT1 single nucleotide polymorphism (rs409119650), could improve sheep profitability. However, both were identified in single reports using small groups of sheep from outside the U.S. We evaluated 896 U.S. sheep from one location to investigate an association of TNXB E2004G and DGAT1 rs409119650 with growth and lifetime production. For TNXB E2004G, glutamic acid homozygotes had greater body weights in spring and fall at ages 3 and 4 (all P ≤ 0.05) and greater milk scores at ages 3 and 4 (P

This study was conducted to identify gene polymorphisms and to investigate the relationships ofpolymorphisms with yield and composition of milk from Holstein cows. Significant relationships ofgrowth hormone (GH), leptin (LEP) and myogenic factor 5 (MYF5) gene polymorphisms with 305-day milk yield were observed (P < 0.01). Signal transducers and activators of transcription 5A(STAT5A) gene polymorphism did not show any significant relationships with investigated traits(P > 0.05). With regard to the 305-day milk yield, GH-LV, LEP-AA and AB and MYF5-GG genotypes were found to be superior over the other genotypes. It was concluded based on present findings that GH, LEP and MYF5 genes could be used as candidate genes for milk yield of Holsteincows.

Scrapie is a transmissible spongiform encephalopathy of sheep and goats, and scrapie eradication programs in many parts of the world rely on strong genetic resistance to classical scrapie in sheep. However, the utility of putative resistance alleles in goats has been a focus of research because goats can transmit scrapie to sheep and may serve as a scrapie reservoir. Prior work showed that disease-free survival time was significantly extended in orally inoculated goats singly heterozygous for prion amino acid substitutions S146 or K222, but average durations were only around 3 years post-inoculation. The aim of this study was to investigate whether extended survival would exceed 6 years, which represents the productive lifetimes of most commercial goats. While all control homozygotes were clinically affected by an average of

In order to identify the effects of nutrients on gene expression and to assess the interactions between genes and nutrition by means of various cutting-edge technologies, the interdisciplinary branch ‘Nutrigenomics’ was created. Therefore, nutrigenomics corresponds to the use of knowledge and techniques of nutrition, genomics, transcriptomics, proteomics, epigenomics, and metabolomics to seek and explain the cross-talk between nutrition and genes in molecular level. Macronutrients are important dietary signals that control metabolic programming of cells and have important roles in maintaining cellular homeostasis by influencing specific gene expression. Recent advancements in molecular genetics studies, for instance, use of next-generation sequencing, microarray and qPCR array to investigate the expression of transcripts, genes, and miRNAs, has a crucial impact on understanding and quantitative measurement of the impact of dietary macronutrients on gene function. This review will shade a light on the interactions and mechanisms how the dietary source of macronutrients changes the expression of specific mRNA and miRNA. Furthermore, it will highlight the exciting recent findings in relation to animal performance characteristics which eventually help us to identify a dietary target to improve animal production.

Background: The major histocompatibility complex (MHC) is an organized cluster of tightly linked vertebrate genes with immunological and non-immunological functions. While the important MHC gene DRB1 has been examined in regard to many sheep infectious disease traits, only one study, based on microsatellite markers, has previously examined DRB1 and sheep production traits. Furthermore, to our knowledge no studies have examined DRB1 relationship with lifetime ewe prolificacy traits. Therefore, we analyzed association between the presence of DRB1 SNP haplotypes with internationally recognized standard names and production traits including growth and lifetime prolificacy in 370 Rambouillet, Columbia, and Polypay sheep., BackgroundThe major histocompatibility complex (MHC) is an organized cluster of tightly linked vertebrate genes with immunological and non-immunological functions. While the important MHC geneDRB1has been examined in regard to many sheep infectious disease traits, only one study, based on microsatellite markers, has previously examinedDRB1and sheep production traits. Furthermore, to our knowledge no studies have examinedDRB1relationship with lifetime ewe prolificacy traits. Therefore, we analyzed association between the presence ofDRB1SNP haplotypes with internationally recognized standard names and production traits including growth and lifetime prolificacy in 370 Rambouillet, Columbia, and Polypay sheep.ResultsTheDRB1*0404 haplotype was associated with increased weaning and mature weights, as well as average daily gain (ŠidákPP=0.084) and number of lambs born alive (ŠidákP=0.084). In contrast, theDRB1*0141 haplotype was associated with decreased mature weight (ŠidákP=0.01).ConclusionsSince the *0404 haplotype was present in all three breeds, these results suggest there is at least one functional mutation in the region that influences growth and prolificacy traits that may be broadly present across several breeds. Furthermore, combined use of the similar *0404 and *0141 multi-marker haplotypes that nonetheless have opposing directions of production trait associations will enhance mutation discovery in this region. If undesirable alleles for underlying mutations can be identified, selective pressure against one or a small number of undesirable alleles may improve production with limited impact on MHC genetic diversity and infectious disease susceptibility.

Scrapie is the transmissible spongiform encephalopathy (TSE) of sheep and goats, and it can be transmitted in both directions between these species. Scrapie eradication programs worldwide have utilized strong genetic resistance to classical scrapie in sheep, but such resistance has not been available for goats until recently. Both S146 (serine at position 146) and K222 (lysine at position 222) amino acid substitutions in the prion protein have been dramatically underrepresented in natural scrapie cases, but exposure times are not known in most natural situations. Therefore, we used a goat classical scrapie homogenate to orally challenge: 1) 5 goat kids with control genotypes [two copies of the most common haplotype], 2) 8 kids with one copy of S146 and one copy of the control haplotype, and 3) 8 kids with one copy of S146 and one copy of the control haplotype. All control goats developed classical scrapie by an average age of 2 years, but there has been no confirmed scrapie in the S146 group by an average age of 7.8 years, or in the K222 group by an average age of 6.8 years (both P

Simple Summary Disease resistance is the ability of animals to inhibit the growth of invading pathogens within the body, which is influenced by the interaction of the host immune system, host genetics, and the pathogens. Resistant animals can be produced by molecular breeding by introducing the genomic marker responsible for disease resistance or immunocompetence. Immunogenomics is an information science that enables the genome-scale investigation of host immune response to pathogenic infection thereby identification of the genomic marker for disease resistance. Once the genomic marker is determined, it could be implemented in producing disease resistance animals by applying the advanced reproductive biotechnology like genome editing. The technical ease and decreasing cost over time might enhance the application of genome editing techniques for producing disease resistance livestock. Abstract Disease occurrence adversely affects livestock production and animal welfare, and have an impact on both human health and public perception of food–animals production. Combined efforts from farmers, animal scientists, and veterinarians have been continuing to explore the effective disease control approaches for the production of safe animal-originated food. Implementing the immunogenomics, along with genome editing technology, has been considering as the key approach for safe food–animal production through the improvement of the host genetic resistance. Next-generation sequencing, as a cutting-edge technique, enables the production of high throughput transcriptomic and genomic profiles resulted from host-pathogen interactions. Immunogenomics combine the transcriptomic and genomic data that links to host resistance to disease, and predict the potential candidate genes and their genomic locations. Genome editing, which involves insertion, deletion, or modification of one or more genes in the DNA sequence, is advancing rapidly and may be poised to become a commercial reality faster than it has thought. The clustered regulatory interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) [CRISPR/Cas9] system has recently emerged as a powerful tool for genome editing in agricultural food production including livestock disease management. CRISPR/Cas9 mediated insertion of NRAMP1 gene for producing tuberculosis resistant cattle, and deletion of CD163 gene for producing porcine reproductive and respiratory syndrome (PRRS) resistant pigs are two groundbreaking applications of genome editing in livestock. In this review, we have highlighted the technological advances of livestock immunogenomics and the principles and scopes of application of CRISPR/Cas9-mediated targeted genome editing in animal breeding for disease resistance.

The primary objective of the present study was to assess the effects of vitamin and mineral premix (VMP) withdrawal from the diets 30 and 60 days ahead of slaughter on carcass and meat quality of Holstein Friesian steers. A total of 45 animals at 16 to 17 months of age were used and the selected animals were divided into three experimental groups: control group (fed with a diet with VMP), VMP withdrawal 30 days ahead of slaughter (VMP30 group), and VMP withdrawal 60 days ahead of slaughter (VMP60 group). Meat samples were taken at 24 h postmortem from the 13th rib section and meat quality was evaluated on the Longissimus dorsi thoracis (LT) muscle. After slaughter, carcass yield and meat drip loss, cooking loss, thawing loss, and shear force traits were determined. Meat pH and color parameters were measured at 24, 48, 72, 96, 120, and 144 h of postmortem. The fatty acid composition in 13th rib section' adipose tissue was determined. The hot and cold carcass weights, carcass yield and chilling loss were not affected by the withdrawal of VMP from the diet. Withdrawal of VMP from the diets 30 and 60 days ahead of slaughter did not have any significant effects on ultimate pH, drip loss, cooking loss, thawing loss, shear force, and meat color. Additionally, dry matter, crude protein, ash, fat contents, moisture-protein ratio of the meat samples, and fatty acid profiles were not affected by VMP30 and VMP60 treatments. It was concluded based on present finding VMP could be withdrawn safely from the diets 30 and 60 days ahead of slaughter without any negative effects on carcass and meat quality traits of feedlot steers. Withdrawal of VMP may reduce feeding costs and environmental damages generated by animal breeding systems.

Despite its essential role in ovulation, oxidative stress (OS) has been found to be cytotoxic to cells, while microRNAs (miRNAs) are known as a major regulator of genes involved in cellular defense against cytotoxicity. However, a functional link between OS and miRNA expression changes in granulosa cells (GCs) remains to be investigated. Here, we investigate the OS modulation of apoptosis-associated miRNAs and their biological relevance in bovine GCs. Following the evaluation of cell viability, accumulation of reactive oxygen species (ROS), cytotoxicity and mitochondrial activity, we used a ready-to-use miRNA PCR array to identify differentially regulated miRNAs. The results showed that exposure to 150 mu M H2O2 for 4 h creates remarkable signs of OS in GCs characterized by more than 50% loss of cell viability, higher nuclear factor erythroid 2-related factor 2 (NRF2) nuclear translocation, significantly (p < 0.05) higher abundance of antioxidant genes, significantly (p < 0.001) higher accumulation of ROS, lower mitochondrial activity and a higher (p < 0.001) number of apoptotic nuclei compared to that of the control group. miRNA expression analysis revealed that a total of 69 miRNAs were differentially regulated in which 47 and 22 miRNAs were up- and downregulated, respectively, in stressed GCs. By applying the 2-fold and p < 0.05 criteria, we found 16 miRNAs were upregulated and 10 miRNAs were downregulated. Target prediction revealed that up- and downregulated miRNAs potentially targeted a total of 6210 and 3575 genes, respectively. Pathway analysis showed that upregulated miRNAs are targeting the genes involved mostly in cell survival, intracellular communication and homeostasis, cellular migration and growth control and disease pathways. Our results showed that OS modulates the expression of apoptosis-associated miRNAs that might have effects on cellular or molecular damages.

An association study between polymorphisms of six genes and boar taint related compounds androstenone, skatole and indole was performed in a boar population (n = 370). Significant association (P < 0.05) was detected for SNP of FMO5 (g.494A>G) with all boar taint compounds, SNP of CYP21 (g.3911T>C) with skatole and indole, and SNP of ESR1 (g.672C>T) with androstenone and indole. mRNA expression of CYP21 and ESR1 was higher in CAB (castrated boar) compared to non-castrated boars; whereas, the expression of FMO5 and ESR1 was higher in LBT (low boar taint) compared to HBT (high boar taint) in liver tissue. FMO5, CYP21 and ESR1 proteins were less detectable in HBT compared with LBT and CAB in liver tissues. These findings suggest that FMO5, CYP21 and ESR1 gene variants might have effects on the boar taint compounds. (C) 2015 Elsevier Ltd. All rights reserved.

The aim of the study was to investigate single nucleotide polymorphisms (SNPs) and expression of SOX-6 to support its candidacy for growth, carcass, and meat quality traits in pigs. The first SNP, rs81358375, was associated with pH 45 min post mortem in loin (pH1L), the thickness of backfat and side fat, and carcass length in Pietrain (Pi) population, and related with backfat thickness and daily gain in Duroc × Pietrain F2 (DuPi) population. The other SNP, rs321666676, was associated with meat colour in Pi population. In DuPi population, the protein, not mRNA, level of SOX-6 in high pH1L pigs was significantly less abundant compared with low pH1L pigs, where microRNAs targeting SOX-6 were also differently regulated. This paper shows that SOX-6 could be a potential candidate gene for porcine growth, carcass, and meat quality traits based on genetic association and gene expression.

The objective of this study was to identify allele and genotype frequencies of the κ-CN and β-LG genes in Anatolian water buffalo. A total of 126 water buffalos from Turkey were genotyped using the PCR-RFLP method. For gene κ-CN, only B allele and BB genotype were observed. And for gene β-LG, two types of alleles (A and B) and three types of genotypes were observed. The genotype frequencies of AA, AB and BB of β-LG in Anatolian water buffalo were 0.254, 0.698 and 0.048, respectively. Surprisingly, the frequency of allele A was higher than that of allele B in contrast to world buffalo breeds.

Follicular development in the mammalian ovary is a complex process which is potentially regulated by an orchestrated action of the pituitary gonadotropins, e.g., follicle stimulating hormone (FSH) and luteinizing hormone (LH), and local ovarian factors, such as peptide growth factors and steroids. Along with hormonal activation it is necessary to have tight coordination of expression of genes during follicular development. This review highlights the structure and function of ovary, follicle and follicular development. This review also confirms the temporal and spatial expression of the specific genes and miRNAs and their involvement in different modulators, the synthesis of active factors, their interactions, and the dynamics of their receptors on the follicular cell surface may be the ultimate determinants of cellular events which are crucial to coordinated growth and differentiation of follicular cells leading to folliculogenesis and ovulation. DOI: http://dx.doi.org/10.3329/ralf.v1i1.22355 Res. Agric., Livest. Fish.1(1): 47-60, Dec 2014

The objective of this study was to examine the allelic and genotypic profiles of four SNPs of bovine Toll-Like receptor 1 (TLR1), Toll-Like Receptor 4 (TLR4) and Solute Linkete Carier 11A1 (SLC11A1) genes in Holstein, two Turkish native breeds and their crossbreds in Turkey. For this purpose, a total of 1023 cattle from Holstein (HL, n= 410), Anatolian Black (AB, n= 106), East Anatolian Red (EAR, n= 84), Anatolian Black Crossbreed (ABC, n= 124) and East Anatolian Red crossbreed (EARC, n= 299) breeds were examined for four SNPs. Samples were genotyped using by the PCR-RFLP method. According to the TLR1 (+1380) SNP, EAR was in Hardy-Weinberg equilibrium (HWE) while the AB, HL and native crossbreeds were deviated from HWE. All cattle breeds were in HWE for the TLR1 (+1596) SNP. In terms of TLR4 (+10) SNP, the CC genotype had the highest frequency in HL and native crossbreeds whereas the AB and EAR breeds were monomorphic and only CC genotype was found these breeds. According to the SLC11A1 (+1066) SNP, the CC genotype had the highest while the GG genotype frequency the lowest in all breeds. In addition EAR breed was in HWE while the AB, HL and native crossbreeds deviated from HWE.

Sulforaphane (SFN) has received a great deal of research attention because of its ability to induce the production of a battery of antioxidant enzymes in certain concentrations through the activation of the Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) signaling pathway, which may effectively neutralize reactive oxygen species (ROS) induced oxidative stress. This study was conducted to investigate the potential of different concentrations of SFN in inducing antioxidative and apoptotic effects in granulosa cells (GCs). For this purpose, bovine GCs were collected from preovulatory antral follicles and cultured with different concentrations of SFN (0-80 μM) and based on phenotypic evaluation three concentrations were selected: 2 μM (low), 10 μM (medium), and 20 μM (high) for further investigations. The results showed that there was a dramatic loss of cell viability and higher cytotoxic effects of SFN on GCs at higher concentrations (15 μM). The expression of NRF2 increased significantly (p 0.05) with fold change ranged 3-8 in SFN treated GCs, whereas Kelch Like ECH Associated Protein 1 (KEAP1) expression was either downregulated or similar as control group under the same conditions. Moreover, the relative expression of the genes (PRDX1, CAT, TXN1and SOD1) downstream to NRF2 activation was found to be highly expressed (fold change ranged from 2 to 5, p 0.05) in SFN treated GCs compared to the untreated control. In addition, ROS accumulation was higher in GCs treated with 20 μM SFN which in turn results in a higher accumulation of lipid droplets. Compared to control, no changes in the mitochondrial activity was observed at 2 and 10 μM SFN concentrations; however, significantly lower mitochondrial activity was found at high concentration (20 μM). The results of this study clearly showed that 10 μM SFN concentration played a crucial role in activating Nrf2 pathway without inducing apoptotic characteristics and this concentration may have beneficial effects in boosting the production of phase II antioxidant enzymes in GCs. However, at high concentration (20 μM), SFN may generate excessive ROS that causes mitochondrial dysfunction and induces cellular stress and eventually leads to apoptosis. These data strongly suggest a concentration dependent antioxidative and apoptotic effects of SFN on GCs.

The objective of this study was to identify allele and genotype frequencies of CSN3 and PRL genes in four cattle breeds in Turkey. For this purpose, a total of 390 cattle of East Anatolian Red (EAR), Zavot, Brown Swiss (BS) and Simmental (SIM) breeds were genotyped by PCR-RFLP method. A 443 bp fragment of CSN3 and a 156 bp fragment of PRL were amplified and digested with HindIII and RsaI restriction enzymes, respectively. For CSN3 and PRL genes, two types of alleles (A and B) and three types of genotypes (AA, BB, and AB) were observed. The highest frequencies for CSN3-A and CSN3-B alleles were estimated for the EAR breed (0.743) and for the BS breed (0.556), respectively. The highest frequency for PRL-A and PRL-B alleles was estimated for the SIM breed (0.801) and for the BS breed (0.315), respectively. The Chi-square test among the investigated cattle breeds showed that only the Zavot breed was in Hardy-Weinberg equilibrium (HWE) for both loci.

To evaluate associations of glucose-6-phosphate dehydrogenase (G6PDH) activity in sheep oocytes with cytoplasmic lipid content, maturational competence, developmental competence to the blastocyst stage, and gene expression of certain molecular markers., Purpose To evaluate associations of glucose-6-phosphate dehydrogenase (G6PDH)activity in sheep oocytes with cytoplasmic lipid content, maturational competence,developmental competence to the blastocyst stage, and gene expression of certainmolecular markers.Methods Before brilliant cresyl blue (BCB) staining test, oocytes were classified ashigh, middle, and low cytoplasmic lipid content (HCLC, MCLC, and LCLC) and after thetest as having low or high G6PDH-activity (BCB+ and BCB-, respectively). Aftermaturation in vitro, a group of oocytes were subjected to IVF followed by in vitroembryo culture and another group was used for evaluation of expression of candidategenes.Results The cleavage and blastosyst rates were lowest (P < 0.05) in LCLC group,intermediate (P < 0.05) in MCLC group and highest (P < 0.05) in HCLC group. More(P < 0.05) oocytes in HCLC group were BCB+, and higher (P < 0.05) maturation,cleavage, and blastocyst rates were seen for BCB+ oocytes than the BCB- oocytes.Our gene expression data indicated that mRNA transcript abundance of ITGB2, pZP3,BMP15, and GDF9 genes were similar between BCB oocytes groups. However, theexpression of ATP1A1 was higher (P < 0.05) for BCB+ oocytes compared to BCBoocytes.In addition, BAX transcript abundance was similar (P > 0.05) among BCB+,BCB-, and control groups, before and after maturation in vitro.Conclusion Activity of G6PDH in sheep oocytes is highly associated with lipid content,and compared with the morphological parameters, might be a more precise andobjective predictor for subsequent developmental competence in vitro.

Toll-like receptors (TLRs), known as the pathogen recognition receptors, recognize the microbial components. The details expression kinetics of TLRs may help to understand the immune responsiveness of pigs, but the expression patterns of all TLRs (TLR1–10) have not yet been studied in pigs. Therefore, this study unravels the expression patterns of the TLR family set of genes (TLR1–10) in different lymphoid tissues collected from pigs at different ages. A total of nine clinically healthy Pietrain pigs at three age groups were selected for this experiment. Each age group consisted of three animals. Cervical lymph node (CLN), thymus, liver, spleen, lung, heart, skin tissue and peripheral blood mononuclear cells (PBMC) were collected for both mRNA and protein isolation. The mRNA expression of TLRs (1−10) was quantified in all tissues. TLR3 mRNA was the most abundant in all tissues. It was expressed highly in thymus, kidney, lungs and liver tissues. Western blot and immunofluorescence was performed for protein expression and localization of selected TLRs (TLR2, 3 and 9) in selected tissues (CLN, spleen and lungs). The Western blot results of TLR2, 3 and 9 in selected tissues appeared to be consistent with the mRNA expression results. Cells in lungs, spleen and CLN were positively immunostained for TLR2, 3 and 9. This study sheds light on the expression patterns of TLR (1–10) genes in important lymphoid tissues in pigs of different ages. The heterogeneous expression of TLRs with ages may contribute to the altered immune responsiveness of pigs with ages.

The aim of the present study was to investigate the age-related production variation of T helper (Th)-type cytokines (IL-2, IL-4, IFN-gamma and IL-10), granulocyte macrophage-colony stimulating factor (GM-CSF) and nitric oxide (NO) by lipopolysaccharide (LPS)-stimulated porcine alveolar macrophages (AMs) in a time-dependent manner. For this purpose, AMs were isolated from 5-days (newborn), 40-days (post-weaned) and 120-days (young) old pigs. Cells were incubated for 24 h in the absence or presence of increasing concentrations of LPS (0.0, 0.01, 1.0, 5.0 and 10.0 mu g/mL)., The aim of the present study was to investigate the age-related production variation of T helper (Th)-type cytokines (IL-2, IL-4, IFN-γ and IL-10), granulocyte macrophage-colony stimulating factor (GM-CSF) and nitric oxide (NO) by lipopolysaccharide (LPS)-stimulated porcine alveolar macrophages (AMs) in a time-dependent manner. For this purpose, AMs were isolated from 5-days (newborn), 40-days (post-weaned) and 120-days (young) old pigs. Cells were incubated for 24 h in the absence or presence of increasing concentrations of LPS (0.0, 0.01, 1.0, 5.0 and 10.0 μg/mL).IL-10, IFN-γ and GM-CSF mRNA expression was upregulated in a dose-dependent manner for all age groups (P < 0.05). Age-related differences included a significantly increased IL-10 mRNA and protein production in newborn piglets compared to post-weaned and young pigs. IL-10 production pattern was similar with a higher peak between 12-36 h post-induction in all age groups. In contrast, IFN-γ mRNA and protein level was significantly elevated in young pigs 12 h and 24 h post-induction, respectively, while the time course production of IFN-γ was mostly consistent in newborn and post-weaned piglets. GM-CSF mRNA expression was significantly lower in newborn piglets than in post-weaned and young pigs. The kinetic GM-CSF expression peaked at 12 h in young and post-weaned pigs and at 24 h in newborn piglets. IL-4 mRNA levels were very low and no apparent change of IL-2 expression was observed following LPS stimulation in all age groups. Only very low levels of NO were detected in the cell supernatants of young pigs. Collectively, these studies suggest age-related differences in time-dependent production of IL-10, IFN-γ and GM-CSF by porcine AMs with potential immunoregulatory consequences to be explored further.

The aim was to study the expression pattern of the porcine TLR family (TLR1-10) genes in gut-associated lymphoid tissues (GALT) of varying ages. A total of nine clinically healthy pigs of three ages group (1 day, 2 months and 5 months old) were selected for this experiment (three pigs in each group). Tissues from intestinal mucosa in stomach, duodenum, jejunum and ileum and mesenteric lymph node (MLN) were used. mRNA expression of TLRs (1-10) was detectable in all tissues and TLR3 showed the highest mRNA abundance among TLRs. TLR3 expression in stomach, and TLR1 and TLR6 expression in MLN were higher in adult than newborn pigs. The western blot results of TLR2, 3 and 9 in some cases, did not coincide with the mRNA expression results. The protein localization of TLR2, 3 and 9 showed that TLR expressing cells were abundant in the lamina propria, Peyer's patches in intestine, and around and within the lymphoid follicles in the MLN. This expressions study sheds the first light on the expression patterns of all TLR genes in GALT at different ages of pigs. (c) 2013 Elsevier Ltd. All rights reserved., The aim was to study the expression pattern of the porcine TLR family (TLR1-10) genes in gut-associated lymphoid tissues (GALT) of varying ages. A total of nine clinically healthy pigs of three ages group (1day, 2months and 5months old) were selected for this experiment (three pigs in each group). Tissues from intestinal mucosa in stomach, duodenum, jejunum and ileum and mesenteric lymph node (MLN) were used. mRNA expression of TLRs (1-10) was detectable in all tissues and TLR3 showed the highest mRNA abundance among TLRs. TLR3 expression in stomach, and TLR1 and TLR6 expression in MLN were higher in adult than newborn pigs. The western blot results of TLR2, 3 and 9 in some cases, did not coincide with the mRNA expression results. The protein localization of TLR2, 3 and 9 showed that TLR expressing cells were abundant in the lamina propria, Peyer's patches in intestine, and around and within the lymphoid follicles in the MLN. This expressions study sheds the first light on the expression patterns of all TLR genes in GALT at different ages of pigs.

The objective of the present study was to investigate LPS and lipoteichoic acid (LTA)-induced TLRs, associated signaling molecules and inflammatory mediators, as well as to compare their combined effect in porcine alveolar macrophages. Macrophages were incubated for 24h with various concentrations of LPS, LTA, LPS+LTA or control. Multiple concentrations of LPS elicited marked up-regulation in mRNA for TLR2 and TLR4, CD14, MD2, MyD88, IRAK-4 and TRAF6 compared with the control. LTA had no effect on TLR4 and MD2; only higher doses up-regulated TLR2, CD14, MyD88, IRAK-4 and TRAF6 mRNA. LPS-activated cells released IL1-, IL12-, TNF-, IL-6, IL-8, IFN- and IL-10 in a dose-dependent manner, while LTA had no effect on IL-1, IL-6 and IFN-. Higher doses of LTA induced IL-12, TNF-, IL-8 and IL-10. Combined stimulation augmented TLR2, CD14 and MyD88 mRNA, and subsequently produced elevated levels of IL-6, TNF- and IL-8 when compared with LPS and LTA alone. Additionally, phagocytosis of macrophages was significantly increased following low concentration of LPS treatment. Only low levels of NO (nitric oxide) were detected in the LPS group. Overall, compared with LPS, LTA was a relatively weak inducer, and co-stimulation accelerated gene and cytokine production associated with pulmonary innate immune function., The objective of the present study was to investigate the lipopolysaccharide (LPS) and lipotechoic acid (LTA)-induced TLRs, associated signaling molecules and inflammatory mediators, as well as to compare their combined effect in porcine alveolar macrophages. Macrophages were incubated for 24 h with various concentrations of LPS, LTA, LPS plus LTA or control. Multiple concentrations of LPS elicited the marked up-regulation in mRNA for TLR-2 and-4, CD-14, MD-2, MyD-88, IRAK-4, TRAF-6, as compared to control. LTA had no effect on TLR-4 and MD-2; only higher doses up-regulated TLR-2, CD-14, MyD-88, IRAK-4 and TRAF-6 mRNA. LPS-activated cells released IL1-ß, IL12-ß, TNF-?, IL-6, IL-8, IFN-? and IL-10 in a dose-dependent manner while LTA had no effect on IL-1ß, IL-6 and IFN-?. Higher doses of LTA induced IL-12ß, TNF-?, IL-8 and IL-10. Combined stimulation augmented TLR-2, CD-14 and MyD-88 mRNA, and subsequently produced elevated levels of IL-6, TNF-? and IL-8 when compared to LPS and LTA alone. Additionally, phagocytosis of macrophages was significantly increased following low concentration of LPS treatment. Only low levels of nitric oxide were detected in LPS group. Overall, compared to LPS, LTA was a relatively weak inducer, and co-stimulation accelerated genes and cytokines production associated with pulmonary innate immune function.

Adipose-derived stem cells are an attractive alternative as a source of stem cells that can easily be extracted from adipose tissue. Isolation, characterization, and multi-lineage differentiation of adipose-derived stem cells have been described for human and a number of other species. Here we aimed to isolate and characterize camel adipose-derived stromal cell frequency and growth characteristics and assess their adipogenic, osteogenic, and chondrogenic differentiation potential. Samples were obtained from five adult dromedary camels. Fat from abdominal deposits were obtained from each camel and adipose-derived stem cells were isolated by enzymatic digestion as previously reported elsewhere for adipose tissue. Cultures were kept until confluency and subsequently were subjected to differentiation protocols to evaluate adipogenic, osteogenic, and chondrogenic potential. The morphology of resultant camel adipose-derived stem cells appeared to be spindle-shaped fibroblastic morphology, and these cells retained their biological properties during in vitro expansion with no sign of abnormality in karyotype. Under inductive conditions, primary adipose-derived stem cells maintained their lineage differentiation potential into adipogenic, osteogenic, and chondrogenic lineages during subsequent passages. Our observation showed that like human lipoaspirate, camel adipose tissue also contain multi-potent cells and may represent an important stem cell source both for veterinary cell therapy and preclinical studies as well.

The aim of the present study was to determine the age-related changes of phagocytic capacity and the kinetic production of cytokines in lipopolysaccharide-stimulated porcine alveolar macrophages. For this purpose, AMs were isolated from 5 (newborn), 40 (post-weaned) and 120 (young) day old pigs. Results of phagocytosis assay showed that AMs from newborn piglets had less phagocytic capacity than those of young pigs (P < 0.05). For the kinetics study, cells and supernatant were collected at 1, 6, 12, 24, 36 and 48 h after LPS stimulation for quantification of cytokine mRNA and protein by quantitative real-time PCR and ELISA, respectively. The kinetics results showed that AMs from newborn piglets were significantly less capable of producing IL1 beta, IL6, IL2 beta, TNF alpha and IL8 than post-weaned piglets or young pigs. IL18 mRNA did not show significant differences between ages. MIP2 and MCP1 mRNA was higher in young pigs. Hence, higher production of cytokines by AMs may be the surfactant factors in the pulmonary host defense system. These results indicate that AMs from newborn piglets might be functionally immature, which may lead to increased susceptibility to lung infections. Future studies of cytokine kinetics in more animals are clearly needed to confirm these results across a wider age range. (C) 2012 Elsevier Ltd. All rights reserved., The aim of the present study was to determine the age-related changes of phagocytic capacity and the kinetic production of cytokines in lipopolysaccharide-stimulated porcine alveolar macrophages. For this purpose, AMs were isolated from 5 (newborn), 40 (post-weaned) and 120 (young) day old pigs. Results of phagocytosis assay showed that AMs from newborn piglets had less phagocytic capacity than those of young pigs (P

Pulmonary alveolar macrophages (AMs) are important in defense against bacterial lung inflammation. Cluster of differentiation 14 (CD14) is involved in recognizing bacterial lipopolysaccharide (LPS) through MyD88-dependent and TRIF pathways of innate immunity. Sulforaphane (SFN) shows anti-inflammatory activity and suppresses DNA methylation. To identify CD14 epigenetic changes by SFN in the LPS-induced TRIF pathway, an AMs model was investigated in vitro. CD14 gene expression was induced by 5 mu g/ml LPS at the time point of 12 h and suppressed by 5 mu M SFN. After 12 h of LPS stimulation, gene expression was significantly up-regulated, including TRIF, TRAF6, NF-B, TRAF3, IRF7, TNF-, IL-1, IL-6, and IFN-. LPS-induced TRAM, TRIF, RIPK1, TRAF3, TNF-, IL-1 and IFN- were suppressed by 5 mu M SFN. Similarly, DNMT3a expression was increased by LPS but significantly down-regulated by 5 mu M SFN. It showed positive correlation of CD14 gene body methylation with in LPS-stimulated AMs, and this methylation status was inhibited by SFN. This study suggests that SFN suppresses CD14 activation in bacterial inflammation through epigenetic regulation of CD14 gene body methylation associated with DNMT3a. The results provide insights into SFN-mediated epigenetic down-regulation of CD14 in LPS-induced TRIF pathway inflammation and may lead to new methods for controlling LPS-induced inflammation in pigs., Pulmonary alveolar macrophages (AMs) are important in defense against bacterial lung inflammation. Cluster of differentiation 14 (CD14) is involved in recognizing bacterial lipopolysaccharide (LPS) through MyD88-dependent and TRIF pathways of innate immunity. Sulforaphane (SFN) shows anti-inflammatory activity and suppresses DNA methylation. To identify CD14 epigenetic changes by SFN in the LPS-induced TRIF pathway, an AMs model was investigated in vitro CD14 gene expression was induced by 5 µg/ml LPS at the time point of 12 h and suppressed by 5 µM SFN. After 12 h of LPS stimulation, gene expression was significantly up-regulated, including TRIF, TRAF6, NF-κB, TRAF3, IRF7, TNF-α, IL-1β, IL-6, and IFN-β. LPS-induced TRAM, TRIF, RIPK1, TRAF3, TNF-α, IL-1β and IFN-β were suppressed by 5 µM SFN. Similarly, DNMT3a expression was increased by LPS but significantly down-regulated by 5 µM SFN. It showed positive correlation of CD14 gene body methylation with in LPS-stimulated AMs, and this methylation status was inhibited by SFN. This study suggests that SFN suppresses CD14 activation in bacterial inflammation through epigenetic regulation of CD14 gene body methylation associated with DNMT3a. The results provide insights into SFN-mediated epigenetic down-regulation of CD14 in LPS-induced TRIF pathway inflammation and may lead to new methods for controlling LPS-induced inflammation in pigs.

The aim of this study was to integrate multi omics data to characterize underlying functional pathways and candidate genes for drip loss in pigs. The consideration of different omics levels allows elucidating the black box of phenotype expression. Metabolite and protein profiling was applied in Musculus longissimus dorsi samples of 97 Duroc × Pietrain pigs. In total, 126 and 35 annotated metabolites and proteins were quantified, respectively. In addition, all animals were genotyped with the porcine 60 k Illumina beadchip. An enrichment analysis resulted in 10 pathways, amongst others, sphingolipid metabolism and glycolysis/gluconeogenesis, with significant influence on drip loss. Drip loss and 22 metabolic components were analyzed as intermediate phenotypes within a genome-wide association study (GWAS). We detected significantly associated genetic markers and candidate genes for drip loss and for most of the metabolic components. On chromosome 18, a region with promising candidate genes was identified based on SNPs associated with drip loss, the protein "phosphoglycerate mutase 2" and the metabolite glycine. We hypothesize that association studies based on intermediate phenotypes are able to provide comprehensive insights in the genetic variation of genes directly involved in the metabolism of performance traits. In this way, the analyses contribute to identify reliable candidate genes.

Hydrogen peroxide (H2O2) and sulforaphane (SFN) are classical reagents to study oxidative stress and antioxidant activity in vitro, respectively. Micromole of H2O2 is sufficient to induce oxidative stress. Similarly, micromole of SFN is able to induce protective activity of cells against oxidative stress. However, higher or lower concentrations of both H2O2 and SFN may lead to aberrant results. Therefore, current study aimed to find out the appropriate concentrations of H2O2 and SFN for granulosa cells in vitro. For this, granulosa cells were collected and cultured with DMEM-F12 media at 37◦ C in a humidified environment until 60–70% confluence was achieved. After that different concentrations of H2O2 (0–400 M) and SFN (0–80 M) were added to the culture media and continue culture for 24 h. The results showed that cell viability decreased in a concentration dependent manner for both H2O2 and SFN treated cells. 50–75 MH2o2 are able to induce morphologically visible stress on cells, whereas 10–15 M SFN showed no effects on cellular morphology and viability. Analysis of candidate transcripts showed that, in H2O2 groups, 50–75 M create significantly higher expression of NRF2 & CASP3 and lower expression of KEAP1. On the other hand, 10–15 M SFN exhibited significant higher expression of NRF2 and lower expression of KEAP1, while expression of CASP3 remains unchanged. It is clear from the morphologic and genetic observations that 50–75 MH2o2 and 10–15 M SFN showed excellent response and are recommended concentrations for granulosa cells to study oxidative stress in vitro.

Toll-like receptors (TLRs) play an important role in non-specific immunity against different infectious agents such as bacterium or parasite. The aim of this work was to investigate the allele and genotype frequencies of three single-nucleotide polymorphisms (SNPs) in bovine TLR1 gene in native Turkish cattle breeds. DNA samples were extracted using the phenol chloroform protocol from 77 Zavot, 60 Turkish Grey, 51 East Anatolian Red, 69 Anatolian Black and 46 South Anatolian Red cattle. Target regions of the TLR1 gene were digested BsII and HpyI88III restriction enzymes. Results showed that the (A) allele frequency had higher in all native Turkish cattle breeds of the TLR1-G1409A locus. The (F) allele frequency was found to be higher compared to (E) allele in the TLR1-G1550A site. The frequencies of both (C) and (T) alleles were close to each other in the TLR1-C632T site. In conclusion genetic polymorphisms exist in Turkish native cattle populations in terms of known TLR1 variants.

ESR2 is involved in oestrogen-related apoptosis in cell cycle spermatogenesis but their effects have not yet confirmed in pig. Therefore, this study was aimed to investigate the association of ESR2 polymorphism with sperm quality and boar fertility traits and to analyse the ESR2 mRNA and protein expressions in boar reproductive tissues. DNA samples from 203 Pietrain (PI) and 100 Pietrain × Hampshire (PIHA) pigs with records of sperm quality [sperm concentration (SCON), motility (MOT), semen volume (VOL), plasma droplet rate (PDR) and abnormal spermatozoa rate (ASR)] and fertility [non-return rate (NRR) and number of piglet born alive (NBA)] traits were available. A SNP in coding region of ESR2 g.35547A>G in exon 5 was associated with MOT and PDR in the PI and with SCON, VOL, MOT and PDR in PIHA population. For mRNA and protein expression study, a total of six boars were divided into two groups with group I (G-I) and group II (G-II) where G-I characterized for relatively a better sperm quality according to the mean of two groups. mRNA expression was higher in brain and testis than that in all parts of epididymis. Both qRT-PCR and western blot analysis revealed that the ESR2 gene expression and protein expression were significantly higher in testis collected from G-II compared with that of G-I boars. Moreover, ESR2 protein localization in germ cell, Leydig and Sertoli cells, epithelial cells and spermatozoa was remarkable, which indicated the important role of ESR2 in spermatogenesis process. These results might shed new light on the roles of ESR2 in spermatogenesis as candidate for boar fertility, but still the lack of association across populations should be considered.

SummaryMicroRNAs (miRNAs) are small endogenous molecules that are involved in a diverse of cellular process. However, little is known about their abundance in bovine oocytes and their surrounding cumulus cells during oocyte development. To elucidate this situation, we investigated the relative expression pattern of sets of miRNAs between bovine oocyte and the surrounding cumulus cells duringin vitromaturation using miRNA polymerase chain reaction (PCR) array. Results revealed that a total of 47 and 51 miRNAs were highly abundant in immature and matured oocytes, respectively, compared with their surrounding cumulus cells. Furthermore, expression analysis of six miRNAs enriched in oocyte miR-205, miR-150, miR-122, miR-96, miR-146a and miR-146b-5p at different maturation times showed a dramatic decrease in abundance from 0 h to 22 h of maturation. The expression of the same miRNAs in preimplantation stage embryos was found to be highly abundant in early stages of embryo development and decreased after the 8-cell stage to the blastocyst stage following a typical maternal transcript profile. Similar results were obtained by localization of miR-205 in preimplantation stage embryos, in which signals were higher up to the 4-cell stage and reduced thereafter. miR-205 and miR-210 were localizedin situin ovarian follicles and revealed a spatio-temporal expression during follicular development. Interestingly, the presence or absence of oocytes or cumulus cells during maturation was found to affect the expression of miRNAs in each of the two cell types. Hence, our results showed the presence of distinct sets of miRNAs in oocytes or cumulus cells and the presence of their dynamic degradation during bovine oocyte maturation.

Among modern western pigs, Duroc (high meat fat ratio) and Pietrain (low meat fat ratio) breeds extensively utilized in commercial pork production differ extremely for their muscle phenotypes. The molecular mechanism, especially the epigenetic mechanism, underlying these breed-specific differences is poorly known. Myogenic factor 6 (MYF6) is the most abundantly expressed myogenic factor in adult muscle. Moreover, MYF6 tends to be expressed more highly in muscle tissue of the lean selection line and is supposed to be one promising candidate gene for growth- and meat quality-related traits in adult pigs. Six months old female Duroc and Pietrain pure breed pigs were used in this study. Protein and mRNA levels of MYF6 in loin eye muscle were determined by Western blotting and quantitative Real-time reverse transcription PCR (qRT-PCR), respectively. The DNA methylation status of the MYF6 5'-regulatory region was determined by bisulfite sequencing PCR (BSP). The global Histone 4 acetylation at lysines 5 (H4K5) and 8 (H4K8) were examined by Western blotting. Pietrain pigs exhibited significant higher expression of MYF6 and hypermethylated E2F1 binding element within MYF6 5'-regulatory region as compared with Duroc pigs. Significant elevation in DNA methyltransferase 1 (DNMT1) expression was observed in Pietrain pigs which are in agreement with hypermethylation of MYF6. Histone acetylation level at neither H4K5 nor H4K8 is significant between two breed pigs. Nevertheless, mRNA and protein expression of E2F1 were significantly elevated in the Pietrain breed. It is thus conceivable that the upregulation of MYF6 transcription in postnatal Pietrain pigs is not associated with cis-activation by epigenetic modification of MYF6 5'-regulatory region, but may be attributed to trans-activation through enriched expression of E2F1. (C) 2011 Elsevier B.V. All rights reserved.

Cytochrome P450 aromatase is the key enzyme in estrogen biosynthesis, encoded by the CYP19 gene. However, little is know about the CYP19 roles in boar spermatogenesis. Therefore, the aim of this work was to investigate the mRNA and protein expression of CYP19 in boar reproductive tissues from boars with different sperm quality. For mRNA and protein expression study, a total of six boars were divided into two groups with Group I (G-I) and Group II (G-II), where G-I is characterized for a relatively better sperm quality. For the expression study between reproductive and non-reproductive tissues by semi-quantitative PCR study, mRNA from all six boars was pooled together according to the tissues. On the other hand, mRNA and protein expression study in different reproductive tissues from two divergent groups of animals were performed by semi-quantitative PCR, qRT-PCR and western blot, respectively. Due to the limitations of fresh samples from G-I and G-II boars, different fresh testis from a healthy breeding boar was collected after slaughtering for protein localization by immunofluorescence. The remarkable CYP19 mRNA expression was detected only in testis. The mRNA expression of CYP19 was not detectable in other reproductive tissue (epididymis and accessory glands) and non-reproductive tissue (brain, liver and muscle) by semi-quantitative PCR.

The aim of this research was to screen for polymorphism and to perform an association study of IFI6 with meat and carcass quality traits. A SNP (g.370A>G) was detected which was associated (P < 0.05) with meat colour, pH 24 h post mortem (p.m.) in ham, conductivity 45 min p.m. in loin and conductivity 24 h p.m. in ham, drip loss and carcass length in Duroc × Pietrain and with meat colour, muscle area and ham percentage in the Pietrain population. Highest expression of IFI6 mRNA was detected in skeletal muscle (longissimus dorsi) by qRT-PCR comparing different tissues. Both qRT-PCR and western blot revealed that the IFI6 gene and protein expressions were significantly (P < 0.05) higher in skeletal muscle with low drip loss compared to that of high drip loss. IFI6 protein was localized in the myocytes membrane. Results suggested that IFI6 might play roles in meat and carcass quality and is a potential positional, physiological and functional candidate gene for improving meat quality traits in pigs.

Phospholipase C zeta (PLCz) and cyclooxygenase isoenzyme type 2 (COX-2) are important in spermatogenesis, but their effect has not yet confirmed in pigs. Therefore, this study was aimed to analyse their association with sperm quality and fertility and to identify the mRNA and protein expression in boars reproductive tissues. DNA samples from 231 Pietrain (PI) and 109 Pietrain × Hampshire (PIHA) pigs with records of sperm quality [sperm concentration (SCON), motility, semen volume, plasma droplet and abnormal spermatozoa rate] and fertility (non-return rate and number of piglet born alive) traits were available. A SNP in non-coding region of PLCz g.158 A > C was associated with SCON (p A in 3' UTR was not associated with any traits. For mRNA and protein expression study, a total of six boars were divided into two groups with G-I and G-II, where G-I was characterized for relatively better sperm quality. Both genes expressed higher in reproductive tissues compared with non-reproductive tissues. Phospholipase C zeta mRNA expressed higher in testis (p < 0.01), all parts of epididymis and spermatozoa from G-I, while COX-2 expressed higher in testis (p < 0.05), head and body of epididymis (p < 0.01), and spermatozoa from G-II boar. Both proteins were localized in Leydig cells and spermatozoa. These results might shed light on roles of these genes in spermatogenesis as candidate for boar sperm quality and fertility, but still the lack of association across populations should be considered.

Osteochondrosis (OC) or leg weakness is an economically important disease of young fast growing pigs and is a concern of animal welfare. The etiology and pathogenesis of osteochondrosis is not fully understood yet, but any abnormalities in the formation of hypertrophic chondrocytes and disrupted blood supply to the growth cartilage are very important predisposing factors. Matrix gla protein (MGP) as a potential calcification inhibitor of extracellular matrix might contribute to the development of OC. Molecular characterization, polymorphisms analysis, methylation at promoter region and expression of MGP gene and protein were performed in both healthy and OC cartilage collected from a Duroc x Pietrain resource population. The porcine MGP gene consists of 4 exons and 3 introns. The full-length MGP cDNA isolated from articular cartilage consists of 606 bp with a 69-bp 5' UTR, a 312-bp open reading frame with a start codon, a 225-bp 3' UTR. Three single-nucleotide polymorphisms (SNP) were detected in the intron 1 (A-115G, C-1073T and C-1135A) and one in the 3'UTR (C-3767T). The relative abundance of MGP mRNA was lower (P

Histone acetylation, regulated by histone deacetylases (HDACs) is a key epigenetic mechanism controlling gene expressions. Although dendritic cells (DCs) are playing pivotal roles in host immune responses, the effect of epigenetic modulation of DCs immune responses remains unknown. Sulforaphane (SFN) as a HDAC inhibitor has anti-inflammatory properties, which is used to investigate the epigenetic regulation of LPS-induced immune gene and HDAC family gene expressions in porcine monocyte-derived dendritic cells (moDCs). SFN was found to inhibit the lipopolysaccharide LPS induced HDAC6, HDAC10 and DNA methyltransferase (DNMT3a) gene expression, whereas up-regulated the expression of DNMT1 gene. Additionally, SFN was observed to inhibit the global HDAC activity, and suppressed moDCs differentiation from immature to mature DCs through down-regulating the CD40, CD80 and CD86 expression and led further to enhanced phagocytosis of moDCs. The SFN pre-treated of moDCs directly altered the LPS-induced TLR4 and MD2 gene expression and dynamically regulated the TLR4-induced activity of transcription factor NF-kappa B and TBP. SFN showed a protective role in LPS induced cell apoptosis through suppressing the IRF6 and TGF-beta 1 production. SFN impaired the pro-inflammatory cytokine TNF-alpha and IL-1 beta secretion into the cell culture supernatants that were induced in moDCs by LPS stimulation, whereas SFN increased the cellular-resident TNF-alpha accumulation. This study demonstrates that through the epigenetic mechanism the HDAC inhibitor SFN could modulate the LPS induced innate immune responses of porcine moDCs.