Effects of propylene glycol used at different doses in Akkaraman lambs rations on metabolism-related parameters and liver gene and protein expression during different feeding periods.

This study aimed to investigate the metabolic effects of propylene glycol (PG) over 60, 90, and 120 days in lambs. Seventy-two weaned male lambs were allocated into three groups: control (Con), PG1.5 (1.5 mL/kg live weight0.75 ), and PG3 (3 mL/kg live weight0.75 ). Blood samples were collected at the beginning and slaughter days. Biochemical parameters (glucose, triglycerides, ALT, AST, LDH, BUN, and insulin) and gene and protein levels of peroxisome proliferator activated receptor gamma (PPARγ), diacylglycerol o-acyltransferase 1 (DGAT1), carbohydrate responsive element binding protein (ChREBP), and sterol regulatory element binding transcription factor 1c (SREBP-1c) in the liver were determined. Glucose in PG1.5 was increased on Day 60, while significant differences were observed in biochemical parameters except for insulin on the 60, 90, and 120 days. Biochemical parameters such as ALT, AST, LDH, and BUN increased over time, while triglycerides decreased. DGAT1 gene and protein levels were lower, while SREBP-1c and PPARγ were higher in PG groups on Day 60. While SREBP-1c was lower in PG1.5, ChREBP was higher in PG3 on Day 90. PPARγ, DGAT1, and ChREBP were upregulated in PG3 on Day 120. Positive correlations were found between proteins. The long-term use of PG in lambs did not have detrimental effects on metabolism. The study provides valuable insights into the molecular mechanisms underlying the metabolic effects of PG in lambs, shedding light on its potential applications in lamb production.

Genome-wide association study of early liveweight traits in fat-tailed Akkaraman lambs.

Small ruminants, especially sheep, are essential for sustainable agricultural production systems, future food/nutrition security, and poverty reduction in developing countries. Within developed countries, the ability of sheep to survive on low-quality forage intake could act as buffer against climate change. Besides sheep's importance in sustainable agricultural production, there has been less ongoing work in terms of sheep genetics in Near East, Middle East and in Africa. For lamb meat production, body weight and average daily gain (ADG) until weaning are critical economic traits that affects the profitability of the industry. The current study aims to identify single nucleotide polymorphisms (SNPs) that are significantly associated with pre-weaning growth traits in fat tail Akkaraman lambs using a genome-wide association study (GWAS). A total of 196 Akkaraman lambs were selected for analysis. After quality control, a total of 31,936 SNPs and 146 lambs were used for subsequent analyses. PLINK 1.9 beta software was used for the analyses. Based on Bonferroni-adjusted p-values, one SNP (rs427117280) on chromosome 2 (OAR2) had significant associations with weaning weight at day 90 and ADG from day 0 to day 90, which jointly explains a 0.8% and 0.9% of total genetic variation respectively. The Ovis aries natriuretic peptide C (NPPC) could be considered as a candidate gene for the defined significant associations. The results of the current study will help to increase understanding of the variation in weaning weight and ADG until weaning of Akkaraman lambs and help enhance selection for lambs with improved weaning weight and ADG. However, further investigations are required for the identification of causal variants within the identified genomic regions.

TLR4, MyD88, and TNF-α Expression in the Lungs of Akkaraman and Romanov Lambs in Response to LPS and LTA.

Toll-like receptors are involved in the recognition of bacterial toxins, which cause infection in the respiratory system. This study aimed to evaluate microanatomical and histological alterations in the lungs of 24 healthy Akkaraman and Romanov lambs after the administration of lipoteichoic acid (LTA), lipopolysaccharide (LPS), and LTA + LPS and investigate the gene, protein, and immune expression levels of TLR4, MyD88, and TNF-α molecules, known to have immune functions. Microanatomical examinations showed thickened peribronchial and alveolar walls in the lungs of groups LTA, LPS, and LTA + LPS of both breeds due to immune cell infiltration. TLR4, MyD88, and TNF-α immunoexpressions were positive to varying degrees in the cytoplasm and nucleus of the bronchial and bronchiolar luminal epithelial cells, alveolar epithelial cells, and alveolar macrophages. TLR4 and TNF-α protein expressions were statistically different in the LPS-treated Romanov lambs, compared to the other groups. Among the Akkaraman lambs, TLR4 gene expression was significantly higher in group LPS, and among the Romanov lambs, TLR4, MyD88, and TNF-α gene expressions were significantly higher in group LTA + LPS. Therefore, TLR4, MyD88, and TNF-α molecules, involved in the immune response, were found to be expressed at different levels against LTA and LPS in the lungs of two different sheep breeds.

LPS- and LTA-Induced Expression of TLR4, MyD88, and TNF-α in Lymph Nodes of the Akkaraman and Romanov Lambs.

Toll-like receptor (TLR)-mediated inflammatory processes play a critical role in the innate immune response during the initial interaction between the infecting microorganism and immune cells. This study aimed to investigate the possible microanatomical and histological differences in mandibular and bronchial lymph nodes in Akkaraman and Romanov lambs induced by lipopolysaccharide (LPS) and lipoteichoic acid (LTA) and study the gene, protein, and immunoexpression levels of TLR4, myeloid differentiation factor 88 (MyD88), and tumor necrosis factor-α (TNF-α) that are involved in the immune system. Microanatomical examinations demonstrated more intense lymphocyte infiltration in the bronchial lymph nodes of Akkaraman lambs in the LPS and LTA groups compared to Romanov lambs. TLR4, MyD88, and TNF-α immunoreactivities were more intense in the experimental groups of both breeds. Expression levels of MyD88 and TNF-α genes in the bronchial lymph node of Akkaraman lambs were found to increase statistically significantly in the LTA group. TLR4 gene expression level in the mandibular lymph node was found to be statistically significantly higher in the LTA + LPS group. In conclusion, dynamic changes in the immune cell populations involved in response to antigens such as LTA and LPS in the lymph nodes of both breeds can be associated with the difference in the expression level of the TLR4/MyD88/TNF-α genes.

Association of GH, STAT5A, MYF5 gene polymorphisms with milk somatic cell count, EC and pH levels of Holstein dairy cattle.

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.

Investigation of TLR1-9 genes and miR-155 expression in dogs infected with canine distemper.

This study aimed to determine the relationship of toll-like receptor (TLR) 1-9 genes and microRNA (miR) -155 expression levels with hematologic parameters in dogs diagnosed with canine distemper. In the study, two groups were used pre-treatment and post-treatment. Infected dogs were diagnosed with canine distemper with the help of a rapid test kit and Real Time-Polymerase Chain Reaction (RT-PCR). Based on the correlation coefficients between the expression levels of the genes examined within the scope of the study and hematologic values, a positive correlation was found between the TLR2 gene and the monocyte (MON) value and between the TLR4 gene and the platelet (PLT) value in the pre-treatment group. A strong positive correlation was identified between TLR3 and TLR9 genes and erythrocyte (RBC) and hemoglobin (HGB) values; between TLR5 gene and RBC, HGB and hematocrit (HCT) values and between TLR9 gene and RBC and HGB values in the post-treatment group, on the other hand, a positive correlation was found between TLR1 gene and MON and neutrophil (GRAN) values; between TLR3 gene and HCT value and between TLR9 gene and MON and HCT values. The study concluded that miR-155 and TLR8 gene were upregulated at a statistically significant level (P < 0.05) Post-treatment in dogs infected with canine distemper and there was a positive correlation between the upregulation of miR-155 and the upregulation of TLR8 in the same period. This result suggests that the upregulated miR-155 expression post-treatment increased TLR8 gene expression. In the light of these findings, it miR-155 may have the potential to be used in clinical practice in the treatment or prognosis of dogs infected with canine distemper.

Differential protein input in the maternal diet alters the skeletal muscle transcriptome in fetal sheep.

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™ 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 ≤ 0.05) in HP vs. SP, LP vs. HP, and SP vs. LP, respectively. Several myogenic genes, including MYOD1, MYH2, MYH1, are significantly upregulated, while genes related to the immune system, such as CXCL11, HLA-E, CXCL10, CXCL9, TLRs, are significantly downregulated in the fetal muscle of the HP group compared to those of SP and LP group. Bioinformatic analysis revealed that the majority of these genes are involved in pathways related to the immune system and diseases. The results of our study demonstrate that both augmented and restricted dietary proteins in maternal diet during pregnancy alter the expression of genes as well as the offspring's genetic marks.

The EDN2 rs110287192 gene polymorphism is associated with paratuberculosis susceptibility in multibreed cattle population.

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.

LEP and SCD polymorphisms are associated with milk somatic cell count, electrical conductivity and pH values in Holstein cows.

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 ≤ 0.05) traits. While the highest EC value was found in SCD-Fnu4HI-CT genotype, the highest milk pH was found in genotype TT. In addition, NR1H3-HpyCH4IV genotypes was found the only associated with pH (p < 0.05) among all studied phenotypes. Based on the present findings, it was concluded that LEP and SCD genes could be used in breeding programs for improved SCC, EC and pH values in Holstein dairy cows.

Oxidative stress modulates the expression of apoptosis-associated microRNAs in bovine granulosa cells in vitro.

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 µM 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.

Polymorphisms in toll-like receptor (TLR) 1, 4, 9 and SLC11A1 genes and their association with paratuberculosis susceptibility in Holstein and indigenous crossbred cattle in Turkey.

Mycobacterium avium subsp. paratuberculosis (MAP) causes major problem in a wide range of animal species. In ruminant livestock including cattle, it causes a chronic disease called Johne's disease, or paratuberculosis (pTB) which is currently considered as potential zoonosis, causing Crohn's disease in humans. MAP infection susceptibility is suspected to be controlled by host genetics. Thus, selecting individuals according to their genetic structure could help to obtain bovine populations that are increasingly resistant to MAP infection. The aim of the present work was to investigate the association between toll-like receptor (TLR) 1 (+1380 G/A), TLR1 (+1446 C/A), TLR4 (+10 C/T), TLR9 (+1310 G/A) and solute carrier family 11 member 1 (SLC11A1) (+1066 C/G) mutations and MAP infection status in 813 cattle comprising East Anatolian Red crossbred, Anatolian Black crossbred and Holstein breed. TLR1 (+1380 G/A) mutation showed an association with bovineMAP (P<0.05). For the TLR1 (+1380 G/A) locus, the odds ratio for AG and AA genotypes versus GG genotypes were 2.31 (1.24-4.30; 95% confidence interval (CI)) and 0<0.001 (<0.001 to >999.999; 95% CI) which indicated that a proportion of AG homozygote was significantly higher in pTB-affected animals as compared with the control. General linear model analysis demonstrated higher MAP antibody response in TLR1 (+1380 AG) genotype as compared with TLR1 (+1380 GG) (P<0.0001). Present findings suggest that selection against TLR1 (+1380 G/A) may reduce the risk of pTB in bovine herds.

Concentration dependent antioxidative and apoptotic effects of sulforaphane on bovine granulosa cells in vitro.

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.