Association of myostatin gene polymorphism with echocardiographic and muscular ultrasonographic measurements in Hungarian thoroughbreds horses.

The g.66493737C/T polymorphism of the myostatin gene (MSTN) majorly influences muscle fiber composition and best race distance of Thoroughbreds. Thus, a better understanding of this process may lead to superior genetic exploitation for maximizing Thoroughbred athletic potential. Our objective is to investigate whether myostatin genotypes are associated with muscular development and cardiac variables of Thoroughbreds. Echocardiography and muscular ultrasonography were performed on three groups having C/C, C/T, and T/T genotypes, respectively. Each group consisted of 22 animals. Homogeneity of variance between the groups was checked by Levene's test. Multivariate analysis of variance was applied to determine differences in measured variables vs. MSTN genotypes. Fascicle length of anconeus and thickness of triceps brachii muscles showed significant differences between C/C and T/T genotypes (pFascicle-length-of-anconeus = 0.004, pthickness-of-triceps-brachii < 0.001). According to the primary outcome, there are associations between myostatin genotypes and cardiac variables. Aortic diameter at the sinus of Valsalva (end-diastole and end-systole) and aortic diameter at the valve (end-systole) indicated significant differences between C/C and T/T genotypes (paortic-diameter-at-the-sinus-of-Valsalva-end-diastole = 0.015, paortic-diameter-at-the-sinus-of-Valsalva-end-systole = 0.011, paortic-diameter-at-the-valve-end-systole = 0.014). Pearson correlation effect sizes were rFascicle-length-of-anconeus = 0.460, rthickness-of-triceps-brachii = 0.590, raortic-diameter-at-the-sinus-of-Valsalva-end-diastole = 0.423, raortic-diameter-at-the-sinus-of-Valsalva-end-systole = 0.450, and raortic-diameter-at-the-valve-end-systole = 0.462. C/C genotypes gave 22.1, 12.2, 6.3, 6.0, and 6.7% higher values compared to T/T genotypes, respectively. Differences regarding aortic diameter between genotype groups support the hypothesis that C/C animals have consequently increased cardiac output and aerobic capacity.

Microbiota Composition of Mucosa and Interactions between the Microbes of the Different Gut Segments Could Be a Factor to Modulate the Growth Rate of Broiler Chickens.

The study reported here aimed to determine whether correlations can be found between the intestinal segment-related microbiota composition and the different growing intensities of broiler chickens. The bacterial community structures of three intestinal segments (jejunum chymus-JC, jejunum mucosa-JM, caecum chymus-CC) from broiler chickens with low body weight (LBW) and high body weight (HBW) were investigated. Similar to the previous results in most cases, significant differences were found in the bacteriota diversity and composition between the different sampling places. However, fewer body weight (BW)-related differences were detected. In the JM of the HBW birds, the Bacteroidetes/Firmicutes ratio (B/F) was also higher. At the genus level significant differences were observed between the BW groups in the relative abundance of Enterococcus, mainly in the JC; Bacteroides and Ruminococcaceae UCG-010, mainly in the JM; and Ruminococcaceae UCG-013, Negativibacillus, and Alistipes in the CC. These genera and others (e.g., Parabacteroides and Fournierella in the JM; Butyricoccus, Ruminiclostridium-9, and Bilophila in the CC) showed a close correlation with BW. The co-occurrence interaction results in the JC revealed a correlation between the genera of Actinobacteria (mainly with Corynebacterium) and Firmicutes Bacilli classes with different patterns in the two BW groups. In the JM of LBW birds, two co-occurring communities were found that were not identifiable in HBW chickens and their members belonged to the families of Ruminococcaceae and Lachnospiraceae. In the frame of the co-occurrence evaluation between the jejunal content and mucosa, the two genera (Trichococcus and Oligella) in the JC were found to have a significant positive correlation with other genera of the JM only in LBW chickens.

Role of genes related to performance and reproduction of Thoroughbreds in training and breeding - A review.

Thoroughbreds have been selected for speed and stamina since the 1700s. This selection resulted in structural and functional system-wide adaptations that enhanced physiological characteristics for outstanding speed of 61-71 kph (38-44 mph) between 1,000 and 3,200 m (5 furlongs - 2 miles). At present, horseracing is still an economically important industrial sector, therefore intensive research is underway to explore genes that allow the utilisation of genetic abilities and are significant in breeding and training. This study aims to provide an overview of genetic research and its applicability related to Thoroughbreds.

The relationship between small heat shock proteins and redox homeostasis during acute heat stress in chickens.

As heat stress is a major emerging issue in poultry farming, investigations on the molecular mechanisms of the heat-triggered cellular response in chickens are of special importance. In the present study, 32-day-old Ross 308 broiler chickens were subjected to 37 °C environmental temperature combined with 50% relative humidity for 4 or 8 h respectively. Following sampling, redox parameters such as malondialdehyde (MDA), reduced glutathione (GSH), protein carbonyl levels as well as glutathione peroxidase activity were assessed in liver, spleen, and kidney homogenates. The concentrations of small heat shock proteins (sHSP-s) HSP27, αA- and αB-crystallins were also investigated. Among these organs, the liver was found the most susceptible to heat-provoked oxidative stress, indicated by enhanced lipid peroxidation and rapid activation of protective pathways, including the definite increase of glutathione peroxidase activity and the excessive utilization of αA- and αB-crystallin proteins. Heat-associated decline of protein carbonylation and GSH content was observed in the liver in correlation with the increased involvement of αA- and αB-crystallins in cellular defense, resulting supposedly in an overcompensation mechanism. These data highlight the hepatic sensitivity to acute heat shock, potential adaptation mechanisms, and the specific role of sHSP-s in the restoration of physiologic cell function.

Effects of Wheat Bran and Clostridium butyricum Supplementation on Cecal Microbiota, Short-Chain Fatty Acid Concentration, pH and Histomorphometry in Broiler Chickens.

Feed additives that can improve intestinal health and maintain a diverse and resilient intestinal microbiota of poultry are of great importance. Thus, the current study investigated the effects of a single strain butyric acid-producing Clostridium (C. butyricum) with (symbiotic) or without wheat bran supplementation on cecal microbiota composition and gut health characteristics of broiler chickens. In total, 384 male Ross 308 day-old chickens were divided into four dietary treatment groups and fed ad libitum until day 37 of life. Cecal samples were taken for Illumina sequencing and pH and short-chain fatty acid analyses, as well as for histological analysis at the end of the experimental period. Neither of the supplemented diets improved chicken growth performance. Caecum was dominated by the members of Bacteroidetes phyla followed by Firmicutes in each dietary group. At the genus level, Bacteroides, Oscillospira, Akkermansia, Faecalibacterium, Ruminococcus and Streptococcus genera exceeded 1% relative abundance. Dietary treatment influenced the relative abundance of the Akkermansia genus, which had a lower relative abundance in the C. butyricum group than in the other groups and in the symbiotic group compared to the wheat bran supplemented group. Dietary treatment also altered cecal crypt depth and had a trend to modify the cecal fermentation profile. Additive effects of wheat bran and C. butyricum supplementation were not detected. Our results suggest that Akkermansia muciniphila colonization in chicken can be influenced by diet composition.

Effects of Acute Heat Stress on a Newly Established Chicken Hepatocyte-Nonparenchymal Cell Co-Culture Model.

Heat stress is one of the most important issues in broiler flocks impairing animal health and productivity. On a cellular level, excess heat exposure can trigger heat shock response acting for the restoration of cell homeostasis by several mechanisms, such as affecting heat shock protein synthesis, redox homeostasis and pro-inflammatory cytokine production. The major aim of this study was to establish a novel avian hepatocyte-nonparenchymal cell co-culture as a model for investigating the cellular effects of heat stress and its interaction with inflammation in chicken liver. Cell fractions were isolated by differential centrifugation from a freshly perfused chicken liver, and hepatocyte mono-cultures as well as hepatocyte-nonparenchymal cell co-cultures (with cell ratio 6:1, hepatocytes to nonparenchymal cells, mimicking a milder hepatic inflammation) were prepared. Isolated and cultured cells were characterized by flow cytometry and immunocytochemistry applying hepatocyte- and macrophage-specific antibodies. Confluent cell cultures were exposed to 43 °C temperature for 1 or 2 h, while controls were cultured at 38.5 °C. The metabolic activity, LDH enzyme activity, reactive oxygen species (H2O2) production, extracellular concentration of heat shock protein 70 (HSP70), and that of the pro-inflammatory cytokines interleukin (IL-)6 and IL-8 were assessed. Shorter heat stress applied for 1 h could strongly influence liver cell function by significantly increasing catabolic metabolism and extracellular H2O2 release, and by significantly decreasing HSP70, IL-6, and IL-8 production on both cell culture models. However, all these alterations were restored after 2 h heat exposure, indicating a fast recovery of liver cells. Hepatocyte mono-cultures and hepatocyte-nonparenchymal cell co-cultures responded to heat stress in a similar manner, but the higher metabolic rate of co-cultured cells may have contributed to a better capability of inflamed liver cells for accommodation to stress conditions. In conclusion, the established new primary cell culture models provide suitable tools for studying the hepatic inflammatory and stress response. The results of this study highlight the impact of short-term heat stress on the liver in chickens, underline the mediatory role of oxidative stress in acute stress response, and suggest a fast cellular adaptation potential in liver cells.

Cellular Effects of T-2 Toxin on Primary Hepatic Cell Culture Models of Chickens.

Trichothecene mycotoxins such as T-2 toxin cause severe problems for agriculture, as well as for veterinary medicine. As liver is one of the key organs in metabolism, the main aim of our study was to investigate the immunomodulatory and cytotoxic effects of T-2 toxin, using primary hepatocyte mono-culture and hepatocyte-nonparenchymal cell (predominantly Kupffer cell) co-culture models of chicken. Cultures were exposed to 10 (T10 group), 100 (T100 group) and 1000 (T1000 group) nmol/L T-2 toxin treatment for 8 or 24 h. Alterations of cellular metabolic activity, the production of reactive oxygen species (extracellular H2O2), heat shock protein 70 (HSP70), and the concentration of different inflammatory cytokines such as interleukin (IL-)6 and IL-8 were investigated. Metabolic activity was intensely decreased by T-2 toxin administration in all of the cell culture models, in every applied concentration and incubation time. Concentrations of HSP70 and IL-8 were significantly increased in hepatocyte mono-cultures exposed to higher T-2 toxin levels (both in T100 and T1000 groups for HSP70 and in T1000 group for IL-8, respectively) compared to controls after 24 h incubation. Similarly, IL-6 levels were also significantly elevated in the T100 and T1000 groups in both of mono- and co-cultures, but only after 8 h of incubation time. In spite of the general harmful effects of T-2 toxin treatment, no significant differences were observed on reactive oxygen species production. Furthermore, the two cell culture models showed different levels of H2O2, HSP70, and IL-8 concentrations independently of T-2 toxin supplementation. In conclusion, the established primary cell cultures derived from chicken proved to be proper models to study the specific molecular effects caused by T-2 toxin. Metabolic activity and immune status of the different examined cell cultures were intensively affected; however, no changes were found in H2O2 levels.

Soluble nondigestible carbohydrates improve intestinal function and increase caecal coliform load in broiler chickens.

Diets rich in various soluble nondigestible carbohydrates (sNDCs) were evaluated on different intestinal characteristics (histological, physico-chemical and microbiological) of chickens and compared with a maize-based diet as a control. A total of 160 Ross 308 male chickens were kept in deep litter pens (n = 40) and fed their appropriate diets from Day 1 to Day 35 of life. Four isocaloric and isonitrogenous diets, differing in their sNDC content, were composed; control (containing maize as the only cereal), maize-wheat-based (M + W) and maize-based supplemented with either 20 g/kg inulin (M + I) or 30 g/kg lactose (M + L). All of the diets tested decreased ileal crypt depth, ileal muscle layer thickness and increased caecal coliform counts relative to the control group. Villus-crypt ratio increased only in the M + L group. Ileal digesta of chickens fed the M + W diet had the highest ileal viscosity and the highest caecal butyrate, valerate and total short-chain fatty acid concentrations while the lowest pH was observed in caecal contents of chickens fed the M + I diet. The diet had no effect on ileal or caecal goblet cell and intraepithelial lymphocyte numbers. Lactobacillus counts in the caecal content remained unchanged. According to the present study, various sNDC sources may have beneficial gut health effects, however, some of the intestinal variables are dependent on the type of sNDCs.

Nutritional modulation of intestinal drug-metabolizing cytochrome P450 by butyrate of different origin in chicken.

Intestinal cytochrome P450 (CYP) enzymes play key role in the first pass metabolism of orally ingested xenobiotics, providing a primary metabolic barrier, being of special importance in maintaining animal health and production. This study was aimed to investigate how intestinal drug-metabolizing CYPs can be modulated by nutritional factors in broiler chicken. We investigated the effects of the natural growth promoter (n-)butyrate of different origin (feed supplementation of protected or non-protected forms and/or inducing caecal microbial production by supporting higher level of dietary non-starch polysaccharides [NSP]) on the activity of duodenal CYPs. To observe the connection between intestinal CYP activity and butyrate concentration, the distribution of differently originated butyrate was also assessed by measuring its concentration in various intestinal segments and different vessels of portal and systemic circulation. Butyrate of different origin showed varying distribution properties as being absorbed from different parts of the gastrointestinal tract. Intestinal CYP1A and CYP2H2 activities were increased by dietary butyrate supplementation and by the increased caecal microbial butyrate production, while CYP3A37 activity was minimally influenced by microbial butyrate only. The present study proved that both dietary and microbial butyrate could alter the activity of CYPs in the duodenal epithelium. Our findings suggest that intestinal CYPs could be induced not only by the intestinal luminal butyrate, but also from basolateral side, by the already absorbed butyrate. Such action of butyrate can be of special importance from food safety and pharmacotherapeutic point of view as it may modify the metabolism and intestinal kinetics of simultaneously applied xenobiotics.

Effects of butyrate on the insulin homeostasis of chickens kept on maize- or wheat-based diets.

The aim of the present study was to investigate the effects of butyrate as a feed supplement on the expression of insulin signalling proteins as potent regulators of metabolism and growth in Ross 308 broiler chickens fed maize- or wheat-based diets. Both diets were supplemented with non-protected butyrate (1.5 and 3.0 g/kg of diet, respectively) or with protected butyrate (0.2 g/kg of diet); the diet of the control groups was prepared without any additives (control). On day 42 of life, systemic blood samples were drawn for analyses of glucose and insulin concentrations, and tissue samples (liver, gastrocnemius muscle and subcutaneous adipose tissue) were taken for Western blotting examinations. The expression of key insulin signalling proteins (IRß, PKCζ and mTOR) was assessed by semiquantitative Western blotting from the tissues mentioned. The type of diet had a remarkable influence on the insulin homeostasis of chickens. The wheat-based diet significantly increased IRß and mTOR expression in the liver as well as mTOR and PKCζ expression in the adipose tissue when compared to animals kept on a maize-based diet. IRß expression in the liver was stimulated by the lower dose of non-protected butyrate as well, suggesting the potential of butyrate as a feed additive to affect insulin sensitivity. Based on the results obtained, the present study shows new aspects of nutritional factors by comparing the special effects of butyrate as a feed additive and those of the cereal type, presumably in association with dietary non-starch polysaccharide- (NSP-) driven enteric shortchain fatty acid release including butyrate, influencing insulin homeostasis in chickens. As the tissues of chickens have physiologically lower insulin sensitivity compared to mammals, diet-associated induction of the insulin signalling pathway can be of special importance in improving growth and metabolic health.

Effects of high ambient temperature on fish sperm plasma membrane integrity and mitochondrial activity - A flow cytometric study.

Local extreme climatic conditions occurring as a result of global climate change may interfere with the reproduction of animals. In the present study fish spermatozoa were incubated at different temperatures (20, 25, 30 and 40 °C) for 10 and 30 minutes, respectively and plasma membrane integrity and mitochondrial membrane potential changes were evaluated with flow cytometry using SYBR-14/PI and Mitotracker Deep Red FM fluorescent dyes. No significant differences were found in plasma membrane integrity at either incubation temperatures or time points. Mitotracker Deep Red FM histogram profiles indicating mitochondrial activity showed significant (p < 0.001) alterations in all cases of higher (25, 30 and 40 °C) temperature treatments as compared to the samples incubated at 20 °C. Our results indicate that fish spermatozoa exposed to high temperatures suffer sublethal damage that cannot be detected with conventional, vital staining techniques.

Effect of feeding different oils on plasma corticosterone in broiler chickens.

A study was conducted to examine the effects of different oils on the plasma corticosterone concentrations of broiler chickens fed ad libitum or deprived of feed for 24 hours. A total of 36 Ross broilers were randomly assigned to one of three dietary treatments at 10 days of age and fed a grower diet supplemented with 60 g/kg soybean oil (rich in linoleic acid, C18:2n-6), linseed oil (rich in a-linolenic acid, C18:3n-3) or fish oil (rich in C14:0, C16:0, C16:1n-7, C20:1n-9; eicosapentaenoic acid and docosahexaenoic acid, EPA, C20:5n-3 and DHA, C22:6n-3), respectively, for 18 days. Dietary supplementation of fish oil resulted in lower (P < 0.05) baseline plasma corticosterone levels of chickens fed ad libitum for 18 days compared to soybean and linseed oil supplementations. Feed deprivation for 24 h induced a significant (P < 0.05) increase in corticosterone concentration in every treatment group compared to the ad libitum-fed birds. The hormone levels of feed-deprived birds did not differ significantly among groups fed diets supplemented with different oils.

A comparison of alternative assays to measure DNA damage in stallion spermatozoa: TUNEL test versus 'Nicoletti assay'.

The aberrations of sperm DNA may cause various problems and have negative consequences on fertility. These influence embryonic development or might lead to early embryo loss. Sperm Chromatin Structure Assay (SCSA) is the flow cytometric method most often used for the detection of DNA lesions; however, some studies using that method reached confusing conclusions. The aim of this pilot study was to adjust and compare two alternative tests, namely the TUNEL test and the Nicoletti assay. The above-mentioned two flow cytometric methods capable of detecting the fragmented DNA of sperm were tested on 12 frozen-thawed stallion semen samples. The TUNEL test demonstrated much higher DNA fragmentation ratio than the Nicoletti assay (mean ± SD: 30.77 ± 13.03% vs. 1.93 ± 0.89%, respectively). A fluorescent microscopic check of the samples showed that TUNEL labelled the plasma membrane and the mitochondria in a nonspecific way, rather than detecting only the fragmented DNA, thus eventually resulting in a false positive sign. The Nicoletti assay is simpler, quicker and does not detect nonspecific binding; however, further analyses are required to determine its diagnostic value.

Influence of rumen-protected choline on liver composition and blood variables indicating energy balance in periparturient dairy cows.

Rumen-protected choline (RPC) was evaluated for effects on the lipid and glycogen content of the liver and metabolic variables in the blood plasma of dairy cows. Thirty-two Holstein cows were allocated into two groups (RPC group with RPC supplementation and control group without RPC supplementation) 28 days before the expected calving. Cows were fed the experimental diet from 21 days before calving until day 60 of lactation. The diet of the RPC group was supplemented with 100 g/day of RPC from 21 days prepartum until calving and 200 g/day of RPC for 60 days postpartum, providing 25 and 50 g of choline, respectively. Liver samples were taken by percutaneous needle biopsy, then analysed for total lipid (TLl), triglyceride (TGl) and glycogen (GLYl) contents on days -21, +7, +35 and +60 relative to calving. Blood was collected on the same sampling days and 21 days after calving. Glucose, non-esterified fatty acid (NEFA), ß-hydroxybutyrate (BHBA), triglyceride (TGp), total cholesterol (TCh), urea, ammonia and aspartate aminotransferase (AST) were determined from blood samples. The TLl and TGl contents were 25.0 ± 4.3 g and 25.3 ± 3.8 g per kg wet weight (mean ± SEM), respectively, lower in the RPC group than in the control animals. No significant differences were observed in the GLYl concentrations between the two groups. However, a lower TGl: GLYl ratio was shown in the liver of cows fed the RPC diet as compared to the controls. RPC supplementation decreased BHBA while increasing TGp concentrations were shown in the blood of cows fed the RPC diet, possibly as a consequence of improved lipoprotein synthesis in, and triglyceride excretion from, the liver, together with a reduced rate of ketogenesis.

Milk production, peripartal liver triglyceride concentration and plasma metabolites of dairy cows fed diets supplemented with calcium soaps or hydrogenated triglycerides of palm oil.

The aim of the study was to test the effect of rumen-inert fat supplements of different chemical forms or containing different unsaturated/saturated (U/S) fatty acid contents on milk production, milk composition and liver and blood metabolic variables of high-yielding dairy cows in the peripartal period. Thirty Holstein-Friesian dairy cows were divided into three equal groups and fed a corn silage-based diet, without fat supplementation (control) or supplemented with 11.75 MJ NEl per day of calcium soaps of palm oil fatty acids (CAS; U/S=61/39) or with 11.75 MJ NEl per day of hydrogenated palm oil triglyceride (HTG; U/S=6/94). Each diet was fed from 25+/-2 d prior to the expected calving to 100+/-5 d post partum. Compared with the control, both CAS and HTG supplementation resulted in an increase of the average milk yield. Milk fat content and fat-corrected milk yield were higher in the HTG group but lower in the CAS group than in the control group. In all groups liver triglyceride concentrations (TGL) increased from 15 d prepartum to 5 d post partum, and then decreased thereafter. At 5 d TGL was lower in the HTG group than control or CAS cows. No significant differences were detected in TGL among dietary treatments at 15 d prepartum and 25 d post partum. Higher plasma glucose and insulin and lower non-esterified fatty acids and beta-hydroxybutyrate concentrations and aspartate aminotransferase activity were measured in the HTG group than in the control or CAS groups at 5 d or 25 d post partum. Our results show that HTG may provide a better energy supply for high-yielding dairy cows in negative energy balance than CAS around calving.

Effect of various dietary fat supplementations on liver lipid and glycogen of high-yielding dairy cows in the peripartal period.

In a model experiment, Holstein-Friesian dairy cows were fed on a corn-silage-based diet supplemented with 11.75 MJ NE1 per day of calcium soaps of palm oil fatty acids (CAS) or hydrogenated triglyceride (HTG) or without fat supplementation (control). All diets were fed to the cows over a period from 21 +/- 3 days (d) prior to the expected calving to d 100 +/- 5 postpartum. On d 25 (basal sample) and d 14 prepartum as well as on d 5 and 25 postpartum liver samples were collected by percutaneous biopsy. Total lipid content, fatty acid composition and glycogen of liver tissues were determined. At d 5 postpartum, both control and CAS cows had higher liver lipid (P < 0.05) and lower glycogen (P < 0.05) concentrations than cows in the HTG group. No significant (P < 0.05) differences were detected in liver fat content among the groups at d 14 prepartum or d 25 postpartum. The glycogen concentration slightly decreased in the liver of cows in each treatment group from d 14 prepartum to d 5 postpartum; however, this decrease was more intensive in both the control and CAS groups than in the HTG group. The variations in liver lipid concentrations were accompanied by significant changes in the proportion of C16:0, C16:1n-7, C18:0, C18:1n-9, C18:2n-6 and C20:4n-6 fatty acids in the liver lipids. The results show that HTG supplementation exerted more advantageous effects on liver lipid and glycogen metabolism than did CAS supplementation.