The effect of succinate on color stability of Bos indicus bull meat: pH-dependent effects during the 14-day aging period.

Bos taurus indicus bulls are very susceptible to pre-slaughter stress, which directly impacts the decline in muscle pH, leading to darker meat. The aim was to investigate the effect of succinate and atmosphere on the color stability of Nellore (Bos taurus indicus) Longissimus lumborum steaks classified by ultimate pH (pHu): normal pHu (5.40 ≤ pHu ≤ 5.79) and high pHu (pHu ≥ 5.80). The experimental treatment systems were: (i) vacuum packaging without succinate injection, (ii) HiOx-MAP (80 % O2 + 20 % CO2), and (iii) HiOx-MAP (80 % O2 + 20 % CO2) enhanced with sodium succinate injection (pH 5.4). Steaks from all treatment systems were stored at 4 °C for 14 days and tested for instrumental color, myoglobin content, oxygen consumption (OC), metmyoglobin-reducing activity (MRA), lipid oxidation, and microbiological analysis. High and normal pHu vacuum-packaged steaks exhibited greater color stability due to higher MRA. High and normal pHu steaks packaged with HiOx-MAP or HiOx-MAP enhanced with succinate showed improved color due to lower deoxymyoglobin content (%DMb) and OC up to the eighth day of storage. Still, succinate injection promoted increased (P < 0.05) lipid oxidation in normal pHu steaks and reduced MRA after 14 days. These findings emphasize the intricate interplay between pHu and packaging systems on Bos taurus indicus meat quality. Further research in this area could contribute to a better understanding of meat color abnormalities and provide insights into potential meat preservation and enhancement strategies.

Transcriptome profile of skeletal muscle using different sources of dietary fatty acids in male pigs.

Pork is of great importance in world trade and represents the largest source of fatty acids in the human diet. Lipid sources such as soybean oil (SOY), canola (CO), and fish oil (FO) are used in pig diets and influence blood parameters and the ratio of deposited fatty acids. In this study, the main objective was to evaluate changes in gene expression in porcine skeletal muscle tissue resulting from the dietary oil sources and to identify metabolic pathways and biological process networks through RNA-Seq. The addition of FO in the diet of pigs led to intramuscular lipid with a higher FA profile composition of C20:5 n-3, C22:6 n-3, and SFA (C16:0 and C18:0). Blood parameters for the FO group showed lower cholesterol and HDL content compared with CO and SOY groups. Skeletal muscle transcriptome analyses revealed 65 differentially expressed genes (DEG, FDR 10%) between CO vs SOY, and 32 DEG for CO vs FO, and 531 DEG for SOY vs FO comparison. Several genes, including AZGP1, PDE3B, APOE, PLIN1, and LIPS, were found to be down-regulated in the diet of the SOY group compared to the FO group. The enrichment analysis revealed DEG involved in lipid metabolism, metabolic diseases, and inflammation between the oil groups, with specific gene functions in each group and altered blood parameters. The results provide mechanisms to help us understand the behavior of genes according to fatty acids.

Brain fatty acid and transcriptome profiles of pig fed diets with different levels of soybean oil.

BACKGROUND: The high similarity in anatomical and neurophysiological processes between pigs and humans make pigs an excellent model for metabolic diseases and neurological disorders. Lipids are essential for brain structure and function, and the polyunsaturated fatty acids (PUFA) have anti-inflammatory and positive effects against cognitive dysfunction in neurodegenerative diseases. Nutrigenomics studies involving pigs and fatty acids (FA) may help us in better understanding important biological processes. In this study, the main goal was to evaluate the effect of different levels of dietary soybean oil on the lipid profile and transcriptome in pigs' brain tissue. RESULTS: Thirty-six male Large White pigs were used in a 98-day study using two experimental diets corn-soybean meal diet containing 1.5% soybean oil (SOY1.5) and corn-soybean meal diet containing 3.0% soybean oil (SOY3.0). No differences were found for the brain total lipid content and FA profile between the different levels of soybean oil. For differential expression analysis, using the DESeq2 statistical package, a total of 34 differentially expressed genes (DEG, FDR-corrected p-value < 0.05) were identified. Of these 34 DEG, 25 are known-genes, of which 11 were up-regulated (log2 fold change ranging from + 0.25 to + 2.93) and 14 were down-regulated (log2 fold change ranging from - 3.43 to -0.36) for the SOY1.5 group compared to SOY3.0. For the functional enrichment analysis performed using MetaCore with the 34 DEG, four pathway maps were identified (p-value < 0.05), related to the ALOX15B (log2 fold change - 1.489), CALB1 (log2 fold change - 3.431) and CAST (log2 fold change + 0.421) genes. A "calcium transport" network (p-value = 2.303e-2), related to the CAST and CALB1 genes, was also identified. CONCLUSION: The results found in this study contribute to understanding the pathways and networks associated with processes involved in intracellular calcium, lipid metabolism, and oxidative processes in the brain tissue. Moreover, these results may help a better comprehension of the modulating effects of soybean oil and its FA composition on processes and diseases affecting the brain tissue.

Differential Gene Expression Associated with Soybean Oil Level in the Diet of Pigs.

The aim of this study was to identify the differentially expressed genes (DEG) from the skeletal muscle and liver samples of animal models for metabolic diseases in humans. To perform the study, the fatty acid (FA) profile and RNA sequencing (RNA-Seq) data of 35 samples of liver tissue (SOY1.5, n = 17 and SOY3.0, n = 18) and 36 samples of skeletal muscle (SOY1.5, n = 18 and SOY3.0, n = 18) of Large White pigs were analyzed. The FA profile of the tissues was modified by the diet, mainly those related to monounsaturated (MUFA) and polyunsaturated (PUFA) FA. The skeletal muscle transcriptome analysis revealed 45 DEG (FDR 10%), and the functional enrichment analysis identified network maps related to inflammation, immune processes, and pathways associated with oxidative stress, type 2 diabetes, and metabolic dysfunction. For the liver tissue, the transcriptome profile analysis revealed 281 DEG, which participate in network maps related to neurodegenerative diseases. With this nutrigenomics study, we verified that different levels of soybean oil in the pig diet, an animal model for metabolic diseases in humans, affected the transcriptome profile of skeletal muscle and liver tissue. These findings may help to better understand the biological mechanisms that can be modulated by the diet.

Weight-loss in obese dogs promotes important shifts in fecal microbiota profile to the extent of resembling microbiota of lean dogs.

BACKGROUND: Among the undesirable changes associated with obesity, one possibility recently raised is dysbiosis of the intestinal microbiota. Studies have shown changes in microbiota in obese rats and humans, but there are still few studies that characterize and compare the fecal microbiota of lean, obese and dogs after weight loss. Thus, this study aimed to evaluate the effects of a weight loss program (WLP) in fecal microbiota of dogs in addition to comparing them with those of lean dogs. Twenty female dogs of different breeds, aged between 1 and 9 years were selected. They were equally divided into two groups: Obese group (OG), with body condition score (BCS) 8 or 9/9, and body fat percentage greater than 30%, determined by the deuterium isotope dilution method, and lean group (LG) with BCS 5/9, and maximum body fat of 15%. Weight loss group (WLG) was composed by OG after loss of 20% of their current body weight. Fecal samples were collected from the three experimental groups. Total DNA was extracted from the feces and these were sequenced by the Illumina methodology. The observed abundances were evaluated using a generalized linear model, considering binomial distribution and using the logit link function in SAS (p < 0.05). RESULTS: The WLP modulated the microorganisms of the gastrointestinal tract, so that, WLG and LG had microbial composition with greater biodiversity than OG, and intestinal uniformity of the microbiota (Pielou's evenness index) was higher in OG than WLG dogs (P = 0.0493) and LG (P = 0.0101). In addition, WLG had values of relative frequency more similar to LG than to OG. CONCLUSION: The fecal microbiota of the studied groups differs from each other. The weight loss program can help to reverse the changes observed in obese dogs.

Evaluation of Serum and Urine Amino Acids in Dogs with Chronic Kidney Disease and Healthy Dogs Fed a Renal Diet.

This observational study aimed to evaluate serum and urinary amino acid (AA) concentrations in healthy dogs and dogs with chronic kidney disease (CKD) fed a commercial therapeutic renal diet with reduced protein and phosphorus levels. Ten dogs with CKD stages 3 or 4 composed the study group and received the renal diet for 180 days (RG T180). A control group (CG T30) composed of seven healthy dogs was fed a renal diet for 30 days. When comparing serum AA between RG T180 and CG T30, histidine, isoleucine, leucine, lysine, phenylalanine, tryptophan, cysteine, citrulline, ornithine, taurine, branched-chain amino acids (BCAA), and total essential amino acids (EAA) were higher in RG T180. Meanwhile, arginine, asparagine, aspartate, glutamine, serine, and tyrosine were higher in CG T30. Serum phenylalanine, tryptophan, and hydroxyproline were higher in RG T0 (dogs with CKD before consuming a renal diet) when compared to RG T180. In addition, the serum ratios of arginine/citrulline, tyrosine/phenylalanine, and serine/glycine were higher in CG T30 than in RG T180. Concerning urinary AA concentrations in CKD dogs, isoleucine, phenylalanine, tryptophan, aspartate, cysteine, and BCAA were higher in RG T180. In urine, the total EAA/total non-essential AA ratio in RG T180 was higher than in CG T30 as well as tyrosine/phenylalanine ratio higher in CG T30. In conclusion, the combination of renal diet and conservative treatment over 6 months in dogs with CKD stages 3 or 4 affected the AAs metabolism when compared to healthy adult dogs.

Influence of number of ingredients, use of supplement and vegetarian or vegan preparation on the composition of homemade diets for dogs and cats.

BACKGROUND: Homemade diets are a worldwide trend, and many recipes are currently available on websites but may not be considered balanced. This study aimed to evaluate if the number of ingredients, supplement, or vegetarian/vegan-only ingredients included in a recipe influence the nutrient content of homemade diets for dogs and cats. Chemical analyses were performed on 75 diets for dogs and 25 for cats prepared according to websites' recipes, and minerals were analyzed by inductively coupled plasma optical emission spectrometry (ICP-OES). RESULTS: Canine diets that met fat requirements had more ingredients than those below recommendations. None of the cat diets met iron requirements, and feline diets that met requirements of manganese had fewer ingredients and those that supplied requirements of protein and sodium had more ingredients than the diets below recommendations (p < 0.05). Non-supplemented canine and feline diets had calcium and calcium:phosphorus ratio below recommendations (p < 0.05). Non-supplemented feline diets had lower sodium and iron, and zinc levels were below recommendation in diets for both species. Diets with animal products for dogs had higher levels of protein and zinc, although zinc was deficient in both groups, and there were higher amounts of crude fiber, magnesium, and manganese in vegetarian/vegan diets (p < 0.05). Diets with animal products for cats had higher levels of protein (p = 0.003), but there was a higher amount of crude fiber (p = 0.014) in vegetarian/vegan diets. CONCLUSION: The number of ingredients and vegetarian/vegan preparation did not guarantee nutritional adequacy of diets, and the presence of supplement did not ensure a balanced diet.

Toxic element levels in ingredients and commercial pet foods.

Nowadays, there is a growing concern about contamination of toxic metals (TM) in pet food due to the great potential for health risks of these elements. TM concentrations in commercial pet foods (n = 100) as well as in ingredients used in their composition (n = 100) were analyzed and compared to the Food and Drug Administration (FDA) maximum tolerable level (MTL), and the TM concentrations found in the different sources of carbohydrate, protein, and fat were compared. The TM concentrations were determined by inductively coupled plasma with optical emission spectrometry (ICP-OES). Concentrations above the MTL for aluminum, mercury, lead, uranium, and vanadium were observed in both dog and cat foods, and the percentage of dog foods that exceeded the MTL of these TM were: 31.9%; 100%; 80.55%; 95.83%; and 75%, respectively, and in cat foods: 10.71%; 100%; 32.14%; 85.71%; 28.57%, respectively. The MTL values of these TMs and the mean values in dog foods (mg/kg dry matter basis) (MTL [mean ± standard deviation]) were: aluminum: 200 (269.17 ± 393.74); mercury: 0.27 (2.51 ± 1.31); lead: 10 (12.55 ± 4.30); uranium: 10 (76.82 ± 28.09); vanadium: 1 (1.35 ± 0.69), while in cat foods were: aluminum: 200 (135.51 ± 143.95); mercury: 0.27 (3.47 ± 4.31); lead: 10 (9.13 ± 5.42); uranium: 10 (49.83 ± 29.18); vanadium: 1 (0.81 ± 0.77). Dry foods presented higher concentrations of most TM (P < 0.05) than wet foods (P < 0.05). Among the carbohydrate sources, there were the highest levels of all TM except cobalt, mercury, and nickel in wheat bran (P < 0.05), while among the protein sources, in general, animal by-products had higher TM concentrations than plant-based ingredients. Pork fat had higher concentrations of arsenic, mercury, and antimony than fish oil and poultry fat. It was concluded that the pet foods evaluated in this study presented high concentrations of the following TM: aluminum, mercury, lead, uranium, and vanadium.

Gene expression of the immunoinflammatory and immunological status of obese dogs before and after weight loss.

Obesity is characterized by a low degree of chronic inflammation state that, along with metabolic modifications, promotes important changes in the animal's organism. Adipose tissue actively participates in inflammation and immunity, and several defense cells of the organism may, therefore, be involved in the diversity found between obese and ideal weight individuals. Studies regarding this subject have shown immune cell changes in humans and rats, however, the literature is scarce in relation to dogs. Thus, the present study aimed to evaluate the gene expression profile of immunoinflammatory response and the lymphoproliferation of obese dogs before and after weight loss. Eight female dogs, neutered, of different breeds, aged between 1 and 8 years (4.74±3.19), obese, with body condition score (BCS) of 9 out of a 9-point scale and body composition determined by the deuterium isotope dilution method were included. The obese dogs were enrolled in a weight loss program and after losing 20% of their initial weight became a second experimental group. A third experimental group consisted of eight female dogs, neutered, aged between 1 and 8 years (3.11±0.78) and with ideal BCS (5 out of a 9-point scale). Gene expression of immunoinflammatory cytokines (resistin, leptin, adiponectin, TNF-α, IL-6, IL-8, and IL-10) was assessed by qRT-PCR and immunity was assessed by lymphoproliferative response using the flow cytometry technique. The data that presented normal distribution was evaluated by analysis of variance by the PROC MIXED of the SAS and when differences were detected, these were compared by the Tukey test. Regarding the gene expression data, the procedure PROC GLIMMIX was adopted and the methodology of generalized linear model was used, in which the Gama distribution proved to be adequate. Values of p<0.05 were considered significant. The mean weight loss period of the animals included in the study was 194.25 ± 28.31 days and the mean weekly weight loss rate was 1.02 ± 0.82%. The average fat mass, both in percentage (P<0.001) and in kilograms (P = 0.012), was higher in the obese group (40.88%; 8.91kg), returning to normal and without difference between the control group (19.16%; 3.01kg) and after weight loss (22.10%; 4.11kg). The weight loss program resulted in an increase in percentage of lean body mass (P = 0.001), 55.50% in obese animals vs 77.90% in obese dogs after weight loss, the latter with no difference when compared to the control group (80.84%). The obese group presented increased gene expression of resistin and IL-8 in relation to the weight loss group (P = 0.002). In adiponectin, the obese group presented increased mRNA gene expression when compared to the weight loss group (P = 0.003). The evaluation of lymphocyte proliferation showed differences between the group of obese animals before and after weight loss (P = 0.004). Weight loss resulted in an increase in the lymphoproliferation rate (18.48%) compared to obese dogs at the beginning of the study (10.71%). These results indicate that weight loss modulates the immunoinflammatory response of obese dogs and may present important benefits to health and longevity of dogs.

Genetic analysis of in-vitro embryo production traits in Dairy Gir cattle.

The potential of dams as oocyte donors can be a selection criterion for animal breeding programs, but also an involuntary driver of the process. In both cases, it is important to determine genetic components influencing the outcome of in vitro embryo production (IVEP). The objective of the present study was to perform a detailed genetic analysis for in vitro embryo production traits in Dairy Gir cows. A dataset containing 11,450 records of ovum pick-up (OPU) and in vitro fertilization (IVF) procedures from 2684 Dairy Gir donors was evaluated. Analyzed traits were number (NOV) and percentage (POV) of viable oocytes; number (NGI) and percentage (PGI) of grade I oocytes; number (NEMB) and percentage (PEMB) of viable embryos. All analyzes were performed using animal models by a Bayesian framework. Heritability estimates varied from 0.16 to 0.32 for count traits and from 0.01 to 0.06 for percentage traits. The proportion of the total variation represented by the additive genetic effect of sire (semen used in IVF) for NEMB and PEMB was 7% and 5% respectively. Associations between estimated breeding values from progeny tested bulls for IVEP traits, milk production, age at first calving and conformation traits were mainly low or close to zero. Results indicate that selection for IVEP traits is possible in Dairy Gir cattle and would not impair genetic progress for traits already considered as selection criteria. The NOV seems to be a promising target trait. However, a selection index could help to avoid the use of sires with negative genetic merit for percentage traits, minimizing possible deterioration in the long term.

Concentrations of macronutrients, minerals and heavy metals in home-prepared diets for adult dogs and cats.

Pet owners often don't acknowledge the need for home-prepared diet formulation by a trained professional and may use recipes from sources such as the internet. Macronutrient and mineral composition of home-prepared diets were analyzed and compared to NRC and FEDIAF recommendations, and heavy metal concentrations were analyzed and compared to FDA maximum tolerable levels (MTL) for dogs and cats. Recipes of home-prepared diets for adult dogs (n = 75) and cats (n = 25) were evaluated. Analyses of protein, fat, and fiber were performed according to AOAC, and mineral and heavy metal analyses were performed using inductively coupled plasma optical emission spectrometry (ICP-OES). None of the diets supplied recommended levels of all nutrients evaluated, and more than 84.0% of diets presented three or more nutrients below recommendations. Nutrients with most levels below recommendations were calcium and potassium in recipes for dogs and iron and zinc in recipes for cats. As for heavy metals, levels of lead, cobalt, mercury, uranium, and vanadium were above MTLs. Results suggest that home-prepared diets may be a health risk to dogs and cats if not properly formulated. Furthermore, the chronic heavy metal intake must be better elucidated in order to understand the full impact of results.

Effects of different sources and levels of copper on lipid metabolism in Merino × Texel lambs.

The study was conducted with 40 lambs Merino × Texel in order to determine the effects of 10 or 30 mg of Cu/kg DM of copper sulphate and copper-methionine on the lipid and cholesterol metabolism. The lambs were randomly distributed into 5 treatments, with 8 animals each. The treatments were: control, without cooper additions; 10 or 30 mg of Cu/kg DM in the form of copper sulphate; 10 or 30 mg of Cu/kg DM in the form of copper-methionine. The hepatic concentration of copper was higher in lambs supplemented with copper. Total cholesterol, HDL cholesterol, LDL cholesterol and triglycerides in the serum, as well as the SFTU did not differ between treatments. The supplementation of copper regardless of source or level reduced the concentration of cholesterol in the LT muscle, and reduced the concentration of GSH and increased the concentration of GSSG in the liver. Additionally, there was a minimal effect on the fatty acid profile of lipid in lambs.

Influence of post-thawing thermal environment on bovine sperm characteristics and in vitro fertility.

Our aim was to evaluate the effects of three thermal environments over time on kinetics, functionality and in vitro fertility of cryopreserved bovine spermatozoa. Four ejaculates from five bulls (n = 20) were cryopreserved. After thawing, semen was evaluated (0 hr), incubated for 4 hr in T36.0 (36.0°C), T38.0 (38.0°C) and T39.5 (39.5°C), and analysed every hour (1 hr, 2 hr, 3 hr, 4 hr). In vitro production of embryos was performed at 0 hr and 4 hr. Sperm motility and cell kinetics (Computer-Assisted Sperm Analysis) were impaired after 2 hr at T38.0 and T39.5 (p < 0.05). Flow cytometry revealed an increase in the cells with injured plasma membrane to 39.5°C and a general reduction in the mitochondrial potential over time (p < 0.05). In vitro fertility was impaired in all temperatures after 4 hr, but there was no difference between 36.0°C and 38.0°C. Our results suggest that the ex situ resilience of semen at 36.0°C after thawing with no major damage to the quality is limited to 3 hr. In normothermia or in thermal stress, sperm cells present a gradual reduction of movement and functionality, which were more significant after 1 hr of incubation. The in vitro production of embryos is impaired when the semen is kept in a thermal environment ≥36.0°C for 4 hr.

Label-free quantitative proteomic analysis reveals muscle contraction and metabolism proteins linked to ultimate pH in bovine skeletal muscle.

The purpose of this research was to investigate the causes and consequences of pHu variations in beef cattle. A group of 176 Nellore beef cattle was evaluated and classified into two different pHu groups: High (≥6.0, N = 17) and Normal (<5.8, N = 159). Plasma concentrations of cortisol and adrenocorticotropic hormone, lactate and glycogen muscular content, meat color, shear force and Longissimus thoracis muscle proteomic profile were evaluated and compared between pHu groups. Muscle glycogen content, meat color and shear force statistically differed between pHu groups. Label-free quantitative proteomic analysis revealed ten differentially abundant proteins between pHu groups, involved in metabolic processes and muscle contraction, which also were significantly correlated with pHu. Thirty-six and 31 proteins were exclusively present in Normal and High pHu group, respectively, which were related to TCA cycle, cortisol production, calcium regulation, and antioxidant function. The MYH7, UGP2, H2AFJ and VDAC3 were identified as potential indicators of pHu variations. CALM and NNT appeared to be interesting proteins to understand the metabolic pathways behind pHu. Data are available via ProteomeXchange with identifier PXD009320.

Sex steroids modulate morphological and functional features of the bovine oviduct.

In cattle, the oviduct plays a major role in the reproductive process; however, molecular control of oviduct receptivity to the embryo is poorly understood. A model for receptivity based on size of the pre-ovulatory follicle (POF) was used to compare oviductal morphology, cellular proliferation, and candidate transcript abundance. Growth of the POF of Nelore (Bos indicus) cows was manipulated to produce two groups: a large POF-large corpus luteum (CL) group (LF-LCL; greater receptivity) and a small POF-small CL group (SF-SCL). Samples of the ampulla and isthmus ipsilateral and contralateral to CL were collected 4 days after GnRH-induced ovulation. Tissues were either embedded in paraffin for Harris-Hematoxylin and Eosin and periodic acid-Schiff staining and KI67 immunostaining, followed by morphological analyses, or stored at -80 °C for RNA extraction, cDNA synthesis, and qPCR analyses. The effects of group (LF-LCL and SF-SCL), region (ampulla and isthmus), and side (ipsilateral and contralateral) were analyzed using three-way nested ANOVA. The ipsilateral ampulla of the LF-LCL group presented more primary mucosal folds, a greater mucosal-folding grade and luminal perimeter, and more secretory cells and proliferating cells when compared with the ampulla of the SF-SCL group and with the contralateral ampulla of both groups. There were no morphological differences in the isthmus between groups and sides. Changes in transcript abundance are suggestive of LF-LCL-stimulated secretory activity. In summary, ovulation of a larger POF generates a periovulatory endocrine milieu that modulates morphological and functional features of the bovine oviduct which may support embryo survival and development.

Uterine lavage is efficient to recover endometrial cytology sample and does not interfere with fertility rate after artificial insemination in cows.

Productivity rates directly depend on the fertility of a herd, which in turn can be influenced by many factors. Semen deposited in the female reproductive tract is foreign to the body and, in response to this invasion, produces an inflammatory reaction, which is characterized by rapid infusion of polymorphonuclear (PMN) cells. Techniques to obtain an endometrial sample are usually invasive and can mask the true inflammatory response. Ultrasound is a noninvasive technique and can contribute to the diagnosis of postartificial insemination (AI) inflammatory response in cattle. The present study was divided into two experiments. The aim of experiment 1 was to compare two methods of endometrial cytology collection, uterine cytobrush (UC) and uterine lavage (UL), and their effects on uterine hemodynamics that provide information about blood flow. The two methods were evaluated by Doppler ultrasound using the spectral and color modes. For that purpose, 19 Nellore cows were synchronized for timed AI and subjected to UC (n = 9) or UL (n = 10). The techniques were performed 4 hours after AI. The results showed that both techniques allow collection of a good quality sample and with enough PMN cells to perform counting. More PMN cells were obtained by UL than UC. There was no difference in uterine blood flow between the UC and UL groups in any of the periods evaluated (34 hours before and 4, 24, and 48 hours after collection of uterine sample). On the basis of results of experiment 1, the effect of UL on fertility was studied in experiment 2. A total of 128 Nellore cows were synchronized for TAI; 35 cows were subjected to endometrial cytology by UL 4 hours after AI, and 93 were not submitted to any procedure (control). Pregnancy diagnosis was performed by transrectal ultrasound 30 days after AI. Pregnancy rates did not differ between UL (54.29%) and control (56.99%) groups. The results of this study showed that UL allows the collection of more representative cells of the surface of the uterus than UC technique and causes no damage to the reproductive tract. Moreover, UL did not affect pregnancy rate when performed 4 hours after AI.

Differential gene expression and developmental competence in in vitro produced bovine embryos.

The embryonic developmental block occurs at the 8-cell stage in cattle and is characterized by a lengthening of the cell cycle and an increased number of embryos that stop development. The maternal-embryonic transition arises at the same stage resulting in the transcription of many genes. Gene expression studies during this stage may contribute to the understanding of the physiological mechanisms involved in the maternal-embryonic transition. Herein we identified genes differentially expressed between embryos with high or low developmental competence to reach the blastocyst stage using differential display PCR. Embryos were analysed according to developmental kinetics: fast cleavage embryos showing 8 cells at 48 h post insemination (hpi) with high potential of development (F8), and embryos with slow cleavage presenting 4 cells at 48 hpi (S4) and 8 cells at 90 hpi (S8), both with reduced rates of development to blastocyst. The fluorescence DDPCR method was applied and allowed the recovery of 176 differentially expressed bands with similar proportion between high and low development potential groups (52% to F8 and 48% in S4 and S8 groups). A total of 27 isolated fragments were cloned and sequenced, confirming the expected primer sequences and allowing the identification of 27 gene transcripts. PI3KCA and ITM2B were chosen for relative quantification of mRNA using real-time PCR and showed a kinetic and a time-related pattern of expression respectively. The observed results suggest the existence of two different embryonic genome activation mechanisms: fast-developing embryos activate genes related to embryonic development, and slow-developing embryos activate genes related to cellular survival and/or death.

Ooplast-mediated developmental rescue of bovine oocytes exposed to ethidium bromide.

Ooplasm transfer has been used successfully to treat infertility in women with ooplasmic insufficiency and has culminated in the birth of healthy babies. To investigate whether mitochondrial dysfunction is a factor in ooplasmic insufficiency, bovine oocytes were exposed to ethidium bromide, an inhibitor of mitochondrial DNA replication and transcription, during in-vitro maturation (IVM). Exposure of immature oocytes to ethidium bromide for 24h during IVM hampered meiotic resumption and the migration of cortical granules. However, a briefer treatment with ethidium bromide during the last 4h of IVM led to partial arrest of preimplantation development without affecting oocyte maturation. Ooplasm transfer was then performed to rescue the oocytes with impaired development. In spite of this developmental hindrance, transfer of normal ooplasm into ethidium bromide-treated oocytes resulted in a complete rescue of embryonic development and the birth of heteroplasmic calves. Although this study unable to determine whether developmental rescue occurred exclusively through introduction of unaffected mitochondria into ethidium bromide-damaged oocytes, e.g. ethidium bromide may also affect other ooplasm components, these results clearly demonstrate that ooplasm transfer can completely rescue developmentally compromised oocytes, supporting the potential use of ooplasm transfer in therapeutic applications.

Placental microstructure and efficiency in cloned bovines: a design-based stereological approach.

Somatic nuclear transfer allows scientists to produce identical copies of individuals (clones) and offers the potential for research in various fields. However, the production of cloned bovines is inefficient, partly because of high rates of embryonic mortality associated with abnormal placental development, although the exact mechanisms are still unclear. In the present study, we have applied design-based stereological methods to quantify placental morphology in cloned and non-cloned Nelore bovines (280-297 days of gestation). We have found no differences for any of the following parameters: number and volume of placentomes, villous volume and surface area, volume and number of star volume units and placental efficiency. Therefore, placental deficiency is unlikely to be attributable to its villous component. Nevertheless, new studies should focus on other placental constituents, e.g. capillaries, trophoblast cells and feto-maternal exchange surface area.

Postnatal-related changes in the size and total number of neurons in the caudal mesenteric ganglion of dogs: total number of neurons can be predicted from body weight and ganglion volume.

Aging is mostly characterized by a progressive decline of neuronal function that involves both the central and the peripheral nervous system. The aging process is accompanied by changes in either the number or the size of neurons. However, these data are controversial and not very well known in the sympathetic ganglia of large mammals. Hence, the present investigation aimed to study the dog's caudal mesenteric ganglion (CMG) in three different periods of postnatal development, searching for qualitative and quantitative alterations. The CMG is responsible for the large intestine, internal anal sphincter, and partially the urogenital system innervations. Nine dead male dogs from the Veterinary Hospital of the College of Veterinary Medicine at University of São Paulo were divided into three well-defined age groups (1-2 months old, 1-2 years old, and 5-10 years old). The stereological study was pursued using the physical disector method combined to the Cavalieri principle. The postnatal development was accompanied by an increase in the nonneuronal tissue amount and in ganglion volume. Additionally, the total number of neurons also increased during aging (from 70,140 to 1,204,516), although the neuronal density showed an opposite trend (from 29,911 to 11,500 mm(-3)). Due to the interrelation between either body weight or ganglion volume and aging in the dogs investigated in this study, it was possible to predict the total number of neurons in CMG using both body weight and ganglion volume in an attempt to verify whether or not size and total number of neurons are both allometrically and aging ruled, i.e., if either the animal's body weight and ganglion volume or aging influence these parameters. The prediction of the total number of neurons was very close to the initially estimated values.