The aim of this study was to evaluate the influence of epidermal growth factor (EGF) on in vitro maturation of canine oocytes at different times of the process. Ovaries were collected from 55 bitches considered healthy and aseptically isolated, immersed in physiological solution (0.9% NaCl) and transported under refrigeration. Grade 1 cumulus-oocyte complexes (COCs) were selected and divided into two groups: control group (CG) and treatment group (TG). In CG 698 grade I COCs were placed in 4-well plates containing TCM-199 medium supplemented with 25 mM HEPES, 100 IU/mL penicillin, 100 mg/mL streptomycin, 26 mM sodium bicarbonate, 1.5 mM sodium pyruvate, 2.9 mM sodium lactate pentahydrate, 0.6 mM cysteine, 0.03 IU/mL hCG, 0.5 μg/mL FSH, 20 μg/mL estrogen at 38.5ºC in a humidified atmosphere of 5% CO2 in times of 24 h, 48 h, and 72 h. In TG 547 COCs received the same maturation medium plus 10 ηg/mL EGF. Logistic regression models (SAS, 2011) were constructed in order to estimate the chances of oocytes being observed at nuclear maturation stages in different culture times (24 h, 48 h, and 72 h). Based on the results found EGF-supplemented medium showed 2.56 times more chances of having an oocyte at metaphase I (M-I) than medium without EGF (p < 0.0001). The results of this study demonstrated that the time of 72 h showed 5.88 times more chances of having an oocyte at metaphase II (M-II) compared to time of 24 h (p = 0.0001) and 7.69 times more chance than time of 48 h (p = 0.0001). The chances of finding an oocyte at M-II were also 9.09 times higher in medium supplemented with EGF than in medium without EGF (p = 0.0001). Thus, these results demonstrated the essential importance of EGF at different moments of oocyte maturation, being a key component for the acquisition of meiotic competence in bitches, increasing the M-I and M-II rates.

Introduction: Canine transmissible venereal tumour (CTVT) is a sexually transmitted tumour affecting dogs worldwide, imposing a financial burden on dog owners. A stable culture cell line in continuous passages for >18 months has only been achieved once. The present study investigated a stable CTVT cell line isolated from a bitch and its potential as a vaccine. Material and Methods: A biopsy from a 2-year-old mongrel bitch with CTVT was obtained for histopathological confirmation and isolation of tumour cells. The isolated cells were cultured to passage 55 and characterised by flow cytometry, with karyotyping by GTG-banding and by PCR detection of myc S-2 and LINE AS1. The isolated CTVT cell line was also used as a preventive vaccine in a canine model. Results: Histopathological analysis of the isolated tumour cells revealed typical CTVT characteristics. Constant proliferation and stable morphological characteristics were observed during culture. Phenotypic analysis determined the expression of HLA-DR⁺, CD5.1⁺, CD14⁺, CD45⁺, CD83⁺, CD163⁺, and Ly-6G-Ly-6C⁺. GTG-banding revealed a mean of 57 chromosomes in the karyotype with several complex chromosomal rearrangements. LINE-c-myc insertion in the isolated CTVT cell line at 550 bp was not detected. However, a 340-bp band was amplified. Isolated CTVT cell line inoculation at a concentration of 1×10⁸ did not induce tumour growth in bitches, nor did a challenge with primary CTVT cells. Conclusion: The present study successfully identified and isolated a stable CTVT cell line that may be useful in CTVT prevention.

Introduction: Significant improvement of sperm motility within one month effected by oral supplementation of selenium and vitamin E was described in four infertile male dogs which failed to conceive in their last three matings with different bitches. Material and Methods: The dogs (a Golden Retriever, an English Cocker Spaniel, and two Tibetan Mastiffs) were supplemented daily with selenium (Se) (0.6 mg/kg organic Se yeast) and vitamin E (vit. E) (5 mg/kg) per os for 60 days. Semen was collected on days 0, 30, 60, and 90. The sperm concentration and motility parameters were evaluated by the CASA system, sperm morphology was explored by Diff-Quick staining, and live and dead spermatozoa were differentiated by eosin/nigrosin staining. The concentrations of Se and vit. E were measured in peripheral blood serum on semen collection days. Results: Before administration, the concentrations of Se in blood plasma were low (86.0–165.0 µg/L). After 30 days of treatment there was an observable improvement in total and progressive sperm motility and kinematic parameters (VAP, VSK, VCL, ALH, BCF, and RAPID). The percentages of live and normal morphology sperm cells were also higher. There was also an observable increase in Se and vitamin E concentrations in blood serum. Bitches were successfully mated and delivered four to six puppies. Conclusion: Supplementation with Se and vit. E improved rapid sperm motility and restored fertility in infertile dogs with low Se status.

The aim of this study was to determine the viability and cardiorespiratory effects of the association of epidural alpha-2 adrenergic agonists and lidocaine for ovariohysterectomy (OH) in bitches. Forty-two bitches were spayed under epidural anesthesia with 2.5 mg/kg body weight (BW) of 1% lidocaine with adrenaline (CON) or in association with 0.25 mg/kg BW of xylazine (XYL), 10 μg/kg BW of romifidine (ROM), 30 μg/kg BW of detomidine (DET), 2 μg/kg BW of dexmedetomidine (DEX), or 5 μg/kg BW of clonidine (CLO). Heart rate (HR), respiratory rate (fR) and arterial pressures were monitored immediately before and every 10 min after the epidural procedure. Blood gas and pH analysis were done before, and at 30 and 60 min after the epidural procedure. Animals were submitted to isoflurane anesthesia if they presented a slightest sign of discomfort during the procedure. Time of sensory epidural block and postoperative analgesia were evaluated. All animals in CON and DEX, 5 animals in ROM and CLO, 4 animals in XYL, and 3 in DET required supplementary isoflurane. All groups, except CLO, showed a decrease in HR. There was an increase in arterial pressures in all groups. Postoperative analgesia lasted the longest in XYL. None of the protocols were totally efficient to perform the complete procedure of OH; however, xylazine provided longer postoperative analgesia than the others.

Breast tumors are the most common tumors in dogs and the study of disease prognostic factors is important for establishing the appropriate treatment protocols. The purpose of this study was to clinically stage mammary tumors of bitches and correlate the stages with histological type and grade. The tumors of 63 dogs were clinically staged based on the findings of tumor sizing, lymph node evaluation, and radiographic examination. After surgical excision, the tumors were classified histologically and graded. The relationship between the tumor grade, stage, and histological type was evaluated using a binomial test. Stage I tumors were the most numerous (31.75%), followed by tumors at stages II, III, IV, and V. Animals with histological grade I carcinomas presented stage I, II, or III tumors more frequently and stage IV and V tumors less frequently. The number of animals with simple carcinomas that were at stage I of the disease was greater than that at stage V. Carcinomas in the mixed tumors were less aggressive; however, the small number of animals in stage V of the disease made any statistical association impossible. The complex carcinomas presented with the invasion of the lymph nodes and less cellular differentiation in a larger number of animals than did simple carcinomas. Histological grading proved to be the best parameter for the prognostic evaluation of the breast carcinomas.

Objective: To determine the effects of hypothyroidism on insulin sensitivity, glucose tolerance, and concentrations of hormones counter-regulatory to insulin in dogs. Animals: 8 anestrous mixed-breed bitches with experimentally induced hypothyroidism and 8 euthyroid control dogs. Procedures: The insulin-modified frequently sampled IV glucose tolerance test and minimal model analysis were used to determine basal plasma insulin and glucose concentrations, acute insulin response to glucose, insulin sensitivity, glucose effectiveness, and disposition index. Growth hormone response was assessed by stimulation and suppression tests. Additionally, basal serum growth hormone (GH) and insulin-like growth factor-1 (IGF-1) concentrations and urine cortisol-to-creatinine concentration ratios were measured and dual energy x-ray absorptiometry was performed to evaluate body composition. Results: Insulin sensitivity was lower in the hypothyroid group than in the euthyroid group, whereas acute insulin response to glucose was higher. Glucose effectiveness and disposition index were not different between groups. Basal serum GH and IGF-1 concentrations as well as abdominal fat content were high in hypothyroid dogs, but urine cortisol-to-creatinine concentration ratios were unchanged. Conclusions and Clinical Relevance: Hypothyroidism appeared to negatively affect glucose homeostasis by inducing insulin resistance, but overall glucose tolerance was maintained by increased insulin secretion in hypothyroid dogs. Possible factors affecting insulin sensitivity are high serum GH and IGF-1 concentrations and an increase in abdominal fat. In dogs with diseases involving impaired insulin secretion such as diabetes mellitus, concurrent hypothyroidism can have important clinical implications.

OBJECTIVE To investigate effects of metoclopramide orally administered to healthy bitches on serum prolactin and milk lactose concentrations, gross energy, and dry matter content and on puppy weight gain during early lactation. ANIMALS 20 client-owned bitches and their 121 puppies. PROCEDURES 10 bitches received metoclopramide (0.2 mg/kg, PO, q 6 h for 6 days; treatment group) starting 10 to 24 hours after birth of the last puppy of the litter (day 0), and 10 bitches served as the control group. Blood and milk samples from all bitches were collected on days 0, 1, 2, 4, and 6. Milk samples for days 1 and 2 and days 4 and 6 were pooled because of small volume. Puppies were weighed twice daily. RESULTS Serum prolactin concentration increased significantly over time in both groups, and no treatment effect was detected. When day-to-day changes were analyzed, the prolactin concentration increased from day 0 to day 1 in the treatment group but not in the control group. Milk lactose concentration increased significantly and was higher in the treatment group than in the control group. Milk dry matter content was unchanged, whereas the time course for milk gross energy content differed significantly between treatment and control bitches. Puppy weight gain was not affected by metoclopramide treatment. CONCLUSIONS AND CLINICAL RELEVANCE Oral administration of metoclopramide to healthy bitches after parturition induced a transient increase in serum prolactin concentration and stimulated milk lactose production. It is likely bitches with insufficient or delayed milk production could benefit from metoclopramide treatment.