Obtaining Wisent early blastocyst in vitro is a basic for protection and creation of biodiversity for this threatened species

Wisent, or European bison (Bison bonasus), is listed as “vulnerable” on the IUCN Red List of Threatened Species and is therefore protected by international law. For the first time, a Wisent embryo has been obtained in vitro. This procedure creates a new opportunity to protect and increase Wisent reproductive potential and thereby opens new possibilities for the establishment of a controlled and broad reserve of the gene pool.     

Cutaneous metastasis of primary appendicular osteosarcoma in a dog

A 6-year-old, neutered male Rottweiler was presented to the University of Illinois Veterinary Teaching Hospital because of a lytic bone lesion involving the distal portion of the right radius and possible pulmonary metastases on thoracic radiographs. Results of serum biochemical analysis were unremarkable. Aspiration and cytologic examination of the bone lesion indicated likely sarcoma with reactive bone. Cutaneous masses were found on the left thigh, interscapular region, and dorsal lumbar region, 4 weeks after initial presentation. Neoplastic spindle cells were found in aspirates from 2 of the masses. The neoplastic cells stained positive for alkaline phosphatase activity using cytochemistry. Re-evaluation of serum biochemical values at this time revealed a marked increase in alkaline phosphatase activity (413 U/L, reference interval 12-110 U/L) compared with the initial value (26 U/L). Due to progressive disease, the dog was euthanized and a necropsy was performed. Histologic findings included primary osteosarcoma of the distal portion of the right radius, with metastases in the lungs, spleen, left fourth and fifth ribs, soft tissue of the right medial thigh, and T1-T3/interscapular region. Cutaneous metastasis of primary appendicular osteosarcoma has been reported rarely in animals and humans. Increased serum alkaline phosphatase activity may be a potential indicator of poor prognosis for this neoplasm.     

Efficiency of Two Enucleation Methods Connected to Handmade Cloning to Produce Transgenic Porcine Embryos

The purpose of our work was to establish an efficient-oriented enucleation method to produce transgenic embryos with handmade cloning (HMC). After 41–42 h oocytes maturation, the oocytes were further cultured with or without 0.4 μg/ml demecolcine for 45 min [chemically assisted handmade enucleation (CAHE) group vs polar body (PB) oriented handmade enucleation (OHE) group respectively]. After removal of the cumulus cells and partial digestion of the zona pellucida, oocytes with visible extrusion cones and/or polar bodies attached to the surface were subjected to oriented bisection. Putative cytoplasts without extrusion cones or PB were selected as recipients. Two cytoplasts were electrofused with one transgenic fibroblasts expressing green fluorescent protein (GFP), while non-transgenic fibroblasts were used as controls. Reconstructed embryos were cultured in Well of Wells (WOWs) with porcine zygote medium 3 (PZM-3) after activation. Cleavage and blastocyst rates were registered on day 2 and day 7 of in vitro culture respectively. Meanwhile, the total blastocyst cell number was counted on day 7. We found that the difference was only observed between blastocyst rates (38.6 ± 2% vs 48.1 ± 3%) of cloned embryos with GFP transgenic fibroblast cells after CAHE vs OHE. With adjusted time-lapse for zonae-free cloned embryos cultured in WOWs with PZM-3, it was obvious that in vitro developmental competence after CAHE was compromised when compared with the OHE method. OHE enucleation method seems to be a potential superior alternative method used for somatic cell nuclear transfer (SCNT) with transgenic fibroblast cells.     

Assessment of corticosteroid-induced alkaline phosphatase as a prognostic indicator in canine lymphoma

OBJECTIVES: To examine the incidence of elevated corticosteroid-induced alkaline phosphatase (sALP) in dogs with lymphoma and to determine if sALP is a reliable prognostic indicator in canine lymphoma. METHODS: The medical records of 62 canine lymphoma patients treated with a combination chemotherapy protocol from 1994 to 2003 at the University of Illinois Veterinary Teaching Hospital were examined. Variables assessed with respect to response rate and remission duration included age, bodyweight, sex, breed, World Health Organization stage (I to V), substage (a or b), pretreatment administration of corticosteroid, and serum levels of alkaline phosphatase, sALP and alanine aminotransferase. RESULTS: sALP was not statistically significant with respect to response rate or duration of remission, nor was preinduction glucocorticoid administration. Stage was significant with respect to achieving remission. CLINICAL SIGNIFICANCE: It was found that sALP is not a useful prognostic indicator for response rate and remission duration in dogs with lymphoma.     

Blood-to-saliva glucose time lag in sedated healthy dogs

The management of diabetes mellitus mandates measurement of blood glucose. Saliva offers an alternative to blood sampling, but measurement of the salivary glucose concentration is difficult, and the blood-to-saliva glucose time lag is uncertain. We aimed to determine the serum–saliva glucose time lag in the saliva of healthy dogs. The combined duct of the mandibular and sublingual salivary glands of 6 dogs was cannulated to collect saliva and prevent glucose degradation by oral bacteria. Following a 0.25 g/kg IV bolus of dextrose, paired serum–saliva samples were collected at baseline and in twelve 5-min blocks over 60 min. Serum and salivary glucose levels were analyzed with a linear mixed model for repeated measures with a compound symmetry error structure. Mean (±SD) saliva production was 10.3 ± 2.9 µL/kg/min, and the area under the curve (AUC_glucose)_saliva/serum ratio was 0.006, which highlights the magnitude of the large difference in glucose concentration between the 2 compartments. The serum–saliva glucose time lag was 30–40 min.     

Mitochondrial Activity, Distribution and Segregation in Bovine Oocytes and in Embryos Produced in Vitro

Contents Bovine oocytes and embryos produced in vitro were studied to determine the mitochondrial pattern of distribution, segregation and activity using DIOC 6 and Jc-1 fluorescence. The highest fluorescence level observed in mature oocytes was taken as 100% activity and six activity levels were estimated as follows: (1) 0%, (2) 1-15%, (3) 16-30%, (4) 31-50%, (5) 51-75% and (6) 76-100%. Three patterns of mitochondrial distribution were found: (1) diffused throughout the cytoplasm in oocytes and embryos, (2) pericytoplasmic in oocytes and embryos, and (3) perinuclear only in embryos. The segregation of mitochondria in blastomeres showed two distinct patterns: (1) symmetrical with an even mitochondrial population, and (2) asymmetrical with different numbers of mitochondria in each blastomere. In immature oocytes, mitochondrial activity was very low and the distribution was diffuse or negligible, while in mature oocytes the activity was high and the distribution was diffuse or pericytoplasmic. Competent embryos up to the 16-cell stage showed intermediate levels of activity (16-50%) but activity decreased thereafter up to the blastocyst stage. Non-competent embryos showed low levels of activity (1-15%) at all stages. These results suggest that mitochondria might play an important role during early development and that a minimum threshold of activity regulates the potential competence for reaching the blastocyst stage.