OBJECTIVE To develop a technique for isolation and culture of canine insulinoma cells and assess expression of cellular hexokinases (glucokinase and hexokinase I) and expression and secretion of insulin from these cells in vitro. SAMPLE Pancreatic insulinomas and normal pancreatic tissue from 4 and 3 dogs, respectively. PROCEDURES Tissues were collected by surgical excision or at necropsy. Insulinoma cells from 2 dogs were cultured for up to 10 weeks with standard techniques; insulin synthesis in vitro was confirmed by immunohistochemical analysis of freshly prepared slides of cultured cells, and insulin secretion was assessed by measurement of insulin concentrations in culture medium with an ultrasensitive mouse insulin ELISA. Expression of cellular hexokinases in insulinomas and adjacent normal (nontumor) pancreatic tissue from the same dog (n = 3) was examined by quantitative reverse transcriptase PCR assay. RESULTS Insulinoma cells survived for up to 10 weeks but did not proliferate in culture. Insulin was detected in isolated cells and secreted into culture medium for up to 10 weeks. Both cellular hexokinases were expressed; glucokinase appeared to be overexpressed in insulinomas, compared with normal pancreatic tissue from the same dogs. CONCLUSIONS AND CLINICAL RELEVANCE Canine insulinomas expressed hexokinases responsible for glucose responsiveness. Insulinoma cells were successfully maintained in short-term culture; cultured cells remained functional for 10 weeks as evidenced by cellular insulin content and had detectable secretion of insulin into the culture medium for ≥ 5 weeks. Apparent glucokinase overexpression by insulinomas suggested a possible mechanism underlying excessive insulin release by these tumors.

OBJECTIVE To assess differences in skin shrinkage between grossly visible tumor and grossly normal marginal skin of dogs for cutaneous mast cell tumors (MCTs) excised with curative intent and to determine an equation to estimate postexcisional gross tumor margins from preexcisional measurements and vice versa. SAMPLE 19 cytologically confirmed and surgically excised cutaneous MCTs obtained from dogs. PROCEDURES Tumors were measured in craniocaudal and dorsoventral directions before excision, immediately after excision, and after fixation in formalin. Both grossly visible tumor and surrounding grossly normal skin that comprised the surgical margin were measured at each time point. Percentage of shrinkage was compared among time points and between the tumor and surrounding grossly normal skin. Patient and histopathologic variables were correlated to skin shrinkage. RESULTS Overall shrinkage was 17.70%. The amount of shrinkage within the grossly visible tumor (4.45%) was less than that within the surrounding grossly normal skin (24.42%). Most of the shrinkage occurred immediately after excision. There was no effect of age, sex, completeness of excision, or degree of edema. Accuracy of an equation to estimate postexcisional margins from preexcisional measurements was only 18.4%. CONCLUSIONS AND CLINICAL RELEVANCE Grossly evident MCTs of dogs shrunk less than did the grossly normal surrounding skin. Although an equation to estimate postexcisional margins from preexcisional measurements could be derived, it likely would need to contain additional variables not included in the study reported here. Until such an equation exists, care must be used when extrapolating surgical margins from histologic margins and vice versa.

The aim of this study was to quantify the longitudinal shrinkage of canine small intestinal specimens after resection and fixation in 10% formalin. Samples were obtained from 12 clinically normal dogs of medium to large breed via ventral midline coeliotomy and enterectomy. The length of each sample was measured before excision, immediately after excision, and after 24 h in 10% formalin. The results were interpreted with the use of single-sample t-tests of the average changes; P-values of less than 0.01 were considered significant. The samples indicated a significant decrease in length after resection and fixation. The mean shrinkage from the pre-excision state was 28.3% immediately after excision (P < 0.0001) and 26.3% after 24 h of fixation (P < 0.0001). There was a small but not significant increase in the length of the specimens between the 2nd and 3rd measurement points. Quantification of the longitudinal shrinkage of resected intestinal specimens may improve interpretation of the distance of surgical margins from abnormal tissue in histopathology reports and allow investigation of the margins required for the clearance of specific tumors.

In a controlled study, malignant murine P815 mastocytoma cells exposed in vitro to distilled and deionized water died as a result of progressive swelling, degranulation, and membrane rupture. A 90% mean cell death occurred when cells obtained directly from culture were exposed to deionized water for 2 minutes. Of 6 cryopreserved malignant murine cell lines, which included Cloudman S91 melanoma, CMT-93 rectum carcinoma, MMT-06052 mammary carcinoma, and S-180 Sarcoma, only P815 mastocytoma and YAC-1 lymphoma were significantly (P < 0.05) affected by hypotonic shock; Cloudman S91 melanoma cells were the most resistant. Mastocytoma cells were selectively killed by hypotonic solution, and lymphoma cells were also killed by isotonic saline solution. Local mast cell tumor (MCT) recurrence and percentage survival were evaluated in 12 cats (21 MCT) and 54 dogs (85 MCT) subjected to surgery alone or local infiltration of deionized water as an adjunct to surgery. Of all 16 incompletely excised MCT in cats, there was no local recurrence following injection. Four mast cell tumors (2 cats) regressed after being injected in situ. In dogs with clinical stage-I MCT, local recurrence was detected in 50% (5/10), but with injection after incomplete excision, local MCT recurrence was significantly (P < 0.05) less (6.6%, 1/15). Percentage recurrence was significantly (P < 0.05) less and survival significantly greater when incompletely excised grade-II MCT were injected. Mean follow-up period after surgery in cats and dogs was 35 and 23.4 months, respectively.

OBJECTIVE To assess the feasibility of a novel technique involving a vessel and tissue–sealing device (VTSD) for ovariectomy in chickens to evaluate the potential application of the procedure to other avian species. ANIMALS 20 domestic laying hens (Gallus domesticus), of which 10 were immature (< 4 months old) and 10 were adults (> 18 months old). PROCEDURES Ovariectomy was performed with a VTSD through a left lateral celiotomy. Birds were allowed to recover for 14 days after the procedure and then were euthanized for necropsy. A board-certified veterinary pathologist performed complete necropsies, with particular attention to identifying any remaining ovarian tissue. RESULTS All birds survived the procedure. For the mature and juvenile birds, the mean ± SD durations of anesthesia (interval from intubation to extubation) were 67.2 ± 7.6 minutes and 50.5 ± 5.1 minutes, respectively, and mean durations of surgery were 45.3 ± 8.5 minutes and 31.6 ± 5.1 minutes, respectively. Three birds had severe hemorrhage during ovariectomy. At necropsy, ovarian tissue was present grossly in 2 mature birds and histologically in 6 additional birds (2 mature and 4 juvenile birds), indicating incomplete excision in 8 (40%) birds. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that the evaluated VTSD can be used to successfully perform ovariectomies in both juvenile and mature chickens, although the procedure was associated with major hemorrhage and incomplete excision of ovarian tissue in some cases. Use of this VTSD for ovariectomy in birds of other species, particularly birds with reproductive tract disease, should be investigated.

OBJECTIVE To determine optimal sample preparation conditions with potassium triiodide (I2KI) and optimal imaging settings for microfocus CT (micro-CT) of excised cat hearts. SAMPLE 7 excised hearts (weight range, 10 to 17.6 g) obtained from healthy adult cats after euthanasia by IV injection of pentobarbital sodium. PROCEDURES Following excision, the hearts were preserved in 10% formaldehyde solution. Six hearts were immersed in 1.25% I2KI solution (n = 3) or 2.5% I2KI solution (3) for a 12-day period. Micro-CT images were acquired at time 0 (prior to iodination) then approximately every 24 and 48 hours thereafter to determine optimal sample preparation conditions (ie, immersion time and concentration of I2KI solution). Identified optimal conditions were then used to prepare the seventh heart for imaging; changes in voltage, current, exposure time, and gain on image quality were evaluated to determine optimal settings (ie, maximal signal-to-noise and contrast-to-noise ratios). Images were obtained at a voxel resolution of 30 μm. A detailed morphological assessment of the main cardiac structures of the seventh heart was then performed. RESULTS Immersion in 2.5% I2KI solution for 48 hours was optimal for sample preparation. The optimal imaging conditions included a tube voltage of 100 kV, current of 150 μA, and exposure time of 354 milliseconds; scan duration was 12 minutes. CONCLUSIONS AND CLINICAL RELEVANCE Results provided an optimal micro-CT imaging protocol for excised cat hearts prepared with I2KI solution that could serve as a basis for future studies of micro-CT for high resolution 3-D imaging of cat hearts.

OBJECTIVE To evaluate effects of anatomic location, histologic processing, and sample size on shrinkage of excised canine skin samples. SAMPLE Skin samples from 15 canine cadavers. PROCEDURES Elliptical samples of the skin, underlying subcutaneous fat, and muscle fascia were collected from the head, hind limb, and lumbar region of each cadaver. Two samples (10 mm and 30 mm) were collected at each anatomic location of each cadaver (one from the left side and the other from the right side). Measurements of length, width, depth, and surface area were collected prior to excision (P1) and after fixation in neutral-buffered 10% formalin for 24 to 48 hours (P2). Length and width were also measured after histologic processing (P3). RESULTS Length and width decreased significantly at all anatomic locations and for both sample sizes at each processing stage. Hind limb samples had the greatest decrease in length, compared with results for samples obtained from other locations, across all processing stages for both sample sizes. The 30-mm samples had a greater percentage change in length and width between P1 and P2 than did the 10-mm samples. Histologic processing (P2 to P3) had a greater effect on the percentage shrinkage of 10-mm samples. For all locations and both sample sizes, percentage change between P1 and P3 ranged from 24.0% to 37.7% for length and 18.0% to 22.8% for width. CONCLUSIONS AND CLINICAL RELEVANCE Histologic processing, anatomic location, and sample size affected the degree of shrinkage of a canine skin sample from excision to histologic assessment.

In an attempt to ablate the laryngeal saccule as an alternative method of sacculectomy (conventially done through a laryngotomy incision for laryngeal hemiplegia) a neodymium:yttrium aluminum garnet laser was used transendoscopically in noncontact fashion in 6 horses. The procedure was easy, quick, and labor-saving, with few complications. Endoscopically, the lased sacculectomy site appeared healed at postsurgical day 42. On postsurgical day 42, microscopic examination revealed mucosal remnants under the granulation bed. Laser energy caused thermal damage to tissues adjacent to the lased saccule. Additional work must be done to refine the technique before it can be recommended for clinical applications.

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.