Objective: To evaluate in vitro effects of gemcitabine alone and in combination with carboplatin on canine transitional cell carcinoma (TCC) cell lines. Sample: In vitro cultures of 5 canine TCC cell lines. Procedures: Cells were treated with gemcitabine, carboplatin, or a combination of both at various concentrations. Cell proliferation was assessed via a fluorescence-based microplate cell proliferation assay. Cell cycle was evaluated via propidium iodide staining, and apoptosis was assessed by measurement of caspase 3 and 7 enzymatic activity. Synergy between gemcitabine and carboplatin was quantified via combination index analyses. Results: Treatment of 5 canine TCC cell lines with gemcitabine or carboplatin decreased cell proliferation, increased apoptosis, and induced cell cycle arrest. Cell cycle arrest and apoptosis were markedly increased when cell lines were treated with both gemcitabine and carboplatin simultaneously or sequentially. Order of administration during sequential treatment did not consistently affect cell proliferation results in TCC cell lines. When TCC cell lines were treated with gemcitabine and carboplatin in combination at therapeutically relevant concentrations (gemcitabine concentration, < 10μM; carboplatin concentration, < 250μM), a significant decrease in cell proliferation was observed, compared with cell proliferation following treatment with gemcitabine or carboplatin alone. In combination, the effects of gemcitabine and carboplatin were synergistic in 3 of 5 cell lines and additive in the other 2. Conclusions and Clinical Relevance: Gemcitabine had antitumor effects on canine TCC cells in vitro, and the combination of gemcitabine and carboplatin had synergistic activity at biologically achievable concentrations.

Objective: To investigate the influence of oxidative stress in terms of antioxidant capacity and lipid peroxidation on the probability of motor neuron disease (MND) in horses. Animals: 88 horses with MND (cases) and 49 controls. Procedures: Blood samples were collected from all horses enrolled, and RBCs and plasma were harvested. Activity of the enzyme erythrocytic superoxide dismutase 1 (SOD1) was determined in the RBCs. Plasma concentrations of α-tocopherols and β-carotenes and activity of glutathione peroxidase were also evaluated. Degree of lipid peroxidation was measured by determining plasma concentrations of lipid hydroperoxides. Differences were evaluated between horse groups. Results: Cases had lower erythrocyte SOD1 activity than did controls, but the difference was not significant. On the other hand, plasma vitamin E concentrations differed significantly between groups, with the cases having lower concentrations. Neither plasma vitamin A concentration nor glutathione peroxidase activity differed between groups; however, cases had significantly higher concentrations of lipid hydroperoxides (18.53μM) than did controls (12.35μM). Conclusions and Clinical Relevance: Horses with MND differed from those without MND by having a lower plasma concentration of vitamin E and higher concentrations of lipid hydroperoxides. Results parallel the findings in humans with sporadic amyotrophic sclerosis and provide evidence supporting the involvement of oxidative stress in the 2 conditions.

The effects of selenium (Se) supplementation and source on equine immune function have not been extensively studied. This study examined the effects of oral Se supplementation and Se source on aspects of innate and adaptive immunity in horses. Fifteen horses were assigned to 1 of 3 groups (5 horses/group): control, inorganic Se (sodium selenite), organic Se (Se yeast). Immune function tests performed included: lymphocyte proliferation in response to mitogen concanavalin A, neutrophil phagocytosis, antibody production after rabies vaccination, relative cytokine gene expression in stimulated lymphocytes [interferon gamma (IFNγ), interleukin (IL)-2, IL-5, IL-10, tumor necrosis factor alpha (TNFα)], and neutrophils (IL-1, IL-6, IL-8, IL-12, TNFα). Plasma, red blood cell Se, and blood glutathione peroxidase activity were measured. Plasma and red blood cell Se were highest in horses in the organic Se group, compared with that of inorganic Se or control groups. Organic Se supplementation increased the relative lymphocyte expression of IL-5, compared with inorganic Se or no Se. Selenium supplementation increased relative neutrophil expression of IL-1 and IL-8. Other measures of immune function were unaffected. Dietary Se content and source appear to influence immune function in horses, including alterations in lymphocyte expression of IL-5, and neutrophil expression of IL-1 and IL-8.

Objective-To determine the effects of increases in dietary intake of polyunsaturated and saturated fatty acids on plasma lipid and lipoprotein concentrations and activity of associated enzymes in healthy domestic cats. Animals-16 healthy adult sexually intact female cats. Procedures-A baseline diet (40% energy from fat) and 4 test diets, with increased amounts of fat (51% and 66% energy from fat) from the addition of polyunsaturated and saturated fatty acids, were fed for 6 weeks each. Plasma cholesterol, triglyceride, and lipoprotein cholesterol concentrations, along with activities of lipoprotein lipase, hepatic lipase, and lecithin-cholesterol acyl transferase, were measured at the end of each feeding period. Results-Diet, amount of fat, or ratio of polyunsaturated to saturated fatty acids had no effect on plasma concentrations of cholesterol, triglycerides, and very–low-density or high-density lipoproteins or the activity of lecithin-cholesterol acyl transferase. Low-density lipoprotein concentrations were significantly lower in cats fed a high-fat diet containing polyunsaturated fatty acids. Lipoprotein concentration and hepatic lipase activity were significantly higher in cats fed the fat-supplemented diets, and this was unrelated to whether diets were enriched with polyunsaturated or saturated fatty acids. Conclusions and Clinical Relevance-Diets containing up to 66% of energy from fat were tolerated well by healthy cats and did not affect plasma lipid concentrations. Therefore, high-fat diets probably will not contribute to hypercholesterolemia or hypertriglyceridemia in cats.

Objective: To determine whether increased gene expression of a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4) in laminae of horses with starch gruel–induced laminitis was accompanied by increased enzyme activity and substrate degradation. Sample: Laminae from the forelimb hooves of 8 healthy horses and 17 horses with starch gruel–induced laminitis (6 at onset of fever, 6 at onset of Obel grade 1 lameness, and 5 at onset of Obel grade 3 lameness). Procedures: Gene expression was determined by use of cDNA and real-time quantitative PCR assay. Protein expression and processing were determined via SDS-PAGE and quantitative western blotting. Protein distribution and abundance were determined via quantitative immunofluorescent staining. Results: ADAMTS-4 gene expression was increased and that of versican decreased in laminitic laminae, compared with expression in healthy laminae. Catalytically active ADAMTS-4 also was increased in the tissue, as were ADAMTS-4–cleavage fragments of versican. Immunofluorescent analyses indicated that versican was depleted from the basal epithelia of laminae of horses at onset of Obel grade 3 lameness, compared with results for healthy laminae, and this was accompanied by regional separation of basal epithelial cells from the basement membrane. Aggrecan gene and protein expression were not significantly affected. Conclusions and Clinical Relevance: Changes in gene and protein expression of ADAMTS-4 and versican in the basal epithelium of laminitic laminae indicated a fundamental change in the physiology of basal epithelial cells. This was accompanied by and may have caused detachment of these cells from the basement membrane.

Objective: To compare mitochondrial complex I and complex IV activity in myocardial mitochondria of clinically normal dogs, clinically normal dogs exposed to inhalation anesthesia, and dogs affected with dilated cardiomyopathy. Sample: Myocardial samples obtained from 21 euthanized dogs (6 clinically normal [control] dogs, 5 clinically normal dogs subjected to inhalation anesthesia with isoflurane prior to euthanasia, 5 dogs with juvenile-onset dilated cardiomyopathy, and 5 dogs with adult-onset dilated cardiomyopathy). Procedures: Activity of mitochondrial complex I and complex IV was assayed spectrophotometrically in isolated mitochondria from left ventricular tissue obtained from the 4 groups of dogs. Results: Activity of complex I and complex IV was significantly decreased in anesthetized dogs, compared with activities in the control dogs and dogs with juvenile-onset or adult-onset dilated cardiomyopathy. Conclusions and Clinical Relevance: Inhalation anesthesia disrupted the electron transport chain in the dogs, which potentially led to an outburst of reactive oxygen species that caused mitochondrial dysfunction. Inhalation anesthesia depressed mitochondrial function in dogs, similar to results reported in other species. This effect is important to consider when anesthetizing animals with myocardial disease and suggested that antioxidant treatments may be beneficial in some animals. Additionally, this effect should be considered when designing studies in which mitochondrial enzyme activity will be measured. Additional studies that include a larger number of animals are warranted.