Ex Vivo Immune Function and Modulatory Effects of Calcitriol in Dogs with Naturally Occurring Diabetes Mellitus.

Human patients with type 1 diabetes mellitus (T1DM) are susceptible to several long-term complications that are related to glycemic control and immune dysregulation. Immune function remains relatively unexplored in dogs with naturally occurring diabetes mellitus (NODM). Calcitriol improves various aspects of immune function in a variety of species, but its effect in diabetic dogs remains unexplored. Therefore, the objectives of this study were to (i) evaluate immune function in dogs with NODM and determine if differences exist based on the level of clinical control and (ii) assess the immunomodulatory effects of calcitriol. Twenty diabetic dogs (clinically controlled, n = ten, not controlled, n = ten) and 20 non-diabetic, healthy control dogs were included in this prospective, case-control study. Whole blood was incubated with calcitriol (10-7 M) or negative control, after which the samples were divided for phagocytosis and leukocyte cytokine response experiments. The phagocytosis of opsonized Escherichia coli (E. coli) was evaluated with flow cytometry. The samples for leukocyte cytokine response evaluations were stimulated with lipopolysaccharide (LPS), lipoteichoic acid (LTA), or phosphate buffer solution (PBS; negative control), and tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-8, and IL-10 were measured in supernatant using a canine-specific multiplex bead-based assay. The leukocytes from diabetic dogs produced higher concentrations of IL-10 (p = 0.01), IL-6 (p < 0.0001), and IL-8 (p < 0.0001) than the control dogs while controlling for the intervention and stimulant. Calcitriol decreased the supernatant concentrations of TNF-α (p < 0.001) and IL-8 (p = 0.04) with concomitant increases in IL-6 (p = 0.005). Diabetic dogs had a lower percentage of leukocytes undergoing phagocytosis (p < 0.0001) but a higher number of bacteria phagocytized per cell (p = 0.001) when compared to the control dogs. Calcitriol had no effect on phagocytic capacity. Lastly, the status of clinical control in diabetic dogs did not yield differences in immune function. These results support that dogs with NODM exhibit immune dysregulation and warrant additional investigation.

The Future of Diabetes Therapies: New Insulins and Insulin Delivery Systems, Glucagon-Like Peptide 1 Analogs, Sodium-Glucose Cotransporter Type 2 Inhibitors, and Beta Cell Replacement Therapy.

As the prevalence of diabetes mellitus increases, so too does the number of available treatment modalities. Many diabetic therapies available in human medicine or on the horizon could hold promise in the management of small animal diabetes. However, it is important to consider how species differences in pathophysiology, management practices and goals, and lifestyle may affect the translation of such treatment modalities for veterinary use. This review article aimed to familiarize veterinarians with the more promising novel diabetic therapies and explore their possible applications in the treatment of canine and feline diabetes mellitus.

Evaluation of Candida spp. and Other Fungi in Feces from Dogs with Naturally Occurring Diabetes Mellitus.

Diabetes mellitus is a common endocrinopathy in dogs and in most cases is analogous to type 1 diabetes mellitus (T1DM) in humans. Candida spp. is a common commensal fungi with higher prevalence and magnitude of growth in humans with T1DM. There is currently no published information about the fungal microbiome in diabetic dogs. Therefore, the objectives of this study were to (i) determine whether diabetic dogs were more likely to have Candida spp. or other types of fungi from feces compared to non-diabetic controls, and (ii) identify variables associated with fungi colonization. Fourteen diabetic dogs and 14 age, sex, and breed matched non-diabetic healthy control dogs were included in this prospective case-control study. Matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was used for fungal identification. Diabetic dogs had greater quantitative fungal growth compared to controls (p = 0.004). Moreover, female dogs were more likely to have fungi colonization than males (p = 0.02). All instances of Candida spp. and Aspergillus spp. colonization were exclusively identified in diabetic dogs. Serum fructosamine concentration was higher in diabetic dogs with fecal colonization of Candida spp. compared to diabetic dogs without growth (p = 0.03). Our results indicate that the fungal microbiome in feces is altered in diabetic dogs, which seem to favor an increased prevalence of Candida spp. and higher quantitative fungal growth. Moreover, female sex and glycemic control could affect the intestinal mycobiome.

Arizona Veterinarians' Perceptions and Consensus Regarding Skills, Knowledge, and Attributes of Day One Veterinary Graduates.

The purpose of this study was to assess Arizona veterinarians' perceptions and consensus regarding the importance of items in the domains of clinical skills, knowledge, and attributes of Day One graduates of veterinary school and to determine demographic predictors for items on which consensus was low. In this survey-based prospective study, respondents were asked to rate the importance of 44 items on a 5-point Likert scale ranging from 1 (not at all important) to 5 (extremely important). Responses were visualized as divergent stacked bar charts and evaluated via summary quantitative and qualitative analyses. Several items had a median score of 5. For clinical skills, items were the ability to formulate a preventive health care plan, the ability to interpret test results, and basic safe handling and restraint of animals; for knowledge, knowledge of pain management and anesthesia; and for attributes, teamwork, problem-solving skills, and client communication skills. The majority of items (80%) had a strong or very strong consensus measure, 18% had a moderate consensus measure, and 2% had a weak consensus measure. Six items (14%) varied by at least one demographic category. We found demographic differences between large and small animal practices in the clinical skill of ability to perform a necropsy, knowledge of large animal theriogenology, and knowledge of canine theriogenology. In conclusion, we found differences in the importance of items and agreement among practitioners, suggesting that critical evaluation of the mapped curriculum, particularly with regard to core curriculum compared with electives and clinical tracks, may benefit students and future employers.

Veterinary Curriculum Transformation at the University of Illinois, 2006-2016.

The organization and delivery of a curriculum is the responsibility of the faculty in educational institutions. Curricular revision is often a hotly debated topic in any college faculty. At the University of Illinois, a 2006 mandate for curriculum modernization from the American Veterinary Medical Association Council on Education provided impetus for a long-discussed curricular revision. After two iterations and a lengthy development process, a new curriculum was gradually implemented at Illinois with the August 2009 matriculation of the Class of 2013. The goals of the revision included earlier clinical exposure for veterinary students through introductions to clinical rotations in years 1 to 3 and an integrated body systems approach in lecture/laboratory courses. A new Clinical Skills Learning Center facilitates development of clinical skills earlier in the curriculum and promotes the development of those skills throughout all 4 years of the curriculum. New outcomes assessments include comprehensive written examinations and Objective Structured Clinical Examinations (OSCEs) in years 2 and 3. Curriculum management, including grading of clinical rotations in all 4 years, is achieved through a commercially available software package. For the past 5 years, when candidates were asked why they chose to apply to Illinois, the new curriculum (27.4%) was the most common answer given during interviews. The Illinois revision has resulted in measurably increased veterinary student self-confidence (p<.001) at graduation.

Altered mRNA and protein expression of filaggrin in the skin of a canine animal model for atopic dermatitis.

BACKGROUND: Filaggrin is a structural protein that has attracted increasing interest over the past decade for its role in the pathogenesis of human atopic dermatitis (AD). Null mutations in its sequence are considered risk factors in the development of AD. HYPOTHESIS/OBJECTIVES: To investigate canine filaggrin mRNA and protein expression in the skin of atopic beagles with experimentally induced AD compared with breed-matched healthy control dogs. METHODS: All dogs were environmentally challenged for 3 days consecutively with allergens to which the atopic dogs had been sensitized. Skin biopsy specimens were taken from six healthy and seven atopic beagles before and after allergen challenge. Canine filaggrin mRNA was measured using quantitative real-time PCR. Indirect immunofluorescence was used to localize the filaggrin protein in canine skin. Analysis of variance with Tukey's multiple comparison test (over-time effect) and unpaired Student's t-test (treatment effect) were used. Values of P ≤ 0.05 were considered significant. RESULTS: Analysis of variance showed a significantly higher expression of filaggrin mRNA in atopic dogs compared with healthy control dogs (P = 0.004 on day 3 and P = 0.01 on day 10) and a decreased mRNA expression on day 3 in healthy control dogs (effect of time, P = 0.006). On blinded evaluation, filaggrin immunofluorescence was distributed homogeneously in the stratum granulosum and the stratum corneum in healthy dogs. Atopic dogs showed a patchy immunofluorescence pattern, which was exacerbated after environmental challenge. CONCLUSIONS AND CLINICAL IMPORTANCE: Altered epidermal filaggrin mRNA expression and protein distribution was detected in this experimental model.

Subcutaneous and gonadal adipose tissue transcriptome differences in lean and obese female dogs.

Canine obesity leads to shortened life span and increased disease incidence. Adipose tissue depots are known to have unique metabolic and gene expression profiles in rodents and humans, but few comparisons of depot gene expression have been performed in the dog. Using microarray technology, our objective was to identify differentially expressed genes and enriched functional pathways between subcutaneous and gonadal adipose of lean and obese dogs to better understand the pathogenesis of obesity in the dog. Because no depot × body weight status interactions were identified in the microarray data, depot differences were the primary focus. A total of 946 and 703 transcripts were differentially expressed (FDR P < 0.05) between gonadal and subcutaneous adipose tissue in obese and lean dogs respectively. Of the adipose depot-specific differences in gene expression, 162 were present in both lean and obese dogs, with the majority (85%) expressed in the same direction. Both lean and obese dog gene lists had enrichment of the complement and coagulation cascade and systemic lupus erythematosus pathways. Obese dogs had enrichment of lysosome, extracellular matrix-receptor interaction, renin-angiotensin system and hematopoietic cell lineage pathways. Lean dogs had enrichment of glutathione metabolism and synthesis and degradation of ketone bodies. We have identified a core set of genes differentially expressed between subcutaneous and gonadal adipose tissue in dogs regardless of body weight. These genes contribute to depot-specific differences in immune function, extracellular matrix remodeling and lysosomal function and may contribute to the physiological differences noted between depots.

Evaluation of canine antimicrobial peptides in infected and noninfected chronic atopic skin.

BACKGROUND: Antimicrobial peptides (AMPs) are small immunomodulatory peptides produced by epithelial and immune cells. ß-Defensins (BDs) and cathelicidins (Caths) are the most studied AMPs. Recently, increased cutaneous expression of AMPs was reported in atopic humans and in beagles with experimentally induced atopy. HYPOTHESIS/OBJECTIVES: Our goal was to analyse mRNA expression and protein levels of canine (c)BD1-like, cBD2-like/122, cBD3-like, cBD103 and cCath in healthy and naturally affected atopic dogs, with and without active skin infection, along with their distribution in the epidermis using indirect immunofluorescence. ANIMALS: Skin biopsies were taken from 14 healthy and 11 atopic privately owned dogs. METHODS: The mRNA levels of cBD1-like, cBD2-like/122, cBD3-like, cBD103 and cCath were quantified using quantitative real-time PCR. The protein levels of cBD3-like and cCath were analysed by relative competitive inhibition enzyme-linked immunosorbent assay, while the distributions of cBD2-like/122, cBD3-like and cCath were detected by indirect immunofluorescence. RESULTS: Dogs with atopic dermatitis had significantly greater mRNA expression of cBD103 (P = 0.04) than control dogs. Furthermore, atopic skin with active infection had a higher cBD103 mRNA expression (P = 0.01) and a lower cBD1-like mRNA expression (P = 0.04) than atopic skin without infection. No significant differences in protein levels (cBD3-like and cCath) or epidermal distribution of AMPs (cBD2-like/122, cBD3-like and cCath) were seen between healthy and atopic dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: Expression of cBD103 mRNA was greater, while expression of cBD1-like mRNA was lower in dogs with atopic dermatitis that had active infections. Work is needed to clarify the biological mechanisms and possible therapeutic options to maintain a healthy canine skin.

Effects of dietary macronutrient composition and feeding frequency on fasting and postprandial hormone response in domestic cats.

The objective was to evaluate the effects of dietary macronutrients and feeding frequency on blood glucose, insulin, total ghrelin and leptin. A total of twelve adult lean neutered male cats were used in three tests, all cross-over studies composed of a 15 d adaptation and blood sampling on day 16. In trial 1, differences between two- and four-meal feeding were tested. On day 16, blood samples were collected every 2 h for 24 h. In trial 2, macronutrient boluses were tested. Instead of the control diet, the morning meal on day 16 was replaced with an isoenergetic bolus of carbohydrate (maltodextrin), protein (chicken meat), fat or water. Fasted and ten postprandial blood samples were collected. In trial 3, diets high in fat (HF), protein (HP), carbohydrate (HC) or a control diet were tested. On day 16, fasted and ten postprandial blood samples were collected. Data were analysed to identify baseline and AUC changes. Cats fed four meals daily had greater (P = 0·03) leptin incremental AUC0-24 h compared with cats fed twice daily. The carbohydrate bolus increased glucose (P < 0·001) and insulin (P < 0·001) incremental AUC0-6 h and tended to increase (P = 0·09) leptin net AUC0-6 h. Cats fed the control and HC diets had greater (P = 0·03) glucose incremental AUC compared with the HF and HP conditions. Circulating hormone data were highly variable and indicated changes due to dietary macronutrients and feeding frequency, but further study is needed to identify impacts on appetite and contributing mechanisms.

Expression and distribution of canine antimicrobial peptides in the skin of healthy and atopic beagles.

Antimicrobial peptides (AMPs) are small immuno-modulatory proteins important in defense against pathogenic organisms. Defensins and cathelicidin are the most frequently studied human AMPs. An increase in AMPs in atopic humans has been reported recently. Our goals were to determine the distribution of AMPs and evaluate their mRNA and protein expression in non-lesional (Day 0), acute lesional skin (Day 3) and post-challenged skin after resolution of skin lesions (Day 10) using a canine model of atopic dermatitis (AD). All dogs were environmentally challenged for three consecutive days with house dust mite. Clinical evaluation of atopic beagles was performed using a CADESI score at each time point before and after environmental challenge. Skin biopsies were taken from six healthy and seven atopic beagles before and after allergen challenge (Day 0, Day 3 and Day 10). The transcription of canine cathelicidin (cCath) and beta-defensins (cBD)-1, -2 and -3 mRNA was quantified using quantitative-RT-PCR while the protein distribution of cBD2, cBD3 and cCath was detected by indirect immunofluorescence. A significant effect, over-time, was seen in CADESI score in AD beagles with an increase score after challenge (Day 3). Quantitative analysis showed a significant difference in mRNA transcript levels between groups (with atopic dogs having more than controls) for all AMPs but cBD2. No effect over time was evident for either group. No significant differences were seen for the AMP protein patterns of distribution (homogenous distribution). Although, these results showed no differences in AMP's localization after allergen exposure in each group; atopic dogs had a higher mRNA expression of AMPs when compared with healthy dogs, a similar finding to humans.

When normal is abnormal: keys to laboratory diagnosis of hidden endocrine disease.

Although veterinary clinicians commonly rely on panels of laboratory tests with individual results flagged when abnormal, care should be taken in interpreting normal test results as well. There are several examples of this in evaluating patients with endocrine disease. The finding of a normal leukogram (absence of a stress leukogram) can be indicative of adrenal insufficiency in dogs, and this disorder can be especially elusive when there are no overt indicators of mineralocorticoid deficiency. Cats with hyperthyroidism can have normal serum thyroid hormone concentrations, normal hematocrits, and normal serum concentrations of creatinine despite the presence of disease that affects these parameters. A normal serum phosphorus concentration, in the face of azotemia, isosthenuria, and hypertension can point a clinician toward a diagnosis of primary hyperaldosteronism rather than primary renal disease. A normal serum parathyroid hormone concentration in the face of hypercalcemia is inappropriate and can indicate the presence of primary hyperparathyroidism. Similarly, hypoglycemia accompanied by a normal serum insulin concentration can be found in cases of hyperinsulinism. These normal findings in abnormal patients, and their mechanisms, are reviewed.

Interpretation of laboratory tests for canine Cushing's syndrome.

Hypercortisolism (HC) is a common disease in dogs. This article will review the laboratory tests that are available for diagnosis of HC and laboratory tests for differentiating between causes of HC. An emphasis will be made on the clinical process that leads to the decision to perform those tests and common misconceptions and issues that arise when performing them. To choose between the adrenocorticotropic hormone (ACTH)-stimulation test and the low-dose dexamethasone suppression test (LDDST), the advantages and disadvantages of both tests should be considered, as well as the clinical presentation. If the index of suspicion of HC is high and other diseases have been appropriately ruled out, the specificity of the ACTH stimulation test is reasonably high with an expected high positive predictive value. Because of the low sensitivity, a negative result in the ACTH stimulation test should not be used to rule out the diagnosis of HC. The LDDST is more sensitive but also less specific and affected more by stress. A positive result on the urine cortisol:creatinine ratio does not help to differentiate HC from other diseases. A negative result on the urine cortisol:creatinine ratio indicates that the diagnosis of HC is very unlikely. The LDDST is useful in differentiating pituitary-dependent HC from an adrenal tumor in about two thirds of all dogs with HC. Differentiation of HC from diabetes mellitus, liver diseases, and hypothyroidism cannot be based solely on endocrine tests. Clinical signs, imaging studies, histopathology, and response to treatment should all be considered.

Adipose tissue transcriptome changes during obesity development in female dogs.

During the development of obesity, adipose tissue undergoes major expansion and remodeling, but the biological processes involved in this transition are not well understood. The objective of this study was to analyze global gene expression profiles of adipose tissue in dogs, fed a high-fat diet, during the transition from a lean to obese phenotype. Nine female beagles (4.09 ± 0.64 yr; 8.48 ± 0.35 kg) were randomized to ad libitum feeding or body weight maintenance. Subcutaneous adipose tissue biopsy, blood, and dual x-ray absorptiometry measurements were collected at 0, 4, 8, 12, and 24 wk of feeding. Serum was analyzed for glucose, insulin, fructosamine, triglycerides, free fatty acids, adiponectin, and leptin. Formalin-fixed adipose tissue was used for determination of adipocyte size. Adipose RNA samples were hybridized to Affymetrix Canine 2.0 microarrays. Statistical analysis, using repeated-measures ANOVA, showed ad libitum feeding increased (P < 0.05) body weight (0 wk, 8.36 ± 0.34 kg; 24 wk, 14.64 ± 0.34 kg), body fat mass (0 wk, 1.36 ± 0.24 kg; 24 wk, 6.52 ± 0.24 kg), adipocyte size (0 wk, 114.66 ± 17.38 µm(2); 24 wk, 320.97 ± 0.18.17 µm(2)), and leptin (0 wk, 0.8 ± 1.0 ng/ml; 24 wk, 12.9 ± 1.0 ng/ml). Microarrays displayed 1,665 differentially expressed genes in adipose tissue as weight increased. Alterations were seen in adipose tissue homeostatic processes including metabolism, oxidative stress, mitochondrial homeostasis, and extracellular matrix. Adipose transcriptome changes highlight the dynamic and adaptive response to ad libitum feeding and obesity development.

Expression and distribution of antimicrobial peptides in the skin of healthy beagles.

Antimicrobial peptides (AMPs) are small proteins involved in defense against pathogenic organisms. Defensins and cathelicidin are the most frequently studied human AMPs. Our goals were to determine the distribution of AMPs and evaluate their mRNA expression in normal canine skin. Skin biopsies were taken from six healthy beagles. The relative transcript level of canine cathelicidin (cCath) and ß-defensin (cBD)-1, cBD2 and cBD3 mRNA was quantified using quantitative real-time polymerase chain reaction. Indirect immunofluorescence (IIF), using polyclonal antibodies against cBD2, cBD3 and cCath, was used to evaluate protein localization in the skin of healthy dogs. The Pfaffl method, using experimentally determined primer efficiencies of amplification, was used to determine the expression level of cCath, cBD1 and cDB3 relative to cDB2. The levels of cCath, cBD3 and cBD1 mRNA were 358, 296 and 177 times higher than those of cBD2, respectively. Using IIF, cBD2 and cBD3 protein fluorescence was detected in all layers of the epidermis, whereas cCath was detected predominantly in the stratum granulosum and corneum. In addition, antimicrobial peptide detection was limited to the infundibular portion of the pilosebaceous units. We have validated useful methods to evaluate AMPs in canine skin. Further studies are needed to compare AMP expression in healthy dogs with that of dogs with inflammatory skin conditions.

Fluoroquinolones: then and now.

Fluoroquinolones were discovered in the 1960s as a derivative of the antimalarial drug chloroquine. Over the past 40 years, many fluoroquinolones have been developed for use in human and veterinary medicine. As with all classes of antibiotics, resistance to fluoroquinolones is a serious concern, and multiple avenues for resistance are being investigated. Resistance-associated point mutations in bacterial DNA and, more recently, plasmid-mediated resistance have been reported in both human and veterinary bacterial isolates. This article reviews the history and most current literature on fluoroquinolones approved for use in dogs and cats and the spectra of activity, mechanisms of action, resistance patterns, and recommendations for appropriate clinical use of these drugs.

Evaluation of the ratio of collagen type III to collagen type I in periurethral tissues of sexually intact and neutered female dogs.

OBJECTIVE: To determine the ratio of collagen type III to collagen type I in the periurethral tissues of sexually intact and neutered female dogs. ANIMALS: 8 neutered and 34 sexually intact female dogs. PROCEDURES: Tissues were obtained from female dogs euthanized for non-urinary tract-related reasons. Indirect immunofluorescent antibody detection of type I and collagen type III was performed by use of confocal microscopy on 2 periurethral samples from each dog, and the ratios of collagen type III to type I area fraction and total area were determined. RESULTS: No significant differences were detected in the collagen ratios of periurethral tissues between sexually intact and neutered female dogs. CONCLUSIONS AND CLINICAL RELEVANCE: In contrast to differences in periurethral collagen content found between pre- and postmenopausal women, such differences may not occur in dogs. This implies that changes in pelvic organ support structures may not play an important role in urinary incontinence in neutered female dogs. Further evaluation is needed to determine the role of age on collagen and pelvic organ support structures in the pathogenesis of canine urinary incontinence.

The effects of body weight, body condition score, sex, and age on serum fructosamine concentrations in clinically healthy cats.

BACKGROUND: Serum fructosamine (SF) concentrations depend on plasma glucose concentrations and are used to evaluate glycemic control in animals with diabetes mellitus (DM). Despite the strong association between obesity and DM, the effects of body weight (BW) and body condition on SF concentrations in clinically healthy cats have not been reported. OBJECTIVE: The aim of the study was to evaluate the effects of BW, body condition score (BCS), sex, and age on SF concentrations in healthy cats. METHODS: BW, BCS, and SF concentrations were determined in 84 clinically healthy client-owned cats (50 neutered males, 33 spayed females, and 1 intact female) of known age. The cats were enrolled prospectively in the study. RESULTS: Mean BW, median BCS, and mean SF concentrations for the 84 cats were 5.4 kg, 5/9, and 268.7 ± 45.5 µmol/L (range 197-399), respectively. BW was weakly but significantly correlated with SF (r=.26; P=.02), whereas BCS was not. Cats weighing >5.4 kg and cats with BCS>5/9 had higher mean SF concentrations compared with cats weighing <5.4 kg and cats with BCS <5/9, respectively. Cats categorized as normal weight to obese by BW (BW ≥ 4.0 kg) had higher mean SF concentrations compared with cats categorized as lean (BW< 4.0 kg). For domestic shorthair cats, the same was true for BCS: cats with BCS ≥ 4/9 had higher mean fructosamine concentrations than those with BCS<4/9. Male cats had significantly higher mean SF concentrations compared with female cats (285.1 ± 45.3 vs 244.5 ± 33.9 µmol/L, P<.001). Age did not affect mean SF concentrations. CONCLUSIONS: BW is positively correlated with SF concentration, and lean cats have lower SF concentrations than normal and obese cats. In contrast to previous reports, mean SF concentrations were higher in male cats than in female cats, even when males and females were matched based on BW, BCS, and age.

Phenobarbital-responsive sialadenosis associated with an esophageal foreign body in a dog.

A 4-year-old Yorkshire terrier was presented for an esophageal foreign body. After removal of the foreign body, clinical signs of gagging, regurgitation, and vomiting continued unabated for >6 weeks. The dog had enlarged submandibular salivary glands that were histologically normal. Treatment with phenobarbital resulted in a rapid and dramatic resolution of clinical signs. After 3 months, the dog was weaned of phenobarbital and was free of any signs of disease 6 months later.