Objective—To assess serum concentrations of adiponectin and characterize adiponectin protein complexes in healthy dogs.
Animals—11 healthy dogs.
Procedures—Sera collected from 10 dogs were evaluated via velocity sedimentation and ultracentrifugation, SDS-PAGE, western immunoblotting, and radioimmunoassay. Visceral adipose tissue (approx 90 g) was collected from the falciform ligament of a healthy dog undergoing elective ovariohysterectomy, and adiponectin gene expression was assessed via a real-time PCR procedure.
Results—Adiponectin gene expression was detected in visceral adipose tissue. Serum adiponectin concentrations ranged from 0.85 to 1.5 μg/mL (mean concentration, 1.22 μg/mL). In canine serum, adiponectin was present as a multimer, consisting of a low–molecular-weight complex (180 kd); as 3 (180-, 90-, and 60-kd) complexes under denaturing conditions; as 2 (90- and 60-kd) complexes under reducing conditions; and as a dimer, a monomer, and globular head region (60, 30, and 28 kd, respectively) under reducing-denaturing conditions. It is likely that adiponectin also circulates as a high–molecular-weight (360- to 540-kd) complex in canine serum, but resolution of this complex was not possible via SDS-PAGE.
Conclusions and Clinical Relevance—After exposure to identical experimental conditions, adiponectin protein complexes in canine serum were similar to those detected in human and rodent sera. Circulating adiponectin concentrations in canine serum were slightly lower than concentrations in human serum. Adiponectin gene expression was identified in canine visceral adipose tissue. Results suggest that adiponectin could be used as an early clinical marker for metabolic derangements, including obesity, insulin resistance, and diabetes mellitus in dogs.
OBJECTIVE To evaluate optimal isolation of endothelial colony-forming cells (ECFCs) from peripheral blood of horses.
SAMPLE Jugular and cephalic venous blood samples from 17 adult horses.
PROCEDURES Each blood sample was divided; isolation was performed with whole blood adherence (WBA) and density gradient centrifugation (DGC). Isolated cells were characterized by uptake of 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocyanine perchlorate–labeled acetylated low-density lipoprotein (DiI-Ac-LDL), vascular tubule formation, and expression of endothelial (CD34, CD105, vascular endothelial growth factor receptor-2, and von Willebrand factor) and hematopoietic (CD14) cell markers by use of indirect immunofluorescence assay (IFA) and flow cytometry.
RESULTS Colonies with cobblestone morphology were isolated from 15 of 17 horses. Blood collected from the cephalic vein yielded colonies significantly more often (14/17 horses) than did blood collected from the jugular vein (8/17 horses). Of 14 cephalic blood samples with colonies, 13 were obtained with DGC and 8 with WBA. Of 8 jugular blood samples with colonies, 8 were obtained with DGC and 4 with WBA. Colony frequency (colonies per milliliter of blood) was significantly higher for cephalic blood samples and samples isolated with DGC. Cells formed vascular tubules, had uptake of DiI-Ac-LDL, and expressed endothelial markers by use of IFA and flow cytometry, which confirmed their identity as ECFCs.
CONCLUSIONS AND CLINICAL RELEVANCE Maximum yield of ECFCs was obtained for blood samples collected from both the jugular and cephalic veins and use of DGC to isolate cells. Consistent yield of ECFCs from peripheral blood of horses will enable studies to evaluate diagnostic and therapeutic uses.
Objective—To determine the isometric responses of isolated intrapulmonary bronchioles from cats with and without adult heartworm infection.
Animals—13 purpose-bred adult cats.
Procedures—Cats were infected with 100 third-stage larvae or received a sham inoculation, and the left caudal lung lobe was collected 278 to 299 days after infection. Isometric responses of intrapulmonary bronchiolar rings were studied by use of a wire myograph. Three cycles of contractions induced by administration of 10μM acetylcholine were followed by administration of the contractile agonists acetylcholine, histamine, and 5-hydroxy-tryptamine. To evaluate relaxation, intrapulmonary bronchiolar rings were constricted by administration of 10μM 5-hydroxytryptamine, and concentration-response curves were generated from administration of sodium nitroprusside, isoproterenol, and substance P.
Results—Compared with tissues from control cats, contractile responses to acetylcholine and 5-hydroxytryptamine were reduced in tissues from heartworm-infected cats. Relaxation to isoproterenol was significantly reduced in tissues from heartworm-infected cats. Relaxation to substance P was increased in tissues from heartworm-infected cats, but relaxation to sodium nitroprusside was unchanged.
Conclusions and Clinical Relevance—Results suggested that despite increased bronchiolar wall thickness in heartworm-infected cats, a hyperreactive response of the bronchiolar smooth muscle is not the primary mechanism of respiratory tract clinical signs. Reduced response of the airway to isoproterenol may indicate refractoriness to bronchiolar relaxation in heartworm-infected cats.
Objective—To characterize adiponectin protein complexes in lean and obese horses.
Animals—26 lean horses and 18 obese horses.
Procedures—Body condition score (BCS) and serum insulin activity were measured for each horse. Denaturing and native western blot analyses were used to evaluate adiponectin complexes in serum. A human ELISA kit was validated and used to quantify high–molecular weight (HMW) complexes. Correlations between variables were made, and HMW values were compared between groups.
Results—Adiponectin was present as a multimer consisting of HMW (> 720-kDa), low-molecular weight (180-kDa), and trimeric (90-kDa) complexes in serum. All complexes were qualitatively reduced in obese horses versus lean horses, but the percentage of complexes < 250 kDa was higher in obese versus lean horses. High–molecular weight adiponectin concentration measured via ELISA was negatively correlated with serum insulin activity and BCS and was lower in obese horses (mean ± SD, 3.6 ± 3.9 μg/mL), compared with lean horses (8.0 ± 4.6 μg/mL).
Conclusions and Clinical Relevance—HMW adiponectin is measurable via ELISA, and concentration is negatively correlated with BCS and serum insulin activity in horses. A greater understanding of the role of adiponectin in equine metabolism will provide insight into the pathophysiology of metabolic disease conditions.