Objective—To evaluate effects of apheresis on mesenchymal stem cells (MSCs) and compare those MSCs with MSCs obtained from adipose tissue or bone marrow (BM).
Sample Population—Samples obtained from 6 adult horses.
Procedures—Samples of blood from a peripheral vein, adipose tissue, and BM aspirate were obtained from each horse. Samples were processed via apheresis of blood and techniques reported elsewhere for adipose tissue and BM. Cultures were maintained until adherence and subsequently were subjected to differentiation protocols to evaluate adipogenic, osteoblastogenic, and chondrogenic potential.
Results—Apheresis product had a significantly higher mononuclear percentage, higher platelet count, and lower RBC count, compared with values for peripheral blood. No cell adherence to the tissue culture plates was detected for the apheresis product. Adherence was detected for 6 of 6 adipose-derived and 4 of 6 BM-derived samples. Variations in efficiency were detected for differentiation of adipose- and BM-derived cells into adipocytes, chondrocytes, and osteoblasts.
Conclusions and Clinical Relevance—Apheresis was able to concentrate mononuclear cells and reduce RBC contamination. However, the apheresis product was unable to adhere to the tissue culture plates. In matched horses, adipose- and BM-derived MSCs were capable of producing lipids, glycosaminoglycan, and mineral. The BM was vastly superior to adipose tissue as a source of MSCs with osteoblastogenic potential in matched horses. Additional studies will be necessary to optimize apheresis techniques for horses before peripheral blood can be considered a suitable source for multipotential cells for use in cell-based treatments.
Objective—To validate a method to assess glomerular filtration rate (GFR) in conscious monkeys via transcutaneous radiation detection after IV injection of technetium Tc 99m pentatate(99mTc-DTPA).
Animals—4 healthy rhesus monkeys.
Procedures—On day 1, each monkey was anesthetized, lothalamate sodium I 125 (125l-iothalamate) was administered via continuous rate infusion (0.0037 MBq/min); blood and urine samples were obtained for determination of 125l-iothalamate plasma clearance variables and estimation of GFR. One dose of 99mTc-DTPA (74 MBq/kg, IV) was also administered during the 125l-iothalamate plasma clearance test, and transcutaneous measurements of technetium 99m-emitted radiation were obtained by use of an ambulatory renal monitor (ARM) applied to a brachium of each monkey. Determination of GFR by use of the ARM was repeated on days 8 and 45 in the same monkeys without anesthesia.
Results—Sensitivity, accuracy, and precision of the 2 methods were similar. By use of the ARM, GFR determined by use of the renal rate constant (κGFR) was calculated; the value obtained on day 1 under anesthesia was similar to values determined via 125l-iothalamate plasma clearance testing on the same day, but was 16% to 23% less than that measured on days 8 and 45 in conscious monkeys.
Conclusions and Clinical Relevance—The ARM method for assessment of GFR was less invasive, faster, and more convenient than the standard clearance method, but yielded comparable results. The need to train animals and size restrictions of the device may limit the use of this technique in other nonhuman animals.
Objective—To determine effects of 2 doses of caffeine on metabolic variables in neonata pigs with peripartum asphyxia
Animals—180 neonatal pigs
Procedures—Neonatal pigs were assigned to 2 groups (groups P and F) on the basis of results for a vitality scale (passed or failed, respectively). Within each group, there were 3 subgroups of 30 pigs each. Within each group, the 3 subgroups received a placebo that consisted of an empty gelatin capsule, a gelatin capsule that contained 20 mg of caffeine, and a gelatin capsule that contained 35 mg of caffeine, respectively; all capsules were administered orally (0 hours). Blood samples were collected immediately before and 24 hours after capsule administration.
Results—Pigs in groups P and F that received 20 or 35 mg of caffeine had significant increases in triglyceride concentrations. All pigs in groups P and F had a significant decrease in lactate concentrations, although the placebo-treated pigs in group F had larger decreases than did the group F pigs treated with 20 or 35 mg of caffeine. Glucose concentrations increased significantly in group F pigs treated with 20 or 35 mg of caffeine (30% and 50%, respectively), whereas glucose concentrations remained unchanged in group P pigs. In pigs treated with 35 mg of caffeine, the final weight obtained for group F was approximately 8% lower than that obtained for group P
Conclusions and Clinical Relevance—Administering caffeine immediately after birth to neonatal pigs with severe oxygen restriction resulted in significant improvements in metabolic variables. (Am J Vet Res 2010;71:1214-1219)
Objective—To determine whether long-distance endurance exercise in sled dogs causes increases in serum concentrations of C-reactive protein (CRP) and whether such increases are correlated with other markers of the exercise-induced acute-phase response
Animals—25 sled dogs.
Procedures—Serum was obtained from 25 sled dogs approximately 48 hours before and immediately after completing a race of 557 km. Serum was analyzed to determine concentrations of CRP and interleukin (IL)-6, and serum biochemical analysis (and iron homeostasis analysis) also was performed.
Results—CRP concentrations increased significantly from a mean ± SD concentration of 22.4 ± 16.3 μg/mL before racing to a mean of 263.3 ± 103.8 μg/mL immediately after racing Serum IL-6 concentrations were unchanged; however, there was a modest but significant correlation (r = 0.50) between the increase in CRP concentration and an overall decrease in serum albumin concentration, which suggested an inverse relationship between hepatic synthesis of the 2 proteins. Differences in CRP concentrations among teams of dogs revealed that concentrations before racing may be influenced by previous episodes of exercise. Serum iron concentration had only a mild decrease, which may have been attributable to iron-rich diets consumed by the dogs.
Conclusions and Clinical Relevance—CRP concentrations may serve as a potential marker for exercise-induced inflammation. The exact amount of exercise required to induce such a response is unknown, but dogs apparently have a more robust acute-phase response than do humans. Clinical evaluation of CRP concentrations must account for physical activity when those concentrations are used as a potential marker for systemic inflammation. (Am J Vet Res 2010;71:1207-1213)
Objective—To evaluate the effects of firocoxib, meloxicam, and tepoxalin administration in healthy cats by measuring the ability of stimulated tissues to synthesize eicosanoids ex vivo.
Animals—8 healthy adult male cats.
Procedures—In a blinded, randomized, crossover study design, cats were treated with firocoxib (1 mg/kg, PO, q 24 h), meloxicam (0.05 mg/kg, PO, q 24 h), tepoxalin (5.0 mg/kg, PO, q 12 h), or a placebo for 8 days. Blood samples and gastric and duodenal mucosal biopsy specimens were collected on days 0 (baseline; immediately before treatment), 3, and 8 of each treatment period. Thromboxane B2 (TXB2) concentrations were measured in serum, and prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) concentrations were measured in plasma. Prostaglandin E1 (PGE1) synthesis, PGE2 synthesis, and LTB4 concentrations were measured in mucosal biopsy specimens. A 21-day minimum washout period was observed between treatments. Repeated-measures analyses were performed.
Results—Firocoxib and meloxicam administration resulted in a lower plasma PGE2 concentration than at baseline on days 3 and 8 of administration, whereas tepoxalin administration did not. Tepoxalin administration resulted in a lower serum TXB2 concentration and pyloric and duodenal PGE1 synthesis on both days, compared with baseline and placebo administration. Neither firocoxib nor meloxicam administration altered pyloric or duodenal PGE1 synthesis on either day, compared with placebo administration. Tepoxalin administration also resulted in lower pyloric mucosal LTB4 concentrations on both days, compared with baseline values.
Conclusions and Clinical Relevance—Firocoxib and meloxicam administration had no effect on cyclooxygenase-1 activity, whereas tepoxalin administration resulted in inhibition of cyclooxygenase-1 and 5-lipoxygenase. (Am J Vet Res 2010;71:1067–1073)
Objective—To determine the effect of IV administration of a bolus of 50% dextrose solution on electrolyte and energy balance and effect of blood collection site on serum electrolyte values in postparturient dairy cows.
Animals—24 clinically normal multiparous cows.
Procedures—A bolus of 50% dextrose solution (0.5 L [n = 8 cows]), 50% dextrose solution (1.0 L ), or saline (0.9% NaCl) solution (1.0 L, control treatment ) was administered via jugular venipuncture 5 to 10 days after parturition. Pretreatment and posttreatment blood samples were analyzed for concentrations of calcium, magnesium, phosphorus, potassium, glucose, insulin, β-hydroxybutyric acid (BHBA), and nonesterified fatty acids. Coccygeal vessel and jugular vein blood samples were obtained prior to treatment, and electrolyte concentrations were compared.
Results—Treatment with 50% dextrose decreased phosphorus concentration in serum, compared with the control treatment. Suppression of BHBA and nonesterified fatty acid concentrations following dextrose treatment lasted for < 12 hours; mean BHBA concentrations in all groups were increased 24 hours after treatment. Mean serum phosphorus concentration in coccygeal vessel blood samples was 0.67 mg/dL greater than the concentration in jugular vein blood samples.
Conclusions and Clinical Relevance—Postpartum cows treated with dextrose solution may be at risk for hypophosphatemia, and 1 treatment with 0.5 or 1 L of 50% dextrose solution is unlikely to prevent or resolve acetonemia (ketosis). The risk of hypophosphatemia may be underestimated when coccygeal vessel blood samples are used for diagnosis. (Am J Vet Res 2010;71:1074–1080)
Objective—To describe the effects of increasing the extracellular fluid (ECF) volume by approximately 20% on acid-base changes and electrolyte concentrations in anesthetized rats.
Animals—18 adult male Sprague-Dawley rats.
Procedures—Rats were assigned to a control group (n = 6 rats) and a treatment group (12). All rats were anesthetized, and instrumentation and bilateral renal pedicle ligation were performed. The treatment group was infused IV with sterile water throughout a 30-minute period. Acid-base variables and concentrations of electrolytes, lactate, albumin, phosphorus, and hemoglobin were measured before (baseline) and 30 and 60 minutes after onset of infusion. Anion gap, strong ion difference, strong ion gap, and contributions of sodium, chloride, albumin, phosphorus, and lactate concentrations to base excess were calculated at each time point.
Results—Infusion of sterile water led to an increase in ECF volume of approximately 18%. This had no effect on acid-base balance, compared with that in control rats. Infusion of sterile water caused a significant decrease in sodium, chloride, ionized calcium, lactate, and albumin concentrations, compared with concentrations in the control group. Anion gap and calculated effects of sodium, chloride, albumin, and lactate concentrations on base excess at 60 minutes differed significantly between infused and control rats.
Conclusions and Clinical Relevance—Infusion of sterile water did not cause clinically relevant dilutional acidosis. The acidotic impact of water administration was offset by generation of new bicarbonate via carbonic acid equilibration and intracellular buffering in combination with the alkalotic effects of decreases in albumin, phosphorus, and lactate concentrations.
Objective—To evaluate N-hydroxysuccinimide (NHS)-biotin labeling of equine RBCs and determine posttransfusion survival of autologous equine RBCs stored in citrate phosphate dextrose adenine-1 (CPDA-1) for 0, 1, 14, and 28 days.
Animals—13 healthy adult Thoroughbreds.
Procedures—Serial dilutions of biotin and streptavidin-phycoerythrin (PE) were evaluated in vitro in blood collected from 3 horses. One horse was used to determine RBC distribution and recovery. Twelve horses were allocated to 4 groups for in vivo experiments in which blood was collected into CPDA-1. Blood was labeled with biotin and reinfused or stored at 4°C for 1, 14, or 28 days prior to labeling with NHS-biotin and reinfusion. Posttransfusion blood samples were collected 15 minutes and 1, 2, 3, 5, 7, 14, 21, 28, and 35 days after reinfusion. Biotin-labeled RBCs were detected via flow cytometry by use of streptavidin-PE. Posttransfusion lifespan of RBCs and RBC half-life were determined.
Results—Optimal biotin concentration was 0.04 pg of biotin/RBC, and the optimal streptavidin-PE ratio was 1.2 μg of streptavidin-PE/1 × 106 RBCs. Posttransfusion lifespan of autologous RBCs was 99, 89, 66, and 59 days after storage for 0, 1, 14, and 28 days, respectively. Storage did not result in significant alterations in RBC lifespan. Mean posttransfusion RBC half-life was 50, 45, 33, and 29 days for 0, 1, 14, and 28 days of storage, respectively.
Conclusions and Clinical Relevance—Biotin can be used to label equine RBCs for RBC survival studies. Posttransfusion survival of equine autologous RBCs was greater than previously reported.
Objective—To determine effects of therapeutic dosages of aspirin, carprofen, deracoxib, and meloxicam on platelet function and systemic prostaglandin concentrations in healthy dogs.
Animals—10 hound-crossbred dogs.
Procedures—Aspirin (10 mg/kg, PO, q 12 h), carprofen (4.4 mg/kg, PO, q 24 h), deracoxib (2 mg/kg, PO, q 24 h), meloxicam (0.1 mg/kg, PO, q 24 h), and a placebo were administered for 7 days in a random order to each of 10 healthy dogs; there was a 21-day washout period between subsequent treatments. One-stage prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen concentration, and plasma concentrations of thromboxane (TX)B2 and 6-keto prostaglandin (PG)F1α were measured before and after treatment administration. Platelet function was assessed by use of a platelet-function analyzer and aggregation.
Results—Aspirin, carprofen, and meloxicam did not significantly affect platelet function. Deracoxib caused a mild decrease in platelet aggregation induced by 50μM ADP. Platelet number, Hct, PT, aPTT, and plasma TXB2 and 6-keto PGF1α concentrations were unchanged after NSAID administration. Meloxicam administration resulted in a significant decrease in fibrinogen concentration, but results remained within the laboratory reference interval.
Conclusions and Clinical Relevance—Oral administration of commonly used NSAIDs at therapeutic dosages in healthy dogs did not alter plasma TXB2 and 6-keto PGF1α concentrations. Deracoxib administration resulted in a minor abnormality in platelet aggregation. Anti-inflammatory doses of aspirin did not affect platelet function as measured by use of optical aggregometry and a platelet-function analyzer. Further evaluation of the effects of aspirin and cyclooxygenase-2–selective inhibitors on hemostasis should be performed.
Objective—To identify expression and localization of cyclooxygenase (COX)-1 and COX-2 in healthy and ischemic-injured left dorsal colon of horses.
Sample Population—Left dorsal colon tissue samples from 40 horses.
Procedures—Tissue samples that were used in several related studies on ischemia and reperfusion were evaluated. Samples were collected during anesthesia, before induction of ischemia, and following 1 hour of ischemia, 1 hour of ischemia and 30 minutes of reperfusion, 2 hours of ischemia, 2 hours of ischemia and 30 minutes of reperfusion, and 2 hours of ischemia and 18 hours of reperfusion. Histomorphometric analyses were performed to characterize morphological injury. Immunohistochemical analyses were performed to characterize expression and localization of COX-1 and COX-2.
Results—COX-1 and COX-2 were expressed in control tissues before ischemia was induced, predominantly in cells in the lamina propria. Ischemic injury significantly increased expression of COX-2 in epithelial cells on the colonic surface and in crypts. A similar significant increase of COX-1 expression was seen in the epithelial cells.
Conclusions and Clinical Relevance—On the basis of information on the role of COX-2, upregulation of COX-2 in surface epithelium and crypt cells following ischemic injury in equine colon may represent an early step in the repair process.