Objective-To assess effects of in vitro meloxicam exposure on metabolism in articular chondrocytes from dogs with naturally occurring osteoarthritis Sample-Femoral head cartilage from 16 dogs undergoing total hip replacement Procedures-Articular cartilage samples were obtained. Tissue sulfated glycosaminoglycan (SGAG), collagen, and DNA concentrations were measured. Collagen, SGAG, chondroitin sulfate 846, NO, prostaglandin E2 (PGE2), and matrix metalloproteinase (MMP)-2, MMP-3, MMP-9, and MMP-13 concentrations in culture medium were analyzed. Aggrecan, collagen II, MMP-2, MMP-3, MMP-9, MMP-13, ADAM metallopeptidase with thrombospondin type 1 motif (ADAMTS)-4, ADAMTS-5, tissue inhibitor of metalloproteinase (TIMP)-1, TIMP-2, TIMP-3, interleukin-1β, tumor necrosis factor-α, cyclooxygenase-1, cyclooxygenase-2, and nducible nitric oxide synthase gene expression were evaluated. Comparisons between tissues cultured without (control) and with meloxicam at concentrations of 0.3, 3.0, and 30.0 μg/mL for up to 30 days were performed by means of repeated-measures analysis. Results-Meloxicam had no effect on chondrocyte SGAG, collagen, or DNA concentrations. Expression of ADAMTS-5 was significantly decreased in all groups on all days, compared with the day 0 value. On day 3, culture medium PGE2 concentrations were significantly lower in all meloxicam-treated groups, compared with values for controls, and values remained low. Culture medium MMP-3 concentrations were significantly lower on day 30 than on day 3 in all meloxicam-treated groups. Conclusions and Clinical Relevance-Results suggested that in vitro meloxicam treatment of osteoarthritic canine cartilage for up to 30 days did not induce matrix degradation or stimulate MMP production. Meloxicam lowered PGE2 release from this tissue, and effects on tissue chondrocyte content and matrix composition were neutral.

Objective: To develop an in vitro model of cartilage injury in full-thickness equine cartilage specimens that can be used to simulate in vivo disease and evaluate treatment efficacy. Sample: 15 full-thickness cartilage explants from the trochlear ridges of the distal aspect of the femur from each of 6 adult horses that had died from reasons unrelated to the musculoskeletal system. Procedures: To simulate injury, cartilage explants were subjected to single-impact uniaxial compression to 50%, 60%, 70%, or 80% strain at a rate of 100% strain/s. Other explants were left uninjured (control specimens). All specimens underwent a culture process for 28 days and were subsequently evaluated histologically for characteristics of injury and early stages of osteoarthritis, including articular surface damage, chondrocyte cell death, focal cell loss, chondrocyte cluster formation, and loss of the extracellular matrix molecules aggrecan and types I and II collagen. Results: Compression to all degrees of strain induced some amount of pathological change typical of clinical osteoarthritis in horses; however, only compression to 60% strain induced significant changes morphologically and biochemically in the extracellular matrix. Conclusions and Clinical Relevance: The threshold strain necessary to model injury in full-thickness cartilage specimens from the trochlear ridges of the distal femur of adult horses was 60% strain at a rate of 100% strain/s. This in vitro model should facilitate study of pathophysiologic changes and therapeutic interventions for osteoarthritis.

Objective: To compare the effects of autologous equine serum (AES) and autologous conditioned serum (ACS) on equine articular chondrocyte metabolism when stimulated with recombinant human (rh) interleukin (IL)-1β. Sample: Articular cartilage and nonconditioned and conditioned serum from 6 young adult horses. Procedures: Cartilage samples were digested, and chondrocytes were isolated and formed into pellets. Chondrocyte pellets were treated with each of the following: 10% AES, 10% AES and rhIL-1β, 20% AES and rhIL-1β, 10% ACS and rhIL-1β, and 20% ACS and rhIL-1β, and various effects of these treatments were measured. Results: Recombinant human IL-1β treatment led to a decrease in chondrocyte glycosaminoglycan synthesis and collagen II mRNA expression and an increase in medium matrix metalloproteinase-3 activity and cyclooxygenase-2 mRNA expression. When results of ACS and rhIL-1β treatment were compared with those of AES and rhIL-1β treatment, no difference was evident in glycosaminoglycan release, total glycosaminoglycan concentration, total DNA content, or matrix metalloproteinase-3 activity. A significant increase was found in chondrocyte glycosaminoglycan synthesis with 20% AES and rhIL-1β versus 10% ACS and rhIL-1β. The medium from ACS and rhIL-1β treatment had a higher concentration of IL-1β receptor antagonist, compared with medium from AES and rhIL-1β treatment. Treatment with 20% ACS and rhIL-1β resulted in a higher medium insulin-like growth factor-I concentration than did treatment with 10% AES and rhIL-1β. No difference in mRNA expression was found between ACS and rhIL-1β treatment and AES and rhIL-1β treatment. Conclusions and Clinical Relevance: Minimal beneficial effects of ACS treatment on proteoglycan matrix metabolism in equine chonrocytes were evident, compared with the effects of AES treatment.

Objective-To determine the pharmacokinetics, pharmacodynamics, and safety of zoledronic acid in horses. Animals-8 healthy horses. Procedures-A single dose of zoledronic acid (0.057 mg/kg, IV) was administered during a 30-minute period. Venous blood was collected at several time points. Zoledronic acid concentration in plasma was measured by liquid chromatography-tandem mass spectrometry, and pertinent pharmacokinetic parameters were determined. Plasma was analyzed for total calcium, BUN, and creatinine concentrations and a marker for bone resorption (C-terminal telopeptides of type I collagen). Results-Zoledronic acid was safely administered IV during a 30-minute period, and no adverse effects were observed. Plasma concentrations of zoledronic acid were consistent with a 2-compartment mammillary model. Plasma concentrations of zoledronic acid were detected for up to 8 hours after administration. Mean total calcium concentrations in plasma were less than the reference range 7 days after zoledronic acid administration. A marker for bone remodeling decreased in concentration after zoledronic acid administration and remained low for the 1-year duration of the study. No changes in BUN and creatinine concentrations were observed after zoledronic acid administration. Conclusions and Clinical Relevance-Zoledronic acid was safely administered in healthy horses. Zoledronic acid is reported as the strongest bisphosphonate presently available, and studies evaluating potential benefits of zoledronic acid in horses with orthopedic conditions are warranted.

Objective: To investigate effects of various imidazoline and nonimidazoline α-adrenergic agents on aggregation and antiaggregation of bovine and equine platelets. Sample: Blood samples obtained from 8 healthy adult cattle and 16 healthy adult Thoroughbreds. Procedures: Aggregation and antiaggregation effects of various imidazoline and nonimidazoline α-adrenergic agents on bovine and equine platelets were determined via a turbidimetric method. Collagen and ADP were used to initiate aggregation. Results: Adrenaline, noradrenaline, or α-adrenoceptor agents alone did not induce changes in aggregation of bovine or equine platelets or potentiate ADP- or collagen-induced platelet aggregation. Adrenaline and the α2-adrenoceptor agonist clonidine had an inhibitory effect on ADP- and collagen-induced aggregation of bovine platelets. The α2-adrenoceptor antagonists phentolamine and yohimbine also inhibited collagen-induced aggregation of bovine platelets. Noradrenaline, other α-adrenoceptor agonists (xylazine, oxymetazoline, and medetomidine), and α-adrenoceptor antagonists (atipamezole, idazoxan, tolazoline, and prazosin) were less effective or completely ineffective in inhibiting ADP- and collagen-induced aggregation of bovine platelets. The imidazoline α2-adrenoceptor agonist oxymetazoline submaximally inhibited collagen-induced aggregation of equine platelets, and the α2-adrenoceptor antagonist idazoxan, along with phentolamine and yohimbine, also inhibited collagen-induced aggregation of equine platelets. The imidazoline compound antazoline inhibited both ADP- and collagen-induced aggregation of equine platelets. Conclusions and Clinical Relevance: Several drugs had effects on aggregation of platelets of cattle and horses, and effective doses of ADP and collagen also differed between species. The α2-adrenoceptor agonists (xylazine and medetomidine) and antagonists (tolazoline and atipamezole) may be used by bovine and equine practitioners without concern for adverse effects on platelet function and hemostasis.

Objective-To determine whether soybean oil emulsion has an in vitro effect on platelet aggregation and thromboelastography in blood samples obtained from healthy dogs. Animals-12 healthy adult dogs. Procedures-Blood samples were collected from each dog into tubes containing EDTA, hirudin, or sodium citrate for a CBC, collagen- and ADP-induced impedance aggregometry, or thromboelastography, respectively. Whole blood platelet aggregation, determined with ADP or collagen agonists, was measured in blood samples containing hirudin and final lipid concentrations of 0, 1, 10, and 30 mg/mL. The thromboelastographic variables R (reaction time), K (clotting time), α angle, and maximum amplitude were evaluated in blood samples containing sodium citrate and final lipid concentrations equivalent to those used for assessment of platelet aggregation. Results-Median maximum ADP- and collagen-induced platelet aggregation in blood samples containing 1, 10, or 30 mg of lipid/mL did not differ significantly from the value for the respective lipid-free blood sample. Maximum amplitude determined via thromboelastography was significantly reduced in blood samples containing 10 and 30 mg of lipid/mL, compared with findings for lipid-free blood samples. Values of other thromboelastographic variables did not differ, regardless of lipid concentrations. Conclusions and Clinical Relevance-Maximum amplitude determined via thromboelastography in canine blood samples was significantly affected by the addition of lipid to final concentrations that are several orders of magnitude higher than clinically relevant lipid concentrations in dogs. Lipid treatment appears to have no significant effect on hemostatic variables in dogs, although clinical studies should be performed to confirm these in vitro findings.

Objective-To investigate histomorphometric changes in the cartilage and subchondral bone of the third carpal bone associated with conditioning exercise in young Thoroughbreds. Animals-Nine 18-month-old Thoroughbreds. Procedures-Both third carpal bones of 9 horses (4 exercised spontaneously at pasture only and 5 given additional conditioning exercise beginning at a mean age of 3 weeks) were evaluated. Histomorphometric variables (hyaline and calcified cartilage thickness and collagen orientation; vascular channel area, number, and orientation; and osteochondral junction rugosity) of the third carpal bone, sampled at 4 dorsopalmar sites in the radial facet, were compared between the exercised and nonexercised groups. Results-The vascular channel area measured at the 4 dorsopalmar sites was larger in the exercised group than in the control group, but none of the variables were significantly different between groups. Both groups had significant site-specific variations in all measured variables. Most importantly, the vascular channel area was highest in the most dorsal aspect. Conclusions and Clinical Relevance-Results suggested that the mild exercise imposed in both groups during the developmental period appeared to be associated with an increase in the vascular channel area beneath the calcified cartilage layer in the third carpal bone. This increased vascular channel area could also be associated with high stress in the dorsal aspect of the radial facet, a region that is known to be vulnerable to osteochondral fragmentation.

Objective-To determine the effects of 2 weeks of intense exercise on expression of markers of pulmonary venous remodeling in the caudodorsal and cranioventral regions of the lungs of horses. Animals-6 horses. Procedures-Tissue samples of the caudodorsal and cranioventral regions of lungs were obtained before and after conditioning and 2 weeks of intense exercise. Pulmonary veins were isolated, and a quantitative real-time PCR assay was used to determine mRNA expression of matrix metalloproteinase-2 and −9, tissue inhibitor of metalloproteinase-1 and −2, collagen type I, tenascin-C, endothelin-1, platelet-derived growth factor, transforming growth factor (TGF)-β, and vascular endothelial growth factor (VEGF). Protein expression of collagen (via morphometric analysis) and tenascin-C, TGF-β, and VEGF (via immunohistochemistry) was determined. Results-Exercise-induced pulmonary hemorrhage was detected in 2 horses after exercise. The mRNA expression of matrix metalloproteinase-2 and −9, tissue inhibitor of metalloproteinase-2, TGF-β, and VEGF was significantly lower in pulmonary veins obtained after exercise versus those obtained before exercise for both the caudodorsal and cranioventral regions of the lungs. Collagen content was significantly higher in tissue samples obtained from the caudodorsal regions of the lungs versus content in samples obtained from the cranioventral regions of the lungs both before and after exercise. Exercise did not alter protein expression of tenascin-C, TGF-β, or VEGF. Conclusions and Clinical Relevance-Results of this study indicated 2 weeks of intense exercise did not alter expression of marker genes in a manner expected to favor venous remodeling. Pulmonary venous remodeling is complex, and > 2 weeks of intense exercise may be required to induce such remodeling.