To assess whether IV regional limb perfusion (IVRLP) and intraosseous regional limb perfusion (IORLP) of ceftiofur sodium resulted in clinically relevant drug concentrations in the synovial fluid of the tibiotarsal-tarsometatarsal joint of chickens (ie, an avian model) and to determine whether one of those techniques was superior to the other.
12 healthy adult hens.
Birds were randomly assigned to receive ceftiofur sodium (2 mg/kg) by the IVRLP (n = 4), IORLP (4), or IM (control; 4) route once daily for 6 consecutive days. Blood and tibiotarsal-tarsometatarsal synovial fluid samples were collected 15 minutes after ceftiofur administration on predetermined days for quantification of ceftiofur concentration. Plasma and synovial fluid ceftiofur concentrations were compared among the 3 groups.
All 4 birds in the IVRLP group developed mild to moderate bruising around the injection site, but this bruising did not prohibit completion of the prescribed treatment regimen. No adverse effects were observed in any of the other birds. The mean plasma and synovial fluid ceftiofur concentrations exceeded the therapeutic threshold for most common bacterial pathogens (> 1.0 μg/mL) at all sample acquisition times for all 3 groups. The mean synovial fluid ceftiofur concentration for the IVRLP group was significantly greater than that for the IORLP and control groups at all sample acquisition times.
CONCLUSIONS AND CLINICAL RELEVANCE
Results suggested that IVRLP may be a safe and effective technique for antimicrobial administration to birds with joint infections, contaminated wounds, pododermatitis, and other musculoskeletal infections of the distal aspect of a limb.
To determine whether differences existed in the viscoelastic properties of synovial fluid samples from the metacarpophalangeal, intercarpal, and distal interphalangeal joints of orthopedically normal athletic horses.
45 warmblood horses and 30 Thoroughbreds (age range, 4 to 16 years).
Synovial fluid samples were aseptically obtained via arthrocentesis from 1 metacarpophalangeal, intercarpal, and distal interphalangeal joint of each horse, and nucleated cell counts were performed. A commercial ELISA was used to measure sample hyaluronic acid concentrations, and full rheological characterization of samples was performed to measure the elastic or storage modulus G' and viscous or loss modulus G“ at 37.5°C (representing the body temperature of horses). Findings were compared among joints and between breed groups by means of ANOVA.
Significant differences in synovial fluid G' and G“ values were identified between Thoroughbreds and warmblood horses for the metacarpophalangeal joint, between the metacarpophalangeal and intercarpal joints of Thoroughbreds, and between the metacarpophalangeal and distal interphalangeal joints and intercarpal and distal interphalangeal joints of warmblood horses. No significant differences were identified between breed groups or among joints in synovial fluid hyaluronic concentrations or nucleated cell counts.
CONCLUSIONS AND CLINICAL RELEVANCE
Viscoelastic properties of the forelimb joints of orthopedically normal Thoroughbreds and warmblood horses differed within and between these 2 groups, mainly as a function of the evaluated joint. To the authors' knowledge, this was the first study of its kind, and additional research is warranted to better understand the viscoelastic properties of synovial fluid in horses to optimize their locomotive function.
OBJECTIVE To evaluate lameness and morphological changes associated with an osteochondral fragment–groove procedure as a means of experimental induction of metacarpophalangeal (MCP) joint osteoarthritis within an 11-week period in horses.
ANIMALS 6 nonlame adult warmbloods.
PROCEDURES The right MCP joint of each horse underwent an osteochondral fragment–groove procedure (day 0). After 1 week of stall rest (ie, starting day 7), each horse was trained daily on a treadmill. Weekly, horses underwent visual and inertial sensor-based assessments of lameness. Both MCP joints were assessed radiographically on days 0 (before surgery), 1, 35, and 77. A synovial fluid sample was collected from the right MCP joint on days 0 (before surgery), 35, 36, 49, 63, and 77 for cytologic and biomarker analyses. On day 77, each horse was euthanized; both MCP joints were evaluated macroscopically and histologically.
RESULTS Right forelimb lameness was detected visually and by the inertial sensor system when horses were moving on a straight line after distal forelimb flexion or circling left on days 14 to 77. Compared with presurgical values, synovial fluid interleukin-6, prostaglandin E2, hyaluronic acid, and interleukin-1 receptor antagonist protein concentrations were increased at 2 or 3 time points, whereas tumor necrosis factor-α and interleukin-10 concentrations were decreased at 1 time point. Gross examination of all right MCP joints revealed synovitis and wear lines; synovitis was confirmed histologically.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that a combined osteochondral fragment–groove procedure can be used to induce clinically and grossly observable early MCP joint osteoarthritis during an 11-week period in horses.
OBJECTIVE: To assess the relationship between histologic degeneration of cranial cruciate ligaments (CCLs) and severity of synovitis and ligament vascularity.
SAMPLE: CCL and synovium from 59 stifle joints (53 dogs). PROCEDURES: CCL and synovium specimens were obtained from stifle joints of juvenile (15 joints; 12 dogs) and adult (25 joints; 22 dogs) dogs with intact CCLs and dogs with CCL rupture (rCCL; 19 joints; 19 dogs). Vascular density and degenerative changes of the CCL core region and severity of synovitis were semiquantitatively evaluated. Relationships were analyzed by use of a random effects model to account for correlated specimens.
RESULTS: Mean ± SD modified Bonar scores (scale, 0 to 9) of adults (4.85 ± 0.44) and dogs with rCCL (5.69 ± 0.49) were significantly higher than scores of juveniles (1.13 ± 0.55). Vascularity scores (scale, 0 to 3) were significantly higher for juveniles (3.00 ± 0.24) than for adults (1.53 ± 0.27) and dogs with rCCL (0.78 ± 0.23). Synovitis scores were not significantly different among groups. There was a significant negative relationship between modified Bonar scores and vascularity scores for juveniles and adults and for adults and dogs with rCCL when controlling for age, but there was not a significant relationship between modified Bonar scores and synovitis scores. There was a significant relationship between modified Bonar scores and body weight of adults.
CONCLUSIONS AND CLINICAL RELEVANCE: Poor blood supply to the core region could be an important underlying condition for spontaneous degeneration of the CCL in at-risk dogs.
OBJECTIVE To compare the effects of 3 equimolar concentrations of methylprednisolone acetate (MPA), triamcinolone acetonide (TA), and isoflupredone acetate (IPA) on equine articular tissue cocultures in an inflammatory environment.
SAMPLE Synovial and osteochondral explants from the femoropatellar joints of 6 equine cadavers (age, 2 to 11 years) without evidence of musculoskeletal disease.
PROCEDURES From each cadaver, synovial and osteochondral explants were harvested from 1 femoropatellar joint to create cocultures. Cocultures were incubated for 96 hours with (positive control) or without (negative control) interleukin (IL)-1β (10 ng/mL) or with IL-1β and MPA, TA, or IPA at a concentration of 10−4, 10−7, or 10−10M. Culture medium samples were collected from each coculture after 48 and 96 hours of incubation. Concentrations of prostaglandin E2, matrix metalloproteinase-13, lactate dehydrogenase, and glycosaminoglycan were determined and compared among treatments at each time.
RESULTS In general, low concentrations (10−7 and 10−10M) of MPA, TA, and IPA mitigated the inflammatory and catabolic (as determined by prostaglandin E2 and matrix metalloproteinase-13 quantification, respectively) effects of IL-1β in cocultures to a greater extent than the high (10−4M) concentration. Mean culture medium lactate dehydrogenase concentration for the 10−4M IPA treatment was significantly greater than that for the positive control at both times, which was suggestive of cytotoxicosis. Mean culture medium glycosaminoglycan concentration did not differ significantly.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that the in vitro effects of IPA and MPA were similar to those of TA at clinically relevant concentrations (10−7 and 10−10M).
OBJECTIVE To characterize the MRI and histologic features of the supraspinatus tendon in nonlame dogs.
ANIMALS 7 cadavers (14 shoulder joints) of nonlame 2-year-old sexually intact male Beagles.
PROCEDURES Multiple MRI fluid-sensitive pulse sequences were obtained for both shoulder joints of each cadaver, and the thickness, volume, and signal intensity of each supraspinatus tendon were assessed. After MRI scanning was complete, the shoulder joints were processed for histologic examination. Tissue specimens were stained with various stains to determine tendon morphology and composition. Histologic and MRI findings were correlated and described.
RESULTS All supraspinatus tendons had a trilaminar appearance on sagittal and transverse MRI images, which was characterized by a thick, hyperintense center layer (central substance) sandwiched between thin hypointense superficial and deep margins. The mean ± SD central substance-to-superficial margin and central substance-to-deep margin thickness ratios were 8.4 ± 1.2 and 9.0 ± 0.9, respectively; supraspinatus tendon-to-triceps brachii muscle signal intensity ratio was 1.3 ± 0.2; and tendon volume was 445 ± 20 mm3. The superficial and deep margins histologically resembled other tendons with highly ordered collagen fibers. The central substance was comprised of water-rich glycosaminoglycans interspersed among haphazardly arranged collagen bundles.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated histologically normal canine supraspinatus tendons have a trilaminar appearance on MRI images. In dogs, a diagnosis of supraspinatus tendinosis should not be based solely on the tendon having a hyperintense signal on MRI images; other MRI evidence of shoulder joint disease and diagnostic findings are necessary to support such a diagnosis.
OBJECTIVE To evaluate gene transfer of recombinant adeno-associated viral (rAAV) vectors with AAV2 or AAV5 capsid and encoding hyaluronic acid (HA) synthase-2 (HAS2) into joints of healthy dogs.
ANIMALS 22 purpose-bred Beagles.
PROCEDURES Plasmid expression cassettes encoding canine HAS2 (cHAS2) were assessed in vitro for concentration and molecular size of secreted HA. Thereafter, rAAV2-cHAS2 vectors at 3 concentrations and rAAV5-cHAS2 vectors at 1 concentration were each administered intra-articularly into the left stifle joint of 5 dogs; 2 dogs received PBS solution instead. Synovial fluid HA concentration and serum and synovial fluid titers of neutralizing antibodies against AAV capsids were measured at various points. Dogs were euthanized 28 days after treatment, and cartilage and synovium samples were collected for vector DNA and mRNA quantification and histologic examination.
RESULTS Cell transfection with plasmids encoding cHAS2 resulted in an increase in production and secretion of HA in vitro. In vivo, the rAAV5-cHAS2 vector yielded uniform genome transfer and cHAS2 expression in collected synovium and cartilage samples. In contrast, rAAV2-cHAS2 vectors were detected inconsistently in synovium and cartilage samples and failed to produce clear dose-related responses. Histologic examination revealed minimal synovial inflammation in joints injected with rAAV vectors. Neutralizing antibodies against AAV capsids were detected in serum and synovial fluid samples from all vector-treated dogs.
CONCLUSIONS AND CLINICAL RELEVANCE rAAV5-mediated transfer of the gene for cHAS2 into healthy joints of dogs by intra-articular injection appeared safe and resulted in vector-derived cHAS2 production by synoviocytes and chondrocytes. Whether this treatment may increase HA production by synoviocytes and chondrocytes in osteoarthritic joints remains to be determined.
OBJECTIVE To use proteomic analysis to determine the protein constituents of synovial fluid samples from the stifle joints of dogs with and without osteoarthritis secondary to cranial cruciate ligament rupture (CCLR).
ANIMALS 12 dogs with clinically normal stifle joints (controls) and 16 dogs with osteoarthritis secondary to CCLR.
PROCEDURES A synovial fluid sample was obtained from all dogs. Synovial fluid total protein concentration was determined by the Bradford assay. Proteins were separated by use of a 1-D SDS-PAGE to detect protein bands that differed between dogs with and without osteoarthritis. Those protein bands then underwent trypsin digestion and were analyzed by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry, the results of which were compared with a curated protein sequence database for protein identification. One of the most frequently identified proteins, apoprotein (apo) A-I, was then quantified in all synovial fluid samples by use of a competitive-inhibition ELISA. Results were compared between dogs with and without osteoarthritis.
RESULTS Median synovial fluid total protein and apo A-I concentrations for dogs with osteoarthritis were significantly greater than those for control dogs. The most abundant proteins identified in the synovial fluid were albumin and apo A-I.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that quantification of synovial fluid total protein and apo A-I concentrations might facilitate diagnosis of osteoarthritis secondary to CCLR in dogs. Further research and validation of synovial fluid apo A-I concentration as a biomarker for osteoarthritis in dogs are necessary before it can be recommended for clinical use.
OBJECTIVE To characterize delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) features of healthy hyaline cartilage of the distal interphalangeal joint (DIPJ) of horses, to determine whether dGEMRIC can be used to differentiate various stages of naturally occurring osteoarthritis of the DIPJ, and to correlate relaxation times determined by dGEMRIC with the glycosaminoglycan concentration, water content, and macroscopic and histologic findings of hyaline cartilage of DIPJs with and without osteoarthritis.
SAMPLE 1 cadaveric forelimb DIPJ from each of 12 adult warmblood horses.
PROCEDURES T1-weighted cartilage relaxation times were obtained for predetermined sites of the DIPJ before (T1preGd) and after (T1postGd) intra-articular gadolinium administration. Corresponding cartilage sites underwent macroscopic, histologic, and immunohistochemical evaluation, and cartilage glycosaminoglycan concentration and water content were determined. Median T1preGd and T1postGd were correlated with macroscopic, histologic, and biochemical data. Mixed generalized linear models were created to evaluate the effects of cartilage site, articular surface, and macroscopic and histologic scores on relaxation times.
RESULTS 122 cartilage specimens were analyzed. Median T1postGd was lower than the median T1preGd for normal and diseased cartilage. Both T1preGd and T1postGd were correlated with macroscopic and histologic scores, whereby T1preGd increased and T1postGd decreased as osteoarthritis progressed. There was topographic variation of T1preGd and T1postGd within the DIPJ. Cartilage glycosaminoglycan concentration and water content were significantly correlated with T1preGd and macroscopic and histologic scores but were not correlated with T1postGd.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that dGEMRIC relaxation times varied for DIPJs with various degrees of osteoarthritis. These findings may help facilitate early detection of osteoarthritis.
OBJECTIVE To determine effects of transforming growth factor (TGF)-β and interleukin (IL)-1β on inflammatory markers in cultured canine chondrocytes to clarify the role of these cytokines in osteoarthritis of dogs.
SAMPLE Pooled chondrocytes isolated from the stifle joints of 4 adult dogs.
PROCEDURES Chondrocytes were isolated, cultured, and frozen at −80°C. Pooled cells were incubated in medium with or without TGF-β (1 or 10 ng/mL) and subsequently stimulated with IL-1β (10 ng/mL). Concentrations of nitric oxide (NO) and prostaglandin (PG) E were measured in culture supernatants. Gene expression of matrix metalloproteinase (MMP)-3, tissue inhibitor of metalloproteinase (TIMP)-2, inducible NO synthase (iNOS), and cyclooxygenase (COX)-2 was quantified by use of real-time quantitative PCR assays.
RESULTS Stimulation with IL-1β increased gene expression of all inflammatory markers, except for TIMP-2. Incubation with TGF-β resulted in a significant decrease in MMP-3 and TIMP-2 mRNA concentrations but had no effect on PGE and NO concentrations. For cells treated with TGF-β followed by IL-1β, concentrations of PGE and NO were lower, compared with concentrations for IL-1β control cells. Furthermore, IL-1β–induced gene expression of iNOS, MMP-3, and COX-2 was downregulated. However, the IL-1β–induced downregulation of TIMP-2 gene expression was partially restored by pretreatment with TGF-β.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that IL-1β increased the expression of inflammatory genes and mediators, and TGF-β largely attenuated the IL-1β–mediated inflammatory response. Therefore, TGF-β might be a novel target for use in the prevention and treatment of cartilage breakdown in dogs with osteoarthritis.