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  • Author or Editor: Alicia L Bertone x
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Abstract

Objective—To compare articular cartilage from horses with naturally developing osteochondrosis (OC) with normal articular cartilage and healing cartilage obtained from horses with experimentally induced osteochondral fractures.

Sample Population—109 specimens of articular cartilage from 78 horses.

Procedure—Morphologic characteristics, proteoglycan (PG), and type II collagen were analyzed in articular cartilage of OC specimens (group 1), matched healing cartilage obtained 40 days after experimentally induced osteochondral fractures (group 2), and matched normal cartilage from the same sites (group 3).

Results—79 specimens of OC cartilage were obtained from horses. Ex vivo PG synthesis was significantly greater in the femoral cartilage, compared with synthesis in the tibial cartilage, and significantly greater for groups 1 and 2, compared with group 3. For groups 1 and 2, femoral fragments had significantly greater PG content, compared with PG content in tibial fragments. Keratan sulfate content was significantly less in group 3, compared with groups 1 and 2. Cartilage from the OC specimens had loss of structural architecture. The OC tissue bed stained positive for chondroitin sulfate and type II collagen, but the fracture bed did not.

Conclusions and Clinical Relevance—Our analyses could not distinguish articular cartilage from horses with OC and a healing fracture. Both resembled an anabolic, reparative process. Immunohistochemical analysis suggested a chondromyxoid tissue in the OC bed that was morphologically similar to fibrous tissue but phenotypically resembled hyaline cartilage. Thus, tissue in the OC bed may be degenerative cartilage, whereas tissue in the fracture bed may be reparative fibrous callus. (Am J Vet Res 2005;66:1881–1890)

Full access
in American Journal of Veterinary Research

Abstract

OBJECTIVE To assess efficiency of gravity filtration to enhance recovery of equine bone marrow elements including stem and progenitor cells.

ANIMALS 12 healthy adult horses.

PROCEDURES Bone marrow aspirates were collected from the fifth sternebral body and filtered by gravitational flow to obtain bone marrow elements. Raw and harvested bone marrow and marrow effluent were evaluated for WBC and platelet counts, automated and cytomorphologic cell differential counts, mesenchymal stem cell CFUs, cell viability, and differentiation capacity. Isolated cells were analyzed for CD90 and major histocompatibility complex (MHC) class I and II antigens.

RESULTS Mean cell viability of harvested bone marrow was 95.9%. Total WBCs and platelets were efficiently captured on the filter (> 95%), and mean recovery in harvested bone marrow was 30%. Cytologic cell differential counts indicated that the percentage of neutrophils was significantly less and the progenitor cell population was significantly higher and concentrated 1.56-fold in harvested bone marrow, compared with results for raw bone marrow. Flow cytometry and cell culture were used to characterize harvested bone marrow cells as positive for expression of CD90 and negative for MHCI and MHCII, which indicated stem cells with a multipotent phenotype that differentiated into chondrocytes, osteocytes, adipocytes, and tenocytes.

CONCLUSIONS AND CLINICAL RELEVANCE Gravitational filtration of bone marrow efficiently yielded platelets and cells and produced a progenitor-enriched, leukocyte-reduced product, compared with raw bone marrow.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine outcome of Standardbred racehorses with moderate to severe midbody suspensory ligament desmitis (MSD) treated by means of ultrasound-guided intralesional injection of a single dose of platelet-rich plasma (PRP) followed by a program of gradually increased exercise.

Design—Nonrandomized clinical trial.

Animals—9 Standardbred racehorses.

Procedures—Following injection of PRP, horses were allowed a controlled, gradual return to exercise. Race records for the year prior to injury and for 3 consecutive years after horses returned to racing were reviewed. For comparison purposes, race records of 9 Standardbred racehorses with no history of MSD racing at the same time were also reviewed.

Results—All 9 horses with MSD returned to racing after treatment; median time to return to racing was 32 weeks. All 9 horses raced at least once during the first and second years after returning to racing, but only 5 raced during the third year. When number of starts, total earnings, and earnings per start were compared between case and comparison horses, the only significant differences were number of starts during the third year after case horses returned to racing and earnings per start during the first year after case horses returned to racing, with values being significantly lower for case horses than for comparison horses.

Conclusions and Clinical Relevance—Results suggested that horses with moderate to severe MSD treated by means of intralesional injection of a single dose of PRP followed by a program of gradually increased exercise had an excellent prognosis for returning to racing.

Full access
in Journal of the American Veterinary Medical Association

Summary

Effects of low-flow ischemia and reperfusion of the large colon on systemic and colonic hemodynamic and metabolic variables were determined in horses. Twenty-four adult horses were randomly allocated to 3 groups: sham-operated (n = 6), 6 hours of ischemia (n = 9), and 3 hours of ischemia and 3 hours of reperfusion (n = 9). Low-flow ischemia was induced in groups 2 and 3 by reducing colonic arterial blood flow to 20% of baseline. Heart rate, arterial blood pressures, cardiac index, pulmonary artery pressure, right atrial pressure, and colonic blood flow were monitored. Arterial, mixed-venous, and colonic venous blood gas and oximetry analyses; PCV; and blood lactate and pyruvate and plasma total protein concentrations were measured. Data were recorded, and blood samples were collected at baseline and at 30-minute intervals for 6 hours; additionally, data were collected at 185, 190, and 195 minutes (corresponding to 5, 10, and 15 minutes of reperfusion in group-3 horses). There were no differences among groups at baseline or across time for any systemic hemodynamic or metabolic variable. Colonic blood flow did not change across time in group-1 horses. Colonic blood flow significantly (P < 0.05) decreased to 20% of baseline at induction of ischemia in horses of groups 2 and 3 and remained significantly decreased throughout the ischemic period in horses of groups 2 (6 hours) and 3 (3 hours). Colonic blood flow significantly (P < 0.05) increased above baseline by 5 minutes of reperfusion in group-3 horses. Colonic oxygen delivery and oxygen consumption, and colonic venous pH, Po2 percentage saturation of hemoglobin, and oxygen content were significantly (P < 0.05) decreased within 30 minutes after induction of ischemia in horses of groups 2 and 3; colonic venous Po2 colonic oxygen extraction ratio, and lactate and pyruvate concentrations were significantiy (P < 0.05) increased by 30 minutes of ischemia. These alterations continued throughout ischemia, but within 5 minutes of reperfusion in group-3 horses, these variables either returned to baseline (pH, Pco2 lactate, pyruvate), significantly (P < 0.05) increased above baseline (Po2 oxygen content, % saturation of hemoglobin), or significantly (P < 0.05) decreased below baseline (colonic oxygen extraction ratio). Colonic oxygen consumption remained decreased during reperfusion in group-3 horses. Colonic mucosal ischemia-reperfusion injury observed in this model of ischemia was associated with local colonic hemodynamic and metabolic alterations in the presence of systemic hemodynamic and metabolic stability. Reactive hyperemia was observed at restoration of colonic blood flow in group-3 horses and persisted during reperfusion. Colonic venous metabolic alterations were corrected at reperfusion, indicating adaptation of the colon to the return of blood flow and oxygen delivery with resultant decrease in anaerobic metabolism. The early alterations in these variables may simply represent a washout of metabolic by-products.

Free access
in American Journal of Veterinary Research

SUMMARY

Light microscopy, morphometry, and scanning electron microscopy were used to examine the mucosal morphologic features of 7 intestinal specimens (3 from the small intestine; 4 from the large intestine) from each of 8 horses 1 year after sham operation (group 1; n = 3) or extensive large-colon resection (group 2; n = 5). Qualitative light microscopic examination did not reveal differences between groups, but morphometry revealed significantly (P < 0.05) greater intercrypt area and distance in horses with colon resection and this was most pronounced in the cecum and remaining right ventral and dorsal colon. Crypt area and depth were similar for horses with colon resection and sham operation (P > 0.05). Qualitative evaluation of the scanning electron micrographs revealed more prominent crypt orifices in the large intestine of horses with colon resection. The larger intercrypt distance in the colon of horses with resection was not an obvious feature of the qualitative evaluation of the surface with scanning electron microscopy. Small intestinal morphologic features were variable and significant differences were not detected between horses with sham operation and colon resection. Horses adapted to extensive large-colon resection within 1 year by increasing the absorptive (intercrypt) surface area of the remaining large intestine.

Free access
in American Journal of Veterinary Research

Abstract

OBJECTIVE To create a bioactive synovium scaffold by infusing decellularized synovial-derived extracellular matrix (synECM) with synovial-derived mesenchymal stem cells (synMSCs).

SAMPLE Synovium from the femoropatellar and medial femorotibial joints of equine cadavers.

PROCEDURES The synMSCs were cultured in monolayer and not treated or cotransduced to enhance expression of green fluorescent protein (GFP) and human bone morphogenetic protein (BMP)-2. The synECM was decellularized with 0.1% peracetic acid and then seeded with synMSCs (0.5 × 106 cells/0.5 mL) by use of a 30% serum gradient. Samples were evaluated on days 0, 3, 7, and 14. Cell migration, differentiation, and distribution into the synECMs were determined by cell surface marker CD90, viability, histologic morphology, and fluorescence microscopy results and expression of GFP, BMP-2, hyaluronan (HA), and proteoglycan (PG).

RESULTS At day 14, synMSCs were viable and had multiplied 2.5-fold in the synECMs. The synECMs seeded with synMSCs had a significant decrease in CD90 expression and significant increases in HA and PG expression. The synECMs seeded with synMSCs cotransduced with GFP, or BMP-2 had a significant increase in BMP-2 expression.

CONCLUSIONS AND CLINICAL RELEVANCE The synECM seeded with synMSCs or synMSCs cotransduced with GFP, or BMP-2 yielded a bioactive synovial scaffold. Expression of BMP-2 by synMSCs cotransduced to enhance expression of BMP-2 or GFP and an accompanying increase in both HA and PG expression indicated production of anabolic agents and synoviocyte differentiation in the scaffold. Because BMP-2 can promote repair of damaged cartilage, such a bioactive scaffold could be useful for treatment of injured cartilage.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate the effects of anti-inflammatory drugs on lipopolysaccharide (LPS)-challenged and -unchallenged equine synovial membrane in terms of production of prostaglandin E2 (PGE2) and hyaluronan, viability, and histomorphologic characteristics.

Sample Population—Synovial membranes were collected from the carpal, tarsocrural, and femoropatellar joints of 6 adult horses.

Procedure—Synovial membranes from each horse were minced and pooled and explants were treated with one of the following: no drug (control), drug, LPS alone, or LPS and drug. Treatment drugs were phenylbutazone (PBZ), flunixin meglumine (FNX), ketoprofen (KET), carprofen (CRP), meloxicam (MEL), low-concentration methylprednisolone (METH), highconcentration METH, dimethyl sulfoxide (DMSO), or an experimental COX-2 inhibitor (dissolved in DMSO). Following 48 hours of culture, medium was assayed for PGE2 and hyaluronan concentration. Synovial explants were assessed for viability and histomorphologic characteristics.

Results—For the LPS-challenged explants, PBZ, FNX, KTP, CRP, MEL, and low-concentration METH suppressed PGE2 production, compared with LPS challenge alone. Only MEL suppressed PGE2 production from LPS-challenged explants, compared with unchallenged explants. Synovial explants maintained > 90% viability and there was no significant difference in viability or hyaluronan production among explants. Histomorphologic scores were significantly decreased for explants treated with low-concentration METH or DMSO.

Conclusions and Clinical Relevance—PBZ, FNX, KTP, CRP, MEL, and low-concentration METH suppressed PGE2 production in LPS-challenged explants. Meloxicam appeared to have more selective suppression of COX-2 activity. Histomorphologic scores suggest detrimental effects of METH, DMSO, and the experimental COX-2 inhibitor. Commonly used nonsteroidal anti-inflammatory drugs suppress induced synovial membrane PGE2 production without detrimental effects on synovial membrane viability and function. ( Am J Vet Res 2001;62:54–60)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate the effects of orally administered phenylbutazone on proteoglycan synthesis and chondrocyte inhibition by IL-1β in articular cartilage explants of horses.

Animals—11 healthy 1- to 2-year-old horses.

Procedure—Horses were randomly assigned to the control (n = 5) or treated group (4.4 mg of phenylbutazone/ kg of body weight, PO, q 12 h; n = 6). Articular cartilage specimens were collected before treatment was initiated (day 0), after 14 days of treatment, and 2 weeks after cessation of treatment (day 30). Proteoglycan synthesis and stromelysin concentration in cartilage extracts were assessed after 72 hours of culture in medium alone or with recombinant human interleukin-1β (IL-1β; 0.1 ng/ml).

Results—On day 0, proteoglycan synthesis was significantly less in cartilage explants cultured in IL-1β, compared with medium alone. Mean proteoglycan synthesis in explants collected on days 14 and 30 was significantly less in treated horses, compared with controls. However, incubation of explants from treated horses with IL-1β did not result in a further decrease in proteoglycan synthesis. Significant differences in stromelysin concentration were not detected between or within groups.

Conclusions and Clinical Relevance—Oral administration of phenylbutazone for 14 days significantly decreased proteoglycan synthesis in articular culture explants from healthy horses to a degree similar to that induced by in vitro exposure to IL-1β. Phenylbutazone should be used judiciously in athletic horses with osteoarthritis, because chronic administration may suppress proteoglycan synthesis and potentiate cartilage damage. (Am J Vet Res 2001; 62:1916–1921)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine the effects of phenylbutazone (PBZ) on bone activity and bone formation in horses.

Animals—12 healthy 1- to 2-year-old horses.

Procedures—Biopsy was performed to obtain unicortical bone specimens from 1 tibia on day 0 and from the contralateral tibia on day 14. Fluorochromic markers were administered IV 2 days prior to and on days 0, 10, 15, and 25 after biopsy was performed. Six horses received PBZ (4.4 mg/kg of body weight, PO, q 12 h) and 6 horses were used as controls. All horses were euthanatized on day 30 and tissues from biopsy sites, with adjacent cortical bone, were collected. Osteonal density and activity, mineral apposition rate (MAR), and percentage of mineralized tissue filling the biopsy-induced defects in cortical bone were assessed. Serum samples from all horses were analyzed for bone-specific alkaline phosphatase activity and concentration of PBZ.

Results—MAR was significantly decreased in horses treated with PBZ. Regional acceleratory phenomenon was observed in cortical bone in both groups but was significantly decreased in horses treated with PBZ. Osteonal activity was similar at all time points in all horses. In control horses, percentage of mineralized tissue filling the cortical defects was significantly greater in defects present for 30 days, compared with defects present for 14 days. Differences in percentage of mineralized tissue were not detected in horses treated with PBZ.

Conclusions and Clinical Relevance—PBZ decreased MAR in cortical bone and appeared to decrease healing rate of cortical defects in horses. (Am J Vet Res 2000;61:537–543)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate the buffy coat and apheresis methods for preparation of platelet concentrates from equine blood by comparing platelet and growth factor concentrations.

Animals—15 mature mixed-breed geldings.

Procedure—Whole blood samples were collected and processed by use of a buffy coat or apheresis method to obtain platelet poor and platelet concentrated fractions. The PCV, WBC count, and platelet count were compared among whole blood samples, platelet poor fractions, concentrates obtained by use of the apheresis method (ie, apheresis platelet concentrates), and concentrates obtained by use of the buffy coat method (ie, buffy coat platelet concentrates). Concentrations of transforming growth factor- β (ie, TGF-β1 and TGF-β2) and insulin-like growth factor were compared between buffy coat and apheresis platelet concentrates.

Results—Platelet concentrations were 8.9-fold and 5.2-fold greater in buffy coat and apheresis platelet concentrates, respectively, compared with whole blood. Platelet concentrations were 13.1-fold greater in filtered apheresis platelet concentrates, compared with whole blood. TGF-β1 concentrations were 2.8- fold and 3.1-fold greater in buffy coat and apheresis platelet concentrates, respectively, and TGF-β1 concentrations were 10.5-fold greater in filtered apheresis platelet concentrates, compared with whole blood. TGF-β2 concentrations were 3.6-fold greater in apheresis platelet concentrates, compared with whole blood. Platelet concentrations correlated with growth factor concentrations across all blood and platelet fractions. White blood cell counts had a significant positive correlation with TGF-β1 concentration in buffy coat platelet concentrates.

Conclusions and Clinical Relevance—Platelets and TGF-β1 can be concentrated reliably from equine blood by use of buffy coat or apheresis methods, without modification of the protocols used for humans. (Am J Vet Res 2004;65:924–930)

Full access
in American Journal of Veterinary Research