Evaluation of three immunological assays to mitigate the risk of transboundary spread of Coxiella burnetii by alpacas.

Coxiella burnetii causes coxiellosis in animals and Q fever in humans, a potentially debilitating zoonotic disease commonly transmitted through domestic ruminants. To prevent transboundary spread of C. burnetii, animals may be tested prior to export. In alpacas, this process is complicated by the lack of scientific evidence for C. burnetii infection in the species, and the unique composition of camelid antibodies, which may cause false-positive results in assays developed for ruminants. We evaluated a complement fixation test (CFT; currently recommended for alpacas in New Zealand), an enzyme-linked immunosorbent assay (ELISA) and an immunofluorescence assay (IFA). Positive analytical control samples were generated through vaccination of alpacas with a human Q fever vaccine, whereas negative analytical control samples were sourced from New Zealand (deemed free of C. burnetii). Immunological assays were conducted on 131 alpaca sera submitted for export testing. Test characteristics (sensitivity, specificity, positive and negative predictive values) for CFT, ELISA and IFA were determined using Bayesian latent class analysis. Due to anticomplementary activity, 37 (28.2%) of the CFT results were inconclusive, making CFT unsuitable for routine use. Of the remaining 94 samples, 10.6%, 0% and 7.4% were positive for C. burnetii antibodies based on CFT, ELISA and IFA, respectively, yielding estimated sensitivities of 58%, 26% and 78%, and estimated specificities of 95%, 98% and 95%, with the estimates for sensitivity being imprecise, as evidenced by wide 95% credible intervals. Positive predictive values were similar across assays, albeit very low at the estimated seroprevalence of 5%. Our results indicate that, of the tests available, IFA appears to be the most appropriate for use in alpacas. Higher sensitivity of antibody detection, use of antigen detection assays and availability of samples from individuals with evidence of infection could provide additional insight into the risk of transboundary spread of C. burnetii by alpacas.

Dermatophilus congolensis associated bronchopneumonia in an alpaca.

A severe, chronic, locally extensive granulomatous bronchopneumonia was diagnosed on post-mortem and histopathological examination of an adult alpaca. Dermatophilus congolensis organisms were isolated from the lungs and genotypic identification of aerobic culture was confirmed by sequence analysis of the entire 16S rDNA gene. This is the first report of D. congolensis-associated bronchopneumonia in any species.

Effects of tendon injury on uninjured regional tendons in the distal limb: An in-vivo study using an ovine tendinopathy model.

Following injury to a tendon little is known about potential for pathology to develop in other regional tendons from overloading or altered function. The aim of this study was to investigate the gene expression and histopathological changes that occur 1) within the deep digital flexor tendon (DDFT) after injury to the superficial digital flexor tendon (SDFT) and 2) within the flexor tendons (SDFT and DDFT) after injury to the extensor tendons. Merino wethers [Ovis aries] (n = 18) were divided into three equal groups and underwent either partial transection of the SDFT, complete transection of the extensor tendons or were left as non-operated controls. Tendons were harvested and sampled regionally for gene expression (real time PCR) and histologic analysis eight weeks after surgery. Transection of the SDFT resulted in increased expression of collagen III, versican, biglycan, lumican and MMP1 (P<0.026 for all genes) within the DDFT. There was no effect of transecting the extensor tendons on the expression of any gene tested in either the SDFT or the DDFT. The DDFT had elevated histopathology scores induced by transection of the SDFT, eight weeks previously. There were minimal histological differences in either the SDFT or DDFT after transection of the extensor tendons. Transection of the SDFT results in a mild, subclinical tendinopathy within the DDFT with potential implications on treatment and rehabilitation of SDFT injuries. Injury to the extensor tendons has minimal measured effect on the SDFT or DDFT.

Size-Selective Sampling Performance of Six Low-Volume "Total" Suspended Particulate (TSP) Inlets.

Several low-volume inlets (flow rates ≤16.7 liters per minute (Lpm)) are commercially available as components of low-cost, portable ambient particulate matter samplers. Because the inlets themselves do not contain internal fractionators, they are often assumed to representatively sample "total" mass concentrations from the ambient air, independent of aerodynamic particle size and wind speed. To date, none of these so-called "TSP" inlets have been rigorously tested under controlled conditions. To determine their actual size-selective performance under conditions of expected use, wind tunnel tests of six commonly used omnidirectional, low-volume inlets were conducted using solid, polydisperse aerosols at wind speeds of 2, 8, and 24 km/hr. With the exception of axially-oriented, isokinetic sharp-edge nozzles operating at 5 and 10 Lpm, all low-volume inlets showed some degree of non-ideal sampling performance as a function of aerodynamic particle size and wind speed. Depending upon wind speed and assumed ambient particle size distribution, total mass concentration measurements were estimated to be negatively biased by as much as 66%. As expected from particle inertial considerations, inlet efficiency tended to degrade with increasing wind speed and particle size, although some exceptions were noted. The implications of each inlet's non-ideal behavior are discussed with regards to expected total mass concentration measurement during ambient sampling and the ability to obtain representative sampling for size ranges of interest, such as PM2.5 and PM10. Overall test results will aid in low-volume inlet selection and with proper interpretation of results obtained with their ambient field use.

Efficacy of administered mesenchymal stem cells in the initiation and co-ordination of repair processes by resident disc cells in an ovine (Ovis aries) large destabilizing lesion model of experimental disc degeneration.

BACKGROUND: Forty percent of low back pain cases are due to intervertebral disc degeneration (IVDD), with mesenchymal stem cells (MSCs) a reported treatment. We utilized an ovine IVDD model and intradiscal heterologous MSCs to determine therapeutic efficacy at different stages of IVDD. METHODOLOGY: Three nonoperated control (NOC) sheep were used for MSC isolation. In 36 sheep, 6 × 20 mm annular lesions were made at three spinal levels using customized blades/scalpel handles, and IVDD was allowed to develop for 4 weeks in the Early (EA) and late Acute (LA) groups, or 12 weeks in the chronic (EST) group. Lesion IVDs received injections of 10 × 106 MSCs or PBS, and after 8 (EA), 22 (LA) or 14 (EST) weeks recuperation the sheep were sacrificed. Longitudinal lateral radiographs were used to determine disc heights. IVD glycosaminoglycan (GAG) and hydroxyproline contents were quantified using established methods. An Instron materials testing machine and customized jigs analyzed IVD (range of motion, neutral zone [NZ] and stiffness) in flexion/extension, lateral bending and axial rotation. qRTPCR gene profiles of key anabolic and catabolic matrix molecules were undertaken. Toluidine blue and hematoxylin and eosin stained IVD sections were histopathologically scoring by two blinded observers. RESULTS: IVDD significantly reduced disc heights. MSC treatment restored 95% to 100% of disc height, maximally improved NZ and stiffness in flexion/extension and lateral bending in the EST group, restoring GAG levels. With IVDD qRTPCR demonstrated elevated catabolic gene expression (MMP2/3/9/13, ADAMTS4/5) in the PBS IVDs and expession normalization in MSC-treated IVDs. Histopathology degeneracy scores were close to levels of NOC IVDs in MSC IVDs but IVDD developed in PBS injected IVDs. DISCUSSION: Administered MSCs produced recovery in degenerate IVDs, restored disc height, composition, biomechanical properties, down regulated MMPs and fibrosis, strongly supporting the efficacy of MSCs for disc repair.

Development of Polydisperse Aerosol Generation and Measurement Procedures for Wind Tunnel Evaluation of Size-Selective Aerosol Samplers.

Accurate development and evaluation of inlets for representatively collecting ambient particulate matter typically involves use of monodisperse particles in aerosol wind tunnels. However, the resource requirements of using monodisperse aerosols for inlet evaluation creates the need for more rapid and less-expensive techniques to enable determination of size-selective performance in aerosol wind tunnels. The goal of recent wind tunnel research at the U.S. EPA was to develop and validate the use of polydisperse aerosols which provide more rapid, less resource-intensive test results which still meet data quality requirements necessary for developing and evaluating ambient aerosol inlets. This goal was successfully achieved through comprehensive efforts regarding polydisperse aerosol generation, dispersion, collection, extraction, and analysis over a wide range of aerodynamic particle sizes. Using proper experimental techniques, a sampler's complete size-selective efficiency curve can be estimated with polydisperse aerosols in a single test, as opposed to the use of monodisperse aerosols which require conducting multiple tests using several different particle sizes. While this polydisperse aerosol technique is not proposed as a regulatory substitute for use of monodisperse aerosols, the use of polydisperse aerosols is advantageous during an inlet's development where variables of sampling flow rate and inlet geometry are often iteratively evaluated before a final inlet design can be successfully achieved. Complete Standard Operating Procedures for the generation, collection, and analysis of polydisperse calibration aerosols are available from EPA as downloadable files. The described experimental methods will be of value to other researchers during development of ambient sampling inlets and size-selective evaluation of the inlets in aerosol wind tunnels.

Spatiotemporal variations in gene expression, histology and biomechanics in an ovine model of tendinopathy.

Flexor tendinopathy is a common problem affecting humans and animals. Tendon healing is poorly understood and the outcomes of conservative and surgical management are often suboptimal. While often considered a localized injury, recent evidence indicates that in the short term, tendinopathic changes are distributed widely throughout the tendon, remote from the lesion itself. Whether these changes persist throughout healing is unknown. The aim of this study was to document gene expression, histopathological and biomechanical changes that occur throughout the superficial digital flexor tendon (SDFT) up to 16 weeks post-injury, using an ovine surgical model of tendinopathy. Partial tendon transection was associated with decreased gene expression for aggrecan, decorin, fibromodulin, tissue inhibitors of metalloproteinases (TIMPS 1, 2 and 3), collagen I and collagen II. Gene expression for collagen III, lumican and matrix metalloproteinase 13 (MMP13) increased locally around the lesion site. Expression of collagen III and MMP13 decreased with time, but compared to controls, collagen III, MMP13 and lumican expression remained regionally high throughout the study. An increase in TIMP3 was observed over time. Histologically, operated tendons had higher pathology scores than controls, especially around the injured region. A chondroid phenotype was observed with increased cellular rounding and marked proteoglycan accumulation which only partially improved with time. Biomechanically, partial tendon transection resulted in a localized decrease in elastic modulus (in compression) but only at 8 weeks postoperatively. This study improves our understanding of tendon healing, demonstrating an early 'peak' in pathology characterized by altered gene expression and notable histopathological changes. Many of these pathological changes become more localized to the region of injury during healing. Collagen III and MMP13 expression levels remained high close to the lesion throughout the study and may reflect the production of tendon tissue with suboptimal biomechanical properties. Further studies evaluating the long-term response of tendon to injury (6-12 months) are warranted to provide additional information on tendon healing and provide further understanding of the mechanisms underlying the pathology observed in this study.

A Histopathological Scheme for the Quantitative Scoring of Intervertebral Disc Degeneration and the Therapeutic Utility of Adult Mesenchymal Stem Cells for Intervertebral Disc Regeneration.

The purpose of this study was to develop a quantitative histopathological scoring scheme to evaluate disc degeneration and regeneration using an ovine annular lesion model of experimental disc degeneration. Toluidine blue and Haematoxylin and Eosin (H&E) staining were used to evaluate cellular morphology: (i) disc structure/lesion morphology; (ii) proteoglycan depletion; (iii) cellular morphology; (iv) blood vessel in-growth; (v) cell influx into lesion; and (vi) cystic degeneration/chondroid metaplasia. Three study groups were examined: 5 × 5 mm lesion; 6 × 20 mm lesion; and 6 × 20 mm lesion plus mesenchymal stem cell (MSC) treatment. Lumbar intervertebral discs (IVDs) were scored under categories (i-vi) to provide a cumulative score, which underwent statistical analysis using STATA software. Focal proteoglycan depletion was associated with 5 × 5 mm annular rim lesions, bifurcations, annular delamellation, concentric and radial annular tears and an early influx of blood vessels and cells around remodeling lesions but the inner lesion did not heal. Similar features in 6 × 20 mm lesions occurred over a 3-6-month post operative period. MSCs induced a strong recovery in discal pathology with a reduction in cumulative histopathology degeneracy score from 15.2 to 2.7 (p = 0.001) over a three-month recovery period but no recovery in carrier injected discs.

Revisiting the size selective performance of EPA's high-volume total suspended particulate matter (Hi-Vol TSP) sampler.

Under the National Ambient Air Quality Standard (NAAQS) for airborne lead, measurements are conducted by means of a high-volume total suspended particulate matter (Hi-Vol TSP) sampler. In the decade between 1973 and 1983, there were 12 publications that explored the sampling characteristics and effectiveness of the Hi-Vol TSP, yet there persists uncertainty regarding its performance. This article presents an overview of the existing literature on the performance of the Hi-Vol TSP, and identifies the reported sampler effectiveness with respect to four factors: particle size (reported effectiveness of 7%-100%), wind speed (-36% to 100%), sampler orientation (7%-100%), and operational state (107%-140%). Effectiveness of the Hi-Vol TSP was evaluated with a solid, polydisperse aerosol in a controlled wind tunnel setting. Isokinetic samplers were deployed alongside the Hi-Vol TSP to investigate three wind speeds (2, 8, and 24 km h-1), three sampler orientations (0°, 45°, 90°), and two operational states (on, off) for aerosols with aerodynamic diameters from 5 to 35 µm. Results indicate that particle diameter was the largest determining factor of effectiveness followed by wind speed. Orientation of the sampler did not have a significant effect at 2 and 8 km h-1 but did at 24 km h-1. In a passive state, the Hi-Vol TSP was collected between 1% and 7% of available aerosol depending on particle size and wind speed. Results of this research do not invalidate results of previous studies but rather contribute to our overall understanding of the Hi-Vol TSP's size-selective performance. While results generally agreed with previous studies, the Hi-Vol TSP was found to exhibit less dependence on these four factors than previously reported.

Chondroitin sulphate glycosaminoglycans contribute to widespread inferior biomechanics in tendon after focal injury.

Both mechanical and structural properties of tendon change after injury however the causal relationship between these properties is presently unclear. This study aimed to determine the extent of biomechanical change in post-injury tendon pathology and whether the sulphated glycosaminoglycans (glycosaminoglycans) present are a causal factor in these changes. Equine superficial digital flexor tendons (SDF tendons) were surgically-injured in vivo (n=6 injured, n=6 control). Six weeks later they were harvested and regionally dissected into twelve regions around the lesion (equal medial/lateral, proximal/distal). Glycosaminoglycans were removed by enzymatic (chondroitinase) treatment. Elastic modulus (modulus) and ultimate tensile strength (UTS) were measured under uniaxial tension to failure, and tendon glycosaminoglycan content was measured by spectrophotometry. Compared to healthy tendons, pathology induced by the injury decreased modulus (-38%; 95%CI -49% to -28%; P<0.001) and UTS (-38%; 95%CI -48% to -28%; P<0.001) and increased glycosaminoglycan content (+52%; 95%CI 39% - 64%; P<0.001) throughout the tendon. Chondroitinase-mediated glycosaminoglycan removal (50%; 95%CI 21-79%; P<0.001) in surgically-injured pathological tendons caused a significant increase in modulus (5.6MPa/µg removed; 95%CI 0.31-11; P=0.038) and UTS (1.0MPa per µg removed; 95%CI 0.043-2; P=0.041). These results demonstrate that the chondroitin/dermatan sulphate glycosaminoglycans that accumulate in pathological tendon post-injury are partly responsible for the altered biomechanical properties.

Focal experimental injury leads to widespread gene expression and histologic changes in equine flexor tendons.

It is not known how extensively a localised flexor tendon injury affects the entire tendon. This study examined the extent of and relationship between histopathologic and gene expression changes in equine superficial digital flexor tendon after a surgical injury. One forelimb tendon was hemi-transected in six horses, and in three other horses, one tendon underwent a sham operation. After euthanasia at six weeks, transected and control (sham and non-operated contralateral) tendons were regionally sampled (medial and lateral halves each divided into six 3 cm regions) for histologic (scoring and immunohistochemistry) and gene expression (real time PCR) analysis of extracellular matrix changes. The histopathology score was significantly higher in transected tendons compared to control tendons in all regions except for the most distal (P ≤ 0.03) with no differences between overstressed (medial) and stress-deprived (lateral) tendon halves. Proteoglycan scores were increased by transection in all but the most proximal region (P < 0.02), with increased immunostaining for aggrecan, biglycan and versican. After correcting for location within the tendon, gene expression for aggrecan, versican, biglycan, lumican, collagen types I, II and III, MMP14 and TIMP1 was increased in transected tendons compared with control tendons (P < 0.02) and decreased for ADAMTS4, MMP3 and TIMP3 (P < 0.001). Aggrecan, biglycan, fibromodulin, and collagen types I and III expression positively correlated with all histopathology scores (P < 0.001), whereas lumican, ADAMTS4 and MMP14 expression positively correlated only with collagen fiber malalignment (P < 0.001). In summary, histologic and associated gene expression changes were significant and widespread six weeks after injury to the equine SDFT, suggesting rapid and active development of tendinopathy throughout the entire length of the tendon. These extensive changes distant to the focal injury may contribute to poor functional outcomes and re-injury in clinical cases. Our data suggest that successful treatments of focal injuries will need to address pathology in the entire tendon, and that better methods to monitor the development and resolution of tendinopathy are required.

Treatment of experimentally induced osteoarthritis in horses using an intravenous combination of sodium pentosan polysulfate, N-acetyl glucosamine, and sodium hyaluronan.

OBJECTIVE: To assess the effects of sodium pentosan polysulfate (PPS), N-acetyl glucosamine (NAG), and sodium hyaluronan (HA) in horses with induced osteoarthritis (OA). STUDY DESIGN: Experimental. ANIMALS: Adult Standard bred horses (n = 16). METHODS: OA was induced arthroscopically in 1 intercarpal joint; 8 horses were administered 3 mg/kg PPS, 4.8 mg/kg NAG, and 0.12 mg/kg HA (PGH), intravenously (IV), weekly and 8 horses were administered an equivalent volume of saline IV until study completion (day 70). Horses underwent a standardized treadmill exercise program. Clinical and radiographic findings and synovial fluid analysis were evaluated throughout the study. Macroscopic, histologic, histochemical, and biochemical findings were evaluated after necropsy. Comparisons of interest included OA and non-OA joints of saline treated horses and OA joints of PGH treated horses and OA joints of saline treated horses. Results were statistically analyzed with significance set at P < .05. RESULTS: OA caused increases in clinical assessment scores, synovial fluid variables, radiographic, macroscopic, and histologic cartilage scores, synovial fluid and cartilage chondroitin sulfate 846-epitope and glycosaminoglycan concentration. Total radiographic scores, total macroscopic joint pathology and macroscopic cartilage pathology scores were significantly reduced in horses treated with PGH compared with saline treated horses. Synovial fluid total protein concentration and white blood cell count were higher in OA joints of PGH treated horses compared with saline treated horses. There were no other significant differences between treatment groups. CONCLUSIONS: Improvements in macroscopic variables were not supported by other outcomes. Further evidence is needed before PGH can be recommended as a therapeutic option for osteoarthritis in horses.

Optimal tension, position, and number of prostheses required for maximum rima glottidis area after laryngoplasty.

OBJECTIVES: To evaluate the effect of 3 laryngeal prostheses alone or in combination on rima glottidis area in horses. STUDY DESIGN: Experimental randomized design. SAMPLE POPULATION: Cadaveric equine larynges (n = 22). METHODS: Three prostheses were preplaced in each of 14 larynges. Rima glottidis area was measured after loading each suture in 5 Newton (N) increments from 0 N to 35 N. In 8 larynges, the 3 prostheses were tied alone or in combination at a fixed load of 15 N and rima glottidis area measured. RESULTS: Rima glottidis cross-sectional area increased as the load on each prosthesis increased with maximum area reached at 20 N for each prosthesis. At a fixed load of 15 N, tying 2 and 3 prostheses in combination resulted in a larger rima glottidis cross-sectional area than achieved with each prosthesis alone. CONCLUSIONS: A combination of 2 or 3 prostheses tied at a fixed load of 15 N optimized rima glottidis cross-sectional area irrespective of the anatomic location of the prosthesis.

Effect of age and prostheses location on rima glottidis area in equine cadaveric larynges.

OBJECTIVE: To determine the effect of horse age and laryngeal prosthesis location on rima glottidis area in cadaveric larynges. STUDY DESIGN: Experimental study. ANIMALS: Cadaveric equine larynges (n = 40). METHODS: Specimens were grouped by age: group 1, ≤5 years (n = 18); group 2, >5 to ≤10 years (n = 12); group 3, >10 years (n = 10). A cranial prosthesis was placed through the dorsal cricoid spine at 70% of the distance of the total cricoid length measured from the caudal rim. A dorsal prosthesis was placed through the caudal rim of the cricoid on the dorsal midline. A lateral prosthesis was placed 1.5 cm lateral to the dorsal prosthesis. All prostheses passed through the muscular process. Rima glottidis area was determined after progressively tightening each suture in 5 N increments from 0 N to 35 N using a tensiometer. RESULTS: There was no significant effect of age on the area of the rima glottidis at any load for any of the three prosthesis locations. CONCLUSIONS: Age did not affect the area of the rima glottidis when prostheses were loaded between 5 N and 35 N.

Altered stress induced by partial transection of the infraspinatus tendon leads to perlecan (HSPG2) accumulation in an ovine model of tendinopathy.

Perlecan is a widely distributed, heparan sulphate proteoglycan with roles in the sequestration of FGFs, PDGF, VEGF through which it promotes cell proliferation and matrix production. Perlecan also stabilises extracellular matrices through interaction with a diverse range of matrix components. This study examined the distribution of perlecan in an ovine partial transection tendinopathy model. In normal tendon, perlecan was immunolocalised to small blood vessels in intrafascicular regions in the tendon-bone and muscle-tendon attachments and to linear arrays of oval shaped tenocytes in the tendon mid-region. Partial transection in the mid-tendon region significantly increased perlecan accumulation within the fascicles, in granulation tissue filling the transection site and in the tendon-bone and tendon-muscle attachments. The accumulation of perlecan in the transected tendon and its known roles in matrix stabilisation and cell proliferation indicate possible roles in tendon remodelling and repair. Perlecan domain-1 has been used as a growth factor delivery vehicle for FGF-2, BMP-2 and BMP-7 in regenerative medicine but has yet to be evaluated in infraspinatus tendon repair. A better understanding of perlecan's contributions to pathobiological processes in remodelling tendon may be useful in such regenerative strategies in the future.

The effect of short- and long-term treatment with manuka honey on second intention healing of contaminated and noncontaminated wounds on the distal aspect of the forelimbs in horses.

OBJECTIVES: To compare the effects of manuka honey and manuka honey gel on second intention healing of noncontaminated distal limb wounds and those contaminated with feces. STUDY DESIGN: Experimental study. ANIMALS: Standardbred horses (n = 10). METHODS: Five full-thickness wounds (2 × 2 cm) were created on both metacarpi. Wounds on 1 forelimb were covered with horse feces for 24 hours. Wounds on the contralateral limb were left uncontaminated. Wounds were assigned to the following 5 different treatments: manuka honey, manuka honey gel or gel applied for 12 days, manuka honey gel applied throughout healing and untreated control. Wound area was measured on day 1 then weekly until day 42 and time to complete healing was recorded. RESULTS: Wounds treated with manuka honey gel throughout healing healed faster than all other wounds (P < .05). Wounds treated with manuka honey and manuka honey gel for 12 days healed faster than gel control and untreated control wounds (P < .05). Wounds treated with manuka honey and manuka honey gel for 12 days and throughout healing were smaller than gel control and untreated control wounds until day 35 (P < .05). Wounds contaminated with feces had greater retraction for 7 days, but healed faster than noncontaminated wounds (P < .05). CONCLUSIONS: Treatment of wounds with manuka honey and manuka honey gel reduced wound retraction and overall healing time compared with gel and untreated control wounds.

Mechanical destabilization induced by controlled annular incision of the intervertebral disc dysregulates metalloproteinase expression and induces disc degeneration.

STUDY DESIGN: An investigation of mechanical destabilization of the lumbar ovine intervertebral disc (IVD) inducing IVD degeneration (IVDD) as determined by multiparameter outcome measures (magnetic resonance imaging [MRI], IVD composition, biomechanical testing, gene profiling, immunohistochemistry, and immunoblotting). OBJECTIVE: To assess the effect of IVD mechanical destabilization on matrix protein and metalloproteinase gene expression to investigate the pathophysiological mechanisms of lumbar IVDD. SUMMARY OF BACKGROUND DATA: Several earlier studies have used annular transection to induce IVDD in sheep, but none have optimized or validated the most appropriate lesion size. METHODS: The annulus fibrosus (AF) incision inducing maximal change in IVD biomechanics was applied to L1-L2, L3-L4, and L5-L6 discs in vivo to compare with a sham procedure at 3 months post operation. IVDs were evaluated by MRI, biomechanics, histopathology, proteoglycan and collagen content, gene expression, and aggrecan proteolysis by Western blotting. RESULTS: Significant changes were observed in lesion (6 × 20 mm(2)) compared with sham IVDs at 3 months post operation: reduced disc height on MRI; increased neutral zone in biomechanical testing; depleted proteoglycan and collagen content in the nucleus pulposus (NP) and lesion half of the AF but not in the contralateral AF; increased messenger RNA for collagen I and II, aggrecan, versican, perlecan, matrix metalloproteinase (MMP)-1 & 13, and ADAMTS-5, in the lesion-site AF and NP but not in the contralateral AF. ADAMTS-4 messenger RNA was increased in the lesion-site AF but decreased in the NP. Despite an upregulation in MMPs, there was no change in MMP- or ADAMTS-generated aggrecan neoepitopes in any region of the IVD in lesion or sham discs. CONCLUSION: Lumbar IVDD was reproducibly induced with a 6 × 20 mm(2) annular lesion, with focal dysregulation of MMP gene expression, cell cloning in the inner AF, loss of NP aggrecan, and disc height. Loss of aggrecan from the NP was not attributable to increased proteolysis in the interglobular domain by MMPs or ADAMTS.

A preliminary study on the effect of manuka honey on second-intention healing of contaminated wounds on the distal aspect of the forelimbs of horses.

OBJECTIVE: To determine the effect of manuka honey on second-intention healing of contaminated, full-thickness skin wounds in horses. STUDY DESIGN: Experimental. ANIMALS: Adult Standardbred horses (n = 8). METHODS: One wound was created on the dorsomedial aspect of the third metacarpus in both forelimbs, contaminated with feces, and bandaged for 24 hours. Bandages were removed and wounds rinsed with isotonic saline solution. Wounds on 1 limb had manuka honey applied daily (n = 8) whereas wounds on the contralateral limb received no treatment (n = 8). Bandages were replaced and changed daily for 12 days, after which treatment stopped, bandages were removed, leaving wounds open to heal. Wound area was measured 24 hours after wound creation (day 1), then weekly for 8 weeks. Overall time for healing was recorded. Wound area and rate of healing of treated and control wounds were compared statistically. RESULTS: Treatment with manuka honey decreased wound retraction and treated wounds remained significantly smaller than control wounds until day 42; however, there was no difference in overall healing time between treatment and control wounds. CONCLUSIONS: Treatment with manuka honey reduced wound area by reducing retraction but did not affect overall healing time of full-thickness distal limb wounds using this wound-healing model.

A preliminary study on the effect of wounding on transforming growth factor-ß1 and cartilage oligomeric matrix protein concentrations in the skin of horses.

OBJECTIVE: To evaluate whether wound type or site influence the production of cartilage oligomeric matrix protein (COMP) and transforming growth factor ß1 (TGF-ß1), and determine if there is a correlation between TGF-ß1and COMP during healing. STUDY DESIGN: Experimental. ANIMALS: Standardbred horses (n=6), 4-8 years old. METHODS: Six, standardized, full-thickness skin wounds (open, straight, and elliptical) were surgically created on the neck (n=3) and metacarpus (3) on each horse. Wounds were randomly allocated to site and side. Tissue samples were collected before creating wounds and on days 7, 14, and 42. COMP concentration (µg/g dry weight of tissue) was determined using a standard competitive ELISA and TGF-ß1 (ng/g dry weight of tissue) was determined using a commercially available sandwich ELISA. RESULTS: COMP concentrations were higher in intact skin on the neck compared with the metacarpus (P=.02). There was no difference in COMP and TGF-ß1 concentrations between the different wound types or sites during healing. There was no correlation between TGF-ß1 and COMP during healing. CONCLUSIONS: Within the limitations of the study design, production of COMP during healing of skin wounds does not appear to be influenced by wound type or anatomic site, nor does it appear to be correlated with TGF-ß1 concentrations.