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Biomedical subjects

L B Jeffcott

Publications and source records attributed to L B Jeffcott.

At least 37 records · Page 2Linked to original sources

Equine dyschondroplasia (osteochondrosis)--histological findings and type VI collagen localization.

This study describes (1) the histological appearance of dyschondroplasia, the primary lesion of osteochondrosis, in articular cartilage of the horse and (2) the localization of type VI collagen which is an important constituent of the extracellular matrix (ECM). Dyschondroplastic cartilage was identified on the basis of the presence of cartilage cores (i.e., cartilage extending into the subchondral bone) and confirmed with subsequent histological examination. Full-thickness cartilage samples from 57 horses were collected and paraffin embedded. Histological examination was used to examine the normal architecture of equine growth cartilage and to determine the presence of various pathological changes in dyschondroplastic lesions. Immunolocalization was used to identify type VI collagen in normal and dyschondroplastic lesions. The abnormalities observed in the dyschondroplastic cartilage fell into two groups. In Group A (n = 18) the lesions were associated with a disruption in the normal sequential transition of the chondrocytes through proliferation and maturation resulting in an accumulation of large numbers of small, rounded chondrocytes. A decrease in type VI collagen immunoreactivity compared with normal animals was detected except around chondrocyte clusters. Group B lesions (n = 9) were characterized by an alteration in the staining pattern of the mineralized cartilage and underlying bone. In these lesions type VI collagen immunoreactivity was increased. In both groups the presence of retained blood vessels, chondrocyte clusters, chondronecrosis and fissure formation was detected. These two histologically-distinct groups suggest that equine dyschondroplasia may be comprised of different pathological entities and that it is associated with alterations in the pattern of distribution of an ECM protein.

Animals↗

Cellular heterogeneity in cathepsin D distribution in equine articular cartilage.

The distribution of cathepsin D in normal equine growth cartilage has been examined immunocytochemically using an antiserum raised against human cathepsin D. The cross-reactivity and specificity of the antiserum for equine cathepsin D was confirmed, and its lysosomal localisation was demonstrated in horse skin fibroblasts by confocal scanning microscopy. Cultured horse chondrocytes were heterogenous in their expression of cathepsin D. Heterogeneity of distribution of the enzyme was also seen in chondrocytes in cartilage from different anatomical sites. A high level of cathepsin D was observed in the deep layer of cartilage from the lateral trochlear ridge of the distal femur. Cathepsin D was absent in the hypertrophic zone of the distal radial growth plate.

Animals↗

Cartilage canals in equine articular/epiphyseal growth cartilage and a possible association with dyschondroplasia.

Cartilage canals have been described in most mammals and contain the vascular elements necessary for the maintenance of epiphyseal growth cartilage. The presence and longevity of cartilage canals in developing articular/epiphyseal cartilage of horses is described for the first time. Growth cartilages from 30 normal horses (from 130 days gestation to age 2 years) and 6 cases of dyschondroplasia (age < 15 months) were examined. Cartilage canals were located in the epiphyses of fetuses as early as 130 days gestation. The secondary centres of ossification had developed by 250 days gestation and, at full term (approximately 340 days), the articular/epiphyseal cartilage was highly vascularised. Cartilage canals were closely surrounded by small round chondrocytes even when present in the zone of hypertrophic chondrocytes. From late gestation until at least age 10 weeks, blood vessels surrounded by cartilage were found extending beyond the zone of hypertrophic chondrocytes into the secondary centre of ossification. Cartilage canals were absent from normal articular/epiphyseal cartilage of animals age > 6 months. Cartilage canals were associated with retained cartilage of dyschondroplastic lesions found in animals age < 15 months. The presence of cartilage canals in association with dyschondroplastic lesions suggests they may have a role in the pathogenesis of this important disease.

Animals↗

Expression of transforming growth factor-beta 1 in normal and dyschondroplastic articular growth cartilage of the young horse.

This study describes the distribution pattern of transforming growth factor-beta 1 (TGF-beta 1) mRNA and protein in normal pre- and post natal growth cartilage and alterations present in lesions of dyschondroplasia (osteochondrosis). TGF-beta 1 expression and immunoreactivity have been investigated by in situ hybridisation and immunolocalisation in the articular/epiphyseal growth cartilage of the lateral trochlear ridge of the distal femur. Cartilage was obtained from 19 normal Thoroughbred horses (5 prenatal and 14 post natal horses) and 15 post natal horses with dyschondroplasia (DCP). TGF-beta 1 mRNA expression and immunoreactivity were detected in the proliferative and upper hypertrophic zones in both pre- and post natal normal articular/epiphyseal cartilage. However, mRNA itself was only detected in the mid- and lower hypertrophic zones. Immunoreactivity was identified intracellularly with some nuclear staining observed. In focal lesions of DCP mRNA expression and immunoreactivity were reduced compared to normal cartilage, but strong mRNA expression was observed in the chondrocyte clusters immediately surrounding a lesion of DCP. The results described in this study demonstrate alterations in TGF-beta 1 dyschondroplastic lesions and indicate that it could be involved in the pathogenesis of this condition in the horse.

Animals↗

Effects of insulin and insulin-like growth factors I and II on the growth of equine fetal and neonatal chondrocytes.

The effects of insulin and insulin-like growth factors (IGFs) I and II on fetal and foal chondrocytes were investigated in vitro. Chondrocytes from the lateral trochlear ridge of the distal femur were obtained from 2 fetuses (280 and 320 days gestation) and one 4-day-old foal and cultured. Membrane proteins consistent with type 1 and type 2 IGF receptors were demonstrated by radioligand cross linking and equilibrium binding analysis. It was demonstrated that both IGF-I and IGF-II acted as mitogens for isolated equine chondrocytes when present as the sole mitogenic factor in monolayer culture. It was further shown that whereas insulin was able to promote the survival and expansion of cell populations of chondrocytes in culture there was significantly reduced mitogenic stimulation compared to the IGFs. These results suggest that the role of insulin in growth cartilage may be to promote chondrocyte survival, or to suppress differentiation/apoptosis. This supports the hypothesis that relative hyperinsulinaemia may be a contributory factor to equine dyschondroplasia (osteochondrosis). Understanding of contributory, and possibly triggering factors such as this may allow the development of modified methods of husbandry which minimise the risk of disease in populations with a known predisposition.

Animals↗

Transverse path of ultrasound waves in thick-walled cylinders.

The path is examined of a beam of ultrasound through the wall and central cavity of a series of thick-walled cylinders made of polymethylmethacrylate (PMMA) and brass used to simulate the cross-section of the cortical shaft of a long long bone. Water is used as a coupling medium for the external ultrasound transducers and to fill the central cavity. Broadband pulse excitation is employed with two focused 2.5 MHz transducers mounted on a digital calliper. The transmission times of flight are measured on a 100 MHz digital cathode ray oscilloscope. Two distinct pulses of ultrasound were consistently observed at the receiver transducer, the first due to a nonlinear path only in the cylinder wall, and the second due to a linear path travelling directly between the transducers through the centre of the cylinder. The nonlinear path can be explained by refraction at the external surface of the cylinder and diffraction around the central cavity. The nonlinear path is influenced by the relative size of the central cavity. The proposed model for the ultrasound propagation in the cylinder wall gives an excellent correlation with the observed speed and attenuation. The technique can be applied to long bones and has considerable potential as a means of measuring bone quality in human and veterinary medicine.

Bone Density↗

Localisation of alkaline phosphatase in equine growth cartilage.

The aim of this study was to localise alkaline phosphatase (ALP) activity in equine growth cartilage both histochemically and ultrastructurally. For histochemical studies, full thickness growth cartilage samples were obtained from 6 anatomical sites from 16 horses and ponies ranging in age from 90 d postconception to 12 years of age. For ultrastructural studies, samples were obtained from the lateral trochlear ridge of the distal femur of 3 animals ranging in age from 157 d postconception to 12 months of age. Alkaline phosphatase in histological sections was localised using a substituted naphthol reaction which revealed enzyme activity around the cell surfaces of hypertrophic chondrocytes. ALP activity was quantified by determining the labelling index of ALP positive chondrocytes. The labelling index depended on (1) the age of the animal (activity being highest in animals < 6 months old), and (2) the anatomical site (activity being lowest in the proximal phalanges and highest in the growth plate, reflecting the degree of long bone growth at each site). In order to demonstrate ALP activity ultrastructurally, two capturing agents were used: cerium and lead. This enzyme was visualised by the precipitation of electron-dense salts at the site of activity. ALP activity was localised at low levels on the surface membranes of chondrocytes in the midproliferative zone. The enzyme activity increased throughout the proliferative zone and was most intense on chondrocytes of the hypertrophic zone. ALP activity was seen to be concentrated on the matrix vesicles, the putative sites of mineralisation. This work demonstrates, for the first time in the horse, that ALP is a useful marker of incipient bone formation.

Aging↗

Surface strain on the dorsal metacarpus of thoroughbreds at different speeds and gaits.

Stacked rectangular rosette strain gages were used to measure surface bone strain on the dorsomedial cortex of the third metacarpal bones in 6 adult Thoroughbred horses during treadmill exercise. The peak principal compressive and tensile strains, angle of the peak strains and strain rate during loading were calculated and compared with speed and gait. At speeds from 4 to 14 m/s loading on the metacarpus resulted in compression of the dorsomedial cortex. Tension only occurred consistently in 1 forelimb of 1 horse at the walk (2 m/s). The angle of peak compression was within 10 degrees of the longitudinal axis of the bone, and the magnitude of compression was proportional to the speed of exercise. Mean maximum compressive strains in the 6 horses at 2 m/s were -668 mu strain, -1,204 mu strain at 4 m/s; -1,617 mu strain at 6 m/s; -2,127 mu strain at 8 m/s, and -2,533 mu strain at 10 m/s. Recordings from 3 horses gave a mean -3,206 mu strain at 12 m/s, and strains at 14 m/s approached -4,000 mu strain. Peak strains were similar in both forelimbs at the canter while strain rate tended to be higher in the leading forelimb.

Animal Husbandry↗

In vivo measurement of bone quality in the horse: estimates of precision for ultrasound velocity measurement and single photon absorptiometry.

The in vivo precision of ultrasound velocity measurement and single photon absorptiometry for the assessment of equine bone quality is discussed. In vivo precisions for ultrasound velocity measurements were less than 0.5 per cent, whereas cortical cross-sectional area, compact bone density and modulus of elasticity were around 1 per cent, and bone mineral content and density were just over 2 per cent. Except for ultrasound velocity, substantial improvements could be achieved by taking the mean of five readings for each measurement. The long-term precision of the techniques was also high, with all precision values being less than 3 per cent. The possible sources of variation in ultrasound velocity measurements were also investigated. The method for combining ultrasonic and photon absorptiometric data for equine bone quality assessment has been proven to be highly accurate and precise. There appears to be no reason why the same principles cannot be applied with equal success to the noninvasive assessment of bone quality in humans.

Absorptiometry, Photon↗

Effects of treadmill exercise on cortical bone in the third metacarpus of young horses.

The effects of exercise and relative inactivity on cortical bone were compared in young horses. Two groups were used; one was given a 14-week programme of exercise (n = 6) and the other kept as unexercised controls (n = 6). The first nine weeks of exercise involved trotting and cantering (2 to 4 km d-1 at speeds up to 12 m s-1) on a treadmill set at an incline of 3 degrees. Over the next five weeks the horses were trained at near maximal speeds (that is, up to 14.5 m s-1) with no incline of the treadmill. At the end of the programme marked differences in cortical porosity and distribution of subperiosteal osteogenesis at the mid-shaft of the third metacarpal bone were found between the groups. Histomorphometrical examination of the dorsal cortex showed minimal bone remodelling in the exercised horses, but extensive modelling as evidenced by the large amount of subperiosteal bone formation. In contrast, the unexercised horses had significantly more bone remodelling and less formation of subperiosteal bone. The histomorphometric and microradiographic findings provided an explanation for changes in the non-invasive bone measurements that occurred during training. Bone mineral content of the mid-metacarpus was found to increase more in the exercised than the unexercised horses despite a lower overall growth in bodyweight. In those horses that completed the full training programme, ultrasound speed increased significantly by the end of the training programme. It remained unchanged in the horse that did not complete the full exercise programme and decreased slightly in the unexercised horses. The difference in ultrasound speed between the groups was considered to reflect differences in intracortical bone porosity, endosteal bone formation and alterations in skin thickness. The stiffness of cortical bone increased significantly in the exercised horses but remained unaltered in the unexercised horses.

Alkaline Phosphatase↗

Pathological aspects of Australian Stringhalt.

Nine horses with clinical signs of Australian Stringhalt were killed and tissues collected for a detailed pathological study. Lesions were limited to peripheral nerves and muscles. The most severely affected nerves were the superficial and deep peroneal, distal tibial, plantar digital, volar and recurrent laryngeal nerve with changes characterised by a selective loss of large diameter myelinated fibres with various degrees of demyelination, fibrosis, Schwann cell proliferation and onion-bulb formation. A routine evaluation of the brain and spinal cord by light microscopy failed to reveal any consistent abnormalities. Morphometric analysis of deep peroneal and recurrent laryngeal nerves confirmed the reduced number of large diameter myelinated axons. Teased fibre preparations of these nerves did not show any abnormalities in internodal distance. The most severe muscle lesions were in the long and lateral digital extensors, cranial tibial, dorsal cricoarytenoid, gracilis and lateral deep digital flexor with extensive atrophy of fibres and diffuse fibrosis. Histochemical evaluation of the long digital extensor from 3 affected horses showed an abnormally wide distribution in fibre size and a reduction in type II fibres compared with controls. These lesions are consistent with a distal axonopathy leading to neurogenic muscle atrophy. The distribution of neuromuscular lesions in Australian Stringhalt may be explained by the susceptibility of longer, larger myelinated nerve fibres to injury, but the cause for this distal axonopathy remains unknown.

Animals↗

Ultrasound speed in the metacarpal cortex--a survey of 347 thoroughbreds in training.

Transmission ultrasound speed was determined in the third metacarpal bone of 347 Thoroughbred racehorses in training. In addition to direct measurement of apparent ultrasound speed and lateral bone diameter, several derived parameters were estimated (cortical ultrasound speed, corticomedullary ratio and cortical cross-sectional area). Multiple regression equations were developed for each ultrasound measurement using the explanatory variables: sex, age, duration and intensity of training, preparation number and previous shin soreness. The horses (114 females, 82 entire males and 151 geldings ranging in age from 20 to 116 months) had been in continuous training for an average of 10.0 weeks when measurements were made. All measurements except corticomedullary ratio were influenced by the sex of the horse. Geldings had significantly lower ultrasound transmission speed than entire males and females, reflecting a lower bone density. However, the geldings had larger cross-sectional areas (ie bone size). We suggest that the relationship between low ultrasound speed (ie low bone density) and large cross-sectional area maintains the overall mechanical integrity of the metacarpus. This relationship (high ultrasound speed and low cross-sectional area) was also found in each sex group. Ultrasound speed and corticomedullary ratio, but not cross-sectional area, were affected by age. Both duration and intensity of training influenced the ultrasound speed measurements. This study indicated that sex, age and state of training affect ultrasound results, but this explained only about 30% of the variance in the data. Other factors, ie genetic variation in bone density and size, differences in soft tissue thickness and distal limb temperature, presumably account for most of the variance.

Age Factors↗

Osteopenic effects of forelimb immobilisation in horses.

Methods for the non-invasive assessment of bone quality were used to monitor the osteopenia induced by immobilising a forelimb in a cast. These techniques included the measurement of ultrasound velocity, single photon absorptiometry and radiographic photodensitometry. Serial measurements were made before, during and after an eight week period of immobilisation of the left forelimb of four adult standardbreds in a light fibreglass cast which included the foot and extended above the carpus. The measurements continued for 12 weeks after the removal of the cast. The results indicated a trend to decreasing bone strength and mineral content in both forelimbs, with the decreases being greater in the limb with the cast. The values tended to recover slowly after the removal of the cast, then more quickly after the horses returned to the paddock four weeks later. The differences between the limb with the cast and the opposite limb were significant at only a few points of comparison.

Absorptiometry, Photon↗

Bone biopsy in the horse. 1. Method using the wing of ilium.

Two hundred and five bone biopsies from the wing of ilium were taken from 52 growing and 70 older horses. The method was initially evaluated in 4 horses at post mortem and then performed in 26 horses under general anaesthesia. The technique was later developed for biopsying horses in the standing position under local anaesthesia (n = 70 adults; n = 22 foals). The core of the bone biopsy consisted of a central cancellous portion between two cortices. This type of sample allowed a complete histomorphometrical analysis. An integral part of the procedure was intravital bone labelling using fluorochromes with a known interval between labels. Potential hazards associated with the use of oxytetracycline labels included the onset of diarrhoea and laminitis in some foals. The method was simple, safe and efficient for both anaesthetised and standing horses. The only difficulties encountered were minor and were associated with blunting of the drill bit, scoring of the internal surface of the drill bit guide and occasional loss of the distal part of the biopsy core. Post operative complications included haematoma formation and skin infections. It was concluded that this technique offered a quick and effective method of collecting bone biopsies in horses, which facilitated the quantitative assessment of skeletal changes.

Animals↗

Bone biopsy in the horse. 2. Evaluation of histomorphometric examination.

The histomorphometrical examination of a series of bone biopsies from the wing of ilium of foals and older horses is reported. Biopsies were obtained from 20 clinically normal horses, which had previously been infused with tetracycline intravenously twice at a 7 day interval at a dose rate of 10 to 15 mg kg-1. The horses were divided into 2 groups. Group 1 consisted of 10 foals (mean age: 4 +/- 1 months) which were biopsied at 3 different sites in the wing of ilium. Group 2 comprised 10 older horses (mean age: 91 +/- 63 months) which were biopsied from only one site in the left ilium. A systematic evaluation of intra-observer, intra-core, intra-bone and inter-bone errors was made to investigate parameter variations that might affect interpretation of bone biopsies. The measurements carried out included cancellous bone volume (Cn-BV/TV), osteoid surface (OS/BS), eroded surface (ES/BS), mean wall thickness (W.Th), mean osteoid thickness (O.Th), distance between labels (Ir.L.Th) and mineralizing surface (MS/BS). The intra-observer variance, the variance between sections, the variance between adjacent biopsies and that between right and left sides proved to be smaller than the inter-individual variance. The estimate of errors associated with the histomorphometrical analysis of bone biopsies is important for correct interpretations of changes due to disease.

Animals↗

Bone biopsy in the horse. 3. Normal histomorphometric data according to age and sex.

Histomorphometrical analysis of bone biopsies from the wing of ilium in 35 clinically normal horses are reported according to age. The biopsies were collected from 20 mixed-bred foals (mean age 6 +/- 1 months) and 15 older horses (mean age 73 +/- 62 months). No gender differences were found in the group of foals examined, but a significant difference (p less than 0.001) in a number of the primary measurements occurred between them and the older horses. The parameters which altered with age were the extent of osteoid surface (OS/BS), the extent of resorption (erosion) surface (ES/BS), the osteoid thickness (O. Th), the mineral appositional rate (MAR), which corresponds to the rate of matrix mineralisation and, if no mineralisation defects exist reflects the amount of bone laid down per day, and the extent of labelled surfaces (MS/BS). These data indicate a much greater rate of bone remodelling in the foals. This was also reflected in a number of the derived parameters (i.e. the extent of quiescent surfaces [QS/BS] and the activation frequency [Ac.f]).

Age Factors↗