Characterisation of key proteoglycans in the cranial cruciate ligaments (CCLs) from two dog breeds with different predispositions to CCL disease and rupture.

Cranial cruciate ligament disease and rupture (CCLD/R) is one of the most common orthopaedic conditions in dogs, eventually leading to osteoarthritis of the stifle joint. Certain dog breeds such as the Staffordshire bull terrier have an increased risk of developing CCLD/R. Previous studies into CCLD/R have found that glycosaminoglycan levels were elevated in cranial cruciate ligament (CCL) tissue from high-risk breeds when compared to the CCL from a low-risk breed to CCLD/R. Our objective was to determine specific proteoglycans/glycosaminoglycans in the CCL and to see whether their content was altered in dog breeds with differing predispositions to CCLD/R. Disease-free CCLs from Staffordshire bull terriers (moderate/high-risk to CCLD/R) and Greyhounds (low-risk to CCLD/R) were collected and key proteoglycan/glycosaminoglycans were determined by semi-quantitative Western blotting, quantitative biochemistry, quantitative reverse transcription polymerase chain reaction, and immunohistochemistry. Gene expression of fibromodulin (P = 0.03), aggrecan (P = 0.0003), and chondroitin-6-sulphate stubs (P = 0.01) were significantly increased, and for fibromodulin this correlated with an increase in protein content in Staffordshire bull terriers compared to Greyhound CCLs (P = 0.02). Decorin (P = 0.03) and ADAMTS-4 (P = 0.04) gene expression were significantly increased in Greyhounds compared to Staffordshire bull terrier CCLs. The increase of specific proteoglycans and glycosaminoglycans within the Staffordshire bull terrier CCLs may indicate a response to higher compressive loads, potentially altering their risk to traumatic injury. The higher decorin content in the Greyhound CCLs is essential for maintaining collagen fibril strength, while the increase of ADAMTS-4 indicates a higher rate of turnover helping to regulate normal CCL homeostasis in Greyhounds.

Changes in the metabolism of chondroitin sulfate glycosaminoglycans in articular cartilage from patients with Kashin-Beck disease.

OBJECTIVES: To identify changes in the expression patterns of enzymes involved in chondroitin sulfate (CS) glycosaminoglycan (GAG) metabolism in articular cartilage proteoglycan (PG) isolated from adolescent patients with Kashin-Beck disease (KBD). METHODS: Samples of articular cartilage were divided into two groups: Control samples (from five normal children), and KBD samples (from five KBD children) aged 3-12 years old. The morphology and pathology of hand joint cartilage were examined by histochemical staining. The localization and expression patterns of enzymes involved in CS GAG metabolism (i.e., PAPS synthetase 2 (PAPSS2), PAPS transporter 1 (PAPST1), Carbohydrate (N-acetylgalactosamine 4-sulfate 6-O) sulfotransferases 15 (CHST15), Arylsulfatase B (ARSB) and N-acetylgalactosamine-6-sulfate sulfatase (GALNS)) were performed using immuno-histochemical analyses. Positive immunostaining in articular cartilage was semi-quantified. RESULTS: Reduced aggrecan staining was observed in KBD samples compared with the control samples. The percentages of positive staining for the anabolic enzymes PAPSS2, PAPST1 and CHST15 in the upper and middle zones of KBD samples were significantly lower than that found in the Controls. In contrast, the percentages of positive staining in KBD samples for the catabolic enzymes ARSB and GALNS were significantly higher than the control samples. However, the staining for all of these GAG metabolism enzymes were hardly observed in the deep zones of KBD cartilage, suggesting that significant cell death and necrosis had occurred in this region. CONCLUSIONS: Our results indicate that alterations of enzymes involved in articular cartilage CS GAG metabolism on PGs in the articular cartilage play an important role in the onset and pathogenesis of KBD in adolescent children.

The recent expansion in the Australian cocaine market: who are the new users and what are the harms?

Cocaine supplies to and within Australia increased after 2006-07, and there is some evidence that cocaine demand may also have risen. However, the extent, nature and public health implications of any changes in cocaine demand remain unclear. Equally unclear, is whether such changes may have been fuelled by declines in two of Australia's other stimulant markets. We examined general population trends in cocaine use and harmful practices and use of related stimulants between 1998 and 2010, and conducted age-period-cohort analyses using five repeated cross-sections of Australia's National Drug Strategy Household Survey. The results indicate past year cocaine use prevalence has increased significantly since 2004, to its highest point in the past 12 years; 2.1% in 2010. But frequency of cocaine use has not increased. Moreover, most harmful practices (injecting, high-quantity use) have remained stable. Changes in the cocaine market appear related to changes in the Australian methamphetamine and ecstasy markets, including declining purity of ecstasy. For example, the cohorts of people most likely to exhibit recent cocaine use were also most likely to have used ecstasy and methamphetamine (those born from 1976 to 1984). The findings indicate that an increase in cocaine demand does not necessarily lead to substantial increases in public health harm: and indeed that the public health implications from the recent increase are likely to be negligible. Moreover, the findings suggest changes to either ecstasy or methamphetamine supply may lead to more shifts in demand for Australia's cocaine market.

A role for adhesion and degranulation-promoting adapter protein in collagen-induced platelet activation mediated via integrin α(2) ß(1).

BACKGROUND: Collagen-induced platelet activation is a key step in the development of arterial thrombosis via its interaction with the receptors glycoprotein (GP)VI and integrin α(2) ß(1) . Adhesion and degranulation-promoting adapter protein (ADAP) regulates α(IIb) ß(3) in platelets and α(L) ß(2) in T cells, and is phosphorylated in GPVI-deficient platelets activated by collagen. OBJECTIVES: To determine whether ADAP plays a role in collagen-induced platelet activation and in the regulation and function of α(2) ß(1). METHODS: Using ADAP(-/-) mice and synthetic collagen peptides, we investigated the role of ADAP in platelet aggregation, adhesion, spreading, thromboxane synthesis, and tyrosine phosphorylation. RESULTS AND CONCLUSIONS: Platelet aggregation and phosphorylation of phospholipase Cγ2 induced by collagen were attenuated in ADAP(-/-) platelets. However, aggregation and signaling induced by collagen-related peptide (CRP), a GPVI-selective agonist, were largely unaffected. Platelet adhesion to CRP was also unaffected by ADAP deficiency. Adhesion to the α(2) ß(1) -selective ligand GFOGER and to a peptide (III-04), which supports adhesion that is dependent on both GPVI and α(2) ß(1), was reduced in ADAP(-/-) platelets. An impedance-based label-free detection technique, which measures adhesion and spreading of platelets, indicated that, in the absence of ADAP, spreading on GFOGER was also reduced. This was confirmed with non-fluorescent differential-interference contrast microscopy, which revealed reduced filpodia formation in ADAP(-/-) platelets adherent to GFOGER. This indicates that ADAP plays a role in mediating platelet activation via the collagen-binding integrin α(2) ß(1). In addition, we found that ADAP(-/-) mice, which are mildly thrombocytopenic, have enlarged spleens as compared with wild-type animals. This may reflect increased removal of platelets from the circulation.

Biotic, temporal and spatial variability of tritium concentrations in transpirate samples collected in the vicinity of a near-surface low-level nuclear waste disposal site and nearby research reactor.

The results of a 21 month sampling program measuring tritium in tree transpirate with respect to local sources are reported. The aim was to assess the potential of tree transpirate to indicate the presence of sub-surface seepage plumes. Transpirate gathered from trees near low-level nuclear waste disposal trenches contained activity concentrations of (3)H that were significantly higher (up to ∼700 Bq L(-1)) than local background levels (0-10 Bq L(-1)). The effects of the waste source declined rapidly with distance to be at background levels within 10s of metres. A research reactor 1.6 km south of the site contributed significant (p < 0.01) local fallout (3)H but its influence did not reach as far as the disposal trenches. The elevated (3)H levels in transpirate were, however, substantially lower than groundwater concentrations measured across the site (ranging from 0 to 91% with a median of 2%). Temporal patterns of tree transpirate (3)H, together with local meteorological observations, indicate that soil water within the active root zones comprised a mixture of seepage and rainfall infiltration. The degree of mixing was variable given that the soil water activity concentrations were heterogeneous at a scale equivalent to the effective rooting volume of the trees. In addition, water taken up by roots was not well mixed within the trees. Based on correlation modelling, net rainfall less evaporation (a surrogate for infiltration) over a period of from 2 to 3 weeks prior to sampling seems to be the optimum predictor of transpirate (3)H variability for any sampled tree at this site. The results demonstrate successful use of (3)H in transpirate from trees to indicate the presence and general extent of sub-surface contamination at a low-level nuclear waste site.

Chondroitin sulphate sulphation motif expression in the ontogeny of the intervertebral disc.

Chondroitin sulphate chains on cell and extracellular matrix proteoglycans play important regulatory roles in developing systems. Specific, developmentally regulated, sulphation motifs within the chondroitin glycosaminoglycan structure may help bind, sequester or present bioactive signalling molecules to cells thus modulating their behaviour. Using monoclonal antibodies 3B3(-), 4C3, 6C3 and 7D4, we have mapped the distribution of different chondroitin sulphation epitopes in a rat intervertebral disc developmental series. The sulphation epitopes had complex, dynamic and specific distributions in the disc and vertebral tissues during their differentiation, growth and ageing. At embryonic day [E]15, prior to disc differentiation, 4C3 and 7D4 occurred within the cellular disc condensations whilst 6C3 was present in the notochordal sheath. At E17, post disc differentiation, 4C3 and 7D4 occurred within the nucleus pulposus, inner annulus and vertebral bodies; 3B3(-) in the nucleus, inner annulus, annulus/vertebral body interface and perichondrium; and 6C3, ventrally, within the perichondrium. At E19, 3B3(-), 4C3 and 7D4 became further restricted to the nucleus, inner annulus, annulus/vertebral body interface and perichondrium. Prior to birth, all four epitopes occurred within the inner annulus and nucleus, with 6C3 and 7D4 also occurring within the future end-plate. Postnatal expression of the sulphation epitopes was more widespread in the disc and also within the growth plate. At 4 months, the epitopes were associated with chondrocyte clusters within the nucleus; and at 24 months, with annular lesions. Overall, our data suggests that differential sulphation of chondroitin correlates with significant events in development, growth and aging of the rat intervertebral disc.

Movement of a tritium plume in shallow groundwater at a legacy low-level radioactive waste disposal site in eastern Australia.

Between 1960 and 1968 low-level radioactive waste was buried in a series of shallow trenches near the Lucas Heights facility, south of Sydney, Australia. Groundwater monitoring carried out since the mid 1970s indicates that with the exception of tritium, no radioactivity above typical background levels has been detected outside the immediate vicinity of the trenches. The maximum tritium level detected in ground water was 390 kBq/L and the median value was 5400 Bq/L, decay corrected to the time of disposal. Since 1968, a plume of tritiated water has migrated from the disposal trenches and extends at least 100 m from the source area. Tritium in rainfall is negligible, however leachate from an adjacent and fill represents a significant additional tritium source. Study data indicate variation in concentration levels and plume distribution in response to wet and dry climatic periods and have been used to determine pathways for tritium migration through the subsurface.

Dietary fatty acids and arthritis.

Musculoskeletal complaints are the second most frequent reason for medical treatments. Within these diseases rheumatoid arthritis (RA) and, especially, osteoarthritis (OA) are common. Although the causes of arthritis are multifactorial and not fully understood, clinical trials have generally shown benefit from dietary n-3 polyunsaturated fatty acids. This has usually been attributed to their anti-inflammatory properties. Recently we have used in vitro model systems to study the molecular mechanism(s) by which n-3 PUFAs may act to alleviate the symptoms of arthritis. These experiments showed that n-3 PUFAs reduce expression of cartilage-degrading proteinases, cyclooxygenase-2 and inflammatory cytokines. Eicosapentaenoic acid (EPA) was more effective than docosahexaenoic acid (DHA) or alpha-linolenic acid. The data provide a scientific rationale for the consumption of n-3 fatty acids as part of a healthy diet and perhaps in treating arthritis.

The novel Syk inhibitor R406 reveals mechanistic differences in the initiation of GPVI and CLEC-2 signaling in platelets.

BACKGROUND: Syk is a key mediator of signaling pathways downstream of several platelet surface receptors including GPVI/FcRgamma collagen receptor, the C-type lectin receptor CLEC-2, and integrin alphaIIbbeta3. A recent study identified the novel small molecule R406 as a selective inhibitor of Syk. OBJECTIVES: The present study evaluates the role of Syk in human platelets using the novel inhibitor R406. METHODS: Agonist-induced GPVI and CLEC-2 signaling were assessed using aggregometry, immunoprecipitation and western blotting to determine the effects of R406 on platelet activation. RESULTS: We demonstrate R406 to be a powerful inhibitor of Syk in human platelets. R406 abrogated shape change and aggregation induced by activation of GPVI and CLEC-2, and reduced platelet spreading on fibrinogen. The inhibitory effect of R406 was associated with inhibition of tyrosine phosphorylation of signaling proteins that lay downstream of Syk for all three receptors, including PLCgamma2. Strikingly, R406 markedly inhibited tyrosine phosphorylation of CLEC-2 and Syk downstream of CLEC-2 activation, whereas phosphorylation of Syk downstream of GPVI and integrin alphaIIbbeta3 was unaffected. CONCLUSIONS: The inhibitory effect of R406 provides direct evidence of a role for Syk in GPVI, CLEC-2 and integrin alphaIIbbeta3 signaling in human platelets. Further, the results demonstrate a critical role for Syk in mediating tyrosine phosphorylation of CLEC-2, suggesting a novel model in which both Src and Syk kinases regulate tyrosine phosphorylation of the C-type lectin receptor leading to platelet activation.

Relative efficacies of omega-3 polyunsaturated fatty acids in reducing expression of key proteins in a model system for studying osteoarthritis.

OBJECTIVE: To assess the relative efficacy of three different omega-3 (n-3) polyunsaturated fatty acids (PUFAs) in suppressing the mRNA levels for important proteins involved in the etiology of osteoarthritis (OA). METHODS: A model cell culture system (bovine chondrocytes) was used. Inflammatory factors and enzymes involved in OA were induced by exposure of the chondrocyte cultures to interleukin-1alpha (IL-1alpha). The effect of pre-incubating cultures with various amounts of exogenous fatty acids on subsequent levels of mRNAs was assessed by reverse transcription-polymerase chain reactions (RT-PCR). RESULTS: Exposure of cultures to IL-1alpha induced expression of the cartilage proteinases A Disintegrin And Metalloproteinase with ThromboSpondin motifs (ADAMTS)-4 and ADAMTS-5, cyclooxygenase (COX)-2, the matrix metalloproteinase (MMP)-3 and the inflammatory cytokines IL-1alpha, interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNF-alpha). n-3 PUFAs were able to reduce the levels of mRNA for ADAMTS-4, ADAMTS-5, MMP-3, MMP-13, COX-2 (but not COX-1), IL-1alpha, IL-1beta and TNF-alpha. Eicosapentaenoic acid (EPA) was the most effective, followed by docosahexaenoic (DHA) and then alpha-linolenic (ALA) acid. The n-6 PUFA, arachidonic acid (AA) had no effect. CONCLUSION: These results show that omega-3 (n-3) PUFAs cause a reduction in the mRNA levels for various proteins known to be important in the pathology of OA. They provide a molecular explanation, at least in part, for beneficial effects of dietary omega-3 PUFAs for the amelioration of symptoms of the disease. The relative efficacy of EPA suggests that this omega-3 PUFA may be especially useful for dietary supplementation in patients with OA.

Molecular basis of platelet activation by an alphaIIbbeta3-CHAMPS peptide.

BACKGROUND: A novel method, known as computed helical anti-membrane protein (CHAMP), for the design of peptides that bind with high affinity and selectivity to transmembrane helices was recently described and illustrated using peptides that bind alphaIIb- and alphav-integrin subunits, which induce selective activation of integrins alphaIIbbeta3 and alphavbeta3, respectively. OBJECTIVES: In the present study, we have investigated the ability of an alphaIIb-CHAMPS peptide (termed integrin-activatory-peptide or IAP) to stimulate protein tyrosine phosphorylation and aggregation in human and mouse platelets. METHODS: The ability of IAP to stimulate platelet aggregation and dense granule secretion was measured in washed preparations of human and mouse platelets. Samples were taken for measurement of tyrosine phosphorylation. RESULTS: IAP stimulates robust tyrosine phosphorylation of the tyrosine kinase Syk and the FcR gamma-chain, but only weak phosphorylation of PLCgamma2. Aggregation to low but not high concentrations of IAP is reduced in the presence of the Src kinase inhibitor, PP1, or by inhibitors of the two feedback agonists, ADP and thromboxane A(2) (TxA(2)) suggesting that activation is reinforced by Src kinase-driven release of ADP and TxA(2). Unexpectedly, aggregation by IAP is only partially inhibited in human and mouse platelets deficient in integrin alphaIIbbeta3. Further, IAP induces partial aggregation of formaldehyde-fixed platelets. CONCLUSIONS: The present study demonstrates that the alphaIIb-CHAMPS peptide induces platelet activation through integrin alphaIIbbeta3-dependent and independent pathways with the former mediating tyrosine phosphorylation of FcR gamma-chain and Syk. The use of the alphaIIb-CHAMPS peptide to study integrin alphaIIbbeta3 function is compromised by non-integrin-mediated effects.

Differential roles for the adapters Gads and LAT in platelet activation by GPVI and CLEC-2.

BACKGROUND: The adapter proteins SLP-76 and LAT have been shown to play critical roles in the activation of PLCgamma2 in platelets downstream of GPVI/FcRgamma and the C-type lectin receptor CLEC-2. SLP-76 is constitutively associated with the adapter Gads in platelets, which also binds to tyrosine phosphorylated LAT, thereby providing a potential pathway of regulation of SLP-76. OBJECTIVE: In the present study, we have compared the role of Gads alongside that of LAT following activation of the major platelet glycoprotein receptors using mice deficient in the two adapter proteins. RESULTS: Gads was found to be required for the efficient onset of aggregation and secretion in response to submaximal stimulation of GPVI and CLEC-2, but to be dispensable for activation following stronger stimulation of the two receptors. Gads was also dispensable for spreading induced through integrin alpha(IIb)beta(3) or the GPIb-IX-V complex. Further, Gads plays a negligible role in aggregate formation on collagen at an arteriolar rate of shear. In stark contrast, platelets deficient in the adapter LAT exhibit a marked decrease in aggregation and secretion following activation of GPVI and CLEC-2, and are unable to form stable aggregates on collagen at arteriolar shear. CONCLUSIONS: The results demonstrate that Gads plays a key role in linking the adapter LAT to SLP-76 in response to weak activation of GPVI and CLEC-2 whereas LAT is required for full activation over a wider range of agonist concentrations. These results reveal the presence of a Gads-independent pathway of platelet activation downstream of LAT.

Globular adiponectin induces platelet activation through the collagen receptor GPVI-Fc receptor gamma chain complex.

BACKGROUND: The adipocyte-derived cytokine, adiponectin (Ad), exerts potent vascular effects, although the direct effects of Ad on blood platelets are unclear. OBJECTIVE: The influence of globular Ad (gAd) on blood platelet function was investigated. RESEARCH DESIGN AND METHODS: We measured platelet aggregation and tyrosine phosphorylation signaling events in human and mouse platelets. The ability of gAd to activate Glycoprotein VI (GPVI) activity was determined with a NFAT luciferase reporter assay. RESULTS: gAd, but not full length Ad, induced rapid aggregation and granule secretion of human and mouse platelets through a pathway that is ablated under conditions of Src kinase inhibition, indicating a tyrosine kinase-dependent mechanism. Consistent with this, gAd stimulates rapid tyrosine phosphorylation of several proteins in human and mouse platelets. The pattern of increase in tyrosine phosphorylation was similar to that induced by collagen, with the tyrosine kinase Syk and PLCgamma2 being identified among the list of tyrosine phosphorylated proteins. As collagen activates platelet through the GPVI-Fc receptor gamma-chain (FcRgamma) complex, we used FcRgamma null platelets (which also lack GPVI) to explore the mechanism by which gAd stimulates platelets. Stimulation of tyrosine phosphorylation and platelet aggregation by gAd was abolished in FcRgamma null platelets and markedly reduced in the absence of PLCgamma2. Further, GPVI was confirmed as a collagen receptor for gAd by increased luciferase activity in Jurkat T-cells transfected with GPVI. CONCLUSIONS: We identify gAd as a novel ligand for GPVI that stimulates tyrosine kinase-dependent platelet aggregation. Our data raise the possibility that gAd may promote unwanted platelet activation at sites of vascular injury.

Articular cartilage metabolism in patients with Kashin-Beck Disease: an endemic osteoarthropathy in China.

OBJECTIVE: The objective of this study was to investigate CD44 and proteoglycan metabolism in patients suffering from Kashin-Beck Disease (KBD), an endemic osteoarthropathy that affects 2.5 million of 30 million people living in the KBD regions of China. METHODS: Immunohistochemical analyses of cluster of differentiation-44 (CD44), BC-13 and 3-B-3(-) expression were performed in cartilage sections harvested from KBD and normal patients. In addition, the serum levels of soluble CD44 (sCD44), interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and matrix metalloproteinase-1 were determined using a sandwich enzyme linked immunosorbent assay. RESULTS: Hematoxylin & eosin and toluidine blue staining indicated that there was cell necrosis and proteoglycan loss in cartilage from both KBD children and adult cartilage. Strong immunohistochemical staining for CD44, BC-13 and 3-B-3(-) occurred in the majority of adult KBD patients and most KBD children. Furthermore, statistically significant elevated levels of sCD44, IL-1beta and TNF-alpha were found in the sera of both adult and child KBD patients when compared to the levels of normal adult and child controls. Interestingly, IL-1beta and TNF-alpha serum levels were all high in normal children from KBD regions when compared to normal children from non-KBD regions suggesting that unidentified factors (e.g., a genetic predisposition) may protect some people from KBD pathology. CONCLUSION: Our results demonstrate that altered CD44, IL-1beta and TNF-alpha metabolism occurs in the pathogenesis of KBD and there is an increased aggrecanase-generated proteoglycan loss from KBD adult and child cartilage. These primary metabolic changes are likely to be significant contributing factor causing pathological joint formation and instability that leads to secondary osteoarthritis in KBD patients.

Intercontinental dispersal prior to human translocation revealed in a cryptogenic invasive tree.

In this study, complementary species-level and intraspecific phylogenies were used to better circumscribe the original native range and history of translocation of the invasive tree Parkinsonia aculeata. Species-level phylogenies were reconstructed using three chloroplast gene regions, and amplified fragment length polymorphism (AFLP) markers were used to reconstruct the intraspecific phylogeny. Together, these phylogenies revealed the timescale of transcontinental lineage divergence and the likely source of recent introductions of the invasive. The sequence data showed that divergence between North American and Argentinean P. aculeata occurred at least 5.7 million years ago, refuting previous hypotheses of recent dispersal between North and South America. AFLP phylogenies revealed the most likely sources of naturalized populations. The AFLP data also identified putatively introgressed plants, underlining the importance of wide sampling of AFLPs and of comparison with uniparentally inherited marker data when investigating hybridizing groups. Although P. aculeata has generally been considered North American, these data show that the original native range of P. aculeata included South America; recent introductions to Africa and Australia are most likely to have occurred from South American populations.

Human Experience Modeler: context-driven cognitive retraining to facilitate transfer of learning.

We describe a cognitive rehabilitation mixed-reality system that allows therapists to explore natural cuing, contextualization, and theoretical aspects of cognitive retraining, including transfer of training. The Human Experience Modeler (HEM) mixed-reality environment allows for a contextualized learning experience with the advantages of controlled stimuli, experience capture and feedback that would not be feasible in a traditional rehabilitation setting. A pilot study for testing the integrated components of the HEM is discussed where the participant presents with working memory impairments due to an aneurysm.

Cytokine induced metalloproteinase expression and activity does not correlate with focal susceptibility of articular cartilage to degeneration.

OBJECTIVE: To determine whether the focal susceptibility to cartilage degeneration in joints is related to a differential response to cytokine stimulation. METHODS: Compare aggrecan and collagen catabolism in in-vitro models of cartilage degradation induced by retinoic acid (RA), interleukin-1 (IL-1), tumor necrosis factor alpha (TNF) and IL-1 plus oncostatin M (OSM). Glycosaminoglycan (GAG) and hydroxyproline (HyPro) quantification and Western immunoblot analyses of aggrecan and collagen degradation products were undertaken in explant cultures of normal cartilage from regions of equine joints with a known high and low susceptibility to degeneration in disease. RNA isolation and semi quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis were performed to determine the expression of aggrecanases, matrix metalloproteinases (MMPs) and their inhibitors. RESULTS: Although the rate of basal cartilage aggrecan turnover was dependent on joint region there was no difference in the response of different cartilages to cytokines. Individual animals did show a significant difference in the response of certain cartilages to cytokines, with both decreased and increased aggrecan loss in cartilage with a low susceptibility to degeneration. Aggrecan release in both short- and long-term cultures from all cartilages was associated with increased cleavage by aggrecanases rather than MMPs. There was a poor correlation between expression of aggrecanases, MMPs or their inhibitors and cytokine induced aggrecan catabolism. IL-1 alone was able to stimulate collagen breakdown in equine articular cartilage and surprisingly, significantly more collagen loss was induced in cartilage from regions less susceptible to degeneration. CONCLUSIONS: Collectively, these studies suggest that a regional difference in response to catabolic cytokines is unlikely to be a factor in the initiation of focal cartilage degeneration in osteoarthritis (OA).

Using radiotracer techniques for coastal hydrodynamic model evaluation.

A three-dimensional (3D) water circulation and contaminant transport model of Manila Bay has been developed with the aim of better understanding the formation and movement of harmful algal blooms. Radiotracer techniques were used to evaluate the model by recording the dispersion of a tracer at depths of 2 and 15 m near the injection point. The selected tracer was 99mTc eluted from a molybdenum/technetium medical generator. The rationale for the choice of the tracer and the location of the injection is discussed. At 2 m the transport was dominated by the prevailing winds, and at 15 m by tidally induced currents. The development of the hydrodynamic model and its experimental evaluation were iterative processes. The experimental study confirmed the need for full 3D modelling of Manila Bay; quantified the impact of the prevailing wind field on contaminant dispersion near the injection point; and allowed the calculation of transverse dispersivity to guide the selection of parameter values used in the overall model.

Recombinant equine growth hormone administration: effects on synovial fluid biomarkers and cartilage metabolism in horses.

REASONS FOR PERFORMING STUDY: Recombinant equine growth hormone (reGH) has recently been evaluated for effects on body condition and wound healing. It has the potential to influence articular cartilage via stimulation of IGF-1. OBJECTIVES: To investigate effects of administration on synovial joint metabolism. METHODS: Six mature horses were given 20 microg/kg bwt reGH daily for 8 weeks by i.m. injection. Three control horses were injected with sterile water. Serum and synovial fluid samples were collected at 6, 8, 11 and 16 weeks for GH and IGF-1 assays. Articular cartilage harvested at week 16 was evaluated by Western analysis using monoclonal antibodies BC-13, BC-4, 8-A-4 and CH-3. RESULTS: Concentrations of IGF-1 in serum and synovial fluid were significantly elevated (P < 0.05) at 6 and 8 weeks in the reGH group. Glycosaminoglycan concentrations in synovial fluid were significantly less than controls at these time points, suggesting that reGH may modulate proteoglycan metabolism in articular cartilage. In the reGH group, there were not any alterations in synovial fluid content of 3B3(-) epitope or aggrecan metabolite, or in aggrecan or link protein catabolites retained within cartilage, that might be expected with development of osteoarthritis. CONCLUSIONS: Intramuscular administration of reGH may be a more efficient means of delivery of IGF-1 to joints for cartilage resurfacing initiatives. POTENTIAL RELEVANCE: We found no alterations in cartilage metabolism indicative of development of osteoarthritis.