Novel roles for syndecan-1 in renal transplantation

Niertransplantatie is een levensreddende behandeling voor patiënten met nierfalen. Op langere termijn is niertransplantatie echter geassocieerd met uitgebreide weefselveranderingen, chronische allo-immuun response tegen de getransplanteerde nier, hoge bloeddruk, verstoringen in de vetstofwisseling en proteïnurie, welke allen zonder behandeling kunnen bijdragen aan interstitiele fibrose en tubulus atrofie en chronisch transplantaatfalen. In dit proefschrift zijn een aantal studies opgenomen die de functies van syndecan-1 bij niertransplantatie beschrijven. Syndecan-1 is een celoppervlak heparan sulfaat proteoglycaan die als co-receptor voor groeifactoren, adhesiemoleculen, cytokines, en lipoproteinen betrokken is bij een aantal belangrijke biologische processen. We bestudeerden de rol van nier en lever syndecan-1 in een ratten model voor niertransplantatie, maar ook onder experimentele proteinurische condities. Naast deze experimentele benadering hebben we ook de rol van syndecan-1 bij niertrnaplantatie patiënten onderzocht. De resultaten die staan beschreven in dit proefschrift laten twee belangrijke functies van syndecan-1 zien, namelijk (1) de rol van syndecan-1 bij de regeneratie en herstel van beschadigd tubulus epitheel; en (2) het effect van een verslechterde nierfunctie op de functie van lever syndecan-1 bij de opname van lipoproteinen en het onstaan van verstoorde vetstofwisseling bij nierpatiënten. Deze bevindingen zijn van belang voor het ontwerpen van nieuwe behandelingsstrategieën om tubulus schade en verstoorde vetstofwisseling bij nierpatienten te kunnen behandelen

The expression of nerve growth factor in healthy and inflamed equine chondrocytes analysed by capillary western immunoassay

Nerve Growth Factor (NGF) is a signalling molecule for pain and inflammation. NGF is increased in synovial fluid from osteoarthritic humans and animals, compared to healthy controls. Monoclonal antibody therapy directed against NGF has been approved to treat pain in osteoarthritic dogs but despite many years of trialling, therapy has not been approved for human use. One reason for this is that adverse reactions with rapidly progressing osteoarthritis has occurred in some individuals. More detailed knowledge of NGF expression in joints is needed. In this study, capillary-based Simple Western was used to analyse NGF in cultured equine chondrocytes. Chondrocytes were collected post mortem from three macroscopically healthy intercarpal joints and three intercarpal joints with mild osteoarthritic changes. The chondrocytes were expanded to passage one and seeded in chondrogenic medium to maintain the phenotype. On day four, cells were either stimulated with LPS or kept untreated in medium. All cells were harvested on day five. Wes analysis of lysates did not show mature NGF but two proforms, 40 and 45 kDa, were identified. Results were confirmed with western blot. The same proforms were expressed in chondrocytes from healthy and osteoarthritic joints. Acute inflammation induced by LPS stimulation did not change the forms of expressed NGF. Capillary Simple Western offers a sensitive and sample -sparing alternative to traditional western blot. However, confirmation of peaks is imperative in order to avoid misinterpretation of findings. In addition, in this case the method did not offer the possibility of quantification advertised by the manufacturers

Biglycan neo-epitope (BGN(262)), a novel biomarker for screening early changes in equine osteoarthritic subchondral bone

Objective: Native biglycan (BGN), which can undergo proteolytic cleavage in pathological conditions, is well known to be involved in bone formation and mineralization. This study aimed to delineate the specific cleavage fragment, a neo-epitope for BGN (BGN(262)), in synovial fluid (SF) from young racehorses in training, osteoarthritic (OA) joints with subchondral bone sclerosis (SCBS), and chip fracture joints.Design: A custom-made inhibition ELISA was developed to quantify BGN(262) in SF. Cohort 1: A longitudinal study comprising 10 racehorses undergoing long-term training. Cohort 2: A cross-sectional study comprising joints from horses (N = 69) with different stages of OA and radiographically classified SCBS. Cohort 3: A cross-sectional study comprising horses (N = 9) with chip fractures. Receiver operating characteristic (ROC) curve analysis was performed (healthy joints vs chip joints) to evaluate BGN(262) robustness.Results: Cohort 1: SF BGN(262) levels from racehorses showed a statistical increase during the first 6 months of the training period. Cohort 2: BGN(262) levels were significantly higher in the SF from severe SCBS joints. Cohort 3: SF BGN(262) levels in chip fracture joints showed a significant increase compared to normal joints. The ROC analysis showed an AUC of 0.957 (95% C.I 0.868-1.046), indicating good separation between the groups.Conclusions: The data presented show that BGN(262) levels increase in SF in correlation with the initiation of training, severity of SCBS, and presence of chip fractures. This suggests that BGN262 is a potential predictor and a novel biomarker for early changes in subchondral bone (SCB), aiming to prevent catastrophic injuries in racehorses. (c) 2022 The Author(s). Published by Elsevier Ltd on behalf of Osteoarthritis Research Society International. This is an open access article under the CC BY license (http://creativecommons.org/ licenses/by/4.0/)

Bone Biomarker in Feedlot Cattle from Two Different Production Systems

The rapid growth rate in feedlot cattle is likely to promote joint abnormalities like osteochondrosis dissecans (OCD) and osteoarthritis (OA) with subsequent lameness. We have identified a small leucine-rich repeat proteoglycan (SLRP) biomarker at cleavage site 262GLGHNQIRM (BGN262) arising from the fragmentation of biglycan (BGN) in subchondral bone associated with OA. With a validated custom-made ELISA, BGN262 has been quantified in serum from cattle. The concentration of BGN262 in serum from cattle raised in the conventional and all-natural production systems increased at harvest compared to the starting period. The limitation of the study is the small sample size. However, the promising results encourage a further evaluation of BGN262 and its potential as a biomarker for subchondral bone pathology in cattle

A randomized, triple-blinded controlled clinical study with a novel disease-modifying drug combination in equine lameness-associated osteoarthritis

ObjectiveThis study aimed to test a novel treatment combination (TC) (equivalent to sildenafil, mepivacaine, and glucose) with disease-modifying properties compared to Celestone® bifas® (CB) in a randomized triple-blinded phase III clinical study in horses with mild osteoarthritis (OA). Joint biomarkers (reflecting the articular cartilage and subchondral bone remodelling) and clinical lameness were used as readouts to evaluate the treatment efficacy.MethodsTwenty horses with OA-associated lameness in the carpal joint were included in the study and received either TC (n = 10) or CB (n = 10) drug intra-articularly-twice in the middle carpal joint with an interval of 2 weeks (visit 1 & 2). Clinical lameness was assessed both objectively (Lameness locator) and subjectively (visually). Synovial fluid and serum were sampled for quantification of the extracellular matrix (ECM) neo-epitope joint biomarkers represented by biglycan (BGN262) and cartilage oligomeric matrix protein (COMP156). Another two weeks later clinical lameness was recorded, and serum was collected for biomarkers analysis. The overall health status was compared pre and post-intervention by interviewing the trainer.ResultsPost-intervention, SF BGN262 levels significantly declined in TC (P = 0.002) and COMP156 levels significantly increased in CB (P = 0.002). The flexion test scores improved in the TC compared to CB (P =0.033) and also had an improved trotting gait quality (P =0.044). No adverse events were reported.ConclusionThis is the first clinical study presenting companion diagnostics assisting in identifying OA phenotype and evaluating the efficacy and safety of a novel disease-modifying osteoarthritic drug

Direct Reprograming to Regenerate Myocardium and Repair Its Pacemaker and Conduction System

The regenerative medicine field has been revolutionized by the direct conversion of one cell type to another by ectopic expression of lineage-specific transcription factors. The direct reprogramming of fibroblasts to induced cardiac myocytes (iCMs) by core cardiac transcription factors (Gata4, Mef2c, Tbx5) both in vitro and in vivo has paved the way in cardiac regeneration and repair. Several independent research groups have successfully reported the direct reprogramming of fibroblasts in injured myocardium to cardiac myocytes employing a variety of approaches that rely on transcription factors, small molecules, and micro RNAs (miRNAs). Recently, this technology has been considered for local repair of the pacemaker and the cardiac conduction system. To address this, we will first discuss the direct reprograming advancements in the setting of working myocardium regeneration, and then elaborate on how this technology can be applied to repair the cardiac pacemaker and the conduction system

Direct Reprograming to Regenerate Myocardium and Repair Its Pacemaker and Conduction System

The regenerative medicine field has been revolutionized by the direct conversion of one cell type to another by ectopic expression of lineage-specific transcription factors. The direct reprogramming of fibroblasts to induced cardiac myocytes (iCMs) by core cardiac transcription factors (Gata4, Mef2c, Tbx5) both in vitro and in vivo has paved the way in cardiac regeneration and repair. Several independent research groups have successfully reported the direct reprogramming of fibroblasts in injured myocardium to cardiac myocytes employing a variety of approaches that rely on transcription factors, small molecules, and micro RNAs (miRNAs). Recently, this technology has been considered for local repair of the pacemaker and the cardiac conduction system. To address this, we will first discuss the direct reprograming advancements in the setting of working myocardium regeneration, and then elaborate on how this technology can be applied to repair the cardiac pacemaker and the conduction system

Incipient renal transplant dysfunction associates with tubular syndecan-1 expression and shedding

Syndecan-1 is a transmembrane heparan sulfate proteoglycan involved in regenerative growth and cellular adhesion. We hypothesized that the induction of tubular syndecan-1 is a repair response to incipient renal damage in apparently stable, uncomplicated renal transplant recipients. We quantified tubular syndecan-1 in unselected renal protocol biopsies taken 1 yr after transplantation. Spearman rank correlation analysis revealed an inverse correlation between tubular syndecan-1 expression and creatinine clearance at the time of biopsy (r = -0.483, P <0.03). In a larger panel of protocol and indication biopsies from renal transplant recipients, tubular syndecan-1 correlated with tubular proliferation marker Ki67 (r = 0.518, P <0.0001). In a rat renal transplantation model, 2 mo after transplantation, mRNA expression of syndecan-1 and its major sheddase, A disintegrin and metalloproteinase-17, were upregulated (both P <0.03). Since shed syndecan-1 might end up in the circulation, in a stable cross-sectional human renal transplant population (n = 510), we measured plasma syndecan-1. By multivariate regression analysis, we showed robust independent associations of plasma syndecan-1 with renal (plasma creatinine and plasma urea) and endothelial function parameters (plasma VEGF-A, all P <0.01). By various approaches, we were not able to localize syndecan-1 in vessel wall or endothelial cells, which makes shedding of syndecan-1 from the endothelial glycocalyx unlikely. Our data suggest that early damage in transplanted kidneys induces repair mechanisms within the graft, namely, tubular syndecan-1 expression for tubular regeneration and VEGF production for endothelial repair. Elevated plasma syndecan-1 levels in renal transplantation patients might be interpreted as repair/survival factor related to loss of tubular and endothelial function in transplanted kidneys