RESUMEN
OBJECTIVE: To delineate the activities of decorin and biglycan in the progression of post-traumatic osteoarthritis (PTOA). DESIGN: Three-month-old inducible biglycan (BgniKO) and decorin/biglycan compound (Dcn/BgniKO) knockout mice were subjected to the destabilization of the medial meniscus (DMM) surgery to induce PTOA. The OA phenotype was evaluated by assessing joint structure and sulfated glycosaminoglycan (sGAG) staining via histology, surface collagen fibril nanostructure and calcium content via scanning electron microscopy, tissue modulus via atomic force microscopy-nanoindentation, as well as subchondral bone structure and meniscus ossification via micro-computed tomography. Outcomes were compared with previous findings in the inducible decorin (DcniKO) knockout mice. RESULTS: In the DMM model, BgniKO mice developed similar degree of OA as the control (0.44 [-0.18 1.05] difference in modified Mankin score), different from the more severe OA phenotype observed in DcniKO mice (1.38 [0.91 1.85] difference). Dcn/BgniKO mice exhibited similar histological OA phenotype as DcniKO mice (1.51 [0.97 2.04] difference vs control), including aggravated loss of sGAGs, salient surface fibrillation and formation of osteophyte. Meanwhile, Dcn/BgniKO mice showed further cartilage thinning than DcniKO mice, resulting in the exposure of underlying calcified tissues and aberrantly high surface modulus. BgniKO and Dcn/BgniKO mice developed altered subchondral trabecular bone structure in both Sham and DMM groups, while DcniKO and control mice did not. CONCLUSION: In PTOA, decorin plays a more crucial role than biglycan in regulating cartilage degeneration, while biglycan is more important in regulating subchondral bone structure. The two have distinct activities and modest synergy in the pathogenesis of PTOA.
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Biglicano/deficiencia , Decorina/deficiencia , Progresión de la Enfermedad , Osteoartritis/patología , Animales , Biglicano/genética , Hueso Esponjoso/patología , Cartílago Articular , Decorina/genética , Modelos Animales de Enfermedad , Meniscos Tibiales/patología , Ratones Noqueados , Osificación Heterotópica/patología , Osteoartritis/genética , Osteofito/patología , Lesiones de Menisco Tibial/patologíaRESUMEN
Biglycan (Bgn) and Fibromodulin (Fmod) are small leucine rich proteoglycans (SLRPs) which are abundant in the extra-cellular matrix (ECM) of mineralized tissues. We have previously generated a Bgn/Fmod double knock-out (DKO) mouse model and found it has a 3-fold increase in osteoclastogenesis compared with Wild type (WT) controls, resulting in a markedly low bone mass (LBM) phenotype. To try and rescue/repair the LBM phenotype of Bgn/Fmod DKO mice by suppressing osteoclast formation and activity, 3- and 26-week-old Bgn/Fmod DKO mice and age/gender matched WT controls were treated with OPG-Fc for 6 weeks after which bone parameters were evaluated using DEXA, micro-computed tomography (µCT) and serum biomarkers analyses. In the appendicular skeleton, OPG-Fc treatment improved some morphometric and geometric parameters in both the trabecular and cortical compartments in Bgn/Fmod DKO female and male mice, especially in the repair module. For many of the skeletal parameters analyzed, the Bgn/Fmod DKO mice were more responsive to the treatment than their WT controls. In addition, we found that OPG-Fc treatment was not able to prevent or ameliorate the formation of ectopic ossification, which are common lesions seen in aged joints and are one of the phenotypical hallmarks of our Bgn/Fmod DKO model. Analysis of skull bones, specifically the occipital bone, showed the treatment recovered some parameters of LBM phenotype in the craniofacial skeleton, more so in the younger rescue module. Using OPG-Fc as treatment alleviated, yet did not completely restore, the severe osteopenia and mineralized tissue structural abnormalities that Bgn/Fmod DKO mice suffer from.
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Biglicano/deficiencia , Huesos/efectos de los fármacos , Fibromodulina/deficiencia , Fragmentos Fc de Inmunoglobulinas/farmacología , Osteoprotegerina/farmacología , Proteínas Recombinantes de Fusión/farmacología , Esqueleto/efectos de los fármacos , Animales , Biomarcadores/sangre , Biomarcadores/metabolismo , Huesos/metabolismo , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Osteoclastos/efectos de los fármacos , Osteoclastos/metabolismo , Osteogénesis/efectos de los fármacos , Fenotipo , Esqueleto/metabolismoRESUMEN
BACKGROUND AND AIMS: Vascular biglycan contributes to atherosclerosis development and increased biglycan expression correlates with increased atherosclerosis. However, mice deficient in biglycan have either no reduction in atherosclerosis or an unexpected increase in atherosclerosis. Biglycan deficient mice have systemically elevated TGF-ß, likely due to lack of sequestration of TGF-ß in the extracellular matrix. The purpose of this study was to determine if prevention of TGF-ß elevations in biglycan deficient mice affected atherosclerosis development. METHODS: Biglycan deficient mice were crossed to Ldlr deficient mice. Diabetes was induced via streptozotocin and all mice were fed a high cholesterol diet. Diabetic biglycan wild type and biglycan deficient Ldlr deficient mice were injected with the TGF-ß neutralizing antibody 1D11 or the irrelevant control antibody 13C4. RESULTS: Biglycan deficient mice had significantly elevated plasma TGF-ß levels, which was further increased by diabetes, and significantly increased atherosclerosis. There was a significant correlation between TGF-ß concentrations and atherosclerosis. However, despite nearly complete suppression of plasma TGF-ß levels in mice treated with the TGF-ß neutralizing antibody 1D11, there was no significant difference in atherosclerosis between mice with elevated TGF-ß levels and mice with suppressed TGF-ß levels. CONCLUSIONS: The increased atherosclerosis in biglycan deficient mice does not appear to be due to elevations in TGF-ß.
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Enfermedades de la Aorta/sangre , Aterosclerosis/sangre , Biglicano/deficiencia , Factor de Crecimiento Transformador beta/sangre , Animales , Anticuerpos Neutralizantes/farmacología , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/patología , Aterosclerosis/genética , Aterosclerosis/patología , Biglicano/genética , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/inducido químicamente , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Placa Aterosclerótica , Receptores de LDL/deficiencia , Receptores de LDL/genética , Estreptozocina , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Factor de Crecimiento Transformador beta/inmunología , Regulación hacia ArribaRESUMEN
Animal studies demonstrate that hyperlipidemia and renal lipid accumulation contribute to the pathogenesis of diabetic nephropathy (DN). We previously demonstrated that renal lipoproteins colocalize with biglycan, a renal proteoglycan. The purpose of this study was to determine whether prevention of renal lipid (apoB) accumulation attenuates DN. Biglycan-deficient and biglycan wild-type Ldlr-/- mice were made diabetic via streptozotocin and fed a high cholesterol diet. As biglycan deficiency is associated with elevated transforming growth factor-ß (TGF-ß), in some experiments mice were injected with either the TGF-ß-neutralizing antibody, 1D11, or with 13C4, an irrelevant control antibody. Biglycan deficiency had no significant effect on renal apoB accumulation, but led to modest attenuation of DN with â¼30% reduction in albuminuria; however, biglycan deficiency caused a striking elevation in TGF-ß. Use of 1D11 led to sustained suppression of TGF-ß for approximately 8 weeks at a time. The 1D11 treatment caused decreased renal apoB accumulation, decreased albuminuria, decreased renal hypertrophy, and improved survival, compared with the 13C4 treatment. Thus, prevention of renal apoB accumulation is protective against development of DN. Furthermore, this study demonstrates that prevention of renal apoB accumulation is a mechanism by which TGF-ß inhibition is nephroprotective.
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Albuminuria/tratamiento farmacológico , Apolipoproteínas B/genética , Diabetes Mellitus Experimental/tratamiento farmacológico , Nefropatías Diabéticas/tratamiento farmacológico , Factor de Crecimiento Transformador beta/genética , Albuminuria/genética , Albuminuria/metabolismo , Albuminuria/patología , Animales , Anticuerpos Neutralizantes/farmacología , Apolipoproteína B-100 , Apolipoproteínas B/metabolismo , Biglicano/deficiencia , Biglicano/genética , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Nefropatías Diabéticas/genética , Nefropatías Diabéticas/metabolismo , Nefropatías Diabéticas/patología , Dieta Alta en Grasa/efectos adversos , Riñón/efectos de los fármacos , Riñón/metabolismo , Riñón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de LDL/deficiencia , Receptores de LDL/genética , Estreptozocina , Análisis de Supervivencia , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
AIMS: Biglycan, a small leucine-rich proteoglycan, has been shown to play an important role in stabilizing fibrotic scars after experimental myocardial infarction. However, the role of biglycan in the development and regression of cardiomyocyte hypertrophy and fibrosis during cardiac pressure overload and unloading remains elusive. Thus, the aim of the present study was to assess the effect of biglycan on cardiac remodeling in a mouse model of left ventricular pressure overload and unloading. METHODS AND RESULTS: Left ventricular pressure overload induced by transverse aortic constriction (TAC) in mice resulted in left ventricular dysfunction, fibrosis and increased biglycan expression. Fluorescence- and magnetic-assisted sorting of cardiac cell types revealed upregulation of biglycan in the fibroblast population, but not in cardiomyocytes, endothelial cells or leukocytes after TAC. Removal of the aortic constriction (rTAC) after short-term pressure overload (3weeks) improved cardiac contractility and reversed ventricular hypertrophy but not fibrosis in wild-type (WT) mice. Biglycan ablation (KO) enhanced functional recovery but did not resolve cardiac fibrosis. After long-term TAC for 9weeks, ablation of biglycan attenuated the development of cardiac hypertrophy and fibrosis. In vitro, biglycan induced hypertrophy of neonatal rat cardiomyocytes and led to activation of a hypertrophic gene program. Putative downstream mediators of biglycan signaling include Rcan1, Abra and Tnfrsf12a. These genes were concordantly induced by TAC in WT but not in biglycan KO mice. CONCLUSIONS: Left ventricular pressure overload induces biglycan expression in cardiac fibroblasts. Ablation of biglycan improves cardiac function and attenuates left ventricular hypertrophy and fibrosis after long-term pressure overload. In vitro biglycan induces hypertrophy of cardiomyocytes, suggesting that biglycan may act as a signaling molecule between cell types to modulate cardiac remodeling.
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Biglicano/deficiencia , Biglicano/metabolismo , Cardiomegalia/etiología , Cardiomegalia/metabolismo , Disfunción Ventricular Izquierda/fisiopatología , Animales , Cardiomegalia/diagnóstico , Modelos Animales de Enfermedad , Ecocardiografía , Femenino , Fibrosis , Hipertrofia Ventricular Izquierda/diagnóstico , Hipertrofia Ventricular Izquierda/etiología , Hipertrofia Ventricular Izquierda/metabolismo , Masculino , Ratones , Ratones Noqueados , Miocitos Cardíacos/metabolismo , Proteoma , Proteómica , Ratas , Remodelación VentricularRESUMEN
OBJECTIVE: Thrombin signaling promotes atherosclerosis by initiating inflammatory events indirectly through platelet activation and directly via protease-activated receptors. Therefore, endogenous thrombin inhibitors may be relevant modulators of atheroprogression and cardiovascular risk. In addition, endogenous thrombin inhibitors may affect the response to non-vitamin K-dependent oral anticoagulants. Here, the question was addressed whether the small leucine-rich proteoglycan biglycan acts as an endogenous thrombin inhibitor in atherosclerosis through activation of heparin cofactor II. APPROACH AND RESULTS: Biglycan concentrations were elevated in the plasma of patients with acute coronary syndrome and in male Apolipoprotein E-deficient (ApoE(-/-)) mice. Biglycan was detected in the glycocalyx of capillaries and the subendothelial matrix of arterioles of ApoE(-/-) mice and in atherosclerotic plaques. Thereby a vascular compartment is provided that may mediate the endothelial and subendothelial activation of heparin cofactor II through biglycan. ApoE and Bgn double-deficient (ApoE(-/-)/Bgn(-/0)) mice showed higher activity of circulating thrombin, increased platelet activation and platelet adhesion in vivo, supporting a role of biglycan in balancing thrombin activity. Furthermore, concentrations of circulating cytokines and aortic macrophage content were elevated in ApoE(-/-)/Bgn(-/0) mice, suggesting a proinflammatory phenotype. Elevated platelet activation and macrophage accumulation were reversed by treating ApoE(-/-)/Bgn(-/0) mice with the thrombin inhibitor argatroban. Ultimately, ApoE(-/-)/Bgn(-/0) mice developed aggravated atherosclerosis. CONCLUSIONS: The present results indicate that biglycan plays a previously unappreciated protective role during the progression of atherosclerosis by inhibiting thrombin activity, platelet activation, and finally macrophage-mediated plaque inflammation.
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Aorta/metabolismo , Enfermedades de la Aorta/metabolismo , Apolipoproteínas E/deficiencia , Aterosclerosis/metabolismo , Biglicano/deficiencia , Inflamación/metabolismo , Trombina/metabolismo , Síndrome Coronario Agudo/sangre , Animales , Antitrombinas/farmacología , Aorta/efectos de los fármacos , Aorta/patología , Enfermedades de la Aorta/sangre , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/prevención & control , Apolipoproteínas E/genética , Aterosclerosis/sangre , Aterosclerosis/genética , Aterosclerosis/prevención & control , Biglicano/sangre , Biglicano/genética , Citocinas/sangre , Modelos Animales de Enfermedad , Genotipo , Cofactor II de Heparina/metabolismo , Humanos , Inflamación/sangre , Inflamación/genética , Inflamación/prevención & control , Macrófagos/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Fenotipo , Placa Aterosclerótica , Activación Plaquetaria , Factores de TiempoRESUMEN
Achilles tendons are a common source of pain and injury, and their pathology may originate from aberrant structure function relationships. Small leucine rich proteoglycans (SLRPs) influence mechanical and structural properties in a tendon-specific manner. However, their roles in the Achilles tendon have not been defined. The objective of this study was to evaluate the mechanical and structural differences observed in mouse Achilles tendons lacking class I SLRPs; either decorin or biglycan. In addition, empirical modeling techniques based on mechanical and image-based measures were employed. Achilles tendons from decorin-null (Dcn(-/-)) and biglycan-null (Bgn(-/-)) C57BL/6 female mice (N=102) were used. Each tendon underwent a dynamic mechanical testing protocol including simultaneous polarized light image capture to evaluate both structural and mechanical properties of each Achilles tendon. An empirical damage model was adapted for application to genetic variation and for use with image based structural properties to predict tendon dynamic mechanical properties. We found that Achilles tendons lacking decorin and biglycan had inferior mechanical and structural properties that were age dependent; and that simple empirical models, based on previously described damage models, were predictive of Achilles tendon dynamic modulus in both decorin- and biglycan-null mice.
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Tendón Calcáneo/fisiología , Biglicano/deficiencia , Decorina/deficiencia , Modelos Biológicos , Tendón Calcáneo/química , Animales , Biglicano/análisis , Biglicano/genética , Fenómenos Biomecánicos/fisiología , Colágeno/fisiología , Colágeno/ultraestructura , Decorina/análisis , Decorina/genética , Femenino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Animales , Estrés MecánicoRESUMEN
INTRODUCTION: Biglycan is an important proteoglycan of the extracellular matrix of intervertebral disc (IVD), and its decrease with aging has been correlated with IVD degeneration. Biglycan deficient (Bgn-/0) mice lack this protein and undergo spontaneous IVD degeneration with aging, thus representing a valuable in vivo model for preliminary studies on therapies for human progressive IVD degeneration. The purpose of the present study was to assess the possible beneficial effects of adipose-derived stromal cells (ADSCs) implants in the Bgn-/0 mouse model. METHODS: To evaluate ADSC implant efficacy, Bgn-/0 mice were intradiscally (L1-L2) injected with 8x104 ADSCs at 16 months old, when mice exhibit severe and complete IVD degeneration, evident on both 7Tesla Magnetic Resonance Imaging (7TMRI) and histology. Placebo and ADSCs treated Bgn-/0 mice were assessed by 7TMRI analysis up to 12 weeks post-transplantation. Mice were then sacrificed and implanted discs were analyzed by histology and immunohistochemistry for the presence of human cells and for the expression of biglycan and aggrecan in the IVD area. RESULTS: After in vivo treatment, 7TMRI revealed evident increase in signal intensity within the discs of mice that received ADSCs, while placebo treatment did not show any variation. Ultrastructural analyses demonstrated that human ADSC survival occurred in the injected discs up to 12 weeks after implant. These cells acquired a positive expression for biglycan, and this proteoglycan was specifically localized in human cells. Moreover, ADSC treatment resulted in a significant increase of aggrecan tissue levels. CONCLUSION: Overall, this work demonstrates that ADSC implant into degenerated disc of Bgn-/0 mice ameliorates disc damage, promotes new expression of biglycan and increased levels of aggrecan. This suggests a potential benefit of ADSC implant in the treatment of chronic degenerative disc disease and prompts further studies in this field.
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Degeneración del Disco Intervertebral/fisiopatología , Degeneración del Disco Intervertebral/terapia , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/citología , Regeneración , Tejido Adiposo/citología , Agrecanos/biosíntesis , Animales , Biglicano/biosíntesis , Biglicano/deficiencia , Biglicano/genética , Células Cultivadas , Enfermedad Crónica , Modelos Animales de Enfermedad , Humanos , Inmunohistoquímica , Disco Intervertebral/diagnóstico por imagen , Disco Intervertebral/fisiopatología , Degeneración del Disco Intervertebral/genética , Imagen por Resonancia Magnética , Masculino , Células Madre Mesenquimatosas/metabolismo , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Radiografía , Trasplante HeterólogoRESUMEN
Proteoglycans of the arterial wall play a critical role in vascular integrity and the development of atherosclerosis owing to their ability to organize extracellular matrix molecules and to bind and retain atherogenic apolipoprotein (apo)-B containing lipoproteins. Prior studies have suggested a role for biglycan in aneurysms and in atherosclerosis. Angiotensin II (angII) infusions into mice have been shown to induce abdominal aortic aneurysm development, increase vascular biglycan content, increase arterial retention of lipoproteins, and accelerate atherosclerosis. The goal of this study was to determine the role of biglycan in angII-induced vascular diseases. Biglycan-deficient or biglycan wildtype mice crossed to LDL receptor deficient (Ldlr-/-) mice (C57BL/6 background) were infused with angII (500 or 1000ng/kg/min) or saline for 28days while fed on normal chow, then pumps were removed, and mice were switched to an atherogenic Western diet for 6weeks. During angII infusions, biglycan-deficient mice developed abdominal aortic aneurysms, unusual descending thoracic aneurysms, and a striking mortality caused by aortic rupture (76% for males and 48% for females at angII 1000ng/kg/min). Histological analyses of non-aneurysmal aortic segments from biglycan-deficient mice revealed a deficiency of dense collagen fibers and the aneurysms demonstrated conspicuous elastin breaks. AngII infusion increased subsequent atherosclerotic lesion development in both biglycan-deficient and biglycan wildtype mice. However, the biglycan genotype did not affect the atherosclerotic lesion area induced by the Western diet after treatment with angII. Biglycan-deficient mice exhibited significantly increased vascular perlecan content compared to biglycan wildtype mice. Analyses of the atherosclerotic lesions demonstrated that vascular perlecan co-localized with apoB, suggesting that increased perlecan compensated for biglycan deficiency in terms of lipoprotein retention. Biglycan deficiency increases aortic aneurysm development and is not protective against the development of atherosclerosis. Biglycan deficiency leads to loosely packed aortic collagen fibers, increased susceptibility of aortic elastin fibers to angII-induced stress, and up-regulation of vascular perlecan content.
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Aneurisma de la Aorta/metabolismo , Aterosclerosis/patología , Biglicano/deficiencia , Angiotensina II/farmacología , Animales , Aorta/metabolismo , Aorta/patología , Aneurisma de la Aorta/patología , Aterosclerosis/metabolismo , Biglicano/metabolismo , Dieta Occidental , Femenino , Genotipo , Proteoglicanos de Heparán Sulfato/metabolismo , Masculino , Ratones Endogámicos C57BL , Rotura , Análisis de SupervivenciaRESUMEN
The corneal stroma is enriched in small leucine-rich proteoglycans (SLRPs), including both class I (decorin and biglycan) and class II (lumican, keratocan and fibromodulin). Transparency is dependent on the assembly and maintenance of a hierarchical stromal organization and SLRPs are critical regulatory molecules. We hypothesize that cooperative interclass SLRP interactions are involved in the regulation of stromal matrix assembly. We test this hypothesis using a compound Bgn(-/0)/Lum(-/-) mouse model and single Lum(-/-) or Bgn(-/0) mouse models and wild type controls. SLRP expression was investigated using immuno-localization and immuno-blots. Structural relationships were defined using ultrastructural and morphometric approaches while transparency was analyzed using in vivo confocal microscopy. The compound Bgn(-/0)/Lum(-/-) corneas demonstrated gross opacity that was not seen in the Bgn(-/0) or wild type corneas and greater than that in the Lum(-/-) mice. The Bgn(-/0)/Lum(-/-) corneas exhibited significantly increased opacity throughout the stroma compared to posterior opacity in the Lum(-/-) and no opacity in Bgn(-/0) or wild type corneas. In the Bgn(-/0)/Lum(-/-) corneas there were abnormal lamellar and fibril structures consistent with the functional deficit in transparency. Lamellar structure was disrupted across the stroma with disorganized fibrils, and altered fibril packing. In addition, fibrils had larger and more heterogeneous diameters with an abnormal structure consistent with abnormal fibril growth. This was not observed in the Bgn(-/0) or wild type corneas and was restricted to the posterior stroma in Lum(-/-) mice. The data demonstrate synergistic interclass regulatory interactions between lumican and biglycan. These interactions are involved in regulating both lamellar structure as well as collagen fibrillogenesis and therefore, corneal transparency.
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Biglicano/metabolismo , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Colágeno/biosíntesis , Opacidad de la Córnea/metabolismo , Sustancia Propia/crecimiento & desarrollo , Sulfato de Queratano/metabolismo , Proteínas/metabolismo , Animales , Biglicano/deficiencia , Proteoglicanos Tipo Condroitín Sulfato/deficiencia , Sustancia Propia/metabolismo , Sustancia Propia/patología , Immunoblotting , Sulfato de Queratano/deficiencia , Proteínas Repetidas Ricas en Leucina , Lumican , Ratones , Ratones Noqueados , Microscopía Confocal , Seudópodos/fisiologíaRESUMEN
Tendons like the flexor carpi ulnaris (FCU) that contain region-specific distributions of proteoglycans (PGs) as a result of the heterogeneous, multi-axial loads they are subjected to in vivo provide valuable models for understanding structure-function relationships in connective tissues. However, the contributions of specific PGs to FCU tendon mechanical properties are unknown. Therefore, the objective of this study was to determine how the location-dependent, viscoelastic mechanical properties of the FCU tendon are impacted individually by PG-associated glycosaminoglycans (GAGs) and by two small leucine-rich proteoglycans (SLRPs), biglycan and decorin. Full length FCU tendons from biglycan- and decorin-null mice were compared to wild-type (WT) mice to evaluate the effects of specific SLRPs, while chondroitinase ABC digestion of isolated specimens removed from the tendon midsubstance was used to determine how chondroitin/dermatan sulfate (CS/DS) GAGs impact mechanics in mature FCU tendons. A novel combined genetic knockout/digestion technique also was employed to compare SLRP-null and WT tendons in the absence of CS/DS GAGs that may impact properties in the mature state. In all genotypes, mechanical properties in the FCU tendon midsubstance were not affected by GAG digestion. Full-length tendons exhibited complex, multi-axial deformation under tension that may be associated with their in vivo loading environment. Mechanical properties were adversely affected by the absence of biglycan, and a decreased modulus localized in the center of the tendon was measured. These results help elucidate the role that local alterations in PG levels may play in processes that adversely impact tendon functionality including injury and pathology.
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Proteoglicanos/metabolismo , Tendones/metabolismo , Animales , Biglicano/deficiencia , Biglicano/metabolismo , Fenómenos Biomecánicos , Sulfatos de Condroitina/metabolismo , Dermatán Sulfato/metabolismo , Módulo de Elasticidad , Femenino , Ratones , Ratones Endogámicos C57BL , Estrés MecánicoRESUMEN
Tendons have complex mechanical properties that depend on their structure and composition. Some studies have assessed the role of small leucine-rich proteoglycans (SLRPs) in the mechanical response of tendon, but the relationships between sophisticated mechanics, assembly of collagen and SLRPs have not been well characterized. In this study, biglycan gene expression was varied in a dose dependent manner using biglycan null, biglycan heterozygote and wild type mice. Measures of mechanical (tension and compression), compositional and structural changes of the mouse patellar tendon were evaluated. Viscoelastic, tensile dynamic modulus was found to be increased in the biglycan heterozygous and biglycan null tendons compared to wild type. Gene expression analyses revealed biglycan gene expression was closely associated in a dose-dependent allelic manner. No differences were seen between genotypes in elastic or compressive properties or quantitative measures of collagen structure. These results suggest that biglycan, a member of the SLRP family, plays a role in tendon viscoelasticity that cannot be completely explained by its role in collagen fibrillogenesis.
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Biglicano/genética , Elasticidad , Expresión Génica , Ligamento Rotuliano/patología , Ligamento Rotuliano/fisiopatología , Animales , Biglicano/deficiencia , Biglicano/metabolismo , Fenómenos Biomecánicos , Colágeno , Modelos Animales de Enfermedad , Módulo de Elasticidad , Femenino , Perfilación de la Expresión Génica , Genotipo , Heterocigoto , Ratones , Ratones Noqueados , Ligamento Rotuliano/metabolismo , Reología/métodos , Estrés Mecánico , Resistencia a la Tracción , ViscosidadRESUMEN
Collagen fiber realignment is one mechanism by which tendon responds to load. Re-alignment is altered when the structure of tendon is altered, such as in the natural process of aging or with alterations of matrix proteins, such as proteoglycan expression. While changes in re-alignment and mechanical properties have been investigated recently during development, they have not been studied in (1) aged tendons, or (2) in the absence of key proteoglycans. Collagen fiber re-alignment and the corresponding mechanical properties are quantified throughout tensile mechanical testing in both the insertion site and the midsubstance of mouse supraspinatus tendons in wild type (WT), decorin-null (Dcn(-/-)), and biglycan-null (Bgn(-/-)) mice at three different ages (90 days, 300 days, and 570 days). Percent relaxation was significantly decreased with age in the WT and Dcn(-/-) tendons, but not in the Bgn(-/-) tendons. Changes with age were found in the linear modulus at the insertion site where the 300 day group was greater than the 90 day and 570 day group in the Bgn(-/-) tendons and the 90 day group was smaller than the 300 day and 570 day groups in the Dcn(-/-) tendons. However, no changes in modulus were found across age in WT tendons were found. The midsubstance fibers of the WT and Bgn(-/-) tendons were initially less aligned with increasing age. The re-alignment was significantly altered with age in the WT tendons, with older groups responding to load later in the mechanical test. This was also seen in the Dcn(-/-) midsubstance and the Bgn(-/-) insertion, but not in the other locations. Although some studies have found changes in the WT mechanical properties with age, this study did not support those findings. However, it did show fiber re-alignment changes at both locations with age, suggesting a breakdown of tendon's ability to respond to load in later ages. In the proteoglycan-null tendons however, there were changes in the mechanical properties, accompanied only by location-dependent re-alignment changes, suggesting a site-specific role for these molecules in loading. Finally, changes in the mechanical properties did not occur in concert with changes in re-alignment, suggesting that typical mechanical property measurements alone are insufficient to describe how structural alterations affect tendon's response to load.
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Envejecimiento , Colágeno/metabolismo , Fenómenos Mecánicos , Proteoglicanos/deficiencia , Manguito de los Rotadores , Tendones/metabolismo , Animales , Biglicano/deficiencia , Biglicano/genética , Fenómenos Biomecánicos , Colágeno/química , Decorina/deficiencia , Decorina/genética , Eliminación de Gen , Ensayo de Materiales , Ratones , Proteoglicanos/genética , Tendones/fisiologíaRESUMEN
Biglycan (BGN), a small leucine-rich proteoglycan, binds the pro-fibrotic cytokine transforming growth factor ß (TGFß) and inhibits its bioactivity in vitro. Nevertheless, it is controversial whether BGN plays an inhibitory role in vivo. Therefore, the purpose of this study was to evaluate the effect of BGN deficiency on TGFß activity in vivo by studying 1-year-old Bgn null and wild-type (WT) mice on an Ldlr-null background. Phenotypic and metabolic characterization showed that the Bgn null mice had lower body weight, shorter body length, and shorter femur length (all p < 0.05). Surprisingly, the Bgn null mice also exhibited a striking reduction in percent body fat compared to WT mice (p == 0.006), but no changes were observed in plasma triglycerides, total cholesterol, or glycohemoglobin. Both total and bioactive TGFß1 concentrations in plasma were markedly elevated in Bgn null mice compared to WT mice (4-fold and 11-fold increase, respectively, both p < 0.001), but no changes were found in hepatic levels of mRNA for Tgfß1 or its receptors. Bgn null mice exhibited elevated expression of hepatic fibronectin protein (p = 0.034) without changes in hepatic or renal histology, and Bgn null mice had decreased urinary albumin/creatinine ratio (p = 0.01). Two key downstream targets of bone morphogenetic protein 4-like signaling, SMAD1/3/5 phosphorylation and Id2 gene expression, were found dramatically reduced in Bgn null livers (p = 0.034). Thus, BGN deficiency decreases body fat in this hyperlipidemic mouse model without changing liver or kidney histology. Overall, we propose that this unexpected phenotype arises from the effects of BGN deficiency in vivo to elevate TGFß levels while decreasing bone morphogenetic protein 4-like signaling.
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Tejido Adiposo/patología , Adiposidad/fisiología , Biglicano/deficiencia , Proteína Morfogenética Ósea 4/metabolismo , Hiperlipidemias/patología , Factor de Crecimiento Transformador beta/sangre , Animales , Composición Corporal , Peso Corporal/fisiología , Femenino , Regulación de la Expresión Génica , Hiperlipidemias/sangre , Hiperlipidemias/genética , Proteína 2 Inhibidora de la Diferenciación/genética , Proteína 2 Inhibidora de la Diferenciación/metabolismo , Riñón/anatomía & histología , Riñón/metabolismo , Pruebas de Función Renal , Hígado/anatomía & histología , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Tamaño de los Órganos , Fosforilación , ARN Mensajero/metabolismo , Transducción de Señal , Proteínas Smad/genética , Proteínas Smad/metabolismo , Factor de Crecimiento Transformador beta/genéticaRESUMEN
Cesarean birth rates are rising. Uterine dysfunction, the exact mechanism of which is unknown, is a common indication for Cesarean delivery. Biglycan and decorin are two small leucine-rich proteoglycans expressed in the extracellular matrix of reproductive tissues and muscle. Mice deficient in biglycan display a mild muscular dystrophy, and, along with mice deficient in decorin, are models of Ehlers-Danlos Syndrome, a connective tissue anomaly associated with uterine rupture. As a variant of Ehlers-Danlos Syndrome is caused by a genetic mutation resulting in abnormal biglycan and decorin secretion, we hypothesized that biglycan and decorin play a role in uterine function. Thus, we assessed wild-type, biglycan, decorin and double knockout pregnancies for timing of birth and uterine function. Uteri were harvested at embryonic days 12, 15 and 18. Nonpregnant uterine samples of the same genotypes were assessed for tissue failure rate and spontaneous and oxytocin-induced contractility. We discovered that biglycan/decorin mixed double-knockout dams displayed dystocia, were at increased risk of delayed labor onset, and showed increased tissue failure in a predominantly decorin-dependent manner. In vitro spontaneous uterine contractile amplitude and oxytocin-induced contractile force were decreased in all biglycan and decorin knockout genotypes compared to wild-type. Notably, we found no significant compensation between biglycan and decorin using quantitative real time PCR or immunohistochemistry. We conclude that the biglycan/decorin mixed double knockout mouse is a model of dystocia and delayed labor onset. Moreover, decorin is necessary for uterine function in a dose-dependent manner, while biglycan exhibits partial compensatory mechanisms in vivo. Thus, this model is poised for use as a model for testing novel targets for preventive or therapeutic manipulation of uterine dysfunction.
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Biglicano/deficiencia , Decorina/deficiencia , Distocia/fisiopatología , Parto/fisiología , Útero/fisiopatología , Alelos , Animales , Biglicano/metabolismo , Fenómenos Biomecánicos/efectos de los fármacos , Decorina/metabolismo , Distocia/patología , Femenino , Masculino , Ratones , Ratones Endogámicos C3H , Ratones Noqueados , Oxitocina/farmacología , Parto/efectos de los fármacos , Embarazo , Factor de Crecimiento Transformador beta/metabolismo , Contracción Uterina/efectos de los fármacos , Contracción Uterina/fisiología , Útero/efectos de los fármacos , Útero/patologíaRESUMEN
Although extracellular control of canonical Wnt signaling is crucial for tissue homeostasis, the role of the extracellular microenvironment in modulating this signaling pathway is largely unknown. In the present study, we show that a member of the small leucine-rich proteoglycan family, biglycan, enhances canonical Wnt signaling by mediating Wnt function via its core protein. Immunoprecipitation analysis revealed that biglycan interacts with both the canonical Wnt ligand Wnt3a and the Wnt coreceptor low-density lipoprotein receptor-related protein 6 (LRP6), possibly via the formation of a trimeric complex. Biglycan-deficient cells treated with exogenous Wnt3a had less Wnt3a retained in cell layers compared with WT cells. Furthermore, the Wnt-induced levels of LRP6 phosphorylation and expression of several Wnt target genes were blunted in biglycan-deficient cells. Both recombinant biglycan proteoglycan and biglycan core protein increased Wnt-induced ß-catenin/T cell-specific factor-mediated transcriptional activity, and this activity was completely inhibited by Dickkopf 1. Interestingly, recombinant biglycan was able to rescue impaired Wnt signaling caused by a previously described missense mutation in the extracellular domain of human LRP6 (R611C). Furthermore, biglycan's modulation of canonical Wnt signaling affected the functional activities of osteoprogenitor cells, including the RUNX2-mediated transcriptional activity and calcium deposition. Use of a transplant system and a fracture healing model revealed that expression of Wnt-induced secreted protein 1 was decreased in bone formed by biglycan-deficient cells, further suggesting reduced Wnt signaling in vivo. We propose that biglycan may serve as a reservoir for Wnt in the pericellular space and modulate Wnt availability for activation of the canonical Wnt pathway.
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Biglicano/metabolismo , Vía de Señalización Wnt/fisiología , Animales , Biglicano/deficiencia , Biglicano/genética , Matriz Extracelular/metabolismo , Células HEK293 , Humanos , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/química , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/genética , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/metabolismo , Ratones , Ratones Noqueados , Mutación Missense , Unión Proteica , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Cráneo/metabolismo , Proteína Wnt3A/genética , Proteína Wnt3A/metabolismo , beta Catenina/metabolismoRESUMEN
Temporomandibular joint osteoarthritis (TMJ OA) is a degenerative disease that affects both cartilage and subchondral bone. We used microarray to identify changes in gene expression levels in the TMJ during early stages of the disease, using an established TMJ OA genetic mouse model deficient in 2 extracellular matrix proteins, biglycan and fibromodulin (bgn(-/0)fmod(-/-)). Differential gene expression analysis was performed with RNA extracted from 3-week-old WT and bgn(-/0)fmod(-/-) TMJs with an intact cartilage/subchondral bone interface. In total, 22 genes were differentially expressed in bgn(-/0)fmod(-/-) TMJs, including 5 genes involved in osteoclast activity/differentiation. The number of TRAP-positive cells were three-fold higher in bgn(-/0)fmod(-/-) TMJs than in WT. Quantitative RT-PCR showed up-regulation of RANKL and OPG, with a 128% increase in RANKL/OPG ratio in bgn(-/0)fmod(-/-) TMJs. Histology and immunohistochemistry revealed tissue disorganization and reduced type I collagen in bgn(-/0)fmod(-/-) TMJ subchondral bone. Early changes in gene expression and tissue defects in young bgn(-/0)fmod(-/-) TMJ subchondral bone are likely attributed to increased osteoclast activity. Analysis of these data shows that biglycan and fibromodulin are critical for TMJ subchondral bone integrity and reveal a potential role for TMJ subchondral bone turnover during the initial early stages of TMJ OA disease in this model.
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Biglicano/fisiología , Remodelación Ósea/fisiología , Huesos/fisiopatología , Cartílago Articular/fisiopatología , Proteínas de la Matriz Extracelular/fisiología , Osteoartritis/metabolismo , Proteoglicanos/fisiología , Trastornos de la Articulación Temporomandibular/metabolismo , Animales , Artritis Experimental/metabolismo , Artritis Experimental/fisiopatología , Biglicano/deficiencia , Huesos/metabolismo , Cartílago Articular/metabolismo , Modelos Animales de Enfermedad , Proteínas de la Matriz Extracelular/deficiencia , Fibromodulina , Perfilación de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Análisis de Secuencia por Matrices de Oligonucleótidos , Osteoartritis/fisiopatología , Osteoclastos/metabolismo , Osteoprotegerina/biosíntesis , Osteoprotegerina/genética , Proteoglicanos/deficiencia , Ligando RANK/biosíntesis , Ligando RANK/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Trastornos de la Articulación Temporomandibular/fisiopatologíaRESUMEN
CXCL13 is a key B cell chemoattractant and marker of disease activity in patients with SLE; however, the mechanism of its induction has not been identified yet. Here, we have shown that the proteoglycan biglycan triggers CXCL13 expression via TLR2/4 in macrophages and dendritic cells. In vivo, levels of biglycan were markedly elevated in the plasma and kidneys of human SLE patients and lupus-prone (MRL/lpr) mice. Overexpression of soluble biglycan in MRL/lpr mice raised plasma and renal levels of CXCL13 and caused accumulation of B cells with an enhanced B1/B cell ratio in the kidney, worsening of organ damage, and albuminuria. Importantly, biglycan also triggered CXCL13 expression and B cell infiltration in the healthy kidney. Conversely, biglycan deficiency improved systemic and renal outcome in lupus-prone mice, with lower levels of autoantibodies, less enlargement of the spleen and lymph nodes, and reduction in renal damage and albuminuria. This correlated with a marked decline in circulating and renal CXCL13 and a reduction in the number of B cells in the kidney. Collectively, our results describe what we believe to be a novel mechanism for the regulation of CXCL13 by biglycan, a host-derived ligand for TLR2/4. Blocking biglycan-TLR2/4 interactions might be a promising strategy for the management of SLE and other B cell-mediated inflammatory disease entities.