RESUMO
Given the wide spread clinical use of ceramic-based bone void fillers, we sought to determine the efficacy of an FDA-approved ß-tricalcium phosphate bone graft substitute (JAX™) in combination with a carboxymethyl cellulose (CMC) handling agent that included a particular heparan glycosaminoglycan (GAG) variant, herein referred to as HS3. Having recently demonstrated efficacy of a combination collagen/HS3 device, we further aimed to determine the support that HS3 could offer a handling agent used to administer a more tissue-relevant bone void filler. This study evaluated the JAX™-HS3 combination device in 1.5 cm critical-sized defects in the ulna bones of 27 male New Zealand White rabbits. Treatment groups consisted of JAX™ applied with CMC alone, or JAX™ with CMC containing either 30 µg or 100 µg of the HS3 GAG. Data based on radiographic, µCT, mechanical, and histological analyses at 4 and 8 weeks post-surgery, clearly demonstrate enhanced new bone formation in the JAX™-HS3 combination treated defects compared to treatment with JAX™ alone. The efficacy of such a combination advocates for inclusion of HS3 in handling agents used in the preparation of various bone void fillers being used in orthopaedic surgery. STATEMENT OF SIGNIFICANCE: Synthetic bone grafts and demineralized bone matrices are gaining prominence as alternatives to autologous and allogeneic bone grafts and are frequently administered in granular form, necessitating their combination with a handling agent. Typical handling agents include glycerol, gelatin, cellulose, hyaluronic acid and lecithin, formulated as hydrogels, which can be further enhanced by the addition of heparan sulfate (HS) glycosaminoglycans that augment the osteostimulatory properties of the graft. Here we assessed the efficacy of ß-TCP granules combined with a hydrogel consisting of carboxymethyl cellulose and the HS variant (HS3) previously shown to enhance osteogenic healing. The data advocates for HS3 to be included during the formulation of hydrogel-based carriers that support the various bone void fillers being used in orthopaedic surgery.
Assuntos
Fosfatos de Cálcio/administração & dosagem , Glicosaminoglicanos/administração & dosagem , Heparitina Sulfato/administração & dosagem , Próteses e Implantes , Ulna/anormalidades , Animais , Masculino , Camundongos , Microtomografia por Raio-XRESUMO
Wear debris generation in total joint replacement remains a concern because of its association with aseptic loosening, osteolysis, and ultimately implant failure. In a quest to develop new implant materials with reduced wear and improved biocompatibility a new composition of oxidized Zr2.5Nb alloy; diffusion hardened oxidized zirconium (DHOxZr) has been developed. In this study, we have determined the in vitro biocompatibility of the wear debris of this new composition and compared it to wear debris particles of Ti6Al4V, Cobalt, and CoCr. The cytotoxicity of these particles on fibroblast-like cells (L929) and osteoblast-like cells (SaOS2) was assessed using lactate dehydrogenase and DNA quantification assays. The inflammatory response to these particles was assessed by release of interleukin-1 beta and tumor necrosis factor from macrophage-like cells. The results showed that wear debris generated from DHOxZr was less cytotoxic and elicited a reduced inflammatory response as compared to that of Cobalt or CoCr and might therefore offer a benefit as a joint prosthesis.
Assuntos
Fibroblastos/metabolismo , Dureza , Macrófagos/metabolismo , Teste de Materiais , Próteses e Implantes/efeitos adversos , Zircônio/química , Animais , Artroplastia de Quadril , Linhagem Celular , Sobrevivência Celular , Ligas de Cromo/efeitos adversos , Ligas de Cromo/química , Fibroblastos/patologia , Humanos , Inflamação/metabolismo , Inflamação/patologia , Interleucina-1beta/metabolismo , Macrófagos/patologia , Camundongos , Oxirredução , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Chronic tendon lesions are degenerative conditions and may represent a failure to repair or remodel the extracellular matrix after repeated micro-injury. Since TGF-beta is strongly associated with tissue repair, we investigated the expression of TGF-beta isoforms (beta1, beta2 and beta3) and their 2 signalling receptors (TGF-betaRI and TGF-betaRII) in normal and pathological Achilles tendons. In all tissues, all 3 TGF-beta isoforms and the 2 receptors were present at sites of blood vessels. Cells in the matrix showed no staining for TGF-beta1 or beta3, while TGF-beta2 was associated with cells throughout the normal cadaver tendon. Tissue from tendons with pathological lesions showed an increase in cell numbers and percentage TGF-beta2 expression. TGF-betaRII showed a wide distribution in cells throughout the tissue sections. As with TGF-beta2, there was an increase in the number of cells expressing TGF-betaRII in pathological tissue. TGF-betaRI was restricted to blood vessels and was absent from the fibrillar matrix. We conclude that despite the presence and upregulation of TGF-beta2, TGF-beta signalling is not propagated due to the lack of TGF-betaRI. This might explain why chronic tendon lesions fail to resolve and suggests that the addition of exogenous TGF-beta will have little effect on chronic tendinopathy.
Assuntos
Tendão do Calcâneo/metabolismo , Doenças Musculares/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Tendão do Calcâneo/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Doença Crônica , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Doenças Musculares/patologia , Isoformas de Proteínas/metabolismoRESUMO
Total collagen assays are often laborious and use large quantities of consumables. We have developed a new method of assaying total 3H-proline-labeled collagen from cultured cells. Cells and media are harvested from 96-well plates directly onto fiberglass filtermats and counted in the Wallac 1205 flat-bed scintillation counter (BetaPlate). The assay was validated by comparison with a traditional total collagen assay. The resulting assay provides a rapid one-step method for quantifying collagen synthesis, which, unlike many collagen assays, does not require extensive dialysis or precipitation of proteins.
Assuntos
Colágeno/análise , Contagem de Cintilação , Radioisótopos de Carbono , Células Cultivadas , Colágeno/biossíntese , Colagenases/metabolismo , Filtração/instrumentação , Hidroxiprolina/análise , Marcação por Isótopo , Prolina/metabolismo , Contagem de Cintilação/instrumentação , TrítioRESUMO
OBJECTIVE: To determine whether human osteoarthritic (OA) cartilage loses its ability to remain avascular when placed into the in-vivo model of angiogenesis, the chick embryo chorio-allantoic membrane (CAM), and to determine specific changes that occur in the cartilage matrix when the cartilage is exposed to an active vasculature. DESIGN: Articular cartilage from OA and non-OA joints was grafted onto the CAM for up to 5 days before fixing and processing for histological, histochemical and immunological examination for specific changes in proteoglycan and collagen. RESULTS: OA cartilage, but not non-OA cartilage, showed invasion of its matrix by blood vessels from the CAM to various extents. Associated with these blood vessels was a loss of staining for proteoglycans and cartilage specific glycosaminoglycans (GAG). A deposition of collagen types I and X was also visualized around the invasive vessels. CONCLUSIONS: OA cartilage loses or has already lost its ability to remain avascular when placed onto the chick CAM. Changes occur in the matrix around the invasive blood vessels, specifically a loss of proteoglycan and GAG, and the deposition of new collagen types, notably I and X.
Assuntos
Cartilagem Articular/irrigação sanguínea , Neovascularização Patológica/patologia , Osteoartrite do Joelho/patologia , Idoso , Animais , Cartilagem Articular/metabolismo , Cartilagem Articular/transplante , Embrião de Galinha , Colágeno/metabolismo , Feminino , Glicosaminoglicanos/metabolismo , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Neovascularização Patológica/metabolismo , Osteoartrite do Joelho/metabolismo , Proteoglicanas/metabolismoRESUMO
The vascularization of developing cartilage rudiments is temporally and spatially defined. By using an in vivo angiogenesis model, the chorioallantoic membrane (CAM) of the chick embryo and chick embryo cartilage rudiments, we conclude that the factors controlling the vascular invasion of cartilage rudiments are intrinsic. Intact rudiments, separate hypertrophic zones and separate rounded cell zones, when grafted onto the CAM, become vascularized in the same temporal and spatial manner as occurs in ovo. When grown as organ cultures prior to CAM grafting, rudiments still become vascularized in the same temporal and spatial manner. The integrity of the extracellular matrix and the presence of the periosteum are two physical factors regulating the control of vascularization. Removal of the periosteum from hypertrophic regions caused a cessation of the invasion. Insults to the matrix via brief enzymatic degradation of extracellular matrix components resulted in invasion and erosion of rounded cell zones at an earlier time than is ordinarily seen both in ovo and on the CAM.