The cytocompatibility and early osteogenic characteristics of an injectable calcium phosphate cement.

In this study, the cytocompatibility and early osteogenic characteristics of rat bone marrow cells (RBMCs) on injectable calcium phosphate (CaP) cement (Calcibon) were investigated. In addition to unmodified CaP cement discs, 2 other treatments were given to the discs: preincubation in MilliQ and sintering at different temperatures. After primary culture, RBMCs were dropwise seeded on the discs and cultured for 12 days. The samples were evaluated in terms of cell viability, morphology (live and dead assays and scanning electron microscopy (SEM)), cell proliferation (deoxyribonucleic acid (DNA) analyses), early cell differentiation (alkaline phosphatase (ALP) activity), and physicochemical analyses (xray diffraction (XRD)). The live and dead, DNA, and SEM results showed that Calcibon discs without any additional treatment were not supporting osteoblast-like cells in vitro. There were fewer cells, and cell layers were detached from the disc surface. Therefore, different preincubation periods and sintering temperatures were evaluated to improve the cytocompatibility of the CaP cement. Preincubating discs in MilliQ for periods of 1, 4, 8, and 12 weeks resulted in the hydrolysis of a-tri calcium phosphate (TCP) into an apatite-like structure with some b-TCP, as shown with XRD, but the material was not cytocompatible. Sintering the discs between 8008C and 11008C resulted in conversion of a-TCP to b-TCP with some hydroxyapatite and an increase in crystallinity. Eventually, the discs sintered at 11008C achieved better cell attachment, more-abundant cell proliferation, and earlier differentiation than other sintered (6008C, 8008C, and 10008C), preincubated, and unmodified specimens. On basis of our results, we conclude that in vivo results with CaP-based cements do not guarantee in vitro applicability. Furthermore, unmodified Calcibon is not cytocompatible in vitro, although preincubation of the material results in a more-favorable cell response, sintering of the material at 11008C results in the best osteogenic properties. In contrast to in vivo studies, the Calcibon CaP cement is not suitable as a scaffold for cellbased tissue-engineering strategies

Bone regenerative properties of injectable PGLA-CaP composite with TGF-beta1 in a rat augmentation model.

The aim of this study was to examine the bone augmentation properties of an injectable composite consisting of PLGA microspheres/CaP cement (20/80), and the additional effect of loading PLGA microspheres with TGF-β1 (200 ng). For this purpose, PLGA/CaP composites (control) and PLGA/CaP composites loaded with TGF-β1 (test group) were injected on top of the skulls of 24 Wistar rats. Each rat received 2 materials from the same experimental group, and in total 48 implants were placed (n = 8). After 2, 4, and 8 weeks the results were evaluated histologically and histomorphometrically. The contact length between the implants and newly formed bone increased in time, and was significantly higher for the TGF-β1-loaded composites after 2 weeks. Also, bone formation was significantly higher for the TGF-β1-loaded composites (18.5% ± 3) compared to controls (7.21% ± 5) after 8 weeks of implantation. Immunohistochemical staining demonstrated massive inflammatory infiltrates in both groups, particularly at 2 weeks, which decreased substantially at 4 and 8 weeks. In conclusion, injectable PLGA/CaP composites stimulated bone augmentation in a rat model. The addition of TGF-β1 to the composite significantly increased bone contact at 2 weeks and enhanced new bone formation at 8 weeks

INFLUENCE OF SURFACE MICROSTRUCTURE AND CHEMISTRY ON OSTEOINDUCTION AND OSTEOCLASTOGENESIS BY BIPHASIC CALCIUM PHOSPHATE DISCS

It has been reported that surface microstructural dimensions can influence the osteoinductivity of calcium phosphates (CaPs), and osteoclasts may play a role in this process. We hypothesised that surface structural dimensions of </= 1 mum trigger osteoinduction and osteoclast formation irrespective of macrostructure (e.g., concavities, interconnected macropores, interparticle space) or surface chemistry. To test this, planar discs made of biphasic calcium phosphate (BCP: 80 % hydroxyapatite, 20 % tricalcium phosphate) were prepared with different surface structural dimensions - either ~ 1 mum (BCP1150) or ~ 2-4 mum (BCP1300) - and no macropores or concavities. A third material was made by sputter coating BCP1150 with titanium (BCP1150Ti), thereby changing its surface chemistry but preserving its surface structure and chemical reactivity. After intramuscular implantation in 5 dogs for 12 weeks, BCP1150 formed ectopic bone in 4 out of 5 samples, BCP1150Ti formed ectopic bone in 3 out of 5 samples, and BCP1300 formed no ectopic bone in any of the 5 samples. In vivo, large multinucleated osteoclast-like cells densely colonised BCP1150, smaller osteoclast-like cells formed on BCP1150Ti, and osteoclast-like cells scarcely formed on BCP1300. In vitro, RAW264.7 cells cultured on the surface of BCP1150 and BCP1150Ti in the presence of osteoclast differentiation factor RANKL (receptor activator for NF-kappaB ligand) proliferated then differentiated into multinucleated osteoclast-like cells with positive tartrate resistant acid phosphatase (TRAP) activity. However, cell proliferation, fusion, and TRAP activity were all significantly inhibited on BCP1300. These results indicate that of the material parameters tested - namely, surface microstructure, macrostructure, and surface chemistry - microstructural dimensions are critical in promoting osteoclastogenesis and triggering ectopic bone formation

Biomaterial Property Effects on Platelets and Macrophages: An in Vitro Study

The purpose of this study was to evaluate the effects of surface properties of bone implants coated with hydroxyapatite (HA) and beta-tricalcium phosphate (beta-TCP) on platelets and macrophages upon implant installation and compare them to grit-blasted Ti and Thermanox used as a control. Surface properties were characterized using scanning electron microscopy, profilometry, crystallography, Fourier transform infrared spectroscopy, and coating stability. For platelets, platelet adherence and morphology were assessed. For macrophages, morphology, proliferation, and polarization were evaluated. Surface characterization showed similar roughness of similar to 2.5 mu m for grit-blasted Ti discs, both with and without coating. Coating stability assessment showed substantial dissolution of HA and beta-TCP coatings. Platelet adherence was significantly higher for grit-blasted Ti, Ti-HA, and Ti-beta-TCP coatings compared to that of cell culture control Thermanox. Macrophage cultures revealed a decreased proliferation on both HA and beta-TCP coated discs compared to both Thermanox and grit-blasted Ti. In contrast, secretion of pro-inflammatory cytokine TNF-alpha and anti-inflammatory cytokine TGF-beta were marginal for grit-blasted Ti and Thermanox, while a coating-dependent increased secretion of pro- and anti-inflammatory cytokines was observed for HA and beta-TCP coatings. The results demonstrated a significantly upregulated pro-inflammatory and anti-inflammatory cytokine secretion and marker gene expression of macrophages on HA and beta-TCP coatings. Furthermore, HA induced an earlier M1 macrophage polarization but more M2 phenotype potency than beta-TCP. In conclusion, our data showed that material surface affects the behaviors of first cell types attached to implants. Due to the demonstrated crucial roles of platelets and macrophages in bone healing and implant integration, this information will greatly aid the design of metallic implants for a higher rate of success in patients.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Radboudumc, Dept Biomat, POB 9101, NL-6500 HB Nijmegen, NetherlandsFed Univ Sao Paulo UNIFESP, Dept Biosci, 136 Silva Jardim St, BR-11015021 Santos, SP, BrazilFed Univ Sao Paulo UNIFESP, Dept Biosci, 136 Silva Jardim St, BR-11015021 Santos, SP, BrazilCAPES: 9424/2014-08Web of Scienc

Complement proteins regulating macrophage polarisation on biomaterials

[EN] One of the events occurring when a biomaterial is implanted in an host is the protein deposition onto its surface, which might regulate cell responses. When a biomaterial displays a compromised biocompatibility, distinct complement pathways can be activated to produce a foreign body reaction. In this article, we have designed different types of biomaterial surfaces to study the inflammation process. Here, we used different concentrations of (3-glycidoxypropyl)-trimethoxysilane (GPTMS), an organically-modified alkoxysilane as a precursor for the synthesis of various types of sol-gel materials functionalizing coatings for titanium implants to regulate biological responses. Our results showed that greater GPTMS surface concentrations induced greater secretion of TNF-alpha and IL-10 on RAW 264.7 macrophages. When implanted into rabbit tibia, osseointegration decreased with higher GPTMS concentrations. Interestingly, higher deposition of complement-related proteins C-reactive protein (CRP) and ficolin-2 (FCN2), two main activators of distinct complement pathways, was observed. Taking all together, inflammatory potential increase seems to be GPTMS concentration-dependent. Our results show that a greater adsorption of complement proteins can condition macrophage polarization.This work was supported by MINECO [MAT2017-86043-R]; Universitat Jaume I [Predoc/2014/25, UJI-B2017-37]; Basque Government [IT611-13, Predoc/2016/1/0141]; University of the Basque Country [UFI11/56]; CIC bioGUNE is supported by Basque Department of Industry, Tourism and Trade (Etortek and Elkartek programs), ProteoRed-ISCIII [PRB3 IPT17/0019]; CIBERehd Network and Severo Ochoa Grant [SEV-2016-0644]. Authors would like to thank Antonio Coso and Jaime Franco (GMI-Ilerimplant) for their inestimable contribution to this study, and Raquel Oliver, Jose Ortega (UJI), René van Rheden, Vicent Cuijpers (Radboudumc) and Iraide Escobes (CIC bioGUNE) for their valuable technical assistance.Araújo-Gomes, N.; Romero-Gavilán, F.; Zhang, Y.; Martínez-Ramos, C.; Elortza, F.; Azkargorta, M.; Martín De Llano, JJ.... (2019). Complement proteins regulating macrophage polarisation on biomaterials. Colloids and Surfaces B Biointerfaces. 181:125-133. https://doi.org/10.1016/j.colsurfb.2019.05.039S12513318

Osmolarity-Induced Altered Intracellular Molecular Crowding Drives Osteoarthritis Pathology

Osteoarthritis (OA) is a multifactorial degenerative joint disease of which the underlying mechanisms are yet to be fully understood. At the molecular level, multiple factors including altered signaling pathways, epigenetics, metabolic imbalance, extracellular matrix degradation, production of matrix metalloproteinases, and inflammatory cytokines, are known to play a detrimental role in OA. However, these factors do not initiate OA, but are mediators or consequences of the disease, while many other factors causing the etiology of OA are still unknown. Here, it is revealed that microenvironmental osmolarity can induce and reverse osteoarthritis-related behavior of chondrocytes via altered intracellular molecular crowding, which represents a previously unknown mechanism underlying OA pathophysiology. Decreased intracellular crowding is associated with increased sensitivity to proinflammatory triggers and decreased responsiveness to anabolic stimuli. OA-induced lowered intracellular molecular crowding could be renormalized via exposure to higher extracellular osmolarity such as those found in healthy joints, which reverse OA chondrocyte's sensitivity to catabolic stimuli as well as its glycolytic metabolism.</p

Osmolarity-Induced Altered Intracellular Molecular Crowding Drives Osteoarthritis Pathology

Osteoarthritis (OA) is a multifactorial degenerative joint disease of which the underlying mechanisms are yet to be fully understood. At the molecular level, multiple factors including altered signaling pathways, epigenetics, metabolic imbalance, extracellular matrix degradation, production of matrix metalloproteinases, and inflammatory cytokines, are known to play a detrimental role in OA. However, these factors do not initiate OA, but are mediators or consequences of the disease, while many other factors causing the etiology of OA are still unknown. Here, it is revealed that microenvironmental osmolarity can induce and reverse osteoarthritis-related behavior of chondrocytes via altered intracellular molecular crowding, which represents a previously unknown mechanism underlying OA pathophysiology. Decreased intracellular crowding is associated with increased sensitivity to proinflammatory triggers and decreased responsiveness to anabolic stimuli. OA-induced lowered intracellular molecular crowding could be renormalized via exposure to higher extracellular osmolarity such as those found in healthy joints, which reverse OA chondrocyte's sensitivity to catabolic stimuli as well as its glycolytic metabolism.</p

Osmolarity-Induced Altered Intracellular Molecular Crowding Drives Osteoarthritis Pathology

Osteoarthritis (OA) is a multifactorial degenerative joint disease of which the underlying mechanisms are yet to be fully understood. At the molecular level, multiple factors including altered signaling pathways, epigenetics, metabolic imbalance, extracellular matrix degradation, production of matrix metalloproteinases, and inflammatory cytokines, are known to play a detrimental role in OA. However, these factors do not initiate OA, but are mediators or consequences of the disease, while many other factors causing the etiology of OA are still unknown. Here, it is revealed that microenvironmental osmolarity can induce and reverse osteoarthritis-related behavior of chondrocytes via altered intracellular molecular crowding, which represents a previously unknown mechanism underlying OA pathophysiology. Decreased intracellular crowding is associated with increased sensitivity to proinflammatory triggers and decreased responsiveness to anabolic stimuli. OA-induced lowered intracellular molecular crowding could be renormalized via exposure to higher extracellular osmolarity such as those found in healthy joints, which reverse OA chondrocyte's sensitivity to catabolic stimuli as well as its glycolytic metabolism.</p

Designing Public Transport To Foster Patronage And Social Inclusion

Institute of Transport and Logistics Studies. Faculty of Economics and Business. The University of Sydne