Implicaciones del calcio extracelular y su receptor en mebrana (CaSR) en la angiogénesis y la osteogénesis. Relevancia en ingeniería tisular.

Bone injury is a major health problem nowadays. Bone tissue engineering is a promising strategy to solve it. In this field many reports pointed out the role of scaffolds containing a calcium phosphate glass, such as G5, in the promotion of bone regeneration. These kinds of biomaterials release Ca2+ during degradation. Moreover, extracellular calcium (Ca2+0) is able to activate a special extracellular receptor called Calcium Sensing Receptor (CaSR), which can modulate migration, chemotaxis and differentiation. In that sense, present work aims to evaluate the role of extracellular calcium and CaSR as key factor in the induction of bone regeneration. We isolated endothelial progenitor cells (EPCs) and Mesenchymal Stromal cells (MSCs) from young rat’s bone marrow and they were stimulated with 10 mM Ca2+0. Results provided strong evidences about the role of Ca2+0 and CaSR in the modulation of chemotaxis, and angiogenic differentiation on EPCs, whilst on MSCs calcium exerted osteoinduction and proliferation through the CaSR. Furthermore, using biodegradable releasing calcium composite (PLA/G5) was measured the ability of that kind of biomaterials in the induction of bone and blood vessels formation in vivo. Finally, using a shell-less chick embryo model was compared the effect of the Ca2+0 and the ionic compound released by PLA/G5, demonstrating that the released Ca2+0 is the key factor in the promotion of angiogenesis and bone formation in vivo. The knowledge developed during this project will be a valuable tool to enhance the efficiency of biomaterials used in bone tissue engineerin

A Step Closer to Elastogenesis on Demand; Inducing Mature Elastic Fibre Deposition in a Natural Biomaterial Scaffold

Elastic fibres play a key role in bodily functions where fatigue resistance and elastic recovery are necessary while regulating phenotype, proliferation and migration in cells. While in vivo elastic fibres are created at a late foetal stage, a major obstacle in the development of engineered tissue is that human vascular smooth muscle cells (hVSMCs), one of the principal elastogenic cells, are unable to spontaneously promote elastogenesis in vitro. Therefore, the overall aim of this study was to activate elastogenesis in vitro by hVSMCs seeded in fibrin, collagen, glycosaminoglycan (FCG) scaffolds, following the addition of recombinant human tropoelastin. This combination of scaffold, tropoelastin and cells induced the deposition of elastin and formation of lamellar maturing elastic fibres, similar to those found in skin, blood vessels and heart valves. Furthermore, higher numbers of maturing branched elastic fibres were synthesised when a higher cell density was used and by drop-loading tropoelastin onto cell-seeded FCG scaffolds prior to adding growth medium. The addition of tropoelastin showed no effect on cell proliferation or mechanical properties of the scaffold which remained dimensionally stable throughout. With these results, we have established a natural biomaterial scaffold that can undergo controlled elastogenesis on demand, suitable for tissue engineering applications

3D silicon doped hydroxyapatite scaffolds decorated with Elastin-like Recombinamers for bone regenerative medicine

The current study reports on the manufacturing by rapid prototyping technique of three-dimensional (3D) scaffolds based on silicon substituted hydroxyapatite with Elastin-like Recombinamers (ELRs) functionalized surfaces. Silicon doped hydroxyapatite (Si-HA), with Ca-10(PO4)(5.7)(SiO4)(0.3)(OH)(1.7)h(0.3) nominal formula, was surface functionalized with two different types of polymers designed by genetic engineering: ELR-RGD that contain cell attachment specific sequences and ELR-SNA15/RGD with both hydroxyapatite and cells domains that interact with the inorganic phase and with the cells, respectively. These hybrid materials were subjected to in vitro assays in order to clarify if the ELRs coating improved the well-known biocompatible and bone regeneration properties of calcium phosphates materials. The in vitro tests showed that there was a total and homogeneous colonization of the 3D scaffolds by Bone marrow Mesenchymal Stromal Cells (BMSCs). In addition, the BMSCs were viable and able to proliferate and differentiate into osteoblasts. Statement of Significance Bone tissue engineering is an area of increasing interest because its main applications are directly related to the rising life expectancy of the population, which promotes higher rates of several bone pathologies, so innovative strategies are needed for bone tissue regeneration therapies. Here we use the rapid prototyping technology to allow moulding ceramic 3D scaffolds and we use different bio-polymers for the functionalization of their surfaces in order to enhance the biological response. Combining the ceramic material (silicon doped hydroxyapatite, Si-HA) and the Elastin like Recombinamers (ELRs) polymers with the presence of the integrin-mediate adhesion domain alone or in combination with SNAI 5 peptide that possess high affinity for hydroxyapatite, provided an improved Bone marrow Mesenchymal Stromal Cells (BMSCs) differentiation into osteoblastic linkage. (C) 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved

Identification of stiffness-induced signalling mechanisms in cells from patent and fused sutures associated with craniosynostosis

Abstract Craniosynostosis is a bone developmental disease where premature ossification of the cranial sutures occurs leading to fused sutures. While biomechanical forces have been implicated in craniosynostosis, evidence of the effect of microenvironmental stiffness changes in the osteogenic commitment of cells from the sutures is lacking. Our aim was to identify the differential genetic expression and osteogenic capability between cells from patent and fused sutures of children with craniosynostosis and whether these differences are driven by changes in the stiffness of the microenvironment. Cells from both sutures demonstrated enhanced mineralisation with increasing substrate stiffness showing that stiffness is a stimulus capable of triggering the accelerated osteogenic commitment of the cells from patent to fused stages. The differences in the mechanoresponse of these cells were further investigated with a PCR array showing stiffness-dependent upregulation of genes mediating growth and bone development (TSHZ2, IGF1), involved in the breakdown of extracellular matrix (MMP9), mediating the activation of inflammation (IL1β) and controlling osteogenic differentiation (WIF1, BMP6, NOX1) in cells from fused sutures. In summary, this study indicates that stiffer substrates lead to greater osteogenic commitment and accelerated bone formation, suggesting that stiffening of the extracellular environment may trigger the premature ossification of the sutures

Translational Studies on the Potential of a VEGF Nanoparticle-Loaded Hyaluronic Acid Hydrogel

Heart failure has a five-year mortality rate approaching 50%. Inducing angiogenesis following a myocardial infarction is hypothesized to reduce cardiomyocyte death and tissue damage, thereby preventing heart failure. Herein, a novel nano-in-gel delivery system for vascular endothelial growth factor (VEGF), composed of star-shaped polyglutamic acid-VEGF nanoparticles in a tyramine-modified hyaluronic acid hydrogel (nano-VEGF-HA-TA), is investigated. The ability of the nano-VEGF-HA-TA system to induce angiogenesis is assessed in vivo using a chick chorioallantoic membrane model (CAM). The formulation is then integrated with a custom-made, clinically relevant catheter suitable for minimally invasive endocardial delivery and the effect of injection on hydrogel properties is examined. Nano-VEGF-HA-TA is biocompatible on a CAM assay and significantly improves blood vessel branching (p < 0.05) and number (p < 0.05) compared to a HA-TA hydrogel without VEGF. Nano-VEGF-HA-TA is successfully injected through a 1.2 m catheter, without blocking or breaking the catheter and releases VEGF for 42 days following injection in vitro. The released VEGF retains its bioactivity, significantly improving total tubule length on a Matrigel(R) assay and human umbilical vein endothelial cell migration on a Transwell(R) migration assay. This VEGF-nano in a HA-TA hydrogel delivery system is successfully integrated with an appropriate device for clinical use, demonstrates promising angiogenic properties in vivo and is suitable for further clinical translation