Clot-entrapped blood cells in synergy with human mesenchymal stem cells create a pro-angiogenic healing response.

Blood clots stop bleeding and provide cell-instructive microenvironments. Still, in vitro models used to study implant performance typically neglect any possible interactions of recruited cells with surface-adhering blood clots. Here we study the interaction and synergies of bone marrow derived human mesenchymal stem cells (hMSCs) with surface-induced blood clots in an in vitro model by fluorescence microscopy, scanning and correlative light and electron microscopy, ELISA assays and zymography. The clinically used alkali-treated rough titanium (Ti) surfaces investigated here are known to enhance blood clotting compared to native Ti and to improve the healing response, but the underlying mechanisms remain elusive. Here we show that the presence of blood clots synergistically increased hMSC proliferation, extracellular matrix (ECM) remodelling and the release of matrix fragments and angiogenic VEGF, but did not increase the osteogenic differentiation of hMSCs. While many biomaterials are nowadays engineered to release pro-angiogenic factors, we show here that clot-entrapped blood cells on conventional materials in synergy with hMSCs are potent producers of pro-angiogenic factors. Our data might thus not only explain why alkali-treatment is beneficial for Ti implant integration, but they suggest that the physiological importance of blood clots to create pro-angiogenic environments on implants has been greatly underestimated. The importance of blood clots might have been missed because the pro-angiogenic functions get activated only upon stimulation by synergistic interactions with the invading cells.

Synergistic interactions of blood-borne immune cells, fibroblasts and extracellular matrix drive repair in an in vitro peri-implant wound healing model.

Low correlations of cell culture data with clinical outcomes pose major medical challenges with costly consequences. While the majority of biomaterials are tested using in vitro cell monocultures, the importance of synergistic interactions between different cell types on paracrine signalling has recently been highlighted. In this proof-of-concept study, we asked whether the first contact of surfaces with whole human blood could steer the tissue healing response. This hypothesis was tested using alkali-treatment of rough titanium (Ti) surfaces since they have clinically been shown to improve early implant integration and stability, yet blood-free in vitro cell cultures poorly correlated with in vivo tissue healing. We show that alkali-treatment, compared to native Ti surfaces, increased blood clot thickness, including platelet adhesion. Strikingly, blood clots with entrapped blood cells in synergistic interactions with fibroblasts, but not fibroblasts alone, upregulated the secretion of major factors associated with fast healing. This includes matrix metalloproteinases (MMPs) to break down extracellular matrix and the growth factor VEGF, known for its angiogenic potential. Consequently, in vitro test platforms, which consider whole blood-implant interactions, might be superior in predicting wound healing in response to biomaterial properties.

The ultrastructure of fibronectin fibers pulled from a protein monolayer at the air-liquid interface and the mechanism of the sheet-to-fiber transition.

Fibronectin is a globular protein that circulates in the blood and undergoes fibrillogenesis if stretched or under other partially denaturing conditions, even in the absence of cells. Stretch assays made by pulling fibers from droplets of solutions containing high concentrations of fibronectin have previously been introduced in mechanobiology, particularly to ask how bacteria and cells exploit the stretching of fibronectin fibers within extracellular matrix to mechano-regulate its chemical display. Our electron microscopy analysis of their ultrastructure now reveals that the manually pulled fibronectin fibers are composed of densely packed lamellar spirals, whose interlamellar distances are dictated by ion-tunable electrostatic interactions. Our findings suggest that fibrillogenesis proceeds via an irreversible sheet-to-fiber transition as the fibronectin sheet formed at the air-liquid interface of the droplet is pulled off by a sharp tip. This far from equilibrium process is driven by the externally applied force, interfacial surface tension, shear-induced fibronectin self-association, and capillary force-induced buffer drainage. The ultrastructural characterization is then contrasted with previous FRET studies that characterized the molecular strain within these manually pulled fibers. Particularly relevant for stretch-dependent binding studies is the finding that the interior fiber surfaces are accessible to nanoparticles smaller than 10 nm. In summary, our study discovers the underpinning mechanism by which highly hierarchically structured fibers can be generated with unique mechanical and mechano-chemical properties, a concept that might be extended to other bio- or biomimetic polymers.

On the cytocompatibility of biodegradable Fe-based alloys.

Biodegradable iron-based alloys are potential candidates for application as temporary implant material. This study summarizes the design strategy applied in the development of biodegradable Fe-Mn-C-Pd alloys and describes the key factors which make them suitable for medical applications. The study's in vitro cytotoxicity tests using human umbilical vein endothelial cells revealed acceptable cytocompatibility based on the alloys' eluates. An analysis of the eluates revealed that Fe is predominantly bound in insoluble degradation products, whereas a considerable amount of Mn is in solution. The investigation's results are discussed using dose-response curves for the main alloying elements Fe and Mn. They show that it is mainly Mn which limits the cytocompatibility of the alloys. The study also supplies a summary of the alloying elements' influence on metabolic processes. The results and discussion presented are considered important and instructive for future alloy development. The Fe-based alloys developed show an advantageous combination of microstructural, mechanical and biological properties, which makes them interesting as degradable implant material.

On the in vitro and in vivo degradation performance and biological response of new biodegradable Mg-Y-Zn alloys.

A design strategy deployed in developing new biodegradable Mg-Y-Zn alloys is summarized and the key factors influencing their suitability for medical applications are described. The Mg-Y-Zn alloys reveal microstructural features and mechanical characteristics expected to be appropriate for vascular intervention applications. The focus of this article lies in the evaluation of the degradation performance and biological response of the alloys with respect to their potential as implant materials (stents). The degradation characteristics analyzed by immersion testing and electrochemical impedance spectroscopy in simulated physiological media reveal slow and homogeneous degradation. In vitro cell tests using human umbilical vein endothelial cells indicate good cytocompatibility on the basis of the alloys' eluates (extracts). Animal studies carried out with pigs on Mg-2Y-1Zn (in wt.%) reveal an auspicious in vivo performance. Evaluation of preparations derived from implants in various types of tissues indicates homogeneous degradation and only limited gas formation during in vivo testing. The characteristics of the tissue reactions indicate good biocompatibility. The new Mg-Y-Zn alloys show an interesting combination of preferred microstructural, mechanical, electrochemical and biological properties, which make them very promising for degradable implant applications.