Percutaneous transluminal angioplasty with stent implantation is used to dilate of arteries narrowed by atherosclerotic plaques and to revascularize coronary arteries occluded by atherothrombosis in myocardial infarction. Commonly applied drug-eluting stents release antiproliferative or anti-inflammatory agents to reduce the incidence of in-stent stenosis. However, these stents may lead to in-stent stenosis and increase the rate late stent thrombosis, an obstacle to Correspondence: Oliver Soehnlein, MD, PhD or Christian Weber, MD, Institute for Cardiovascular Prevention, Pettenkoferstr. 9, 80336 Munich, Phone +49-(0)89-5160-4350, Fax +49-(0)89-5160-4352, oliver.soehnlein@med.uni-muenchen.de or christian.weber@med.uni-muenchen.de. * These authors contributed equally. Competing interests:The authors do not declare any competing financial interests. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript optimal revascularization possibly related to endothelial recovery. Here we examined the contribution of neutrophils and neutrophilic granule proteins to arterial healing after injury. We found that neutrophil-born cathelicidin (mouse CRAMP, human LL-37) promoted reendothelization and thereby limited neointima formation after stent implantation. We then translated these findings, generating a neutrophil-instructing biofunctionalized miniaturized Nitinol stent coated with LL-37. This stent reduced in-stent stenosis in a mouse model of atherosclerosis, suggesting that LL-37 may promote vascular healing after interventional therapy.

The results of the present study demonstrated that ethanol precipitation is a simple and effective method for isolation of fibrinogen and a suitable alternative to cryoprecipitation. This technique allows minimization of the necessary blood volume for fibrinogen isolation, particularly important for pediatric applications, and also has no negative influence on microstructure, mechanical properties, cell proliferation, or tissue development.

Chronic wounds represent a serious problem in daily medical routine requiring improved wound care. Silk of the domesticated silkworm (Bombyx mori) has been used to form a variety of biomaterials for medical applications. We genetically engineered B. mori to produce silk functionalized with growth factors to promote wound healing in vitro. In this study FGF-, EGF-, KGF-, PDGF- or VEGF-functionalized silk membranes were compared to native B. mori silk membranes without growth factors for their ability to support wound healing in vitro. All silk membranes were cytocompatible and supported macrophage secretion of neutrophil recruiting factor CXCL1 and monocyte chemoattractant protein 1 (MCP-1). VEGF-functionalized silk significantly outperformed other growth factor-functionalized silk membranes, but not native silk in angiogenesis assays. In addition, EGF- and VEGF-functionalized silk membranes slightly enhanced macrophage adhesion compared to silk without growth factors. In wound healing assays in vitro (reduction of wound lesion), dermal equivalents showed a higher wound healing capacity when covered with EGF-, FGF- or VEGF-functionalized silk membranes compared to native, KGF- or PDGF-functionalized silk membranes. Keratinocyte migration and growth is overstimulated by KGF- and VEGF-functionalized silk membranes. In conclusion, growth factor-functionalized silk membranes prepared from genetically engineered silk worm glands are promising wound dressings for future wound healing therapies.

A successful strategy to enhance the in vivo survival of engineered tissues would be to prevascularize them. In this study, fabricated silk fibroin scaffolds from mulberry and non-mulberry silkworms are investigated and compared for supporting the co-culture of human umbilical vein endothelial cells and human foreskin fibroblasts. Scaffolds are cytocompatible and when combined with fibrin gel support capillary-like structure formation. Density and interconnectivity of the formed structures are found to be better in mulberry scaffolds. ELISA shows that levels of vascular endothelial growth factor (VEGF) released in co-cultures with fibrin gel are significantly higher than in co-cultures without fibrin gel. RT PCR shows an increase in VEGFR2 expression in mulberry scaffolds indicating these scaffolds combined with fibrin provide a suitable microenvironment for the development of capillary-like structures.

The guidance of vessel growth within tissue-engineered constructs can be achieved using biofunctionalized PLLA microfibers. Further methods are warranted to perform specified spatial positioning of fibers within 3D formative scaffolds to enhance the applicability of the concept.

Organotypic full-thickness skin grafts (OTSG) are already an important technology for treating various skin conditions and are well established for skin research and development. These obvious benefits are often impaired by the need of laborious production, their noncomplete autologous composition, and, most importantly, their lack of included vasculature. Therefore, our study focused on combining a prevascularized dermal layer with an epidermis to cultivate full-thickness skin grafts incorporating capillary-like networks. It has been shown that prevascularization accelerates ingrowth of tissue-engineered grafts, and it is a prerequisite to circumvent diffusion limits due to graft thickness. To obtain such a graft, we chose a dermal layer incorporating human umbilical vein endothelial cells (HuVEC) amid human dermal fibroblasts within a fibrin-based scaffold, seeded apically with human foreskin keratinocytes (hfKC). Our research investigated the used concept's feasibility, as well as the effect of hfKC addition on the development of a well-connected capillary-like network after approximately 21 days. In addition, we evaluated the utilization of a custom-made constant flow bioreactor for simplified cultivation of these grafts, therefore possibly easing graft production and presumably increasing their cost effectiveness. Skin grafts were assessed by conventional two-dimensional histology. In addition, software-assisted three-dimensional evaluation of the capillary-like structure networks was performed by twophoton laser scanning microscopy (TPLSM) and subsequent image processing was done with ImagePro Ò Analyzer 7.0 software, thereby evaluating its platform technology power in the field of prevascularized skin grafts. All samples showed a capillary-like structure network, but we could report a significant reduction of its total length after 14 days of tri-culture with 5 · 10 5 /cm 2 seeded hfKC, possibly indicating nutritional deficiencies for this particular high cell density experimental setup. Lower concentrations of hfKC did not affect the formation of the capillary-like structures significantly. The developed bioreactor simplified cultivation of prevascularized OTSG. However, a flow-dependent reduction of capillary-like structures in 1 and 5 mL/min flow conditions occurred. We conclude that our technique for creating prevascularized OTSG is feasible. In addition, TPLSM is well suited for analyzing the prevascularization process. We hypothesize that the handling benefits of our bioreactor can be preserved by using considerably lower flow rates while not impairing the forming of capillary-like structure networks.

The generation of complex three-dimensional bioengineered scaffolds that are capable of mimicking the molecular and topographical cues of the extracellular matrix found in native tissues is a field of expanding research. The systematic development of such scaffolds requires the characterisation of cell behaviour in response to the individual components of the scaffold. In the present investigation, we studied cell-substrate interactions between purified populations of Schwann cells and three-dimensional fibrin hydrogel scaffolds, in the presence or absence of multiple layers of highly orientated electrospun polycaprolactone nanofibres. Embedded Schwann cells remained viable within the fibrin hydrogel for up to 7 days (the longest time studied); however, cell behaviour in the hydrogel was somewhat different to that observed on the two-dimensional fibrin substrate: Schwann cells failed to proliferate in the fibrin hydrogel, whereas cell numbers increased steadily on the two-dimensional fibrin substrate. Schwann cells within the fibrin hydrogel developed complex process branching patterns, but, when presented with orientated nanofibres, showed a strong tendency to redistribute themselves onto the nanofibres, where they extended long processes that followed the longitudinal orientation of the nanofibres. The process length along nanofibre-containing fibrin hydrogel reached near-maximal levels (for the present experimental conditions) as early as 1 day after culturing. The ability of this three-dimensional, extracellular matrix-mimicking scaffold to support Schwann cell survival and provide topographical cues for rapid process extension suggest that it may be an appropriate device design for the bridging of experimental lesions of the peripheral nervous system.

The BioStent concept is a platform technology offering a novel opportunity to generate a viable, self-expanding stent structure with a functional endothelial cell lining. This platform technology can be transferred to different applications depending on the luminal cell lining required.