Volume 16, Issue 4 - Special issue: Biofunctionalization

Resorbable polymers have been established for several decades in biomedical applications. The most frequently used resorbable polymers are still the aliphatic polyesters polylactides (PLA), polyglycolid (PGA) and polycaprolactone (PCL) homo- and copolymers. However, inherent pH dropping during degradation of some biomaterials may provoke inflammation and, thus, hamper the healing process. In this study we investigate the manufacturing method of microgel functionalised PLA Fibres in a dry-spinning process and the buffering effect of the poly(N-vinylcaprolactam-co-acetoacetoxyethyl methacrylate) vinylimidazole (VCL/AAEM/Vlm) microgels during the degradation of the fibres. Furthermore we examine the biocompatibility of the produced fibres and established a mathematical model to describe and analyse the pH level in the vicinity of the PLA fibre.

Polyelectrolyte multilayer (PEM) films, established by layer-by-layer deposition, are attractive for drug delivery systems as drug delivery can be systematically controlled. However, one limitation of this coating method is the time-consuming character. The adsorption time of one layer typically takes minutes and commonly, investigated multilayers loaded with drugs contain more than 80 layers. Within our study we systematically evaluate parameters allowing the generation of a PEM-based drug delivery system on poly(L-lactide) (PLLA) with a reduced time consuming construction process and a controlled delivery of physiological amounts of active drugs. Therefore, several PEM based on PLLA with only five tetralayers consisting of polycation/polyanion/drug/polyanion are established and investigated regarding the release profiles of avidin fluorescein isothiocyanate as model substance and vascular endothelial growth factor (VEGF). We demonstrate that the polyelectrolyte nature and molecular weight of polyanions (hyaluronic acid, polyacrylic acid) and polycations (poly-L-lysine, chitosan) strongly influence the drug release profiles. Furthermore, in vitro cell culture studies show that the released VEGF amount and the cover layer from the favorite coating systems can beneficially enhance human umbilical vein endothelial cell (HUVEC) proliferation.

Functionality of implant surfaces is considerably determined by the conformations of immobilized biomolecules adjustable by the applied surface modification approach. Moreover, surface modification of drug eluting systems often needs to be thoroughly optimized with regard to possible drug losses associated with a loss of functionality. Here, we thoroughly investigate biomolecule surface loading, functionality, morphology and stability in dependence of modification conditions at the example of a site-selective immobilization of anti-CD34 antibodies (CD34-Ab) to drug-eluting stents with the aim of providing general information on the biofunctionalization of drug eluting systems. In this context, we demonstrate that the reaction time of biomolecule immobilization defines achievable surface loads but also drug loss. We could moreover show that the used site-selective immobilization procedure elevated the surface functionality considerably in comparison to surfaces modified by random physisorption. Furthermore investigated bionfunctional stability gives important indications for storage conditions of biofunctionalized implants.

Background: Optimizing endothelialization of medical implants requires deep mechanistic insight into cellular adhesion, cell junction and physiological basement membrane development at the endothelial cell-to-scaffold substrate interface. Methods and results: We employed and standardized endothelial cells and fibrin hydrogel for simultaneous cell-plus-fibrin (EC-Fib) spray application using the Maslanka ® spray pen. Quality assessment illustrated excellent structural integrity of EC-Fib. Acellular SynerGraft ® showed improved intimal endothelial recellularization after short-term physical and chemical preconditionings including dry-freezing, sodium desoxycholate, Triton-X, acetic acid and collagenase treatments. Artificial substrates poly-L-lactic acid (PLLA), and polyamide-6 (PA-6) were tested regarding their mechanical appropriateness, revealed complete endothelialization following EC-Fib application and showed partially physiological basement membrane development. Additional laminin 1 biofunctionalization on PA-6 moderately enhanced basement membrane gene and protein expression. However, scanning electron microscopy, immunohistology and PCR analyses underlined immature endothelialization, basement membrane and cell junction development on all substrates with clear substrate-dependent differences. Conclusions: Direct sprayed endothelialization outlined the necessity for preconditioning acellular SynerGraft ® prostheses for adequate recellularization. Laminin 1 biofunctionalization could slightly improve endothelialization of PA-6 substrates whereas physiological organization of the microenvironment remained immature and seemed highly substrate dependent.

One step to enable long-term use of extracorporeal membrane oxygenation devices or even the development of an artificial fully implantable lung is the endothelialization of oxygenator membranes in order to present a physiological and anti-thrombogenic surface to the blood flow. Since cell seeding decreases the gas transfer of oxygenation devices, a way to reincrease gas exchange performance by other means has to be found. In this study membrane assemblies suitable for endothelialization were developed, which profit from a secondary flow arrangement to increase gas transfer rates, the so-called Bellhouse effect. Therefore textiles that allow a homogeneous gas flow between the membranes and provide a structure that can be used as mold for hollow imprinting onto the membrane surfaces are combined with flat membranes to a sandwich structure. On top of that two approaches for hollow imprinting are generated and their results compared. The furrowed membrane assemblies are seeded with HUVECs and regularly inspected over 3 days cultivation. A surface characterization of the applied membranes is performed by contact angle measurement to identify reasons for inhomogeneous cell growth. In general first important results to develop a biohybrid lung assist device could be achieved in this study.

Currently used synthetic vascular grafts bear a high infection risk due to insufficient microvascularization of the graft wall disabling the infiltration of immune cells. Tissue-engineered grafts with a functional pre-vascularization thus would be desirable. However, autologous tissue sources for capillary forming cells need to be evaluated. Here, peripheral blood outgrowth endothelial cells (PB-OEC) from 17 healthy donors and pericyte-like mesenchymal stem cells derived from adipose tissue (ASC) of 17 patients scheduled for visceral surgery were characterized and investigated regarding their ability to form capillary-like networks in plasma-derived fibrin gels. To obtain proliferating PB-OEC with endothelial cell-specific properties (CD31-, VE-cadherin-expression, ac-LDL uptake and three-dimensional (3D)-tube formation in fibrin gels) both enrichment of CD34 + blood cells and young donor age was necessary (7/17, age≤24 years). In contrast, all isolated ASC revealed the expression of surface antigens expressed on pericytes [human neural/glial antigen-2 (hNG2), platelet-derived growth factor receptor β (PDGF-Rβ)] and showed mesodermal differentiation capacity. Moreover, co-culture of PB-OEC and ASC in fibrin gels resulted in highly branched capillary-like networks with significantly increased tube length (2.9-fold, p<0.0001) and number of junctions (8-fold, p<0.0001). In conclusion, successful cell isolation from autologous tissues for pre-vascularization of vascular grafts has been demonstrated although certain limitations for autologous EC require further strategies to enable the use of allogeneic cells.