Daniela Arbeiter

VEGF-releasing suture material for enhancement of vascularization: Development, in vitro and in vivo study

As it has been demonstrated that bioactive substances can be delivered locally using coated surgical suture materials, the authors developed a vascular endothelial growth factor (VEGF)-releasing suture material that should promote vascularization and potentially wound healing. In this context, the study focused on the characterization of the developed suture material and the verification of its biological activity, as well as establishing a coating process that allows reproducible and stable coating of a commercially available polydioxanone suture material with poly(l-lactide) (PLLA) and 0.1μg and 1.0μg VEGF. The in vitro VEGF release kinetics was studied using a Sandwich ELISA. The biological activity of the released VEGF was investigated in vitro using human umbilical vein endothelial cells. The potential of the VEGF-releasing suture material was also studied in vivo 5days after implantation in the hind limb of Wistar rats, when the histological findings were analyzed. The essential results, enhanced cell viability in vitro as well as significantly increased vascularization in vivo, were achieved using PLLA/1.0μg VEGF-coated suture material. Furthermore, ELISA measurements revealed a high reproducibility of the VEGF release behavior. Based on the results achieved regarding the dose-effect relationship of VEGF, the stability during its processing and the release behavior, it can be predicted that a bioactive suture material would be successful in later in vivo studies. Therefore, this knowledge could be the basis for future studies, where bioactive substances with different modes of action are combined for targeted, overall enhancement of wound healing.

Novel approach for a PTX/VEGF dual drug delivery system in cardiovascular applications—an innovative bulk and surface drug immobilization

The successive incorporation of several drugs into the polymeric bulk of implants mostly results in loss of considerable quantity of one drug, and/or the loss in quality of the coating and also in changes of drug release time points. A dual drug delivery system (DDDS) based on poly-l-lactide (PLLA) copolymers combining the effective inhibition of smooth muscle cell proliferation while simultaneously promoting re-endothelialization was successfully developed. To overcome possible antagonistic drug interactions and the limitation of the polymeric bulk material as release system for dual drugs, a novel concept which combines the bulk and surface drug immobilization for a DDDS was investigated. The advantage of this DDDS is that the bulk incorporation of fluorescein diacetate (FDAc) (model drug for paclitaxel (PTX)) via spray coating enhanced the subsequent cleavable surface coupling of vascular endothelial growth factor (VEGF) via the crosslinker bissulfosuccinimidyl suberate (BS3). In the presence of the embedded FDAc, the VEGF loading and release are about twice times higher than in absence. Furthermore, the DDDS combines the diffusion drug delivery (FDAc or PTX) and the chemical controlled drug release, VEGF via hydrolysable ester bonds, without loss in quantity and quality of the drug release curves. Additionally, the performed in vitro biocompatibility study showed the bimodal influences of PTX and VEGF on human endothelial EA.hy926 cells. In conclusion, it was possible to show the feasibility to develop a novel DDDS which has a high potential for the medical application due to the possible easy and short modification of a polymer-based PTX delivery system.

Nanofibrous polyamide 6 scaffolds promote adhesion of endothelial cells

Abstract The usage of synthetic scaffolds is a promising approach in development of implant materials. In this study we fabricated nanofibrous nonwovens of polyamide 6 (PA-6) by means of electrospinning and performed a systematic characterization with regard to the mechanical and biological performance of scaffold materials. Mechanical strength was assessed by uniaxial tensile testing and biological performance was evaluated by measuring cell viability and qualitative analysis of cellular morphology when human umbilical vein endothelial cells (EA.hy926) or human fibroblasts (HT-1080) were grown on polymeric substrates. While all polymeric materials exhibited an excellent biocompatibility with respect to cell viability, their surface topography promoted the adhesion of endothelial but not fibroblast cells. A better understanding of the physicochemical and morphological material properties with a selective impact on cell adhesion will help to further improve biocompatibility of nonwovens for biomedical applications.

Systematic analysis about residual chloroform removal from PCL films

Abstract For fabrication of polymeric implants and drug carriers volatile organic solvents, such as chloroform, are widely used. In order to remove solvents for patient safety several processes have to be carried out to avoid changes in the thermal characteristics of the polymers, as well as in the drug release profile and mechanical properties. We analyzed films of the polymer poly(ε−caprolactone) (PCL), a common commercial medical grade polymer which is widely used in implants and drug carriers. Considering the influence on the morphology and thermal properties we screened different post treatment processes. Acceptable chloroform contents according to recommendations of the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) without changes in morphology and crystallinity were achieved via a simple annealing process for 24 h at 40°C and 40 mbar.

Influence of additives on physico-chemical properties of electrospun poly(L-lactide)

Abstract Electrospun poly(L-lactide) (PLLA) nonwoven represent potential options for biodegradable medical implants. They can be manufactured with high reproducibility and do offer the potential for chemical modification to alter matrix properties. In our study, we investigate the mechanical, thermal and morphological properties of PLLA fiber matrices. Fibrous nonwovens were fabricated from polymer solution by needle electrospinning. The polymer solutions were loaded with Triton X-100 (TX- 100), formic acid and tetraethyl ammonium chloride (TEAC) with respect to polymer weight. Morphology of the PLLA nonwoven scaffolds was examined with SEM. We performed uniaxial tensile tests and differential scanning calorimetry (DSC). Different concentrations of the additives TEAC, formic acid and Triton X-100 lead to strong changes regarding mechanical and thermal properties of the electrospun PLLA fiber matrices. In comparison with mechanical properties of established biological tissue materials, the results indicate the suitability for medical applications.

Electrospinning for polymeric implants in cardiovascular applications

Abstract Electrospinning is a method for producing fibrous polymer scaffolds that can be applied in drug delivery systems as well as for polymer-based implants. Biodegradable polymers for the purpose of cardiac tissue engineering are often applied as fibrous scaffolds for morphological mimikry of natural matrices but also drugeluting approaches are very promising. Hydrolytic degradation is one of the key parameters for successful application. The focus of our investigations is on monitoring accelerated in vitro degradation of electrospun nonwoven scaffolds. In the presented study degradation of poly(Llactide) is accelerated by alkaline hydrolysis. The process is characterized by weight loss, loss of molecular mass, surface morphology and thermal behavior of nonwoven samples, showing a fast degradation of the fibrous material within two weeks.

Polymer drug release system for biofilm inhibition in medical application

Abstract Bacterial biofilm formation on surfaces is still a critical challenge regarding the application of implants. Generally, in order to avoid this, an additional systemic administration of antibiotics is given, which can lead to side effects, such as the reduction of the intestinal flora. Continuous treatment may lead to antibiotics resistance. Within this study we investigated the local drug delivery of N-acetyl-L-cysteine (NAC) from a Poly-L-lactide (PLLA) coating, an ished biodegradable polymer and a polyetherurethane (PEU) coating, a promising representative non-degradable polymer for cardiovascular applications as alternative to the administration of antibiotics. The incorporation of NAC influenced the surface properties of PEU in contrast to that of PLLA. The in vitro NAC release is almost completed after 24 h for PEU. For PLLA only small amounts of incorporated NAC, depending on the NAC loading, is released after a short time. Both systems are rather useful as local NAC delivery system directly after implantation.

Optimization of manufacturing processes for biodegradable polymeric stents regarding improved mechanical properties

Abstract Although current drug eluting stents show low risks of in-stent restenosis and stent thrombosis, the presence of a permanent foreign body inside the vessel represents a major limitation. In order to overcome this limitation, stents made of biodegradable polymeric materials are currently being developed. The present work describes an optimized fabrication process for tubular semi-finished products for manufacturing of stents made of poly-L-lactide (PLLA). The impact of the haul-off speed as a major parameter during extrusion processing on the cross-sectional area of tubular specimens was analyzed. It could be shown that the crosssection of the extrudate, in particular the tubing diameter and wall thickness, can be adjusted by varying haul-off speed. In a subsequent blow molding process the influence of the holding time on polymer cold crystallization was analyzed. Thermal properties of the polymeric material after processing were examined by differential scanning calorimetry (DSC). The results showed that there is almost no cold crystallization using a holding time of at least 20 minutes. The investigations showed that semi-finished products with variable geometry and improved mechanical properties can be produced with the described extrusion and blow molding process.

Cyclic stress-strain behavior of polymeric nonwoven structures for the use as artificial leaflet material for transcatheter heart valve prostheses

Abstract Xenogenic leaflet material, bovine and porcine pericardium, is widely used for the fabrication of surgically implanted and transcatheter heart valve prostheses. As a biological material, long term durability of pericardium is limited due to calcification, degeneration and homogeneity. Therefore, polymeric materials represent a promising approach for a next generation of artificial heart valve leaflets with improved durability. Within the current study we analyzed the mechanical performance of polymeric structures based on elastomeric materials. Polymeric cast films were prepared and nonwovens were manufactured in an electrospinning process. Analysis of cyclic stress-strain behavior was performed, using a universal testing machine. The uniaxial cyclic tensile experiments of the elastomeric samples yielded a non-linear elastic response due to viscoelastic behavior with hysteresis. Equilibrium of stress-strain curves was found after a specific number of cycles, for cast films and nonwovens, respectively. In conclusion, preconditioning was found obligatory for the evaluation of the mechanical performance of polymeric materials for the use as artificial leaflet material for heart valve prostheses.

Influence of bulk incorporation of FDAc and PTX on polymer properties

Abstract In the last decades PLLA-based copolymers have been among the most attractive polymeric candidates used to fabricate devices for drug delivery and stent applications in the cardiovascular system. PLLA is biocompatible and biodegradable, exhibits a wide range of erosion times and has tunable mechanical properties. Therefore, the influence of drug incorporation on the physicochemical properties of biodegradable PLLA copolymers were examined in this study using Fluorescein diacetate (FDAc) and Paclitaxel (PTX). A percental amount of these drugs (17.5 %) were incorporated into poly(L-lactide-co-glycolide) (P(LLA-co-GA)) and poly(L-lactide-co-ε-caprolactone) (P(LLA-co-CL)) made via spray coating. The polymer surface properties, such as surface morphology and hydrophilicity were also examined and remained rather unchanged for both polymers after drug loadings. Furthermore, also the contact angle changed rather marginally. However, both polymers have already different thermal properties without the drug embedded, especially the glass transition temperature (TG) is for P(LLA-co-CL) under 37°C and for P(LLA-co-GA) considerable above with around 66°C. An rather high increase in TG achieved by addition of FDAc or PTX, crucial influences the drug release profiles for P(LLA-co-CL) in contrast to P(LLA-co-GA). Besides these results preliminarily experiments of additional coupling of other drugs on the polymer surface were performed and we obtained an influence of FDAc or PTX. The drug incorporation and physicochemical characterization data obtained in this study is relevant in optimizing the incorporation or coupling of further drugs on the polymer surface and delivery properties of these potential multi drug delivery coatings.