Human serum albumin in electrospun PCL fibers: structure, release, and exposure on fiber surface

Abstract

Human serum albumin (HSA) introduced to the fibers produced by electrospinning from HSA and polycaprolactone (PCL) solutions in hexafluoroisopropanol has been studied in terms of its structure, release from the fibers, stability of interaction with basic polymer, accessibility for protease attack, and cellular receptors, as well as dependence of the studied parameters on the protein concentration in fibers. A limited part of the protein leaves the fibers right after soaking with water, whereas the remaining protein stays tightly bound to fibers for a long time because protein nanoparticles are tightly integrated with PCL, as shown by small‐angle X‐ray scattering. As has been demonstrated, the proteins leave the fibers in complexes with PCL. X‐ray photoelectron spectroscopy demonstrates that the protein concentration on the fiber surface is higher than the concentration in electrospinning solution. The surface‐exposed protein is recognized by cell receptors and is partially hydrolyzed by proteinase K. The data on pulse protein release, presence of PCL in the protein released from matrixes, overrepresentation of the protein on the fiber surface, and tight interaction of protein with PCL may be useful for rational design of electrospun scaffolds intended for drug delivery and tissue engineering. Copyright © 2016 John Wiley & Sons, Ltd.

General Study and Gene Expression Profiling of Endotheliocytes Cultivated on Electrospun Materials

“…The microstructure of the matrix surface was studied using scanning electron microscopy (SEM), (JSM-6460 LV, Jeol, Tokyo, Japan) as described previously [20] and atomic force microscopy (AFM) with Multimode 8 (Bruker, Germany) using NSG10-DLC diamond-like carbon NSG series AFM cantilever (NT-MDT, Zelenograd, Russia) in tapping mode. Fiber diameter and pore size were calculated from SEM images according to ISO 7198:1998 using AxioVision 4.8 software (Carl Zeiss, Gottingen Germany).…”

“…Polytetrafluoroethylene rings were also installed in control wells to make the free bottom surface area even across all the wells. Following 48 h cultivation, the viability of endothelial cells was assessed using Alamar Blue (Invitrogen, Eugene, Oregon, USA) as described previously [20]. Matrices not seeded with cells were used as control for dye sorption on the material, and the surface of the 3D matrices (after O-ring installation) was taken into account in the further estimations.…”

“…3D matrices seeded with HUVEC were prepared for SEM as follows: after 48 h cultivation, culture medium was removed from the wells; matrices were washed twice with phosphate buffer, fixed with 2% formaldehyde in physiological saline solution for 30 min, washed thrice with H 2 O, and air-dried. Matrices were removed from the wells, sputter-coated with 10-nm gold film, and examined by SEM as described previously [20].…”

“…XPS demonstrated that the concentration of Gl on the surface of the fibers in these matrices was no less than 21%, and most of the surface-exposed Gl was tightly bound to the fibers [44]. It was previously shown that, in 3D matrices produced from PCL with 10% HSA (w:w), 20% of the protein was exposed at the surface while only 34.7% of HSA was exposed at the surface of the fibers produced from PCL with 30% HSA [20]. Excess surface Gl can dissociate from the fibers, potentially negatively affecting cell adhesion.…”

“…Polycaprolactone (PCL), in particular, was extensively studied in this context, both as a sole component of matrices and blended with different proteins [17][18][19]. In the latter case, the proteins were exposed at the fiber surface, free to interact with cellular receptors, thus facilitating cell adhesion and promoting growth [20].…”