pH-Dependent Ordered Fibrinogen Adsorption on Polyethylene Single Crystals

Abstract

Nanostructured surfaces have the potential to influence the assembly as well as the orientation of adsorbed proteins and may, thus, strongly influence the biomaterials’ performance. For the class of polymeric (bio)­materials a reproducible and well-characterized nanostructure is the ordered chain folded surface of a polyethylene single crystal (PE-SC). We tested the hypothesis that the trinodal-rod-shaped protein human plasma fibrinogen (HPF) adsorbs on the (001) surface of PE-SCs along specific crystallographic directions. The PE-SC samples were prepared by isothermal crystallization in dilute solution and characterized by atomic force microscopy (AFM) before as well as after HPF adsorption at different concentrations and pH values. At a physiological pH of 7.4, connected HPF molecules, or e.g., fibrils, fibril networks, or sponge-like structures, were observed on PE-SC surfaces that featured no preferential orientation. The observation of these nonoriented multiprotein assemblies was explained by predominant protein–protein interactions and limited surface diffusion. However, at an increased pH of 9.2, single HPF molecules, e.g., spherical-shaped and trinodal-rod-shaped HPF molecules as well as agglomerates, were observed on the PE-SC surface. The presence of single HPF molecules at increased pH was explained by decreased protein–protein interactions. These single trinodal-rod-shaped HPF molecules oriented preferentially along crystallographic [100] and [010] directions on the PE-SC surface which was explained by an increased amount of intermolecular bonds along these crystallographic directions with increased surface atom density. The study established that HPF molecules can align on chemically homogeneous surface topographies one order of magnitude smaller than the dimension of the protein. This advances the understanding of how to control the assembly and orientation of proteins on nanostructured polymer surfaces. Controlled protein adsorption is a crucial key to improve the surface functionality of future implants and biosensors.

Principles of Fibrinogen Fiber Assembly In Vitro

“…Single nanofibers as well as networks of fibril‐like structures were found on the hydrophobic PE‐SC crystals. [ 83 ] Interestingly, the fibrous networks resembled the fibrinogen structures previously observed on HOPG nanosteps [ 18 ] although much lower protein concentrations were used. Overall, increasing protein concentrations yielded fibers with larger dimensions and a higher network density: 0.5 µg mL −1 of fibrinogen yielded single nanofibers with an average width of 30 ± 5 nm and 2 µg mL −1 resulted in a nanofiber network with nanofiber widths of 45 ± 10 nm.…”

“…In a similar study, the assembly of fibrous fibrinogen structures on nanostructured, hydrophobic polyethylene single crystals (PE‐SC) was reported. [ 83 ] Hydrophobic PE‐SC nanocrystals were fully covered with fibrinogen solution and stored at 37 °C for 2 h prior to rinsing with PBS or CB and drying with compressed air. Single nanofibers as well as networks of fibril‐like structures were found on the hydrophobic PE‐SC crystals.…”

“…Overall, increasing protein concentrations yielded fibers with larger dimensions and a higher network density: 0.5 µg mL −1 of fibrinogen yielded single nanofibers with an average width of 30 ± 5 nm and 2 µg mL −1 resulted in a nanofiber network with nanofiber widths of 45 ± 10 nm. [ 83 ] With a concentration of 5 µg mL −1 fibrinogen sponge‐like structures with interconnected fibers were observed. [ 83 ]…”

Controlling Protein Adsorption through Nanostructured Polymeric Surfaces

“…These findings were corroborated by work of Helbing et al who studied the adsorption behavior of fibrinogen on a more fundamental PE surface, namely the surface of polyethylene single crystal, PE-SC, with surface features one order of magnitude smaller than the dimension of the protein. [13] The crystallographic directions of high surface chain densities at the PE-SC surface were found to steer the orientation of single molecules. The observed adsorption behavior of fibrinogen was explained by the similarity of the α-helix structure dimensions (width, pitch length, and angle) to the PE unit cell (Figure 6a).…”

“…Adapted with permission. [13] Copyright 2016, American Chemical Society. b) Model of a nanostructured melt-drawn UHMWPE surface (left) and AFM image (right) displaying fibrinogen assembly guided by lamellar crystals (protein concentration of 10 mg L −1 ).…”