Improved Hemocompatibility on Superhemophobic Micro-Nano-Structured Titanium Surfaces

Abstract

Blood-contacting titanium-based implants such as endovascular stents and heart valve casings are prone to blood clotting due to improper interactions at the surface level. In complement, the current clinical demand for cardiovascular implants is at a new apex. Hence, there is a crucial necessity to fabricate an implant with optimal mechanical properties and improved blood compatibility, while simultaneously interacting differentially with cells and other microbial agents. The present study intends to develop a superhydrophobic implant surface with the novel micro-nano topography, developed using a facile thermochemical process. The surface topography, apparent contact angle, and crystal structure are characterized on different surfaces. The hemo/blood compatibility on different surfaces is assessed by evaluating hemolysis, fibrinogen adsorption, cell adhesion and identification, thrombin generation, complement activation, and whole blood clotting kinetics. The results indicate that the super-hemo/hydrophobic micro-nano titanium surface improved hemocompatibility by significantly reducing fibrinogen adsorption, platelet adhesion, and leukocyte adhesion. Thus, the developed surface has high potential to be used as an implant. Further studies are directed towards analyzing the mechanisms causing the improved hemocompatibility of micro/nano surface features under dynamic in vitro and in vivo conditions.

Keywords: hemocompatible; micro–nano surface topography; superhemophobic; superhydrophobic; titanium implant surface.

Figure 1

(a) Representative SEM images of different surfaces. Images were taken at 500×, and image inserts depict 5000× magnification. (b) Apparent contact angle measurements using Milli-Q water, PRP, and blood on different surfaces. (c) XRD intensity peaks of different surfaces (n_min = 9).

Figure 2

Fibrinogen adsorption from PRP on different surfaces measured using a microplate reader. (* p < 0.05; n_min = 9.)

Figure 3

(a) Fluorescence images of live cells (platelets and WBCs) adhered to different surfaces. (b) Percentage area covered by adhered platelets and WBCs on different surfaces. (c) Fluorescence images of adhered platelets (red) and WBCs (purple) on different surfaces. (d) Percentage of the areas covered by adhered platelets (red) on different surfaces. (e) Percentage area covered by adhered WBCs (blue) on different surfaces. (* and ** p < 0.05; n_min = 9.)

Figure 4

Representative SEM images of adhered platelets and WBCs (purple—Photoshop used for better visibility) on different surfaces. Images were taken at 500× and 2000× magnification. (n_min = 9.)

Figure 5

Hemoglobin release from erythrocytes solution incubated with different surfaces was measured using a microplate reader. (n_min = 9.)

Figure 6

Highest thrombin generation velocity of plasma incubated with different surfaces between two points. (* p < 0.05, n_min = 9).

Figure 7

Complement activation of plasma incubated with different surfaces was measured as activation of complement convertase C5a. The lines indicate the regions of inactive/low (≤0.2), medium (>0.2 and ≤0.6), and high (>0.6) reactivity. (n_min = 9.)

Figure 8

Whole blood clotting on different surfaces for up to 45 min. The dotted line represents the absorbance of free hemoglobin in un-clotted blood. (* p < 0.05; n_min = 9.)