Effect of PEG grafting density on surface properties of polyurethane substrata and the viability of osteoblast and fibroblast cells

Abstract

The surface of Tecoflex SG-80A Polyurethane (PU) films was modified by grafting polyethylene glycol (PEG) chains at three different molar amounts (0.05, 0.10, and 0.15 mmol). The resulting substrata were characterized by FTIR-ATR, TGA, AFM, SEM and contact angle to assess the surface modifications occurred during the grafting reactions. Osteoblasts and fibroblasts were cultured with PU extracts for 24 h, and their cell viability and morphology were evaluated by CellTiterBlue assay, Crystal Violet staining and Live/Dead assay. FTIR and TGA results indicated that PEG chains were successfully grafted onto PU surfaces, specifically in the hard segment of PU forming allophanate groups as the PEG grafting density increased. SEM and AFM images suggest that PU substrata were partially covered by PEG, increasing the dispersive and basic components of the PU surface energy. It was found that extracts from PEG-grafted polyurethanes increased the osteoblast viability, although fibroblasts viability remained constant regardless PEG grafting density; in spite of this both cells presented a more spread morphology at the lower PEG grafting density. Our results showed that surface energy of PU substrata can be tuned by PEG grafting density; also, the PEG leached tends to increase the pH of culture medium which leads to a higher viability of osteoblasts; nevertheless, PEG grafting density should be optimized to promote a healthy cell morphology as alterations in its morphology were detected at higher concentrations. Graphical abstract.

Graphical abstract

Fig. 1

Grafting reaction of PEG onto PU substrata. (a) One isocyanate group from end groups of HMDI reacts with the secondary amine of the urethane group to yield allophanate linkages remaining an isocyanate group unreacted (first step); (b) hydroxyl group from PEG reacts with the free isocyanate group obtained in the previous stage to yield a new-urethane linkage (second step)

Fig. 2

FTIR-ATR spectra of the PU films and PEG. As the graft density is increased the spectra of the films look more similar to PEG spectra, indicating that there is a higher amount of PEG chains present

Fig. 3

Deconvolution of carbonyl bands from 1800 to 1600 cm^–1. The signals at 1719 cm⁻¹ (A) are assigned to non-hydrogen-bonded carbonyls, 1695 cm⁻¹ (B) to hydrogen-bonded carbonyls, and 1663 cm⁻¹ (C) to carbonyls from allophanate groups

Fig. 4

Raman spectra of the PU films and PEG

Fig. 5

SEM images of the grafted films. A PEG covering is formed on the surface of the substrata

Fig. 6

AFM images of untreated PU and PEG-grafted PU films. a PU, (b) PU-PEG 0.05, (c) PU-PEG 0.10, (d) PU-PEG 0.15

Fig. 7

Cell viability of osteoblast and fibroblasts cells by indirect contact with PU substrata for 24 h. CWE: cells without contact with extracts of the films. P < 0.001, one-way ANOVA with Tukey’s multiple comparison test, n = 5 in each group

Fig. 8

Microscopic images of Crystal Violet-stained osteoblasts in contact with extracts of the films for 24 h. a CWE, (b) PU, (c) PU-PEG 0.05, (d) PU-PEG 0.10, (e) PU-PEG 0.15

Fig. 9

Microscopic images of Crystal Violet-stained fibroblasts in contact with extracts of the films for 24 h. a CWE, (b) PU, (c) PU-PEG 0.05, (d) PU-PEG 0.10, (e) PU-PEG 0.15

Fig. 10

Confocal laser images of LIVE/DEAD staining of osteoblasts in contact with extracts of the films for 24 h