PEG-penetrated chitosan-alginate co-polysaccharide-based partially and fully cross-linked hydrogels as ECM mimic for tissue engineering applications

Abstract

The emerging strategy of tissue engineering for the management of end-stage organ failure and associated complications mainly relies on ECM mimicking scaffolds for neo-tissue genesis. In the current study, novel polyethylene glycol interpenetrated cross-linked hydrogel scaffold based on a co-polysaccharide (PIAC) synthesized from two marine heteropolysaccharides, alginate and chitosan, was designed. Partially cross-linked (PIAC-P) and fully cross-linked hydrogels (PIAC-F) were prepared. The physiochemical evaluations of both the hydrogels revealed the presence of alginate fraction and extensive -OH groups on the surface, sufficient water content and water holding capacity. The porosity and bulk density were also appreciable. The scaffolds were hemocompatible and were able to adsorb appreciable plasma proteins on to the surface. MTT assay on hydrogel extracts and direct contact assay showed the nontoxic effects of fibroblast cells upon contact with the hydrogel. Live/dead assay using ethidium bromide/acridine orange cocktail on fibroblast cells grown on the hydrogels after 5 days of initial seeding displayed green nucleus revealing the non-apoptotic cells. PIAC-P hydrogels were superior to certain aspects due to the availability of free functional groups than PIAC-F where most of these groups were utilized for cross-linking. The biological evaluations confirmed the healthy being and 3D growth of fibroblasts on the porous networks of both the hydrogels. The present hydrogel can form an ECM mimic and can form a potent candidate for various tissue engineering applications.

Keywords: Alginate; Chitosan; Cross-linked hydrogels; ECM mimic; Tissue engineering.

Fig. 1

Synthesis of partially cross-linked PIAC-P hydrogel scaffolds

Fig. 2

Synthesis of fully cross-linked PIAC-F hydrogel scaffolds

Fig. 3

IR spectral analysis of PIAC-P- and PIAC-F-based hydrogel scaffolds

Fig. 4

Water holding capacity and profile of PIAC-P-and PIAC-F-based hydrogel scaffolds. The values of time zero indicate the dry weights of the scaffolds

Fig. 5

RBC aggregation studies showing the absence of aggregation in PIAC-P-based (b) and PIAC-F-based (c) hydrogel scaffolds in comparison with saline control (a) and positive control polyethyleneimine (d)

Fig. 6

SDS-PAGE analysis of plasma protein adsorption on to PIAC-P- and PIAC-F-based hydrogel scaffolds showing thick bands corresponding to that of albumin

Fig. 7

Cytocompatibility evaluations of hydrogels. Direct contact assay on L929 cells—control (a), PIAC-P-based (b) and PIAC-F-based (c) hydrogel scaffolds. Live/dead assay on L929 cells—control (d) PIAC-P-based (e) and PIAC-F-based (f) hydrogel scaffolds