Three-Dimensional Printed Cell Culture Model Based on Spherical Colloidal Lignin Particles and Cellulose Nanofibril-Alginate Hydrogel.

Xue Zhang,Maria Morits,Ari Ora,Orlando Rojas,Mika Henrikki Sipponen,Monika Österberg,Rubina Ajdary,Markus Linder,Siqi Huan,Christopher Jonkergouw,Muhammad Farooq,Juan José Valle-Delgado

doi:10.1021/acs.biomac.9b01745

Abstract

Three-dimensional (3D) printing has been an emerging technique to fabricate precise scaffolds for biomedical applications. Cellulose nanofibril (CNF) hydrogels have attracted considerable attention as a material for 3D printing because of their shear-thinning properties. Combining cellulose nanofibril hydrogels with alginate is an effective method to enable cross-linking of the printed scaffolds in the presence of Ca2+ ions. In this work, spherical colloidal lignin particles (CLPs, also known as spherical lignin nanoparticles) were used to prepare CNF-alginate-CLP nanocomposite scaffolds. High-resolution images obtained by atomic force microscopy (AFM) showed that CLPs were homogeneously mixed with the CNF hydrogel. CLPs brought antioxidant properties to the CNF-alginate-CLP scaffolds in a concentration-dependent manner and increased the viscosity of the hydrogels at a low shear rate, which correspondingly provide better shape fidelity and printing resolution to the scaffolds. Interestingly, the CLPs did not affect the viscosity at high shear rates, showing that the shear thinning behavior typical for CNF hydrogels was retained, enabling easy printing. The CNF-alginate-CLP scaffolds demonstrated shape stability after printing, cross-linking, and storage in Dulbecco’s phosphate buffer solution (DPBS +) containing Ca2+ and Mg2+ ions, up to 7 days. The 3D-printed scaffolds showed relative rehydration ratio values above 80% after freeze-drying, demonstrating a high water-retaining capability. Cell viability tests using hepatocellular carcinoma cell line HepG2 showed no negative effect of CLPs on cell proliferation. Fluorescence microscopy indicated that HepG2 cells grew not only on the surfaces but also inside the porous scaffolds. Overall, our results demonstrate that nanocomposite CNF-alginate-CLP scaffolds have high potential in soft-tissue engineering and regenerative-medicine applications.

Highlights

There is a growing interest to manufacture three-dimensional (3D) cell culture models.[1]
A variety of materials have been used for the preparation of hydrogels, such as synthetic polymers like poly(ethylene glycol), poly(vinyl alcohol), and poly(2-hydroxy ethyl methacrylate) and natural polymers like collagen, fibrin, hyaluronic acid, chitosan, alginate, and cellulose nanofibrils (CNFs).[4,7]
Colloidal Lignin Particles (CLPs) have been used for the first time to tune the properties of hydrogels for 3D printing

Summary

Introduction

There is a growing interest to manufacture three-dimensional (3D) cell culture models.[1] The reasons behind this interest are the many advantages of 3D cell culture over conventional twodimensional culture. Hydrogels have attracted interest as materials for producing 3D cell culture models.[4,5] The physical properties of the hydrogels, such as mechanical properties, dimensional stability, and rehydration degree play vital roles in the potential applications.[4] Mechanical properties, stiffness, regulate cellular processes such as proliferation, cell spreading, and cell differentiation via different mechanotransduction mechanisms.[6] A variety of materials have been used for the preparation of hydrogels, such as synthetic polymers like poly(ethylene glycol), poly(vinyl alcohol), and poly(2-hydroxy ethyl methacrylate) and natural polymers like collagen, fibrin, hyaluronic acid, chitosan, alginate, and cellulose nanofibrils (CNFs).[4,7] CNF is a nanomaterial derived from wood (and other plants) by mechanical disintegration It has micrometerlong flexible fibrils with a thickness in the nanometer range.[8] CNF hydrogels are attractive materials for biomedical applications because they are nontoxic, xeno-free, biocompatible, and abundant.[9,10] The high water-binding ability and structural resemblance to the natural extracellular matrix make CNF an interesting material for 3D cell cultures.[7,11,12]

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Conclusion