Enhancing the printability of low-concentration GelMA through viscosity modulation and integration of hydroxyapatite for bone tissue engineering bioinks

Abstract

In recent years, there has been a significant focus on developing hydrogel-based scaffolds for reconstructing and repairing damaged tissues. Despite these efforts, the selection of appropriate hydrogel formulation tailored to specific clinical applications remains a primary challenge. Gelatin methacryloyl (GelMA) has been widely investigated as a baseline biomaterial in the realm of tissue engineering. Through comprehensive experimentation and quantitative analysis, we explore the intricate interplay among various biophysical properties (uniaxial compression behavior, scaffold microstructure, swelling properties, and enzymatic degradation kinetics), viscoelastic properties, printability, and cellular responses of a range of GelMA compositions. The experimental data were comprehensively analyzed to establish an empirical relationship between biophysical properties and molar crosslinking density. In particular, the viscoelastic properties were tailored for low-concentration GelMA, containing biomineralized bone-specific biomaterial ink by tailoring the addition of methacrylated carboxymethyl cellulose (mCMC), and nanocrystalline hydroxyapatite (nHAp). The resulting hybrid hydrogel demonstrates significantly higher stiffness (∼7-fold), improved yield stress (∼17-fold), reduced swelling (∼1.3-fold), and diminished degradation (∼4-fold) properties compared to pristine GelMA. To assess the bone mimetic tissue matrix development, we conducted 2D cultures of human patient-derived primary bone marrow mesenchymal stem cells (hBMSCs) and human osteoblasts (hOBs) on hydrogel scaffolds in standard growth media and differentiation media. Our results qualitatively and quantitatively indicate robust proliferation of both cell types on all biomaterial scaffolds over 21 days in culture. Furthermore, an analysis of alkaline phosphatase (ALP) activity reveals a ∼3.1-fold and ∼5.8-fold increase in ALP expression for hBMSCs-seeded nHAp-loaded hydrogels, cultured in non-differentiation media and differentiation media, respectively. Taken together, our findings suggest that the nHAp-incorporated GelMA/mCMC matrix holds promise as a potential biomaterial ink for bone tissue regeneration applications.