PH Modification of High-Concentrated Collagen Bioinks as a Factor Affecting Cell Viability, Mechanical Properties, and Printability.

Jana Stepanovska,Roman Matejka,Martin Otahal,Karel Hanzalek,Monika Supova

doi:10.3390/gels7040252

Abstract

The 3D bioprinting of cell-incorporated gels is a promising direction in tissue engineering applications. Collagen-based hydrogels, due to their similarity to extracellular matrix tissue, can be a good candidate for bioink and 3D bioprinting applications. However, low hydrogel concentrations of hydrogel (<10 mg/mL) provide insufficient structural support and, in highly concentrated gels, cell proliferation is reduced. In this study, we showed that it is possible to print highly concentrated collagen hydrogels with incorporated cells, where the viability of the cells in the gel remains very good. This can be achieved simply by optimizing the properties of the bioink, particularly the gel composition and pH modification, as well as by optimizing the printing parameters. The bioink composed of porcine collagen hydrogel with a collagen concentration of 20 mg/mL was tested, while the final bioink collagen concentration was 10 mg/mL. This bioink was modified with 0, 5, 9, 13, 17 and 20 μL/mL of 1M NaOH solution, which affected the resulting pH and gelling time. Cylindrical samples based on the given bioink, with the incorporation of porcine adipose-derived stromal cells, were printed with a custom 3D bioprinter. These constructs were cultivated in static conditions for 6 h, and 3 and 5 days. Cell viability and morphology were evaluated. Mechanical properties were evaluated by means of a compression test. Our results showed that optimal composition and the addition of 13 μL NaOH per mL of bioink adjusted the pH of the bioink enough to allow cells to grow and divide. This modification also contributed to a higher elastic modulus, making it possible to print structures up to several millimeters with sufficient mechanical resistance. We optimized the bioprinter parameters for printing low-viscosity bioinks. With this experiment, we showed that a high concentration of collagen gels may not be a limiting factor for cell proliferation.

Highlights

We showed that it is possible to print highly concentrated collagen hydrogels with incorporated cells, where the viability of the cells in the gel remains very good
The maintenance of the bioink temperature at 10 ◦ C before printing followed by heating to 37 ◦ C after extrusion, as well as adjusting the printing speed, was found to be crucial
With a final collagen concentration of 10 mg/mL, these collagens were considered unsuitable for cell growth, as the cells had low proliferation

Summary

Introduction

Bioprinting of cell-incorporated gels is a promising direction for producing “customized” tissues and organs for patients. Polymers used for bioprinting vary in their properties and, suitability for this application [1]. The ideal polymer should achieve high mechanical integrity and stability, insolubility in culture medium and ideal biodegradability, while being non-toxic to cells, non-immunogenic and promoting cell adhesion and proliferation [2]. Ease of workability, suitability for bioprinting, affordability and commercial availability are essential for selecting a suitable gel [2]. Synthetic gels are characterized by excellent mechanical resistance, whereas natural gels, on the other hand, achieve exceptionally high biocompatibility [3].

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Results

Conclusion