Development of an on-demand foaming printhead for biofabrication of constructs with heterogeneous porosity

Abstract

Three-dimensional bioprinting stands out as a distinctive subset of additive manufacturing, characterized by the precise deposition of biomaterials to create intricate 3D structures with embedded living cells. This technology enables the biofabrication of complex constructs capable of mimicking natural tissues. However, for effective 3D bioprinting, bioinks—cellladen biomaterials—must meet stringent rheological and mechanical properties to ensure high printability and structural integrity. The intrinsic porosity of bioinks is often insufficient for biological applications, posing significant challenges in tissue engineering. Adequate porosity is crucial for supporting cellular processes such as proliferation, differentiation, and extracellular matrix deposition. Enhancing the porosity of bioinks is therefore essential to improve their functionality and effectiveness in tissue engineering and regenerative medicine. This thesis addresses the challenge in the development of tunable porous 3D printable biomaterials with a wide range of pore sizes for the biofabrication of heterogeneous constructs. The approach of this research minimizes the use of additives and processing steps for porosity generation. To achieve this, a foaming printhead was designed and developed. The printhead proves to be a superior alternative to bioprinting a passively foamed solution, due to its higher control over the tunability of foam characteristics while bioprinting. This foaming method involves the mechanical agitation of albumin (the foaming agent) within the printhead, which leads to the denaturation of its protein structure. Bubbles in the foam are created by trapping air with the hydrophilic and hydrophobic groups of the denatured protein. Sodium alginate is used alongside albumin to leverage its biocompatibility and rapid ionic crosslinking properties. Gelatin is added to the bioink to enhance cell attachment. The produced foam is then crosslinked upon exiting the needle using a crosslinking module capable of delivering a calcium chloride mist. It was hypothesized that changing the foaming speed would result in changing the pore size of the constructs and affect cell activities and drug delivery properties. Therefore, the foaming printhead includes a foaming module with a DC motor to change the rotation speed of a pinion in an enclosed space with an internal gear design to obtain a wide range of pore sizes. The developed printhead enables on-demand mixing of bioink and air at high speeds in a foaming module, followed by combining the resulting foam with cells in a mixing unit. Finally, the cell-laden foam is crosslinked by delivering the crosslinker in mist form. The biofabricated scaffolds were characterized by assessments of printability, porosity, water absorption, degradation, and drug release capabilities. Our studies revealed that these properties can be tuned by controlling the foaming speed, which affects the porosity size. Foaming speeds of 1500, 2500, and 3500 rpm generated average pore sizes of 96, 84, and 53 μm, respectively. Increasing the foaming speed induces higher shear forces in the solution, breaking larger bubbles into smaller ones. Samples produced at lower foaming speeds exhibited higher rates of water absorption, degradation, and drug release. The biocompatibility of the porous constructs and the bioprinting system was assessed using the Live/Dead assay, which showed over 90% cell viability after 1 day. The results of this study highlight the capability of biofabricating heterogeneous structures for tissue engineering and drug delivery applications, with the option to regulate pore size on demand by adjusting the foaming speed.

Date	26 Aug 2024
Original language	American English
Awarding Institution	École de technologie supérieure
Supervisor	Ali Ahmadi (Supervisor) & Sophie Lerouge (Co-supervisor)