Implementation of Functional Microfluidic Dispensing Tools Enabling Enhanced Control in Extrusion Bioprinting

Inspired from additive manufacturing's success, the structuring of biocompatible materials through bioprinting has become a growing field of research in the past fifteen years. It opened the door to a plethora of applications ranging from the fabrication of tissue models for research to regenerative medicine. This field of research has mainly been tackled from a biological view point to study the culture of cells ''ex-vivo'' as well as through a materials science lens as to synthesize extracellular matrices. Per contra, the tools currently used for bioprinting do not allow advanced manipulations that are needed to reproduce the complex structures found in tissues. This sets strict limitations on what can be achieved by bioprinting. This thesis proposes an engineering perspective on bioprinting by providing new tools which improve the level of control over deposited biomaterials through extrusion bioprinting. The advancements in microfluidic technology are leveraged to micromanufacture extrusion tools serving as printing heads for bioprinters.

Successful implementation of a microfluidic mixer combined with a heating element and temperature sensor in an extrusion tool, enabled the printing of cryogels. These macroporous materials display remarkable properties, such as high pore interconnection, large surface to volume ratio and high mechanical stability. A collagen coating that is covalently bonded to the cryogel has been developed to enable the culture of adherent cells within the printed scaffold. Using the heating element, the pores' size of the printed cryogel can be controlled during printing allowing the selection of the position of the seeded cells.

A cell concentrator capable of working as a printing head was implemented. The ability to concentrate cells just before printing presents remarkable advantages such as reduced risk of cell lysis in tubings and reduced cell losses in dead volumes of bioprinters. As evidence of the beneficial effects of printing at a high cell concentration were found, such tools provide the opportunity to work at high cellular concentrations without the complications usually associated with it. The implementation of a microfluidic mixer downstream of the cell concentrator enables the addition of extracellular matrix, providing a complete platform for tissue printing at high cell concentration. Using this tool, fibroblasts were printed at high concentration in a collagen matrix. Moreover, using the concentrator, primary uroepithelial cells were printed in a basement membrane extract. The control over the cellular concentration allowed for the optimization of the amount of printed cells which has been demonstrated to have a major impact on bladder organoid formation.

Finally, as many extracellular matrices take several minutes to crosslink, multi-layer 3D printing of these materials can be challenging. Hence solutions for the printing of liquid materials are explored through the creation of a composite biomaterial, co-printing of rigid support materials and liquid materials using a dedicated extrusion head and the printing of liquid materials within a supporting gel. These printing techniques allowed to cope with the long curing time of certain bioinks and to shape them in 3D.

The tools developed in this work set the ground for the development of a new generation of bioprinters providing more freedom for the design and printing of functional tissues through enhanced control over the printed materials.