NSF/FDA/SIR: Evaluating Cellular Integrity as a Function of Bioprinter Nozzle Geometry

Three dimensional bioprinting is an emerging technique for the fabrication of tissue models mimicking native human tissues, and these synthetic tissues have been proposed as physiological models for preclinical testing of drugs and biologics in vitro. In the widely-used extrusion-based bioprinting systems, cells are often damaged (e.g., altered metabolic activity, reduced cell adhesiveness, cell lysis, changes in phenotype, increased susceptibility to toxic leachables) by the shear stress imposed by the high-speed dispensing of the cell laden hydrogel or bioink through a straight micronozzle. As micronozzle geometry evolves to provide greater resolution of the formed structure, the impact of this geometry on the dispensed cellular species has not been adequately considered. As the cells move from a reservoir into the nozzle they experience shear stress and undergo deformations in response. Bioprinter nozzle geometry (e.g., size and shape) is a crucial determinant of the shear stress distributions generated on the cells along the length of the capillary tube of nozzle. While cross-sectional diameter critically governs the magnitude of shear stresses generated on the surfaces of the cells, the length-scale of the micronozzle determines the duration of exposure to the shear stresses. These parameters of micronozzle geometry can be used to adjust shear stress exposure regimes on cells. The use of different nozzle shapes (e.g., conical vs cylindrical) affords an additional level of control of the shear stress distributions on cells while fine-tuning bioprinter print resolution. For example, whereas a cylindrical “capillary tube” geometry imposes a shear stress on cells along the entire length of the nozzle, a conical shaped nozzle would introduce a gradient of shear stress with the highest shear stresses being imposed on cells for shorter durations at the bioprinter outlet. Currently, at FDA, as a part of an ongoing NSF-SIR project – Brad Berron (PI; University of Kentucky), Hainsworth Shin (FDA Sponsor; CDRH/OSEL/DBCMS), Katherine Vorvolakos (FDA Consultant; CDRH/OSEL/DBCMS), and Anuhya Gottipati (Scholar; University of Kentucky) are quantifying the protection afforded by hydrogel coatings to the integrity of individual cells under bioprinting shear. Here, we propose to use a combined tissue engineering, chemical engineering, and cell mechanics approach (using computational simulations) to (1) characterize the cellular impact of complex fluid mechanical force distributions derived from different micronozzle shapes on bioprinted cells and (2) evaluate the impact of single cell encapsulation on the permeability of the cells to leachables during the print process and on preservation of cell integrity and phenotype. The proposed study to evaluate the cellular damage from different (e.g., cylindrical, conical) bioprinter nozzle holds great significance for developing cell-based platforms (e.g., organ-on-a-chip) to facilitate preclinical testing of devices. The results from this study can also be used to develop standardized approaches for reviewing and evaluating the efficacy of bioprinting approaches for medical product applications.