Creating Novel High-Throughput Microfluidic Platform for Osteocyte Mechanotransduction Studies
Xu, Liangcheng 1 ; You, Lidan 1, 2
1. Institute of Biomaterials & Biomedical Engineering, University of Toronto; 2. Department of Mechanical Engineering, University of Toronto
Bone remodelling is an important process that is responsible for bone growth and recovery from injuries to the bone. Osteocytes are the major bone cells embedded in bone matrix with important mechaosensing functions. Current research has focused on observing bone cell mechanotransduction under different simulated physiological conditions (e.g., shear stress, strain, pressure, etc.) using macro-scale devices. However, these devices often require large sample volumes and extensive setup protocol, as well as very limited designs only suitable for general cell culture. On the other hand, in vitro microfluidic devices provide an optimal tool to better understand this biological process with its flexible design, physiologically-relevant dimensions, cell-medium volume ratio, and high-throughput capabilities. There still lacks a robust system where multi-physiological flow conditions are combined with the high-throughput advantages of microfluidic devices for bone research. In this study, we aim to combine the development of a high throughput microfluidic platform with novel OCY 454 osteocyte-like cell line to create the next generation of in vitro platforms to study osteocyte mechanobiology.
In this project, we are determined to design and fabricate a novel microfluidic device that can induce oscillatory fluid flow (OFF) in a high-throughput manner, with multi-level shear stress for mechanobiology study of osteocytes. Initial design of the device allows for three identical experiments to be run simultaneously. Devices are fabricated using PDMS, and initial testing with OFF showed similar RANLK protein expression levels from all three channels. The experiment also demonstrated that concentration of excreted RANKL is detectable using standard ELISA kits despite having to dilute the initial sample solution due to limited volumes. This prototype device will provide experimental foundation for future devices with varying shear stresses. Currently, our experiments in traditional macro-scale flow chambers have validated that differentiated OCY454 cells excrete significantly more sclerostin than undifferentiated cells in response to mechanical loading.
Design of future devices will be based on common microfluidic principles and utilize channel resistance as the varying factor to adjust for different shear stress. By connecting multiple channels with different path length to a common perfusion source, each channel experiences a shear stress dependent solely on its own geometry. Preliminary experimental results using devices built with this design concept shows differential RANKL expression from different-length channels connected to the same inlet.
Ultimately, the novel high-throughput platform developed in this study can be used to measure extracellular sclerostin levels of OCY454 and establish it as an ideal cell line for in vitro osteocyte studies, as well as help deepen the understanding of mechanical stimulus and their role in bone diseases.