LymphangionChip Experimental Platform Supports Coculture and Signaling Between Lymphatic Endothelial and Mural Cells
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abstract
The pathophysiology of several lymphatic diseases, such as lymphedema, depends on the function of lymphangions that drive lymph flow. Even though the signaling between two main cellular components of a lymphangion, Lymphatic Endothelial Cells (LECs) and Lymphatic Muscle Cells (LMCs), is responsible for crucial lymphatic functions, there are no in vitro models that have included both cell types in a physiologically relevant environment. Here, a fabrication technique (Gravitational Lumen Patterning or GLP) is developed to create LymphangionChip. This organonchip consists of coculture of a monolayer of endothelial lumen surrounded by multiple and uniformly thick layers of muscle cells. The platform allows construction of a wide range of luminal diameters and muscular layer thicknesses, thus providing a toolbox to create variable anatomy. In this device, lymphatic muscle cells align circumferentially while endothelial cells aligned axially under flow, as only observed in vivo in the past. This model further demonstrated a robust sensitivity of this relative alignment of the two cells with respect to the presence or absence of coculture and mechanical forces (shear stress), thus suggesting that LECs and LMCs are biologically and functionally active within the chip. This system successfully characterizes the dynamics of cell size, density, growth, alignment, and intercellular gap due to coculture and shear. The timedependent decrease in the subendothelial gap strongly suggests proactive LECLMC signaling as LECs and LMCs grow and proliferate within the device. Further exposure to proinflammatory cytokines reveals that the device could produce the regulation of endothelial barrier function through the lymphatic muscle cells. Finally, measurement of intracellular calcium of LECs and LMCs while exposed to step and oscillatory flow profiles with various shear amplitudes revealed the bidirectional mechanism of calcium signaling within lymphatic vessel. Therefore, this organchip technology allows researchers to include essential lymphatic vascular components in a tunable 3D microphysiological system that is suitable for use in preclinical research of lymphatic and blood mechanobiology, inflammation, and translational outcomes.
