Paper BM+MN+MS+TF+BI-ThA9
Microfluidic Models of Endothelial Cell Sprouting in Response to Biomechanical and Biochemical Microenvironments

Thursday, November 12, 2009, 4:40 pm, Room A8

A novel microfluidic device was designed in order to generate stable, quantifiable concentration gradients of biomolecules in a cell culture chamber for 2-D and 3-D studies of shear-sensitive cell types such as endothelial cells. Endothelial cells form the inner lining of blood vessels and initiate a critical step in angiogenesis (the sprouting of new blood vessels) during wound healing and cancerous tumor growth. Therefore, a deeper understanding of the critical biomechanical and biochemical factors regulating endothelial cell sprouting can lead to improved clinical therapies for a multitude of diseases. Concentration distribution of soluble growth factors inside the microfluidic cell culture chamber was determined by simulation and experiment, and the stability of the gradient was verified over multiple hours. This device allows independent tuning of the matrix rigidity, the growth factor absolute concentration, and the growth factor concentration gradient steepness within a single experimental platform. Sprout formation of dermal microvascular endothelial cells was studied within collagen gels of varying density (0.3 - 2.7 mg/mL, corresponding to shear moduli of 8 – 800 Pa) that contained stable gradients of soluble vascular endothelial growth factor (VEGF). These experiments revealed that endothelial sprouting into multi-cellular, capillary-like structures is optimized at an intermediate collagen matrix density (G’~100 Pa). At lower matrix densities, cells were more likely to lose their coordinated motion and migrate as individual cells through the matrix; while at higher matrix densities, the cells formed broad cell clusters that rarely elongated into a sprout. Sprout thickness directly correlated with matrix rigidity, with thicker and less frequent sprouts present in gels with the highest shear moduli. Intriguingly, our 3D experiments also found that endothelial sprouts alter their sensitivity to VEGF depending on the matrix density, suggesting a complex interplay between biochemical and biomechanical factors. As matrix stiffness increases, steeper VEGF gradients and higher VEGF absolute concentrations are required to induce directional sprouting. In more compliant gels, endothelial sprouts that originally misaligned were able to turn and properly reorient parallel to the VEGF gradient; however, this turning phenomenon was only rarely observed in stiffer gels. These results demonstrate that matrix stiffness is an effective factor in stabilization and orientation of endothelial cells during sprouting and suggests new anti-angiogenic strategies for potential cancer treatment as well as pro-angiogenic strategies for regenerative medicine scaffolds.