3D ECM-inflammation model on a microfluidic chip for neutrophil transmigration from whole blood investigations.

Abstract

Neutrophils have been linked to several inflammation diseases. To study the role of neutrophils in inflammation diseases and conditions, <i>in vitro</i> inflammation assays have been developed. Two drawbacks of these assays include the reliance on pre-processing techniques to isolate neutrophils and 2D migration analysis. These assays limit the physiological relevance of <i>in vivo</i> neutrophil migration which involves other blood components and the transmigration of 3D extracellular matrix-tissue environments. Extracellular matrices regulate neutrophil activation and deformation - important factors in the study of neutrophil migration behavior. To address these limitations, we have successfully created a microfluidic chip that recreates an inflammation event and directly isolates neutrophils from a small volume of whole blood using a 3D extracellular matrix. We optimized our platform by adjusting the extracellular matrix collagen, chemoattractant, and blood concentrations to maximize neutrophil yield. Six individual blood samples showed a range of 30-70 isolated neutrophils per mm² from whole blood with 100% viability and purity using 2 mg mL^-1 extracellular matrix collagen and 150 nM fMLP concentrations. Using this preliminary data, we performed a regression analysis to examine the effect of blood component quantities - white blood cells, red blood cells, neutrophils, and platelets - on the number of isolated neutrophils. The regression analysis revealed that the number of platelets possibly affects the number of transmigrated neutrophils conforming to a non-linear second-degree polynomial function, with an <i>R</i>² of 0.88. Our findings highlight the potential of our platform to facilitate and improve the understanding of neutrophil migration and invasion in inflammation resolution, diseases, and treatments.