Microcontact printing of P-selectin increases the rate of neutrophil recruitment under shear flow.

The local variation of P-selectin expression on inflamed endothelial layers affects leukocyte recruitment in vivo. As an initial study of the spatially heterogeneous presentation of P-selectin in vitro, the influence of microcontact printing (microCP) of P-selectin on a planar surface in neutrophil recruitment was investigated using a parallel-plate flow chamber. Microline patterned and nonpatterned P-selectin were prepared using PDMS, Poly(dimethylsiloxane) (PMDS) stamps and isolated neutrophils perfused over the surface to quantify the level of neutrophil recruitment. We first found a significant increase in cell rolling flux and a decrease in cell rolling velocity on the microcontact printed P-selectin-surfaces compared with a randomly adsorbed P-selectin-surface. However, the increase in rolling adhesion under shear on the surfaces prepared by microCP was not proportional to the number of functional sites of P-selectin transferred using immunofluorescent labeling. Interestingly, the relative immunofluorescent intensities of the nonfunctional regions of microcontact printed P-selectin-surfaces were substantially lower than that that of randomly adsorbed P-selectin. Taken together, these data indicate that the microCP of selectin increases the transfer rate of the adhesion molecule on a surface in the functionally correct orientation and, consequently, improves the recruitment of leukocytes to the selectin surface under flow. It is concluded that microCP may be a more general technique to control protein orientation on a substrate.