Directional control of cell motility through focal adhesion positioning and spatial control of Rac activation.

Local physical interactions between cells and extracellular matrix (ECM) influence directional cell motility that is critical for tissue development, wound repair, and cancer metastasis. Here we test the possibility that the precise spatial positioning of focal adhesions governs the direction in which cells spread and move. NIH 3T3 cells were cultured on circular or linear ECM islands, which were created using a microcontact printing technique and were 1 microm wide and of various lengths (1 to 8 microm) and separated by 1 to 4.5 microm wide nonadhesive barrier regions. Cells could be driven proactively to spread and move in particular directions by altering either the interisland spacing or the shape of similar-sized ECM islands. Immunofluorescence microscopy confirmed that focal adhesions assembled preferentially above the ECM islands, with the greatest staining intensity being observed at adhesion sites along the cell periphery. Rac-FRET analysis of living cells revealed that Rac became activated within 2 min after peripheral membrane extensions adhered to new ECM islands, and this activation wave propagated outward in an oriented manner as the cells spread from island to island. A computational model, which incorporates that cells preferentially protrude membrane processes from regions near newly formed focal adhesion contacts, could predict with high accuracy the effects of six different arrangements of micropatterned ECM islands on directional cell spreading. Taken together, these results suggest that physical properties of the ECM may influence directional cell movement by dictating where cells will form new focal adhesions and activate Rac and, hence, govern where new membrane protrusions will form.

Combined microfluidic-micromagnetic separation of living cells in continuous flow.

This paper describes a miniaturized, integrated, microfluidic device that can pull molecules and living cells bound to magnetic particles from one laminar flow path to another by applying a local magnetic field gradient, and thus selectively remove them from flowing biological fluids without any wash steps. To accomplish this, a microfabricated high-gradient magnetic field concentrator (HGMC) was integrated at one side of a microfluidic channel with two inlets and outlets. When magnetic micro- or nano-particles were introduced into one flow path, they remained limited to that flow stream. In contrast, when the HGMC was magnetized, the magnetic beads were efficiently pulled from the initial flow path into the collection stream, thereby cleansing the original fluid. Using this microdevice, living E. coli bacteria bound to magnetic nanoparticles were efficiently removed from flowing solutions containing densities of red blood cells similar to that found in blood. Because this microdevice allows large numbers of beads and cells to be sorted simultaneously, has no capacity limit, and does not lose separation efficiency as particles are removed, it may be especially useful for separations from blood or other clinical samples. This on-chip HGMC-microfluidic separator technology may potentially allow cell separations to be carried out in the field outside of hospitals and clinical laboratories.