Cell migration in the context of immobilised and soluble cues

Abstract

Cells respond to adhesive and soluble cues in the extracellular matrix by modulating their behaviour including cell adhesion, migration speed and direction. In vivo, cells are exposed to multiple, even conflicting directional cues, however, how cell migration is influenced by multiple spatial cues is still unclear. The effects of adhesive and soluble cues on cell adhesion and migration have often been studied separately. In this thesis, surface-patterning techniques and microfluidics are combined to independently present adhesive and soluble cues. Microcontact printing was used to pattern adhesive cues onto a layer of polyethylene glycol (PEG) that passivated the underlying surfaces. A microfluidic device was employed to deliver a gradient of soluble cues to migrating cells. Migration of Hela cells derived from human cervical cancer and J774 cells, immortalized mouse macrophages, was recorded with live-cell fluorescence microscopy and analysis by single cell tracking. In the first chapter reporting results (Chapter 3), surface passivation and surface patterning of fibronectin via microcontact printing were established and cell migration was observed with a modified wound assay. It is shown that HeLa cells required fibronectin for adhesion and migration and did not migrate into passivated PEG regions of the surface. Only when the cell density on the fibronectin tracks was high did individual HeLa cells migrate into PEG regions. Conversely, J774 cells migrated faster on PEG regions than on fibronectin tracks. The distinct cell morphology indicated that J774 may adopt different migrating modes on fibronectin and PEG surfaces. In Chapter 4, the modified surfaces from Chapter 3 were integrated with a microfluidic device to create a gradient of fetal bovine serum (FBS) or complement 5a (C5a) as a soluble cue for chemotaxis. Hence this chapter describes the role of patterned surfaces and directional cue of soluble gradients on chemotaxis. HeLa cells were found to migrate towards higher concentrations of FBS, but only when a fibronectin track was accessible in that direction. It was concluded that chemotaxis of HeLa cells is strongly adhesion dependent. In contrast, J774 cells migrated faster and more directed towards the source of C5a on fibronectin tracks than on PEG regions, suggesting that chemotaxis of macrophages is enhanced on adhesive cues.In Chapter 5, two competing directional cues were presented to migrating cells. These were (i) gradients of arginine-glycine-aspartic acid (RGD) peptides immobilised onto streptavidin tracks printed on PEG passivated surfaces and (ii) soluble chemoattractant gradient introduced with the microfluidic device used in Chapter 4. When RGD gradients and FBS gradients opposed each other, a greater fraction of HeLa cells was found to migrate towards the source of FBS rather than towards higher concentrations of RGD peptides. Hence for HeLa cells, in the presence of any adhesive cues, soluble cues were dominant in determining the direction of migration. J774 cells on RGD gradients and PEG regions were found to migrate randomly despite the presence of a chemoattractant gradient. In conclusion, combining microcontact printing and microfluidics described in this thesis revealed novel insights of how HeLa and J774 cells integrate multiple spatial cues environment and modulate their migration behaviour accordingly