Quantitative analysis of immune cell motility and mechanics on hydrogel substrates

Cell migration is central to the development and regulation of immune function, yet the biophysical and molecular basis of motility and force generation in immune cells is not well understood. Previous studies examining immune cell behavior have either been conducted in vivo or in non-physiologic tissue culture systems such as glass or plastic. Few studies have been completed to date to investigate the behavior of immune cells on compliant hydrogel surfaces that more closely resemble in vivo environments. The use of hydrogel substrates offers a number of advantages in the investigation of immune cell behavior as the mechanical and chemical properties of hydrogels are easily controlled and quantitative information on cell behavior can be obtained. In this thesis, we develop a novel system to enable chemical gradients to be generated over hydrogel surfaces using microfluidic technology. Using this system, we investigate the motility and force generation of neutrophils, a key cellular component of the innate immune response. Through systematic studies of neutrophil motility on compliant hydrogel substrates, we develop a quantitative understanding of how neutrophils respond to chemical and mechanical cues. We find that neutrophil adhesion, migration and force generation are influenced by both chemical and mechanical properties of their surrounding matrix. In response to changes in substrate mechanics, neutrophils can rapidly modulate their adhesive area, directionality, and traction forces. We are the first to show that traction force generation can influence neutrophil polarity and the response to chemotactic signals. Lastly, using inhibitors and in vivo mouse models, we show that both integrin and RhoA signaling are implicated in force generation in neutrophils