Research Projects: Phagocytosis and Therapeutic Delivery


**Page is still under development.**

One goal of cell and tissue engineering is the targeted delivery of therapeutic agents to cells, and toxins to unwanted cells.  Unfortunately, our understanding of phagocytosis, or particle uptake, is situational and empirical, hence difficult to generalize.  Indeed, the kinetics and general phenomenology of phagocytosis are poorly characterized, largely complicated by its speed and unpredictability of occurence.  We have circumvented these limitations by monitoring individual particles being taken up by living cells.  For the macrophage-like cell, Dictyostelium discoideum, laser-tracking microrheology (LTM) reveals that forces during uptake jump >30-fold and are fast (<10s).  Complementary real-time fluorescence measurements provide the opportunity to resolve the signalling events and systematically explore factors that affect particle uptake.  The implications for cell and tissue engineering are obvious.Our studies of pathogenic bacteria provides a powerful synergy to understanding phagocytosis.  Pathogenic bacteria have evolved exquisite mechanisms to enhance their uptake and subsequent vacuolar/lysosomal escape into cytoplasm.1) What are the kinetic events during uptake? Of the signalling molecules involved, which ones precede uptake?2) Which factors enhance particle uptake?3) Can mechanisms of pathogen escape into cytoplasm be incorporated into artificial particles?4) What controls the routing of particles once in the cytoplasm?

We plan to use optical tweezers to micromanipulate both bacteria and particles onto cells, thus precisely initiating particle uptake. Subsequent monitoring using both real-time fluorescence microscopy and LTM will quantify their uptake kinetics and forces. By comparing wild type and mutant bacteria, we should be able to identify the factors that control uptake. Because most candidate proteins have been cloned, we can attach these factors onto particles and determine if their uptake is also enhanced. Listeria, Shigella, and enteropathogenic E. coli all use different mechanisms of entry and would provide complementary information.