While the goal of this project was to understand the mechanisms controlling CIL, we realized that we needed a better understanding of the processes governing normal cell migration. For example, in Aim 1 our goal was to dissect how actin flows are involved in cell repulsion. However, we have little understanding of how these flows are controlled during normal migration, nor do we have good analytical approaches to quantify and describe these flows. We therefore took a step back and developed techniques to understand the biomechanics of actin flows during normal Drosophila macrophage migration with the goal of then extending these new techniques to CIL and cell motility in other cell types. We have now developed novel approaches to visualize and quantify actin dynamics in migrating macrophages using novel image analysis tools and computational packages. This software allowed us to automatically track the speed and direction of the flowing actin network in macrophages as well as mammalian cell types. The enabled us to precisely dissect the regulation of actin dynamics in cells during migration, which resulted in several publications. We have subsequently used these tools to highlight a number of novel regulators of actin flows in Drosophila macrophages, resulting in a manuscript that is currently in preparation.
We have also unveiled an unexpected physiological function of CIL in Drosophila macrophages. We revealed that macrophages are the primary producers of extracellular matrix (ECM) in developing embryos and that these cells need to rapidly and efficiently spread throughout the animal to evenly assemble the ECM. We subsequently revealed that the embryonic ECM is surprisingly dynamic during early development in terms of its self-assembly and its rapid turnover (the ECM was once thought to be incredibly long-lived and stable).
An additional achievement that I would like to highlight is related to extrapolating to model systems and physiological processes outside of flies, which has led to one published manuscript and a second in preparation. First, we developed an approach to easily screen for CIL behaviours in cultured mammalian cells. We discovered that melanoma cells invade through epithelial monolayers upon collision rather than simply ceasing motion, suggesting that loss of CIL may indeed be related to enhanced metastatic capacity. However, we discovered that cells of a second cancer type, fibrosarcoma, showed a robust repulsion response revealing that not all cancer cells lose CIL capacity as was originally assumed. In a second project we investigated the behaviour of fibrotically active fibroblasts (keloids). It is well known that fibrotic fibroblasts produce a highly aligned ECM, which is thought to alter the mechanical properties of the tissue. In a manuscript that is currently in preparation, we revealed that keloid dermal fibroblasts (KDF), unlike normal dermal cells (NDF), develop a highly aligned supracellular network in culture, which subsequently drives the alignment of the ECM (see attached image). We subsequently revealed that KDF cells show an enhanced CIL behaviour which we hypothesise is helping to drive or maintain their alignment. Additionally, we highlighted the cellular mechanisms that allow these fibrotic cells to align, which we hypothesise may allow for clinical intervention.
