Date of Award
microfluidics, Drosophila, neurons, glia, microfabrication, migration
More than 172 million people are influenced by a retinal disorder that stems from either age-related or developmental causes. Of those, 1.5 million people endure a developmental retinal disorder. In the developing retina, neural cells undergo a series of highly complicated differentiation and migration process. A main cause of these diseases is abnormal collective migration of neural progenitors hindering the retinogenesis process. However, our grasp of collective migration and signaling molecules, critical to the developing retina, is incompletely understood. Understanding the molecular mechanisms, such as the fibroblast growth factor pathway, that regulate glial and neuronal migration provides decisive insights in retinal dysfunctions due to abnormal neural (neuronal and glial cell) migration. Since retinal glia did not migrate chemotactically, as individual cells or glial clusters, it is quintessential to examine the collective migration of glial cells in concert with neuronal cells as well. Consequently, additional research efforts are needed to combat retinal dysfunctions from a genetic and cellular standpoint, and to create effective therapy. In this thesis, to observe the underlying problems involved in abnormal collective neural cell migration, an animal model was selected to recreate the retinogenesis process. The animal model of interest is the Drosophila Melanogaster, commonly known as the fruitfly. These invertebrates are very similar to vertebrates in terms of their protein mutations. To emulate the microenvironment of the developing eye, a highly controlled system is required. We developed a novel and efficient microfluidic system called the micro-optic stalk (μOS) that geometrically approximates the microenvironment of the optic stalk in Drosophila Melanogaster. After several iterations, the system was simulated by COMSOL MultiPhysics, created by two-layer photolithography and soft-lithography, and validated with FITC-dextran. The μOS was coated with Concanavalin A (ConA), seeded with 15 dissociated brain complexes from third instar larvae, the prime retina development phase for Drosophila. A fibroblast growth factor-8 (FGF-8) gradient (100ng/mL) was applied and images of the cell migration were taken every 30 seconds for 4 hours. Theresults showed that clusters experienced a gradient dependent system, indicating that as the gradient increased, net displacement increased as well. The small and large clusters had significant increase in average net displacement from low to high gradients of 20.9±5.2 to 79.4±19.2 μm and 26.0±9.6 to 106.2±12.4 μm, respectively. The clusters also expressed a chemotactic relationship with FGF-8 from the average directionality being greater than 0.5. Future work will focus on introducing a knockdown system for FGF receptor (FGFR), a mammalian homolog to Drosophila breathless (btl) gene, to observe the migration of glia- and neuron-specific btl knockdown.
Zhang, Stephanie, "Collective Chemotaxis of Retinal Neural Cells from Drosophila Melanogaster in Controlled Microenvironments" (2018). CUNY Academic Works.