Structured illumination microscopy applications towards liver sinusoidal endothelial cell fenestrations and HIV-1 cell-to-cell transmission

3D structured illumination microscopy (SIM) was used to image pores in liver sinusoidal endothelial cells that separate blood plasma from cells and debris before the plasma encounters the underlying hepatocytes. These structures have previously only been accessible to electron microscopy, but somewhat hampered by its low throughput, sample fixation, and the difficulty to simultaneously label multiple structures for electron and atomic force microscopy. 3D-SIM helped us gain novel insights in the structure and function of these fragile cellular structures. In another application of 3D-SIM, the problem of direct cell-to-cell transmission of HIV-1 via T cell virological synapses was investigated. In this concept, an HIV-1 infected CD4+ T lymphocyte can engage another CD4+ T lymphocyte and form a stable, connective synapse between the cells which promotes targeted assembly of new virus at the point of cell contact leading to rapid passage of large quantities of viral particles into the target cell. Previous studies using a natively fluorescent HIV-1 clone termed HIV Gag-iGFP and spinning disk confocal microscopy allowed seamless tracking of HIV-1 cell-to-cell transmission, but was hindered by the diffraction-limited resolution, which did not allow us to distinguish individual virus . To study this process in finer detail we employed 3D-SIM and gained several structural insights that will be discussed.