Understanding adeno-associated virus (AAV) trafficking is critical to advance our knowledge of AAV biology and exploit novel aspects of vector development. Similar to the case for most DNA viruses, after receptor binding and entry, AAV traverses the cytoplasm and deposits the viral genome in the cell nucleus. In this study, we examined the role of the microtubule (MT) network in productive AAV infection. Using pharmacological reagents (e.g., nocodazole), live-cell imaging, and flow cytometry analysis, we demonstrated that AAV type 2 (AAV2) transduction was reduced by at least 2-fold in the absence of the MT network. Cell surface attachment and viral internalization were not dependent on an intact MT network. In treated cells at 2 h postinfection, quantitative three-dimensional (3D) microscopy determined a reproducible difference in number of intracellular particles associated with the nuclear membrane or the nucleus compared to that for controls (6 to 7% versus 26 to 30%, respectively). Confocal microscopy analysis demonstrated a direct association of virions with MTs, further supporting a critical role in AAV infection. To investigate the underling mechanisms, we employed single-particle tracking (SPT) to monitor the viral movement in real time. Surprisingly, unlike other DNA viruses (e.g., adenovirus [Ad] and herpes simplex virus [HSV]) that display bidirectional motion on MTs, AAV2 displays only unidirectional movement on MTs toward the nuclei, with peak instantaneous velocities at 1.5 to 3.5 μm/s. This rapid and unidirectional motion on MTs lasts for about 5 to 10 s and results in AAV particles migrating more than 10 μm in the cytoplasm reaching the nucleus very efficiently. Furthermore, electron microscopy analysis determined that, unlike Ad and HSV, AAV2 particles were transported on MTs within membranous compartments, and surprisingly, the acidification of AAV2-containing endosomes was delayed by the disruption of MTs. These findings together suggest an as-yet-undescribed model in which after internalization, AAV2 exploits MTs for rapid cytoplasmic trafficking in endosomal compartments unidirectionally toward the perinuclear region, where most acidification events for viral escape take place.

Our data demonstrate that ITF(l) but not ITF(s) delays the development of T1D via modulation of gut-pancreatic immunity, barrier function, and microbiota homeostasis.

He, T. (2015). A sacrificial-layer approach to fabricate polysulfone support for forward osmosis thin-film composite membranes with reduced internal concentration polarisation. Journal of Membrane Science, 481 106-114.A sacrificial-layer approach to fabricate polysulfone support for forward osmosis thin-film composite membranes with reduced internal concentration polarisation AbstractThis study demonstrates a sacrificial-layer approach by co-casting, which is the simultaneous casting of two layers, to prepare a polysulfone support (denoted as PSfco) layer with open bottom surface morphology for fabricating thin-film composite forward osmosis (FO) membranes. In the co-casting process, polyetherimide (PEI), used as the sacrificial layer, was co-cast beneath the PSf layer. After the PEI layer was peeled off, PSfco was yielded with an open-bottom structure. Results showed that under the same operating condition, the FO membrane prepared by co-casting (denoted as PSfco-TFC) demonstrated a 10% higher water flux using 0.5 M NaCl draw solution and 30% higher water flux using 4 M NaCl draw solution in the AL-FS mode in comparison to membranes prepared in a single layer casting technique (denoted as PSfs-TFC). The PSfco-TFC exhibits a lower average structural parameter (S, 167 μm) than that of the PSfs-TFC (241 μm), while the water and salt permeability coefficients of both membranes are similar. Results reported here demonstrate that the co-casting technique can be used to fabricate FO membranes with significantly improved performance compared to the conventional approach. Results showed that under the same operating condition, the FO membrane prepared by cocasting (denoted as PSf co -TFC) demonstrated a 10% higher water flux using 0.5 M NaCl draw solution and 30% higher water flux using 4 M NaCl draw solution in the AL-FS mode in comparison to membranes prepared in a single layer casting technique (denoted as PSf s -TFC).The PSf co -TFC exhibits a lower average structural parameter (S, 167 microns) than that of the PSf s -TFC (241 microns), while the water and salt permeability coefficients of both membranes are similar. Results reported here demonstrate that the co-casting technique can be used to fabricate FO membranes with significantly improved performance compared to the conventional approach.