Browsing by keyword "degradation"
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Inhibition or Stimulation of Autophagy Affects Early Formation of Lipofuscin-Like Autofluorescence in the Retinal Pigment Epithelium CellThe accumulation of lipofuscin in the retinal pigment epithelium (RPE) is dependent on the effectiveness of photoreceptor outer segment material degradation. This study explored the role of autophagy in the fate of RPE lipofuscin degradation. After seven days of feeding with either native or modified rod outer segments, ARPE-19 cells were treated with enhancers or inhibitors of autophagy and the autofluorescence was detected by fluorescence-activated cell sorting. Supplementation with different types of rod outer segments increased lipofuscin-like autofluorescence (LLAF) after the inhibition of autophagy, while the induction of autophagy (e.g., application of rapamycin) decreased LLAF. The effects of autophagy induction were further confirmed by Western blotting, which showed the conversion of LC3-I to LC3-II, and by immunofluorescence microscopy, which detected the lysosomal activity of the autophagy inducers. We also monitored LLAF after the application of several autophagy inhibitors by RNA-interference and confocal microscopy. The results showed that, in general, the inhibition of the autophagy-related proteins resulted in an increase in LLAF when cells were fed with rod outer segments, which further confirms the effect of autophagy in the fate of RPE lipofuscin degradation. These results emphasize the complex role of autophagy in modulating RPE autofluorescence and confirm the possibility of the pharmacological clearance of RPE lipofuscin by small molecules.
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Microglia jointly degrade fibrillar alpha-synuclein cargo by distribution through tunneling nanotubesMicroglia are the CNS resident immune cells that react to misfolded proteins through pattern recognition receptor ligation and activation of inflammatory pathways. Here, we studied how microglia handle and cope with alpha-synuclein (alpha-syn) fibrils and their clearance. We found that microglia exposed to alpha-syn establish a cellular network through the formation of F-actin-dependent intercellular connections, which transfer alpha-syn from overloaded microglia to neighboring naive microglia where the alpha-syn cargo got rapidly and effectively degraded. Lowering the alpha-syn burden attenuated the inflammatory profile of microglia and improved their survival. This degradation strategy was compromised in cells carrying the LRRK2 G2019S mutation. We confirmed the intercellular transfer of alpha-syn assemblies in microglia using organotypic slice cultures, 2-photon microscopy, and neuropathology of patients. Together, these data identify a mechanism by which microglia create an "on-demand" functional network in order to improve pathogenic alpha-syn clearance.
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Sterilization of Polymeric Implants: Challenges and OpportunitiesDegradable and environmentally responsive polymers have been actively developed for drug delivery and regenerative medicine applications, yet inadequate consideration of their compatibility with terminal sterilization presents notable barriers to clinical translation. This Review discusses industry-established terminal sterilization methods and aseptic processing and contrasts them with innovative approaches aimed at preserving the integrity of polymeric implants. Regulatory guidelines, fiscal considerations, and potential pitfalls are discussed to encourage early integration of sterility regulatory considerations in material designs.
