Lysosomes play important roles in catabolism, nutrient sensing, metabolic signaling, and homeostasis. NPC1 deficiency disrupts lysosomal function by inducing cholesterol accumulation that leads to early neurodegeneration in Niemann-Pick type C (NPC) disease. Mitochondria pathology and deficits in NPC1 deficient cells are associated with impaired lysosomal proteolysis and metabolic signaling. It is thought that activation of the transcription factor TFEB, an inducer of lysosome biogenesis, restores lysosomal-autophagy activity in lysosomal storage disorders. Here, we investigated the effect of trehalose, a TFEB activator, in the mitochondria pathology of NPC1 mutant fibroblasts
This study introduces a rapid, volumetric live-cell imaging technique for visualizing autofluorescent sub-cellular structures and their dynamics by employing high-resolution Fourier light-field microscopy. We demonstrated this method by capturing lysosomal autofluorescence in fibroblasts and HeLa cells. Additionally, we conducted multicolor imaging to simultaneously observe lysosomal autofluorescence and fluorescently-labeled organelles such as lysosomes and mitochondria. We further analyzed the data to quantify the interactions between lysosomes and mitochondria. This research lays the foundation for future exploration of native cellular states and functions in three-dimensional environments, effectively reducing photodamage and eliminating the necessity for exogenous labels.
more » « less- Award ID(s):
- 2145235
- NSF-PAR ID:
- 10434849
- Publisher / Repository:
- Optical Society of America
- Date Published:
- Journal Name:
- Biomedical Optics Express
- Volume:
- 14
- Issue:
- 8
- ISSN:
- 2156-7085
- Page Range / eLocation ID:
- Article No. 4237
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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in vitro and in mouse developmental Purkinje cellsex vivo . We found that in NPC1 mutant fibroblasts, serum starvation or/and trehalose treatment, both activators of TFEB, reversed mitochondria fragmentation to a more tubular mitochondrion. Trehalose treatment also decreased the accumulation of Filipin+cholesterol in NPC1 mutant fibroblasts. However, trehalose treatment in cerebellar organotypic slices (COSCs) from wild-type andNpc1 nmf164 mice caused mitochondria fragmentation and lack of dendritic growth and degeneration in developmental Purkinje cells. Our data suggest, that although trehalose successfully restores mitochondria length and decreases cholesterol accumulation in NPC1 mutant fibroblasts, in COSCs, Purkinje cells mitochondria and dendritic growth are negatively affected possibly through the overactivation of the TFEB-lysosomal-autophagy pathway. -
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null (Ed.)Multicolor carbon dots (CDs) have been developed recently and demonstrate great potential in bio-imaging, sensing, and LEDs. However, the fluorescence mechanism of their tunable colors is still under debate, and efficient separation methods are still challenging. Herein, we synthesized multicolor polymeric CDs through solvothermal treatment of citric acid and urea in formamide. Automated reversed-phase column separation was used to achieve fractions with distinct colors, including blue, cyan, green, yellow, orange and red. This work explores the physicochemical properties and fluorescence origins of the red, green, and blue fractions in depth with combined experimental and computational methods. Three dominant fluorescence mechanism hypotheses were evaluated by comparing time-dependent density functional theory and molecular dynamics calculation results to measured characteristics. We find that blue fluorescence likely comes from embedded small molecules trapped in carbonaceous cages, while pyrene analogs are the most likely origin for emission at other wavelengths, especially in the red. Also important, upon interaction with live cells, different CD color fractions are trafficked to different sub-cellular locations. Super-resolution imaging shows that the blue CDs were found in a variety of organelles, such as mitochondria and lysosomes, while the red CDs were primarily localized in lysosomes. These findings significantly advance our understanding of the photoluminescence mechanism of multicolor CDs and help to guide future design and applications of these promising nanomaterials.more » « less
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