Single-organelle visualization tracking natural glycosaminoglycans within mitochondria-lysosome crosstalk for inflammatory homeostasis.

Glycosaminoglycans (GAGs), as natural products with diverse biological activities, play a significant role in regulating inflammatory homeostasis. Nevertheless, the mechanism underlying their intracellular anti-inflammatory properties remains unclear. Herein, we propose a single-organelle visualization tracking framework, leveraging an advanced fluorescent imaging technology combined with labeling methods to dynamically trace the subcellular regulatory mechanisms of GAGs in eliminating inflammatory markers, such as reactive oxygen species (ROS). By utilizing conventional fluorescein isothiocyanate (FITC)-labeled GAGs, we successfully achieved in situ single-organelle visualization of the subcellular localization and intracellular activities of GAGs. Our findings revealed that GAGs enter lysosomes and increase their number and activity, with chondroitin sulfate (CS) exhibiting particularly prominent effects. Significantly, we visually depict that CS-loaded lysosomes selectively cleave ROS-enriched terminal mitochondria, driving mitochondrial fission and reprogramming. These results corroborate that CS regulate mitochondria-lysosome crosstalk to control mitochondrial quality, thereby maintaining intracellular inflammatory homeostasis. Collectively, our work presents an evidence on the single-organelle visualization and regulatory mechanism of GAGs, thereby offering novel perspectives and avenues for researching other natural products.