Tracking and recording of intracellular oxygen concentration changes in cell organelles: preparation and function of azide-modified fluorescent probes

Abstract

Tracking hypoxic environments and changes in oxygen levels contribute to the elucidation of pathological mechanisms. In this study, we attempted to design molecular probes that can be activated to show fluorescence under hypoxic conditions and that can move to specific cell organelles. Considering that azide groups were selectively reduced to primary amines by reductases under hypoxic conditions, we prepared Hoechst and fluorophore Cy-5 derivatives with azide groups (Hoechst-N₃ and Cy-N₃) as hypoxia probes. Hoechst-N₃ and Cy-N₃ showed weak fluorescence, but once activated in the cytosol of hypoxic cells, they exhibited robust fluorescence and then moved to their target organelles, the cell nucleus and mitochondria. In addition, when these probes were administered to the cells in the proper sequence, each probe was activated in response to the intracellular oxygen concentration at that point and exhibited oxygen concentration-dependent fluorescence at the target organelle. By measuring the fluorescence intensity of the cell nucleus and mitochondria, we successfully traced the history of changes in intracellular oxygen levels. Thus, we achieved tracking and recording of oxygen status in the cells.