Fluorescent Probes Used to Discover to Enzymes in Lysosomes

A recent study from scientists at Zhejiang Sci-Tech University in Hangzhou, China studied how hypochlorous acid (HOCl), an enzyme that can be found in lysosomes, can be found using fluorescent probes. Their findings were later published in Spectrochima Acta Part A: Molecular and Biomolecular Spectroscopy (1).

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Lysosomes are acidic organelles within eukaryotic cells that contain vast amounts of highly catalytic digestive enzymes in mildly acidic microenvironments. They act as cellular degradation centers, primarily recycling damaged organelles and digest substances like nucleic acids, polysaccharides, lipids, and proteins. Furthermore, scientists are beginning to discover that lysosomes can be critical in various pathological processes, such as neurodegenerative diseases, inflammation, and cancer, while influencing key cellular physiological processes. The concentration and distribution of active molecules in lysosomes (enzymes, reactive nitrogen species, reactive oxygen species [ROS]) can significantly impact lysosomal biological functions, including hypochlorous acid (HOCl), one of the most important endogenous ROS involved in various physiological and pathological processes.

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HOCl, which primarily resides in the acidic lysosomes of phagocyes, is vital to helping the immune system’s defense against invading germs and harmful microorganisms. If its concentration is disrupted, various diseases, such as cardiovascular disorders, neurodegeneration, and inflammation, can occur. As such, real-time tracking of HOCl content in lysosomes can be critical to researching the physiological and pathological processes of lysosomes, as well as for the early diagnosis of diseases associated with HOCl.

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For this study, the scientists created a bodipy-based fluorescent probe, which, in addition to using bodipy fluorescent dyes, was derived from pyridine and phenyl selenide, for specifically detecting HOCl in aqueous solutions.

The scientists then leveraged the probe’s sensitive photoinduced electron transfer effect from phenyl selenide to the fluorophore, which then exhibited satisfactory high sensitivity to hypochlorous acid. Specifically, the process had a limit of detection (LOD) of 5.2 nM and a response time of 15 seconds. Further biological experiments showed that introducing pyridine moiety enabled the probe molecule to selectively target lysosomes. Overall, the probe was successful in facilitating real-time HOCl monitoring in cell models stimulated by N-acetylcysteine (NAC) and lipopolysaccharide (LPS), in addition to zebrafish models. This new method can be a universal means of dynamically sensing HOCl in lysosomes.

Fluorescent probes have been used in various purposes related to spectroscopy, such as mercury detection. In a recent study from the same journal, Young-A Son of Chungnam National University used fluorescent probes to detect and quantify mercury ions, which proved beneficial for environmental and health monitoring (2). Scientists have also used fluorescent sensors to detect permanganate ions in water, which can lead to toxicity and harm aquatic life if left unchecked (3).

Bodipy fluorescent dyes are widely used in fluorescence imaging because they have excellent photophysical properties; this includes high molar extinction coefficient, high quantum yield, wavelength tunability, and good biocompatibility. These dye molecules have been used in various fields, such as ion probes, fluorescent probes for biological imaging research, and photovoltaic power generation (4).

References

(1) An, K.; Fan, J.; Lin, B.; Han, Y. A Lysosome-Targeted Fluorescent Probe for Fluorescence Imaging of Hypochlorous Acid in Living Cells and In Vivo. Spectrochim. Acta Part A: Mol. Biomol. Spectrosc. 2024, 316, 124316. DOI: 10.1016/j.saa.2024.124316

(2) Novel Fluorescent Probe Unveiled for Mercury Detection. MJH Life Sciences 2024. https://www.spectroscopyonline.com/view/novel-fluorescent-probe-unveiled-for-mercury-detection (accessed 2024-5-10)

(3) Wetzel, W. Novel Fluorescent Sensor Revolutionizes Detection of Permanganate Ions in Water. MJH Life Sciences 2024. https://www.spectroscopyonline.com/view/novel-fluorescent-sensor-revolutionizes-detection-of-permanganate-ions-in-water (accessed 2024-5-10)

(4) Introduction of BODIPY Fluroescent Dyes. BOC Sciences 2022. https://www.bocsci.com/blog/introduction-of-bodipy-fluorescent-dyes/ (accessed 2024-5-9)