Development of Targetable Two-Photon Fluorescent Probes to Image Hypochlorous Acid in Mitochondria and Lysosome in Live Cell and Inflamed Mouse Model

Abstract

Hypochlorous acid (HOCl), as a highly potent oxidant, is well-known as a key "killer" for pathogens in the innate immune system. Recently, mounting evidence indicates that intracellular HOCl plays additional important roles in regulating inflammation and cellular apoptosis. However, the organelle(s) involved in the distribution of HOCl remain unknown, causing difficulty to fully exploit its biological functions in cellular signaling pathways and various diseases. One of the main reasons lies in the lack of effective chemical tools to directly detect HOCl at subcellular levels due to low concentration, strong oxidization, and short lifetime of HOCl. Herein, the first two-photon fluorescent HOCl probe (TP-HOCl 1) and its mitochondria- (MITO-TP) and lysosome- (LYSO-TP) targetable derivatives for imaging mitochondrial and lysosomal HOCl were reported. These probes exhibit fast response (within seconds), good selectivity, and high sensitivity (<20 nM) toward HOCl. In live cell experiments, both probes MITO-TP and LYSO-TP were successfully applied to detect intracellular HOCl in corresponding organelles. In particular, the two-photon imaging of MITO-TP and LYSO-TP in murine model shows that higher amount of HOCl can be detected in both lysosome and mitochondria of macrophage cells during inflammation condition. Thus, these probes could not only help clarify the distribution of subcellular HOCl, but also serve as excellent tools to exploit and elucidate functions of HOCl at subcellular levels.

Lighting the Way to See Inside Two-Photon Absorption Materials: Structure–Property Relationship and Biological Imaging

“…There is now a strong demand for efficient 2PA dyes for a wide range of applications, including microscopy [4,5,6,7,8,9,10,11,12,13], micro-fabrication [14,15,16], three-dimensional data-storage [17,18], up-converted lasing [19,20], and photodynamic therapy [21,22,23]. Recently, this demand has been matched by rapid advances in the design and synthesis of 2PA dyes [24,25,26,27,28,29,30,31,32,33,34].…”

A lysosome-targeted and ratiometric fluorescent probe for imaging exogenous and endogenous hypochlorous acid in living cells

“…41 Compared with nanoprobe, small molecule fluorescent probes are much easier in membrane permeable and diffuse into cells, and then, in 2015, three small molecules based, lysosome-targeted fluorescent probes were reported with a turn-on response model. [42][43][44] However, to the best of our knowledge, there is no report yet describing a small molecule-based ratiometric fluorescent probe for successful detection of intracellular HOCl in lysosome. Therefore, it is urgent to develop a new effective small molecule-based and ratiometric response model fluorescent probe for imaging endogenous and exogenous HOCl in lysosomes with good membrane permeable, fast response and excellent biocompatibility.…”

A specific fluorescence probe for hypochlorous acid based on rhodamin101 thiosemicarbazides and its application for living cells imaging

“…In addition, in conjunction with microscopy, bio-imaging applications can be used in living cells. Based on the reported ClO -probes [5][6][7][8][9] , they have the drawbacks like unsuitable pH range for cells [7,9] , lack of selectivity between ClO -and Cu…”