Superoxide Anion Two-photon Fluorescence Imaging In Organelles

Reactive Oxygen Species(ROS)participate in a series of physiological and pathological processes,including cell signaling,cell death,aging and so on.Superoxide anion(O2·-)is the first produced ROS,the precursor of other ROS.The changing of O2·-concentration is connected to many biological processes,as well as the onset and progress of related diseases.Hence,it is of great importance to monitor the O2·-level in live cells and in vivo.In live cells,different kinds of organelles are cooperating with each other to finish the metabolisms.Mitochondria is the main site of O2·-production,to study the O2·-concentration in mitochondria will be useful for uncovering the effect of O2·-in connected processes.Proteins are mainly synthesized and modified in endoplasmic reticulum(ER),O2·-is produced in the reduction and oxidation of proteins.However,the connection between O2·-level and ER related diseases is still unknown.The development of effective monitoring methods of O2·-in organelles is of great importance.Fluorescence imaging offers a powerful method to monitor the levels of active molecules in cell,owing to its high sensitivity and selectivity.In recent years,Two-Photon Excited Fluorescence Microscopy(TPM),is developing fast.TPM displays distinguished advantages including increased penetration depth and reduced specimen photodamage.It is urgent to design and synthesize two-photon fluorescence probes with larger two-photon absorption across section,high stability,sensitivity,selectivity,and organelles location.For these reasons,two new organelle-targeting two photon fluorescent probes for O2·-are designed and synthesized:ER-BZT and Mito-CA.In this paper,the main contents were shown as follows:1.A new endoplasmic reticulum-targeted two-photon fluorescent probe termed ER-BZT is designed and synthesized for imaging of O2·-.The probe ER-BZT shows high sensitivity,selectivity,stability,and low cytotoxicity.The rise of O2·-levels in endoplasmic reticulum induced with different stimuli is visualized by one-and two-photon fluorescence imaging.By utilizing ER-BZT,the two-photon fluorescence imaging results demonstrate that the endogenous O2·-concentration in abdominal or hepatic tissue of diabetic mice is higher than that in normal mice.2.A new mitochondria-targeted two-photon fluorescent probe termed Mito-CA is designed and synthesized for imaging of O2·-The probe consists of two moieties:the potent two-photon fluorophore and O2·-recognition unit caffeic acid;the mitochondria-targetable moiety triphenylphosphine.