Two-Photon Fluorescence Imaging Of Malondialdehyde And Formaldehyde In The Brains Of Depressed Mice

Depression is a common mental disease,which seriously affects the physical health and daily life of patients and causes great mental and economic burden to the family and society.However,due to the complex pathogenesis of depression,there is still a lack of effective diagnosis and treatment methods.Therefore,it is necessary to have a detailed understanding of the molecular mechanism of the occurrence and development of depression.Oxidative stress and lipid peroxidation have been shown to play a crucial role in the development of depression.Active carbonyl species（RCS）,as end-products of lipid peroxides,play an important role in a variety of biological events and are closely related to the occurrence and development of a variety of oxidative stress-related diseases.Malondialdehyde（MDA）and Formaldehyde（FA）,as two key active carbonyl species,are involved in regulating gene expression and cell signal transduction within the physiological concentration range.However,high concentration of MDA and FA can cause mitochondrial DNA damage and protein misfolding,interfere with normal functional activities of cells and cause degeneration of neurone,thus leading to the occurrence of a variety of diseases.Therefore,the development of a method that can simultaneously monitor the changes of MDA and FA levels in depression is of great biological significance to relevant molecular mechanisms of depression and the diagnosis and treatment of depression.Due to the advantages of simple operation and real-time imaging,fluorescence imaging technology has become a powerful tool for real-time and in situ study of bioactive molecules in cells and in vivo.In particular,two-photon fluorescence imaging has deeper penetration depth and lower light damage than traditional single-photon fluorescence imaging.Importantly,two-photon fluorescence imaging is more conducive to multi-signal detection in the same region due to its high fixed-point region excitation performance.Therefore,in recent years,many two-photon fluorescence probes have been developed for imaging RCS in cells and in vivo.However,the two-photon fluorescence probe used for simultaneous imaging detection of MDA and FA content changes in cells and living bodies has not been reported.This greatly hinds further exploration of the synergistic regulation of MDA and FA on physiological and pathological processes.Based on the above reasons,this paper designed and synthesized a two-photon fluorescence probe for simultaneous detection of MDA and FA,and realized the simultaneous and in situ fluorescence imaging analysis of endogenous MDA and FA in cells and the brain of depressed mice,and preliminarily explored the synergistic relationship between MDA and FA in pathological process.The main contents of this paper are as follows:1.Synthesis and characterization of a two-photon fluorescence probe for simultaneous detection of MDA and FAA two-photon fluorescence probe TFCH,which can detect MDA and FA simultaneously,was construct.TFCH uses coumarin as the chromogenic fluorophore and hydrazine group as the recognition group,the fat-soluble trifluoromethyl group（-CF₃）is introduced into the fluorophore to facilitate the crossing of the blood brain barrier.TFCH is prone to photoinduced electron transfer（PET）effect and fluorescence quenching due to the presence of active lone pair electrons in the terminal N atom of hydrazine group.After the reaction with MDA,the hydrazine group in the probe structure was condensed,cyclized and dehydrated with MDA to form pyrazole group,emitting strong blue fluorescence at 440 nm.However,after the reaction with FA,the hydrazine group in the probe formed hydrazone with FA,and the PET effect disappeared,emitting strong green fluorescence at 510 nm.Spectral resolvable properties of TFCH after reaction with MDA and FA,simultaneous dichromatic imaging analysis of MDA and FA in cells and in vivo under the same excitation was achieved.2.The content of MDA and FA in the brain of depressed mice were analyzed by two-photon fluorescence imagingThe content changes of MDA and FA and their real-time and synergistic changes in the depression model were further explored.TFCH was used for two-photon fluorescence imaging,and it was found that MDA and FA contents in the depressed model increased compared with the normal group,and the changes of MDA and FA levels are correlated.In vivo brain imaging of depressed mice,the concentration of MDA and FA in the brain of depressed mice was significantly higher than that of normal mice.During the course of depression,the concentration of MDA and FA in the brain of mice increased synergistically.The development of this work could help advance the discovery of potential biomarkers for antidepressants.