Construction And Application Of MOF Fluorescent Probe For Detection Of ROS And Phosphorylation/pH In Atherosclerosis

Atherosclerosis（AS）is a chronic inflammatory cardiovascular disease,which is one of the main causes of myocardial infarction and stroke.Excessive accumulation of low-density lipoprotein（LDL）in the development of AS triggers the formation of lipid droplets（LDs）,and the level of protein phosphorylation in lipid droplets is closely related to the pathogenesis of AS.Studies have shown that hypochlorous acid（HClO）can promote the formation of AS through several ways.AS inflammation will also be accompanied by the imbalance of acid-base in the body.Therefore,it is of great significance to explore the HClO,protein phosphorylation and pH levels in the occurrence and development of AS disease for the study of early AS pathogenesis and disease diagnosis.Until now,most probes have been designed for individual analytes.However,many important physiological or pathological problems are related to various biological species/parameters.The design of probes that can simultaneously monitor multiple species and study the horizontal changes of multiple biomolecules in the course of disease development and the interaction between molecules is of great significance for the study of disease pathogenesis and disease diagnosis.Based on this,two fluorescent probes PCN@FL and Al-MOF@TCFL were designed and synthesized in this thesis to simultaneously detect HClO/protein phosphorylation and pH/HClO level changes,respectively.Finally,the probes were applied to study the relationship between HClO and protein phosphorylation,pH and HClO levels during the development of early AS mice.The details are as follows:1.We designed a nano-fluorescent probe PCN@FL using dimethylthiocarbamate and Zr（Ⅳ）as active centers,which can recognize both HClO and phosphorylation sites.The addition of HClO led to the cleavage of the spironolactone structure with inhibited fluorescence and the fluorescence recovery,achieving the specific response of the probe to HClO.The coordination between the metal ion Zr⁴⁺in the probe and the ligand weakened the fluorescence of porphyrin.After the addition of phosphate,P in the phosphate formed a coordination bond with Zr⁴⁺,resulting in the weakening of the coordination between Zr and the porphyrin ligand,and the fluorescence recovered,achieving the specific response of the probe to phosphate.In vitro and cell experiments verified the fluorescence luminescence performance of the probe.Then,combining the advantages of two-photon imaging,the prepared nanoprobes were applied to fluorescence detection imaging of HClO and phosphorylation levels in serum and tissues of early AS mice.The results showed that the levels of HClO and protein phosphorylation in early AS mice were significantly higher than those in normal mice,indicating that oxidative stress and protein phosphorylation in early AS were more serious.This study provides a new analysis method for further revealing the level of HClO and protein phosphorylation in early AS.2.Using diaminoterephthalic acid and dimethylthiocarbamate as active centers,we constructed a nano-fluorescent probe Al-MOF@TCFL that can recognize pH and HClO at the same time.Due to the protonation of 2-amino terephthalic acid due to the addition of H⁺,the conjugate system was weakened,thus weakening the fluorescence emission peak of ligand molecules at 430 nm.The specific response of the probe to pH was realized.The addition of dimethyl thiocarbamate,as the recognition unit of HClO,led to the cracking of spironolactone structure with fluorescence inhibition,which led to the fluorescence recovery of lucifcein at 513nm,and realized the specific response of the probe to pH.The morphologic characteristics of the probe were determined by TEM and SEM experiments,and the fluorescence experiments confirmed that the probe had good fluorescence response and selectivity to pH/HClO.Then,we applied the prepared nanoprobe to fluorescence detection imaging of pH and HClO levels in serum and tissue of early AS mice.The experimental results showed that the pH level decreased and the HClO level increased in early AS mice.This study provides a good research method for further revealing the pH and HClO levels in early AS disease and understanding the disease mechanism of AS,and provides a new analytical method for the diagnosis of early arteriosclerosis.