| Golgi apparatus(Golgi) and endoplasmic reticulum(ER) are two important organelles,which play a key role in the modification,classification and transportation of biomacromolecules such as proteins and lipids.The stability of microenvironment(such as polarity and pH) in Golgi and ER maintains their normal physiological functions.Once the microenvironment homeostasis is broken,the protein processing and transportation process will be disturbed,causing abnormal glycosylation or misfolding of the protein,and accumulate in the Golgi and the ER.It is not conducive to the normal function of Golgi and ER.Since proteins regulate a variety of life activities in cells,most diseases are related to abnormal changes in proteins.Therefore,the influence of microenvironment disorders in the Golgi and ER on protein is likely to be an important cause of the development of many diseases.Studies have shown that the polarity and pH value of Golgi and ER often deviate from normal physiological indicators in cancer,diabetes,liver injury,depression and Alzheimer,et al.disease.However,it is not easy to accurately detect changes in these microenvironmental indicators in cells and in vivo.Therefore,accurate,real-time and quantitative monitoring of polarity and pH changes in Golgi and ER are necessary for early diagnosis and timely prevention of diseases.Unfortunately,only a few small molecule fluorescent probes have been reported to detect polarity and pH in Golgi and ER.In this paper,in order to explore the relationship between disease and microenvironment in Golgi and ER,we designed and synthesized two small molecule fluorescent probes for quantitative imaging the polarity of Golgi and the pH of ER.Both probes have good selectivity and biocompatibility,and have been successfully applied to the imaging of Golgi polarity and ER pH in cells or in vivo,respectively.The main contents of this paper are as follows:1.Two photon fluorescence probe has the advantages of low phototoxicity,deep tissue penetration,weak background interference and high resolution.Therefore,we developed a new two-photon ratio imaging probe TP-Golgi for quantitative calibration of Golgi polarity.TP-Golgi uses p-aminobenzene sulfonamide as the Golgi targeting group and indole hemicyanine derivative as the fluorophore.It can be used not only to quantitatively detect the polarity of solution in vitro,but also to sensitively detect the Golgi polarity changes in cell and in vivo.By using TP-Golgi,we found that the polarity of Golgi in cancer cells was lower than that in normal cells.We also successfully detected the increase of Golgi polarity in the kidney of mice with acute kidney injury(AKI) induced by lipopolysaccharide(LPS) and acetaminophen(APAP) using TP-Golgi.In addition,through the use of TP-Golgi,we also found that the excessive use of bleomycin hydrochloride,a drug for cancer such as cervical cancer,can lead to AKI.This work reveals the close relationship between the level of AKI and the increase of Golgi polarity,and provides a new and effective detection parameter for clinical diagnosis of AKI.2.We designed and synthesized a ratiometric fluorescent probe pH-ER that can quantitatively detect pH changes in the ER.The probe pH-ER uses p-toluene sulfonamide as the ER targeting group,indole hemicyanine derivative as the fluorophore,and phenolic hydroxyl group as the pH response group.In vitro experiments showed that pH-ER had a sensitive and reversible ratio response to pH,and it showed different color phenomena in PBS buffer solutions with different pH values,suggesting that it had the potential to develop into pH test paper.Cell experiments showed that pH-ER had good ER targeting,and could be used for quantitative detection of ER pH in cells.We successfully applied the probe to detect the changes of ER pH in cells under ER stress,and the results showed that ER pH decreased to 6.30 during ER stress.Next,we plan to use pH-ER to detect the changes of ER pH during apoptosis,and then further study the pH changes in inflammatory tissues. |
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