Design,Synthesis And Imaging Application Of Fluorescent Probes For Monitoring Of Apoptosis

As an effective way to maintain the number of cells,apoptosis plays a very important role in maintaining the balance of biological system.Studies have found that the human body produces billions of cells every day,and in order to maintain the balance in the body,an equal number of cells die and are devoured by phagocytes.As a bioactive sulfur substance,sulfur dioxide is involved in various physiological and pathological pathways.Peroxynitrite(ONOO~-)is an active oxygen species,which plays a very important role in both physiology and pathology.Fluorescent probes are used in detecting apoptosis because of their high specificity and sensitivity.The construction of fluorescent probes for apoptosis is mostly monochrome fluorescent probes,and there are few two-color fluorescence probes.Monochrome fluorescent probes often have false positive results and uneven staining,which limits the detection.Fluorescent probes for the detection of apoptosis are very common,while fluorescence probes for the detection of ONOO~-and apoptosis are very few,which makes it more difficult to study the association between apoptosis and ONOO~-.Similarly,there are many kinds of fluorescence probes that respond to apoptosis,sulfur dioxide and ONOO~-separately,but the fluorescence probes that monitor all three at the same time are rarely reported.Based on the above research status of fluorescent probes,three kinds of fluorescent probes were constructed in this paper.The specific contents are as follows:(Ⅰ)Due to the great polarity difference between mitochondrial inner membrane and RNA,two polar response probes CMST and CMSQ can be constructed according to the above characteristics.Coumarin was selected as the polar response fluorophones in the probes.A cationic thiazole salt derivative was modified based on the coumarin structure of the CMST probe,and a cationic quinoline salt derivative was modified by the CMSQ probe.Piperazine was selected as the binding group for both probes.Spectroscopic experiments showed that both probe CMST and probe CMSQ can response to polarity,RNA and viscosity.Meanwhile,in biological imaging,the probe CMST targeted mitochondria in living cells,and showed obvious fluorescence signals in both blue and red channels.After apoptosis,the probe moved to the nucleus and only emitted red light,which realized the two-color monitoring mode of apoptosis.(Ⅱ)Using the FRET and PET mechanisms,a novel fluorescence probe,NIML,was constructed based on a naphthalanhydride fluorophore,which was then combined with an indole salt via a piperazine connector.The carbon-carbon double bond in the fluorescent probe can be used as the response site of sulfur dioxide and ONOO~-at the same time,and different emission peaks were generated after the reaction,which can be effectively distinguished.The spectral results showed that NIML can response to both viscosity and RNA.At the same time,the fluorescence signal of NIML was significantly changed in apoptosis-related imaging experiments.(Ⅲ)In this project,two probes,namely probe NAI and probe NABI,were designed to respond to sulfur dioxide,ONOO~-,viscosity and RNA.The structure of the two probes was very similar,and both were constructed on the basis of 6-dimethylamino-2-naphthalene formaldehyde.Probe NAI modified an indole salt on the group;NABI modified a benzoindole salt.Because the structural design of probe NAI and probe NABI can distinguish the situation after the reaction of sulfur dioxide and ONOO~-,respectively.In addition,NABI can perform imaging observation on cell apoptosis.