Highly Efficient Red/Nir-Emissive Fluorescent Probe With Polarity-Sensitive Character For Visualizing Cellular Lipid Droplets And Determining Their Polarity

Lipid droplet is a kind of spherical organelle ubiquitous in eukaryotic cells.The traditional study thought that lipid droplet function is relatively simple,mainly used to store lipid.However,recent studies have shown that lipid droplets play a very complex role in membrane formation,protein storage,signaling and other important cellular processes.And with obesity,fatty liver,diabetes,arteriosclerosis and other diseases closely related.Therefore,the study of lipid droplets has gradually become one of the most active directions in the field of cell biology.Fluorescence imaging is a very powerful tool for observing and studying lipid droplets.However,the existing lipid droplet fluorescence probes are limited,and there are many deficiencies in lipid droplet labeling selectivity,photostability,fluorescence band and polarity sensitivity.Therefore,this paper intends to design and synthesize a new red/near-infrared emission polarsensitive lipid droplet fluorescence probe,and use it for fluorescence imaging to observe lipid droplet and determine its polarity.Specific work contents are as follows:a)A donor-receptor-donor fluorescence probe(LDs-Red)was synthesized and its photophysical properties were characterized in detail.The fluorescence probe consists of benzodithiophene tetroxide in the middle as electron acceptor and diethylamino at both ends as electron donor,forming a donor-acceptor-donor molecular structure.The probe has efficient red/near-infrared emission and significant polarity sensitivity(i.e.,the fluorescence emission wavelength is gradually redshifted with increasing solvent polarity).By density functional theory calculations,it is found that HOMO and LUMO orbitals of the fluorescent probe are partially separated,which leads to a large Stokes shift and significant polarity sensitivity.b)Fluorescent probe LDs-Red is used for fluorescence imaging of cell lipid droplets to observe the distribution of lipid droplets and determine the polarity of lipid droplets through spectral scanning imaging.Fluorescent probe LDs-Red has the advantages of high fluorescence brightness,high lipid droplet staining selectivity and highlight stability,which enable it to achieve high quality 3D confocal imaging.The red light/near infrared emission characteristics of the probe enable it to combine with other short wavelength commercial probes to achieve four-color confocal imaging of living cells.In the spectroscopic scanning imaging experiment,the polarity sensitive emission characteristic of the probe makes it have different emission wavelength in different polar lipid droplets.Therefore,lipid droplet polarity in different cells can be accurately measured using the probe,and it was experimentally confirmed that the lipid droplet polarity in cancer cells(Hep G2,He La and Panc02)was lower than that in normal cells(L929,U251 and HT22).To sum up,a novel polar-sensitive lipid droplet fluorescence probe LDs-Red with high efficiency red/near-infrared emission was designed and synthesized based on donor-acceptor-donor skeleton in this paper.The probe has the advantages of high lipid droplet staining specificity,high light stability and high biocompatibility.High quality3 D confocal imaging of fixed cells and four-color confocal imaging of living cells have been achieved by using the probe.The significant polarity sensitivity of the fluorescence probe enables the quantitative determination of lipid droplet polarity by spectroscopic scanning imaging.Therefore,the fluorescence probe can provide not only the morphology/location information of lipid droplets,but also the chemical polarity information,which is of great significance for the biological study of lipid droplets.