Construction Of Rare Earth Upconversion Fluorescence Nanoplatform And Its Photodynamic Targeted Synergistic Therapy For Liver Cancer Cell

Objective To synthesize a nanoplatform composed of lanthanide-doped upconversion nanomaterials modified with polyethyleneimine(PEI)connected with the photosensitizer dihydroporphyrin E6(Ce6)and hepatocellular carcinoma-specific targeting antibody(anti-Ep CAM),and to explore the feasibility of its application in the field of hepatocellular carcinoma treatment by the synergistic effect of near-infrared-excited photodynamic therapy(PDT)and targeted therapy.Methods:(1)lanthanide-doped upconversion nanoparticles Na YF4:20%Yb,2%Er-PEI(UCNPs)coated on the surface of PEI were first synthesized by hydrothermal method using rare-earth chloride and ammonium fluoride as the main raw materials,hydroxyethylcellulose as crystal growth inhibitor,and polyethyleneimine(PEI)with abundant amino groups as the surface modifier in one step.(2)Then UCNPs-Ce6 and anti-Ep CAM-UCNPs-Ce6 were further synthesized by using EDC/NHS as a carboxyl activator to promote the amide bonding between the photosensitizer dihydroporphyrin E6(Ce6),the epithelial cell adhesion molecule anti-Ep CAM,and UCNPs.(3)The above nanoparticles were characterized by systematic analysis using XRD,TEM,IR,zeta potential,PL,UV-Vis,etc.(4)Toxicity of the synthesized nanoparticles was assessed using cck8 kit.(5)Observe the specific targeting ability of the above nanoparticles on hepatocellular carcinoma cells using fluorescence inverted microscopy,Image J fluorescence semi-quantitative analysis,etc.(6)Evaluate the in vitro therapeutic effect of the synthesized nanoparticles in photodynamic under NIR light irradiation using cck8 kit.Results:(1)XRD and TEM showed that the above three nanoparticles were hexagonal structures with high upconversion fluorescence efficiency,based on β-Na YF4 isomerization,and the nanoparticle sizes were all less than 32 nm.(2)The IR patterns of UCNPs-Ce6 and anti-Ep CAM-UCNPs-Ce6 showed the characteristic absorption peaks of amide N-H bending vibration,P=O stretching vibration and P-O-C stretching vibration,respectively,while their zeta potentials gradually decreased with the surface modification of Ce6 and anti-Ep CAM,indicating that both Ce6 and anti-Ep CAM were modified with The surface modification of Ce6 and anti-Ep CAM was successfully modified by amide bonding to the nanoparticle surface with good chemical stability.(3)The aqueous solutions of the above three nanoparticles produced visible fluorescence at the wavelengths of 522 nm,541 nm and 657 nm under 980 nm IR laser excitation,and their emission spectra were similar to the upconversion fluorescence spectra of Na YF4:Yb,Er reported in many literatures,but the fluorescence intensity gradually decreased with the coating of Ce6 and anti-Ep CAM,which further confirmed the effect of Ce6 and anti-Ep CAM on the surface of UCNPs were successfully modified.(4)UV-Vis test showed that the loading of Ce6 on the surface of the nanoion was about2.9% and they could be excited by upconversion visible fluorescence to generate reactive oxygen species.(5)In vitro cell experiments confirmed the biological safety of the synthesized nanoparticles,and the good specific binding ability of the coupled hepatocellular carcinoma-targeting antibody nanoplatform and hepatocellular carcinoma cells was verified by inverted fluorescence microscopy and Image J fluorescence semi-quantitative analysis.(6)With the increase of anti-Ep CAM-UCNPs-Ce6 concentration under 980 nm NIR laser irradiation,good photodynamic therapeutic effect was demonstrated at the cellular level.(7)Conclusion:A lanthanide-doped upconversion nanoplatform with simultaneous modification of Ce6 and anti-Ep CAM was successfully constructed,which had a small particle size,the better dispersion,water solubility and chemical stability,and the better biocompatibility;its surface had an appropriately sized surface charge,which both avoids nanoparticle agglomeration and gives it the ability to enter cells while reducing the damage to cell membranes;The platform combines the upconversion fluorescence property of UCNPs,the photosensitizing activity of Ce6 and the specific antibody effect of anti-Ep CAM,which makes it potentially promising and translatable in the field of NIR PDT and targeted synergistic therapy.