IR780 Loaded Hollow HMnO₂ Nanoparticles For Dual-mode Imaging And Enhanced Photodynamic Therapy Of Oral Squamous Cell Carcinom

PurposeA kind of hollow mesoporous HMnO₂multifunctional nanoparticles loaded with IR780 was prepared,and its general properties were characterized.The bimodal imaging ability in vitro,the effect of oxygen enrichment in vitro,the uptake ability of tongue cancer squamous cell and the photodynamic killing ability of tongue cancer cells in vitro were evaluated.Content and resultIn the first part,hollow mesoporous MnO₂nanoparticles（HMnO₂-IR780）coated with photosensitizer IR780 were prepared.Firstly,hollow mesoporous manganese dioxide nanoparticles（HMnO₂）were prepared by inverse microemulsion method,and then loaded with photoacoustic imaging and fluorescence imaging photosensitizer IR780,to obtain self-oxygen-producing photodynamic therapy nanoparticles（HMnO₂-IR780）.The appearance,dispersion and structure of the nanoparticles were observed by optical microscope and transmission electron microscope,and the particle size and surface potential were detected by Malvern particle size meter.The ultraviolet absorbance of nanoparticles（HMnO₂-IR780）,HMnO₂and IR780 were determined by ultraviolet spectrophotometry,which proved the successful entrapment of photosensitizer IR780.The experimental results show that the prepared nanoparticles are regular and well dispersed and spherical.Malvern particle size analyzer shows that theaverage diameter of HMnO₂-IR780 nanoparticles is about 242.7±22.2nm,which can pass through capillaries,which is a suitable size for tumor aggregation through EPR effect.The average zeta potential of HMnO₂-IR780 nanoparticles is-38.9±0.9 m V.This shows that HMnO₂-IR780 nanoparticles have considerable stability and are beneficial to the later in vivo photodynamic therapy.By comparing the UV-vis spectra of IR780,HMnO₂and HMnO₂-IR780,it is proved that IR780 is successfully supported on HMnO₂nanostructures.The second part confirmed the ability of multifunctional nanoparticles to promote photodynamic therapy of tumor hypoxia inhibition in vitro,and evaluated the photoacoustic imaging and fluorescence imaging effects of nanoparticles in vitro.The dissolved oxygen in solution was measured by portable dissolved oxygen meter and the fluorescence intensity of ROS was detected by SOSG（singlet oxygen detection kit）.The ability of HMnO₂-IR780NPs to produce oxygen and ROS in the presence of H₂O₂was verified,and it was preliminarily proved that HMnO₂-IR780NPs can enhance the ability of photodynamic therapy of tumor hypoxia inhibition.The photoacoustic imaging model was established and the photoacoustic images of nanoparticles at different concentrations were detected by photoacoustic imager excited by pulsed laser with wavelength 780nm.The corresponding photoacoustic signal values（PA）;quantitative fluorescence and bioluminescence imaging system（IVISlumina）were used to observe the fluorescence imaging of nanoparticles at different concentrations.The photoacoustic signal of HMnO₂-IR780 nanoparticles increased with the increase of the concentration of nanoparticles,and at the concentration of2.Very obvious photoacoustic signals can be detected in 00mg/ml.The fluorescence signal of nanoparticles also increased with the increase of nanoparticles concentration（P<0.05）.The third part is to study the effect of HMnO₂-IR780 nanoparticles on human tongue squamous cell carcinoma.HMnO₂-IR780 nanoparticles were labeled with fluorescent dye Di I,and the nuclei of HN-6 cells（human tongue squamous cell carcinoma cells）were stained with DAPI.The phagocytosis of HMnO₂-IR780 nanoparticles was observed by laser confocal microscope.The results showed that HMnO₂-IR780 nanoparticles could be absorbed by HN-6 cells,and the uptake reached the peak at about4 h with the increase of time.After the photosensitizer was excited by laser irradiation,the photodynamic killing effect of HMnO₂-IR780 nanoparticles on HN-6 was observed by living dead cell double staining kit and laser confocal microscope.The results showed that HMnO₂-IR780 nanoparticles had good phototoxicity and high biocompatibility,which proved that HMnO₂-IR780 nanoparticles was an effective photodynamic therapy nanoparticles for tongue squamous cell carcinoma.ConclusionIn summary,we have successfully prepared a multifunctional nano-platform consisting of hollow MnO₂containing IR780 for OSCC photoacoustic and fluorescent molecular imaging and oxygenated photodynamic therapy.In vitro experiments show that HMnO₂-IR780NPs has the ability to catalyze H₂O₂to produce oxygen and then increase the yield of ROS.It has high fluorescence intensity and strong PA signal,which shows the potential to become an excellent PA/FL contrast agent.In addition,HMnO₂-IR780 NPs not only improves the stability of IR780and prolongs its blood circulation,cell experiments show that HMnO₂-IR780 NPs can be well absorbed by cells,and HMnO₂-IR780 NPs can effectively kill HN-6 cells after laser irradiation.This study provides more inspiration for overcoming tumor hypoxia,improving the clinical effect of OSCC therapy and OSCC nano-imaging diagnosis.