Study On The Preparation Of Membrane Bionic Polypyrrole Coated Fe₂O₃ NPs And The Photothermal Treatment Of Prostate Cancer Guided By Dual-mode Imaging

Objective:Prostate cancer is the most common malignant tumor of male urogenital system.Compared with traditional cancer treatment methods（such as radiotherapy and chemotherapy）,photothermal therapy（PTT）has shown great prospect in tumor ablation,and it has become a research focus to develop an integrated diagnosis and treatment nano platform with high targeting and high photothermal conversion efficiency.Therefore,we have developed biomimetic cell membrane-modified Fe₂O₃nanoclusters implanted in polypyrrole（CM-LFPP）,achieving photoacoustic/magnetic resonance dual-modal imaging-guided photothermal therapy of prostate cancer.Methods:1 Preparation and characterization of CM-LFPPFe₂O₃ NPs was prepared by the thermal decomposition method reported in the literature.Fe₂O₃ NPs was coated with PVP through the surface coating of hydrophobic interaction.The surface coating of Fe₂O₃ NPs on the polymer made many Fe₂O₃ NPs coated in the polymer,which further modified the photothermal agent PPy,and then modified CM to form CM-LFPP（Cell Membrane-modified Lipids-Fe₂O₃-PVP nanoclusters implanted in Polypyrrole）.Transmission electron microscopy（TEM）was used to observe the morphology of nanoparticles,EDS mapping was used to measure the composition of nanoparticles,dynamic light scattering（DLS）was used to measure the size of nanoparticles,and zeta potential was used to evaluate the surface electrical properties of nanoparticles.Absorption spectra of nanoparticles in near infrared（NIR）were analyzed by UV-Vis-NIR.2 In vitro performance of CM-LFPPNano particles with different concentrations were prepared and the photoacoustic imaging signal performance was evaluated by 3D photoacoustic imaging system.The T2 imaging performance was evaluated by 3.0T small animals magnetic resonance imaging system.The temperature rise of aqueous solution under 1064 nm laser irradiation was measured to evaluate its photothermal properties,and the temperature change was recorded by thermal imager.3 Biosafety assessment and in vivo performance of CM-LFPPMouse brain microvascular endothelial cells b End.3 cells and human prostate cancer PC-3 cells were used as experimental subjects to evaluate the cytotoxicity of nanoparticles by CCK-8 assay.Meanwhile,in order to evaluate the safety of the material in vivo,whole blood analysis and blood biochemical analysis were performed on mice 0,1,4,and 7 days after injection of the material,and H&E staining was performed on major organs.A subcutaneous tumor model was established in nude mice,and PAI and MRI scans were performed on the tumor sites before and after administration,respectively.In vivo imaging performance was evaluated by intravenous and intratumoral injection.The treatment group and the control group were set up.Nano particles were injected into the tumor of the treatment group and laser irradiation was given to evaluate the effect of photothermal treatment in vivo.The tumor temperature rise of mice in each group was photographed by infrared camera,and the subcutaneous tumors in each group were dissected for H&E staining and TUNEL staining to evaluate the therapeutic effect.Results:1 It can be observed by TEM that many small Fe₂O₃ NPs are wrapped up to form large spherical structures,and the size of each large nanoparticle is roughly uniform,about 100 nm.Further EDS mapping analysis of the elemental composition of nanoparticles shows that elements C,N and O are evenly dispersed in the particles.Fe element is evenly distributed inside the nanoparticles,which preliminarily proves the successful surface coating PVP of Fe₂O₃ NPs.DLS results show that the nanoparticles have spherical structure and uniform particle size.The particle size of FP,LFPP and CM-LFPP were respectively tested for 138.3 nm,154.8 nm and 146.8 nm.It can be found that the particle size of FP modified PPy was larger,and then the particle size was slightly smaller when the membrane was modified by mechanical extrusion for several times.The changes of the potential of LFPP,CM-LFPP and CM were-12.9m V,-20.2 m V and-28 m V,respectively,indicating that the nanoparticles tended to the potential of the cell membrane after modification.The UV-Vis-NIR absorption spectra of FP and CM-LFPP were tested,and it was found that the synthesized nanoparticles had strong absorption in the range of 600-1100 nm,while FP had almost no absorption in this range,which proved that the nanoparticles successfully modified PPy.2 The analysis of the imaging performance of nanoparticles with different concentrations showed that even at a low concentration,they could show higher PAI and MRI signals,and the signal intensity increased with the increase of the concentration,indicating that the nanoparticles had good PAI and MRI properties.T2relaxation rates of FP and CM-LFPP were measured at 53.1 s^-1 m M^-1 and 48.7 s^-1 m M^-1,respectively.The correlation between photoacoustic signal and concentration can reach R=0.9991.Under the irradiation of 1064 nm laser,the solution temperature of nanoparticles showed a concentration-dependent heating trend,and the solution temperature increased with the increase of laser power density,indicating that the nanoparticles have good photothermal properties.3 CCK-8 of b End.3 cells and PC-3 cells showed that the cell activity remained above 80%even at high concentration,indicating low toxicity and good biocompatibility.Hematological indexes,blood biochemistry and main organ tissue section structure showed that the nanoparticles had no toxicity to mice,which preliminarily proved that the biological safety of nanoparticles was high.PAI and MRI scans of the tumor site showed that the tumor signal was significantly stronger after administration than before administration,and intratumoral injection had better imaging effect than intravenous injection.The study on the effect of tumor treatment showed that the highest temperature rise in the treatment group was compared with the control group.Tissue H&E and TUNEL staining showed apoptosis in the treatment group.Conclusion:Firstly,Fe₂O₃ NPs was synthesized by thermal decomposition method reported in the literature.The synthesized Fe₂O₃ NPs is hydrophobic.In order to apply it in biomedicine,amphiphilic polymer PVP was modified on its surface through hydrophobic interaction to transform it into water soluble,that is,FP.Then Py polymerization on the surface of PVP further form PPy in inducing agent Fe Cl₃·6H₂O,namely FPP,after using thin-film hydration method liposome is modified on the surface of nanoparticles to form LFPP,finally in the surface of the liposome through mechanical extrusion method is modified prostate cancer PC-3 pieces of the cell membrane,The aim is to modify the membrane proteins on the cell membrane onto the surface of the nanoparticles to form the final composite nanoparticles,namely CM-LFPP.Due to the modification of PPy,as a photothermal agent,has the ability of photoacoustic imaging（PAI）and in the near infrared（NIR）1064 nm laser irradiation can increase the temperature for photothermal therapy（PTT）,its core Fe₂O₃ NPs has the ability of magnetic resonance imaging（MRI）,so it can achieve dual-mode imaging guided photothermal therapy,The modified cell membrane can then be actively targeted to the tumor.To sum up,this topic is mainly about cell membrane bionic polypyrrole coated iron oxide for PAI/MRI dual-mode imaging guided photothermal therapy.In addition,the TEM data,particle size,potential and NIR absorption tested can preliminically prove the synthesis of nanoparticles,the photoacoustic signal capability of the particles tested by 3D photoacoustic imaging system and the T2 signal of the particles tested by 3.0T magnetic resonance imaging system,and it is found that the signal is obviously darkened,showing good T2magnetic resonance imaging capability.The good photothermal properties of the particles were further tested under the irradiation of 1064 nm laser.After safety evaluation,the synthesized nanoparticles are non-toxic and have good biological safety.In vivo performance test of nanoparticles showed that they had good PAI and MRI capabilities.Finally,photothermal treatment was tested.Compared with the control group,the temperature of the experimental group could be raised to 57.2℃,which was sufficient for tumor ablation and photothermal treatment could be carried out.