| Objectives:Oral squamous cell carcinoma（OSCC）is the sixth most common malignant tumor worldwide.It threatens human life seriously due to the limited treatment methods and poor prognosis.In order to improve the shortcomings of conventional treatment methods effectively and provide a new treatment strategy for oral cancer,the piezoelectric catalytic material of sodium bismuth titanate（BNT）with high catalytic performance is selected based on the characteristics of the material that generated a large number of reactive oxygen species（ROS）under ultrasound（US）irradiation.The BNT-c RGD nanoplatform which killed tumor cells through piezoelectric catalysis combined with active tumor targeting was constructed based on BNT materials and evaluated its antitumor effectiveness.In order to further enhance the antitumor effect,a drug-loaded BNT nanoplatform BNT-TK-DOX was constructed which was responsive to ROS,the nanoplatform generated ROS to kill tumor cells under ultrasound stimulation,while excessive ROS broke the thioketal bond（TK）and releases doxorubicin（DOX）to exert chemotherapy effects.Through the dual effects of piezoelectric catalysis and chemotherapy,it killed tumor cells,significantly improving the anti-tumor effect.In order to better diagnose and treat oral cancer,a composite nanoplatform BNT-Mo S2 that included diagnosis and treatment has been constructed.This nanoplatform not only could diagnose tumors through CT imaging and photoacoustic（PA）imaging,but also had the properties of piezoelectric catalysis and photothermal effects due to graft of Mo S2 nanodots on the surface of BNT material with excellent photothermal properties.This platform could generate a large amount of ROS through piezoelectric catalysis under ultrasound stimulation to kill tumor cells.At the same time,the platform converted energy into thermal energy,rising local temperature to kill tumors under the irradiation of 808nm laser through photothermal therapy（PTT）,the construction and application of a composite nanoplatform with both diagnostic and excellent anti-tumor effects were achieved.Finally,the mechanism of piezoelectric catalysis of BNT to kill oral squamous cell carcinoma cells under US irradiation was explored through bioinformatics methods,laying a foundation for future research.Methods:1.BNT nanomaterials were synthesized by hydrothermal method,and c RGD was grafted onto the surface of BNT by covalently bound.The shape,particle size,potential and other general characteristics of BNT-c RGD nanoparticles were detected by means of material testing.The effects of BNT-c RGD nanoparticles to treat oral squamous cell carcinoma were evaluated in vitro and in vivo.2.After the surface of BNT nanomaterials was modified by polyethylene glycol（PEG）,the-COOH-TK-PEG-DOX was grafted onto the surface of BNT by esterification reaction.The characteristics with BNT-TK-DOX nanoparticles were detected by material testing methods.The effect of BNT-TK-DOX nanoparticles to treat oral squamous cell carcinoma under US irradiation was evaluated in vitro and in vivo.3.BNT nanomaterials were synthesized by hydrothermal method,sulfhydryl（-SH）modification was performed on the surface of the material,and Mo S2 nanodots were covalently bound to the surface of BNT.The characteristics with BNT-Mo S2nanoparticles were detected by material testing methods.The effect of BNT-Mo S2nanoparticles to treat oral squamous cell carcinoma under US irradiation was evaluated in vitro and in vivo.4.The biological processes,cell components,molecular functions,mechanism pathways,subdivisions of cell localization and protein domain enrichment were analysed with significantly different proteins.The key proteins were screened through bioinformatics analysis,then experimental validation was conducted.Results:1.BNT-c RGD nanoparticles were synthesized,and their active tumor targeting properties were confirmed through in vitro cell uptake experiments and in vivo small animal imaging technology.In vitro cytotoxicity experiment,hemolysis experiment,H&E staining of tissues and organs in vivo,blood routine and biochemical routine experiments have verified the biological safety of BNT-c RGD nanoparticles.In vitro CCK-8 experiment,staining of living dead cells,and in vivo anti-tumor treatment of nude mouse models of oral squamous cell carcinoma demonstrated the effectiveness of BNT-c RGD nanoparticles in killing tumors under ultrasound irradiation.2.BNT-TK-DOX nanoparticles were synthesized,and in vitro cell uptake experiment confirmed the characteristics of BNT-TK-DOX nanoparticles in response to ROS to drug release.The biological safety and in vitro anti-tumor effect of BNT-TK-DOX were verified through in vitro cytotoxicity experiment,live/dead cells staining.3.BNT-Mo S2nanoparticles were synthesized.The photothermal characteristics of material was verified through thermal imaging.The biological safety of BNT-Mo S2was verified through in vitro cytotoxicity experiment.The anti-tumor effect of BNT-Mo S2 was evaluated in vitro through live/dead cells staining and apoptosis flow cytometry detection.In vivo experiments were conducted to validate the CT imaging and PA imaging functions of BNT-Mo S2 nanoparticles using a subcutaneous tumor bearing model of oral cancer in nude mice.The biological safety and good anti-tumor effect of BNT-Mo S2 nanoparticles were also confirmed.4.The bioinformatics analysis of differential proteins showed that compared with the control group,the proteins associated with Ca2+binding and transportation were significantly overexpressed in the US irradiation group（P<0.05）.At the same time,the proteins associated with mitochondrial respiratory chain complexes were also overexpressed in the US group（P<0.05）.Through in vitro experiments,the changes of Ca2+concentration and mitochondrial membrane potential of two type cells of human oral squamous cell carcinoma under US irradiation were compared.The results showed that compared with the control group,intracellular Ca2+concentration increased in BNT+US group（P<0.001）and mitochondrial membrane potential decreased（P<0.001）,the results were statistically significant.Conclusions:1.BNT-c RGD nanoplatform has the function of targeting actively oral squamous cell carcinoma cells,it plays the role of piezoelectric catalysis to produce a large number of ROS to kill tumor cells under US stimulation.The nanoplatform has good biocompatibility and antitumor effect.The successful formulation of BNT-c RGD nanoplatform provides a strategy for the combination of piezoelectric catalysis and active targeted treatment of tumors.2.BNT-TK-DOX is a composite nanoplatform with ROS response,piezoelectric catalytic performance and chemotherapy effect.The piezoelectric catalytic effect generates a large number of ROS under US irradiation to kill tumor cells.At the same time,with the increase of ROS,the ROS-responsive thioketal bond（TK）breaks,releasing doxorubicin（DOX）,and playing the role of chemotherapy.The nanoplatform has good biocompatibility and antitumor effect.The successful formulation of BNT-TK-DOX nanoplatform provides a reference for the treatment of tumor with ROS response and piezoelectric catalysis combined with chemotherapy.3.BNT-Mo S2 is a composite nanoplatform with integrated diagnosis with treatment.BNT-Mo S2 has the functions of CT and PA imaging.At the same time,BNT-Mo S2 plays the role of piezoelectric catalysis and PT property under the irradiation of US and NIR,producing a large number of ROS,increasing the intracellular temperature,and killing tumor cells.BNT-Mo S2 provides a design idea to formulate a nanoplatform that includes diagnosis and combines piezoelectric catalysis with photothermal treatment of tumors.4.The piezoelectric catalysis mediated by BNT nanoparticles would have increased the ROS content in cells under US irradiation,resulted in intracellular Ca2+overload,and changed in mitochondrial membrane potential to cell death. |
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