Study On The Photodynamic Effect Of Tanshinone ⅡA On Choroidal Melanoma MUM-2B Cells

ObjectiveMultimodal combination therapy is a trend of tumor therapy.Seeking a variety of tumor treatment methods will provide a choice space for tumor combination therapy.Choroidal melanoma is a primary ocular malignant tumor with high mortality and complex pathogenesis.In view of the fact that choroidal melanoma is also a superficial tumor and the unique advantages of photodynamic therapy（PDT）,this paper attempts to study the theory of PDT for choroidal melanoma.In recent years,in the field of PDT,natural photosensitive molecules have attracted more and more attention because of their low toxicity,green environmental protection and other characteristics.Therefore,the main goal of this paper is to evaluate whether tanshinoneⅡA,the main active extract of Salvia miltiorrhiza,can be used as a natural photosensitizer in PDT for choroidal melanoma,and to explore the related mechanismsMethodIn this paper,laser flash photolysis was used to investigate the transient photochemistry of TanshinoneⅡA and to evaluate whether TanshinoneⅡA can be used as a photosensitizer in PDT.Then,¹O₂probe and high performance liquid chromatography technology were utilized to detect the generation of ¹O₂ in the process of TanshinoneⅡA photosensitization.The effect of TanshinoneⅡA photosensitization on the growth and migration of choroidal melanoma MUM-2Bcell was analyzed by MTT,wond healing assay and monoclonal formation rate assay.In order to explore related mechanism,the effect of TanshinoneⅡA photosensitization on cell apoptosis and necrosis,cell cycle arrest,intracellular ROS and mitochondrial membrane potential was analyzed by confocal laser scanning microscope and flow cytometry.Finally,in order to solve the problem of poor tissue penetration and water solubility of TanshinoneⅡA,mesoporous silica coated upconversion luminescence nanocomposites was designed and synthesized.Results（1）355 nm and 532 nm laser flash photolysis of nitrogen saturated TanshinoneⅡA acetonitrile solution can induce the photoexcitation of TanshinoneⅡA and generate the corresponding excited triplet state.The triplet state of TanshinoneⅡA has two characteristic absorption peaks centered at 340 nm and 680 nm,respectively.The triplet state of TanshinoneⅡA can also be generated by energy transfer between TanshinoneⅡA and the triplet state of benzophenone or tetramethylbenzoquinone.The energy levels of triplet state of TanshinoneⅡA are determined ranging from159 k J/mol to 176 k J/mol.（2）The growth inhibitive effect of TanⅡA photosensitization on human choroidal melanoma cell MUM-2B was investigated.The generation of ¹O₂ by TanⅡA photosensitization was firstly confirmed in this study.TanⅡA photosensitization could efficiently inhibit cell survival and migration,enhance intracellular ROS levels and decrease theΔψ_m in MUM-2B cells.Besides,TanⅡA photosensitization could induce cell apoptosis and necrosis and cause cell arrest in G2/M phase.（3）UCNPs@m Si O₂ nanocomposite was successfully prepared which has obvious core-shell structure.The dispersion of TanshinoneⅡA in aqueous phase was improved by the load of drugs into the nanocomposite.At the same time,980 nm laser-induced TanshinoneⅡA photodynamic action on MUM-2B cells was realized by utilizing UCNPs@m Si O₂ nanocomposite.ConclusionTanshinoneⅡA can be effectively excited by its absorption light to form corresponding the excited triplet state which can be effectively quenched by oxygen to form ¹O₂.¹O₂ is the most important effectors in PDT process.At the cellular level,TanshinoneⅡA can be used as a photosensitizer for photodynamic therapy on choroidal melanoma MUM-2B.The photodynamic effect of TanshinoneⅡA on MUM-2B cells may be induced by increasing intracellular ROS or causing DNA damage,and thereby inducing cell apoptosis and necrosis.In addition,nanotechnology based on upconversion luminescence can overcome the problems of poor tissue penetration and low bioavailability of tanshinoneⅡA.This study provides a new idea for the"new use of old drugs"for tanshinoneⅡA,and also enriches the compound library of PDT photosensitizer from nature source.