Application Of Fullerene As Photosensitizer And Free Radical Scavenger In The Field Of Biomedicine

Reactive oxygen species?ROS?is a category of oxygenated species with higher oxidation reactivity,The ROS category in living systems also includes hypochlorous acid?HClO?,singlet oxygen?¹O₂?,lipid peroxide?ROOH?,and ozone?O₃?.The key to human health is the balance between oxidation and anti-oxidation.If this balance is broken,Oxidative stress would occur in the human body.Excessive ROS in the human body will damage macromolecules such as proteins,nucleic acids and lipids,which implies that some diseases are happening,mostly related to aging,such as Alzheimer's disease.Therefore,antioxidants,enzymes and nanoparticles have become potential therapeutic agents,which can relieve oxidative stress by scavenging or neutralizing ROS and reduce the downstream substances of ROS.Compared with other antioxidants,nano-particles can increase bioavailability by increasing absorption,and they can be targeted after functionalization to improve their scavenging activity.These nanoparticles include mesoporous silica,yttrium oxide,fullerenes,et al.On the other hand,the excessive ROS can cause cell death,which can also be utilized in the treatment of the certain diseases.Cancer cells,for example,can be killed by local excess ROS.Photodynamic therapy is using photosensitizers which can react with light environment and produce reactive oxygen species to kill cancer cells.Fullerene,as a third carbon allotrope,is a potentially valuable material that can be applied to biomedicine.On the one hand,fullerenes can be excited by ultraviolet light to become long life triple excited state and then,because of their unique delocalization conjugation system,they transfer energy or electrons to molecular oxygen,producing high reactive ¹O₂ or O₂^·-,which can be applied to photodynamic treatment.On the other hand,fullerene can quench all kinds of free radicals,because of which it is called"free radical sponge".Therefore,fullerene can be applied to scavenging free radicals,blocking oxidative stress pathways,and protecting cells and tissues.The aim of this study is investigating this unique property of fullerenes.?1?Photodynamic therapy of cervical cancer cells by fullerene derivative C₇₀-OEGFullerene can generate singlet oxygen by irradiation of ultraviolet light,but,due to its poor water solubility,its solubility in biological media is not good.Therefore,the functional groups of fullerenes need to be modified to make them hydrophilic or amphiphilic.In this study,hydrophilic fullerene derivatives were obtained by functional group modification of fullerenes using oligo ethylene glycol which has hydrophilicity,flexible structure and good biocompatibility.HeLa cells were divided into 6 groups,one of which was dark-treated and the others were light-treated.The concentration gradients of C₇₀-OEG added to the dark-treated group were same as the four light-treated groups,which were 0,1,1.5,2,2.5,5,and 10?mol/L.Cells from the dark group were incubated with C₇₀-OEG for 24 h,and then the cell viability was measured directly with CCK-8;the cells in the four light-treated groups,was incubated with C₇₀-OEG for 3 h and treated with 20 mW/cm² white light for 0 min,5min,10 min,and 20 min respectively.In the end,CCK-8 was used to measure cell viability.The 5th light-treated groups received 2?mol/L Ce6 and C₇₀-OEG in different concentrations,ie,0,0.1,0.5,1,1.5,2,and 2.5?mol/L.After irradiation with white light at20 mW/cm² for 10 min,CCK-8 was used to detect cell viability.The nuclei,cell membranes,mitochondria,and lysosomes were stained,and the cell morphology was observed under a confocal microscope.The fullerene derivative C₇₀-OEG,as a photosensitizer,can effectively kill Hela cells by irradiating with white light can generate singlet oxygen,which can be applied to photodynamic therapy.?2?Application of the PVP coated fullerenes(PVP@C₆₀?PVP@C₇₀)in oxidative stress and anti-inflammatoryC₆₀ and C₇₀ were coated with polyvinylpyrrolidone to get water-soluble PVP@C₆₀and PVP@C₇₀,which could improve their bioavailability.Fullerenes,knew as"free radical sponges",can scavenge active oxygen.When the balance of oxidation and oxidation is broken in the body,excess ROS can be cleared by fullerenes to protect cells and tissues.We used hydrogen peroxide to stimulate HEK cells to establish an oxidative stress model.HEK cells were incubated with different concentrations of PVP@C60 and PVP@C70 for 24 h,the activity of cells was detected by CCK-8;PVP@C₆₀ and PVP@C₇₀at different concentrations were respectively associated with HEK cells in oxidative stress,and the activity of cells was detected by CCK-8.LPS stimulated mouse macrophages RAW 264.7 to establish oxidative stress and inflammation models.With or without LPS stimulation,different concentrations of PVP@C₆₀0 and PVP@C₇₀ were incubated with RAW 264.7 for 20 h,and DCFH-DA was used to detect the production of reactive oxygen species.In the oxidative stress and inflammatory model of LPS stimulating RAW264.7,different concentrations of PVP@C₆₀ and PVP@C₇₀ were incubated with RAW 264.7 for20 h.ELISA was used to detect IL-2,TNF-?and iNOS mRNA's expression,and Western blot was used to detect the protein expression of p-ERK1/2,ERK1/2,p-AKT and AKT.In the oxidative stress model,it was verified that PVP@C₆₀ and PVP@C₇₀ have the ability to scavenge free radicals and protect cells from damage.In the inflammatory model,PVP@C₆₀ and PVP@70 effectively inhibit the release of TNF-?and iNOS,increase the release of IL-2,and effectively inhibit the phosphorylation of ERK1/2.