High-Voltage Pulsed Electric Field Plus Photodynamic Therapy Kills Breast Cancer Cells By Triggering Apoptosis

Irreversible Electroporation(IRE)is a new tumor ablation technology,which use high voltage pulse to produce nano-pore on the cell membrane and cause cell apoptosis.As a non-thermal,minimally invasive ablation technology,IRE allows tissue selectivity,preserving key structures like large vessels and nerves in the ablation area,avoiding heat sink effects,reducing inflammation and scar formation after treatment,shortening treatment cycles and enabling real-time monitoring.IRE ablation has many strengths compared with current ablation technology and holds promising potential for clinical application.Despite the potential of IRE in treating solid tumors,the residual tumor boundary in IRE ablation affects cure rates and can cause recurrences.When using IRE,it is common to see over or insufficient treatment,so solely relying on IRE has clear shortcomings.Photodynamic Therapy(PDT)is a new tumor treatment method.In theory,the controllable illumination area of PDT can compensate for the incomplete ablation boundary of IRE,and the electroporation of cell membranes can improve cellular ingestion of the photosensitizer.Therefore,we proposed IRE+PDT combined therapy to improve the effect of IRE and try to provide a new solution to solve the problem of residual tumor boundary in IRE ablation.ObjectiveTo evaluate the effect of combining irreversible electroporation(IRE)and Photodynamic Therapy(PDT)on breast cancer cells in vitro and in vivo and explore their combined mechanism of action.MethodsIn vitro,Jin's formula was used to assess the total efficacy of IRE+PDT therapy on killing MCF-7 cells at different doses;assays including flow cytometry,high content imaging and confocal laser scanning microscopy were used to qualitatively and quantitatively detect apoptosis in the IRE+PDT group,and real-time PCR and western blotting were used to detect the expression of apoptosis-related genes and proteins.In vivo,different doses of IRE+PDT combined therapy were applied to BALB/C mice bearing breast cancer tumors,and tumor size was used to assess treatment efficacy,and the killing mechanism was analyzed using transmission electron microscopy to observe ultrastructural changes and immunohistochemistry to analyze pathological changes and changes in VEGF,CD31,TGF-?,KI-67 and TNF-? expression.Results 1.IRE+PDT combined therapy showed significant synergic effects on killing breast cancer cells,with the synergy index of the IRE(380 V)+ PDT(10 ?g/m L)group q=1.32.2.Apoptosis was observed during this process,with early apoptosis rates up to 16% in the IRE+PDT group,significantly higher than the IRE-alone(7.6%)and PDT-alone(4.6%)groups(p<0.05).3.Real-time PCR showed higher Caspase1,3,5,6,7,8,9 and TNFRSF1 A expression in the IRE+PDT group than in controls,and western blots showed increased cleaved Caspase3,7,9 and PARP levels.4.IRE+PDT also effectively suppressed tumor growth in vivo,with a tumor suppression rate of IRE(1200 V)+ PDT(10 mg/kg)up to 68.3%.5.The results of transmission electron microscopy and HE found apoptosis occurring in tumor tissues,and the most obvious apoptosis effect was found in the combined therapy group.6.Compared with controls,the new blood vessel indicators VEFG and CD31,metastasis indicator TGF-? and cell proliferation indicator KI-67 were all lower 1 day after treatment in the IRE+PDT group,while the inflammation indicator TNF-? was higher.Conclusion IRE+PDT therapy significantly improves the therapeutic effects on breast cancer compared with IRE or PDT alone.