Acitretin Enhances The Cytotoxic Effect Of ALA-PDT Treatment On SCL-1 Cells And Its Mechanism

Background and objectivePhotodynamic therapy(PDT)has been increasingly accepted as a new choice for Cutaneous squamous cell carcinoma(cSCC),owing to its minimally invasive feature and good cosmetic effect.Our previous study demonstrated that PDT using 5-aminolevulinic acid(ALA)as photosensitizer(ALA-PDT)could significantly increase the cytotoxic effect of acitretin on SCL-1 cells,but whether acitretin can enhance the effect of ALA-PDT on SCL-1 cells remains unclear.In this study,the cell viability including cell morphology,death,including apoptosis of SCL-1 cells were studied with combined treatment of acitretin followed by ALA-PDT.The possible mechanism of the action was also studied by detecting the reactive oxygen species(ROS)level and cell membrane permeability.This study provides a theoretical basis for the combined treatment of cSCC with ALA-PDT,as well as new ideas for the treatment of refractory skin tumorsMethods1.SCL-1 cell lines were treated with acitretin of different concentrations,and combined treatment of acitretin followed by ALA-PDT.The morphological alteration of SCL-1 cells was observed under microscope after treatment to study the effect of the treatments on SCL-1 cells.2.Trypan blue exclusion assay was used to detect the cell death effect on SCL-1 cell lines after combined treatment with acitretin followed by ALA-PDT.3.Confocal microscopy and flow cytometry were used to detect the effects of cell apoptosis and ROS production on SCL-1 after acitretin treatment with different concentrations,and combined treatment of acitretin followed by ALA-PDT.4.The membrane permeability of SCL-1 cells was detected using Confocal microscope and flow cytometry methods after treatment with acitretin of different concentrations.Results1.SCL-1 cells grew more slowly,become round and smaller,and some cells experienced karyopyknosis after acitretin treatment,manifesting an obvious inhibitor effect by acitretin in a dose-dependent manner within the concentration of 1.6×10-4mg/mL to 1.6×10-1mg/mL.2.Acitretin had a death promoting effect on SCL-1 cells,and a stronger effect was observed at a acitrretin concentration of 3.2×10-3mg/mL than that of 6.4×10-4mg/mL.Acitretin significantly promoted the cell death induced by ALA-PDT.3.Acitretin have a pro-apoptotic effect on SCL-1 cells,in an acitretin concentration-dependant manner within 1.28×10-4mL to 3.2×10-3mg/mL.Early apoptosis was the main type of apoptosis,and the late apoptosis increased with the increase of acitretin concentration.Besides,the pro-apoptotic effect of ALA-PDT was significantly promoted by acitretin.4.Acitretin could promote the production of ROS in SCL-1 cells,in an acitretin concentration-dependant manner within 6.4×10-4mg/mL to 3.2×10-3 mg/mL.Similarly,the effect of ALA-PDT on ROS production in SCL-1 cells was significantly enhanced by acitretin in a concentration-dependent manner in the range of 2.56×10-5mg/mL to 3.2×10-3mg/mL.5.Acitretin increased the membrane permeability of SCL-1 cells in a acitretin concentration-dependent manner in the range of 1.6×10-4 mg/mL to 1.6×10-2mg/mL.Conclusion1.At given concentrations,the growth,morphology and viability of SCL-1 cells are significantly inhibited,and apoptosis was promoted by actretin,in a n acitretin concentration-dependant manner.Combined treatment with acitretin followed by ALA-PDT can enhance these effects compared to ALA-PDT treatment alone.2.Acitretin may increase the permeability of SCL-1 cells,inducing more ROS when treated with ALA-PDT,and eventually enhance the effect of ALA-PDT on SCL-1 cells.