The effect of autophagy and crosstalk with apoptosis induced by Ultraviolet in A375 / 实用医学杂志

Objective To investigate the autophagy effect and the crosstalk with apoptosis by UV in A375 cells.Methods GFP-RFP-LC3 lentivirus were used to evaluate the effect of autophagy after being irradiated with UV of different doses (0、10、30、50 mJ/cm2).After being treated with 30 mJ/cm2 irradiation,the apoptosis rate of A375 with or without autophagy inducer was evaluated by annexin V-FITC/PI with flow cytometry.Western blot was used to distinguish the biomarker of autophagy (BECN,LC3) and apoptosis (Caspase 3,9).Results After being irradiated with 10 mJ/cm2 or 30 rnJ/cm2 UV,the autophagosome was observed at 6 h and rich at 9 h.However,the dots of autophagy had been abundant continually from 3 h with 50 mJ/cm2.The ability of inducing autophagy of UVB is stronger than UVA.UVA and UVB showed synergistic effect in autophagy with the dose of 30 mJ/cm2.The Caspase 3 and Caspase 9 proteins were downregulated after 30 mJ/cm2 UV irradiation with autophagy biomarkers increasing,whereas the apoptosis biomarkers were enriched with the inhibition of autophagy.Conclusions UV can induce autophagy with more significant effect of UVB.Autophagy paly protective role by delaying apoptosis after 30 mJ/cm2 UV irradiation in A375.

Effects of 14-3-3σ on UVB-induced radiation damage in HaCaT cells / 中华放射医学与防护杂志

Objective To explore the role of 14-3-3σ in cell cycle arrest,proliferation inhibition of HaCaT cells after UVB exposure.Methods Crystal violet assay was used to determine the viable density of HaCaT,HaCaTKD cells after being irradiated with UVB of different doses (10,20,30,40,50,60 and 80 mJ/cm2) for 48 h.After HaCaT and HaCaTKD being treated with 30 mJ/cm2irradiation,cell growth and cell cycle distribution were detected by CCK-8 assay and PI staining combined with flow cytometry,respectively.Western blot was used to evaluate the protein expression of 14-3-3σ,Cdc2,Cdc25c and Cyclin B1.Results After 48 h,the survival rate of both HaCaT and HaCaTKD decreased in a dosedependent manner.Especially,HaCaTKD cells had drastically low proliferation rate compared with normal HaCaT at 10 mJ/cm2 (t =8.83-49.63,P ＜ 0.05).The proliferation rate of HaCaTKD cells was significantly lower than that of HaCaT cells (F =32.89,P ＜ 0.05).After treatment with 30 mJ/cm2 UV irradiation,the ability of proliferation in normal HaCaT cells was recovered after 24 h while there was no proliferation in HaCaTKD cells within 72 h after the same treatment.The distribution of cell cycle has little change in HaCaTKD (P ＞ 0.05).UVB treatment led to cell cycle arrest from 6 to 18 h in HaCaT cells(t =7.41,9.22,9.16,P ＜0.05)while no cell cycle arrest could be observed in the HaCaTKID cell.Western blot detection indicated that the expression of 14-3-3σ in HaCaT cells was upregulated(t =5.42-9.57,P ＜0.05)while the Cdc25c and Cyclin B1 proteins were downregulated in both HaCaT and HaCaTKD cells (t =3.95-11.21,P ＜0.05).Specifically,Cdc2 protein decreased in HaCaT cells(t =4.93-5.37,P ＜ 0.05)but there was no change in HaCaT~ cells.Conclusions 14-3-3σ protein affects the proliferation and cell cycle of HaCaT cells after UVB irradiation.14-3-3oσ co-activates the expression of Cdc2,Cdc25cand Cyclin B1 to mediate UVB-induced G2/M arrest in HaCaT cells.