Inhibition Effect Of GA On The EMT Of RPE Cells Via CYLD/NF-κB Signaling Pathway Under HG Circumstance

Objective To studay the effect of gambogic acid(GA)on the epithelial-mesenchymal transition(EMT)of retinal pigment epithelium(RPE)cells in high glucose environmeng and its regulation mechanism via the deubiquitinating enzyme cylindromatosis/nuclear factor-kappa B(CYLD/NF-κB)signaling pathy and.Methods First ARPE-19 cells line was cultured in vitro,and logarithmic phase cells with good growth were used in the experiment.The cells were divided into four groups by induction of high glucose and GA at different concentrations:(1)normal control group(glucose concentration: 5.5 mmol·L-1);(2)high glucose group(glucose concentration: 30 mmol·L-1);(3)hypertonic control group(glucose concentration: 5.5 mmol·L-1+mannitol: 30 mmol·L-1);(4)high glucose +CYLD overexpression group,HG+GA of different concentrations group(glucose concentration: 30 mmol·L-1+2、4、8μmol·L-1 GA);The proliferation and viability of RPE cells in all groups were evaluated by Cell Counting Kit-8,the expression levels of α-smooth-muscle-actin(α-SMA)and CYLD were detected by immunofluorescence staining and the expression levels of α-SMA,CYLD and phosphorylation nuclear factor-κB(p-NF-κB)were measured by Western blot.The statistical data were analyzed by SPSS26.0 software and expressed as sx ?.One-way ANOVA was used for comparison between groups and LSD test was used for pairwise comparison,and P < 0.05 was defined as statistically different.Results Compared with the control ARPE-19 cells,the proliferation level was significantly up-regulated,the expression of CYLD protein was decreased,but the expression of α-SMA was increased,and the NF-κB pathway was activated upon the stimuli of HG.However,hyperosmotic stimulation did not affect the proliferation level of RPE cells.When GA treatment was included in the HG group,the proliferation level of RPE cells was significantly down-regulated,the expression of CYLD was increased,the expression of α-SMA was decreased,and the activation of NF-κB pathway was significantly inhibited in a GA dose-dependent manner.Conclusions In RPE cells,GA treatment can significantly inhibit their proliferation and epithelial-mesenchymal transition,which is mainly through up-regulating CYLD,inhibiting the activation of NF-κB signaling pathway,and then preventing the inflammatory response.This study suggests that GA may alleviate diabetic retinopathy and is expected to be a potential therapeutic drug.