The Inhibition Studies Of HSP90 Targeting In The Treatment On PCO

Background Posterior capsule opacification(PCO)is the most common complication after cataract extraction with intraocular lens(IOL)implantation.It is the main factor influencing the postoperative visual acuity.PCO is caused by the lens epithelial cells retained in the capsular bag following surgery,which proliferate,migrate and epithelial-mesenchymal transition,collagen deposition,and lens fiber generation.Advances in surgical techniques,intraocular lens materials and designs have reduced the PCO rate,but it is still a significant problem.The only effective treatment for PCO at this stage is the Nd:YAG laser.This method is not only expensive to treat,but also causes various complications after surgery,such as crystal damage,retinal detachment,cystic macular edema,endophthalmitis,corneal edema,Increased intraocular pressure,dislocation or subluxation of the lens,and depression of the lens.At present,there is no drug for treating or preventing PCO that is safe for clinical trials.In the study,we found that heat shock protein 90(HSP90)inhibitor 17 AAG can inhibit the proliferation of the lens epithelial cell lines.HSP90 is an ATP?dependent molecular chaperone which plays an important role in protein stabilization and degradation.Inhibition of HSP90 function results in the degradation of its target protein.17 AAG is an HSP90 N-terminal inhibitor,which inhibits the binding of ATP and HSP90,and inhibits the activity of HSP90 ATPase.It has been in clinical phase II study.EGFR is a target protein of HSP90.EGFR signaling pathway is regulated by HSP90.17 AAG can inhibit cell proliferation by inhibiting EGFR degradation and inhibiting its downstream pathway.The lens epithelial cell line is the simplest experimental model for the study of PCO,but it does not consider the effect of cell adhesion on the lens capsule and fails to truly reflect the pathophysiological changes of PCO.Therefore,we prepared in vitro rat PCO model and PCO model in rabbits,so as to investigate the inhibitory effect and molecular mechanism of 17 AAG on PCO.Purpose To investigate the inhibitory effect and molecular mechanism of HSP90 specific inhibitor-17 AAG on PCO.Method 1.The PCO model of rat lens in vitro was successfully prepared,and the expression of HSP90,EGFR and pERK in serum-free cultured 6h,12 h,24h and normal tissue epithelial cells were detected by Western blotting.2.The inhibitory effect of 17 AAG on lens epithelial cell line was verified by cell proliferation assay using mouse lens epithelial cell line(m LEC)and human lens epithelial cell line(SRA01/04,HLE-B3).3.Using a rat lens PCO model,the inhibitory effect of 17 AAG on residual epithelial cells on the capsular bag was detected by culturing and inverted microscope observation,and the inhibitory effect was used verified by Ed U labeling method.4.Using a rat lens PCO model,the effect of 17 AAG on the expression of EGFR and downstream pathways in the residual epithelial cells was examined by Western blotting.5.Using a rat lens PCO model,TUNEL staining and Western blotting assays were used to determine whether 17 AAG induced apoptosis in the epithelial cells.6.Using a rat lens PCO model,the inhibitory effect of 17 AAG and HSP90 non-N-terminal inhibitor PEITC on residual epithelial cells on the capsular bag was detected by culturing and inverted microscope observation,and the inhibitory effect was used verified by Ed U labeling method.7.Successfully constructed a rabbit PCO model in vivo to detect the effect of 17 AAG in preventing PCO.Result 1.High expression of HSP90 in residual epithelial cells on the capsular bag,and the EGFR signaling pathway was activated.2.17 AAG inhibits proliferation of the lens epithelial cell line in a time and concentration dependent manner.3.17 AAG inhibits proliferation of the residual epithelial cells on the capsular bag.4.17 AAG promotes degradation of EGFR in the residual epithelial cells and inhibits its downstream pathway 5.17 AAG induced the epithelial cell apoptosis on the capsular bag.6.PEITC,a non-N-terminal inhibitor of HSP90,did not inhibit the residual epithelial cells on the capsular bag.7.17 AAG delay the occurrence of PCO in rabbits.Conclusion 1.The expression of HSP90 increased during PCO,and the EGFR signaling pathway was activated,and it was involved in the proliferation of capsular residual epithelial cells.2.17 AAG inhibits the proliferation of rat capsular bag epithelial cells and the occurrence of PCO in rabbits by inhibiting HSP90-EGFR signaling pathway.3.HSP90 is a new target molecule for preventing the occurrence of PCO.