Cathespsin L-mediated Resistance Of Paclitaxel And Cisplatin In A549 Cells Is Under The Control Of Distinct Regulatory Mechanisms

Objective:The aim of this study is to preliminarily investigate the possible mechanism of paclitaxel and cisplatin resistance mediated by CTSL via using human lung cancer cell lines A549 cells as model.Methods:A549 cells,A549/TAX(Paclitaxel-resistant A549 cells)and A549/DDP(Cisplatin-resistant A549 cells)were introduced to performe the following study.(1)CCK8 was performed to detect the drug resistance of cells to cisplatin or paclitaxel,western blot and immunofluorescence were used to detect the expression level of CTSL and TGF-β/smad associated proteins;silence TGF-βor smad3 by transfecting A549/TAX cells with siRNA,and then detected associated protein expression,and CCK8 was used to detect the change in drug resistance to paclitaxel of cells;Luciferase assay and CHIP method were performed to detect the interaction between CTSL promoter and smad3.(2)Western blot was used to detect the expression level of CTSL,Egr-1 and CREB of A549/DDP cells;use siRNA to silence the expression of Egr-1 or CREB in A549/DDP cells,CCK8 was adopted to detect the change in drug resistance to ciaplatin of cells;Luciferase assay was used to detect the interaction between CTSL promoter and CREB.(3)Western blot and immunohistochemical was adopted to detect the correlation between CTSL and samd3 in vivo and human lung cancer.Results:(1)Compared with normal A549 cells,the expression of TGF-β,CTSL and TGF-β/smads associated proteins were higher in A549/TAX cells;silence of TGF-P or smad3 in A549/TAX cells significantly decreased the expression of CTSL and enhanced sensitivity to paclitaxel;CTSL was induced after treated with TGF-P or paclitaxel in A549 cells;smad3 promoted CTSL transcription and CTSL protein expression by binding to CTSL promoter in A549/TAX cells.(2)The expression of CTSL,Egr-1 and CREB in A549/DDP cells were higher than A549 cells;silence of Egr-1 or CREB in A549/DDP cells significantly decreased the expression of CTSL and enhanced sensitivity to cisplatin;CTSL,Egr-1 and CREB were induced after treated with cisplatin in A549 cells;CREB raised CTSL promoter activity and elevated the expression of CTSL by binding to CTSL promoter in A549/DDP cells.(3)Smad3 was positively correlated with CTSL in vivo and in human lung tissues.Conclusion:(1)In A549/TAX cells,TGF-β promoted the expression of CTSL through TGF-β/smads signaling pathway.On the other hand,TGF-β promoted CTSL expression through the binding of smad3 with CTSL promoter to promote CTSL transcription,and then mediated paclitaxel resistance.(2)In A549/DDP cells,Egr-1 and CREB mediated the expression of CTSL so as to mediate cisplatin resistance,and CREB increased the transcriptional activity of CTSL through binding with CTSL promoter.