Study On Molecular Mechanism Of Resveratrol's Inhibition Of Tumor Cell Adhesion Via Regulating PP2A-Erk1/2 Pathway

The present study,using cellular and molecular biology techniques and methods including cell culture,cell adhesion assay,RNA interference,Western blotting,etc.,and employing Rhl,Rh30.HT29 and HeLa cells as experimental objects,synthetically studied the molecular mechanism of resveratrol's inhibition of tumor cell adhesion via PP2A-ERK1/2 signaling pathway.The detailed results were summarized as follows:1 Resveratrol inhibits adhesion in tumor cellsRh1,Rh30,HT29 and HeLa cells were treated with different concentrations of resveratrol(10-100 ?M)for 4 h and then stimulated with/without IGF-1(10 ng/ml)for 1 h?Tumor cell adhesion was evaluated using CN IV(0.2 ?g/ml),fibronectin(0.5?g/ml)or laminin(0.5 ?g/ml)coating assay,respectively,and cell viability was detected by trypan blue exclusion and MTS assay.The results showed that resveratrol inhibited tumor cell adhesion in concentration-dependent manner,but resveratrol didn't suppress cell viability in same time.The data suggest that resveratrol inhibits tumor cell adhesion.2 Resveratrol inhibits Erkl/2-mediated adhesion in tumor cellsRhl,Rh30,HT29 and/or HeLa cells,Rh30 and HeLa cells infected with lentiviral shRNA to Erkl/2 or GFP,or Rh30 and HeLa cells infected with Ad-MKK1-R4F,Ad-MKK1-K97M or Ad-GFP,respectively,were treated with/without resveratrol(10-100 ?M or 100 ?M)for 4 h and then stimulated with/without IGF-1(10 ng/ml)for 1 h,or with resveratrol(100 ?M)for 4 h and then stimulated with/without IGF-1(10 ng/ml)for 1 h following pretreatment with/without U0126(5 ?M)for 1 h.Expression of relative proteins was analyzed using Western blotting,and tumor cell adhesion was evaluated using CN IV coating assay.The results showed that resveratrol inhibited IGF-1-stimulated Erkl/2 phosphorylation in the cells;Inhibition of Erkl/2 by U0126 or downregulation of Erkl/2 potentiated resveratrol's inhibition of IGF-1-stimulated Erkl/2 phosphorylation and cell adhesion;Expression of MKK1-K97M or MKK1-R4F affected resveratrol's inhibition of IGF-1-stimulated Erkl/2 phosphorylation and cell adhesion.The findings indicate that resveratrol inhibits Erkl/2-mediated adhesion in tumor cells.3 Resveratrol blocks Erkl/2-dependent tumor cell adhesion by activating PP2ARh1,Rh30,HT29 and/or HeLa cells,Rh30 and HeLa cells infected with Ad-PP2A-wt,Ad-dn-PP2A or Ad-GFP,respectively,were treated with/without resveratrol(10-100 ?M or 100 ?M)for 4 h and then stimulated with/without IGF-1(10 ng/ml)for 1 h,or with/without resveratrol(100 ?M)for 4 h and then stimulated with/without IGF-1(10 ng/ml)for 1 h following pretreatment with/without okadaic acid(OA,100 nM)or U0126(5 ?M)for 1 h.Expression of relative proteins was analyzed using Western blotting,and tumor cell adhesion was evaluated using CN IV coating assay.The results showed that resveratrol activated IGF-1-inhibited PP2A activity;Inhibition of PP2A by OA or expression of dn-PP2A resisted resveratrol's inhibition of IGF-1-activated Erkl/2-dependent tumor cell adhesion,whereas overexpression of PP2A strengthened the above events.The evidences underscore that resveratrol blocks Erkl/2-dependent tumor cell adhesion by activating PP2A.