title: Enhanced TLR4 Expression on Colon Cancer Cells After Chemotherapy Promotes Cell Survival and Epithelial-Mesenchymal Transition Through Phosphorylation of GSK3β.
abstract: BACKGROUND
  Phosphorylation of glycogen synthase kinase 3β (GSK3β) by phosphatidyl-inositide 3-kinase (PI3K)/protein kinase B (AKT) or inhibition of GSK3β with small-molecule inhibitor attenuates cell survival and proliferation and increases apoptosis in most cancer cell lines. In this study, we investigated the role of phosphorylated GSK3β activated by enhanced toll-like receptor 4 (TLR4) expression in drug-treated colon cancer cells as a model of post-chemotherapy cancer cells.
  
  
  METHODS
  The effect of TLR4 stimulation on metastasis and apoptosis in drug-exposed colon cancer cells was determined by real-time polymerase chain reaction (PCR) and immunoblotting.
  
  
  RESULTS
  Despite the induction of apoptosis after treatment with oxaliplatin and 5-fluorouracil, lipopolysaccharide (LPS) stimulation via increased TLR4 in drug-treated cancer cells effectively inhibited apoptosis through up-regulation of expression of anti-apoptosis-related B-cell lymphoma 2 (BCL2) family proteins [X-linked inhibitor of apoptosis protein (XIAP), BCL2, and survivin] and drug-resistance proteins [multidrug-resistance protein 1 (MDR1), multidrug resistance-associated protein (MRP)1/2/3]. LPS-mediated signaling in drug-treated cancer cells elevated the expression of phosphorylated GSK3β, extracellular signal-regulated kinase (ERK), and the p65 subunit of nuclear factor kappa-light-chain-enhancer of activated B-cells (NF-κB). Pharmacological inhibition of GSK3β (using SB216763) reduced phosphorylation of GSK3β, re-activated caspase-dependent apoptosis, and blocked the expression of cancer stem cell markers and invasive characteristics in LPS-stimulated drug-treated cells. In addition, the ERK-specific inhibitor, PD98059, triggered the apoptosis of TLR4-activated drug-exposed colon cancer cells, whereas there was no effect on the expression of epithelial-mesenchymal transition markers or GSK3β phosphorylation.
  
  
  CONCLUSIONS
  These results suggest that TLR4-induced GSK3β and ERK phosphorylation independently controls cancer cell survival and regulation of GSK3β and ERK after chemotherapy, making TLR4 a critical target for reducing drug resistance and metastasis in patients with colon cancer.
authors.affiliation: Department of Anatomy, Chung-Ang University College of Medicine, Seoul, Republic of Korea.
authors.forename: Yoon Hee
authors.initials: YH
authors.lastname: Chung
authors.affiliation: Department of Anatomy, Inje University College of Medicine, Busan, Republic of Korea kimdj@inje.ac.kr.
authors.forename: Daejin
authors.initials: D
authors.lastname: Kim
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects / Q000502:physiology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005472:Fluorouracil / Q000494:pharmacology; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism*; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D009944:Organoplatinum Compounds / Q000494:pharmacology; D000077150:Oxaliplatin; D010766:Phosphorylation / Q000187:drug effects; D051197:Toll-Like Receptor 4 / Q000096:biosynthesis*
chemical_list: D008070:Lipopolysaccharides; D009944:Organoplatinum Compounds; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D000077150:Oxaliplatin; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D048049:Extracellular Signal-Regulated MAP Kinases; D005472:Fluorouracil
keywords: EMT; GSK-3β; TLR4; colorectal cancer; epithelial–mesenchymal transition