title: Comparative secretome analysis of cholangiocarcinoma cell line in three-dimensional culture.
abstract: Cholangiocarcinoma (CCA) is a lethal malignancy which occurs with relatively high incidence in Thailand. This cancer is often difficult to diagnose and associated with high mortality. The secretome, containing the secreted proteins from cells, are potentially useful as biomarkers of cancers. Since three-dimensional (3D) cell culture may mimic growth characteristics and microenvironment of solid tumors in vivo better than monolayer culture, we have developed culture of CCA in natural collagen-based scaffold, to enable analysis of the secretome by 2DE. Our results indicated that CCA growth in 3D environment alters cell shape significantly and enhances extracellular matrix deposition. Interestingly, more secreted proteins were detected from 3D culture compared to monolayer culture. Secretome analysis using 2DE coupled with LC-MS/MS demonstrated 10 secreted proteins uniquely found in 3D culture. Moreover, 25 proteins were enriched in 3D culture compared to monolayer culture, including 14-3-3 σ, triosephosphate isomerase, phosphoglycerate mutase 1, α-enolase, and L-plastin. Immunoblotting was used to confirm the presence of L-plastin in conditioned media of CCA and of hepatocellular carcinoma (HCC) cell lines. The results revealed that L-plastin, an actin bundling protein, was uniquely expressed only in the CCA cell line and could be a promising biomarker for differential diagnosis of CCA compared to HCC.
authors.affiliation: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok 10210, Thailand.
authors.forename: Phanthakarn
authors.initials: P
authors.lastname: Tit-Oon
authors.affiliation: Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok 10210, Thailand.
authors.forename: Daranee
authors.initials: D
authors.lastname: Chokchaichamnankit
authors.affiliation: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok 10210, Thailand.
authors.forename: Amnart
authors.initials: A
authors.lastname: Khongmanee
authors.affiliation: Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok 10210, Thailand.
authors.forename: Phannee
authors.initials: P
authors.lastname: Sawangareetrakul
authors.affiliation: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok 10210, Thailand.
authors.forename: Jisnuson
authors.initials: J
authors.lastname: Svasti
authors.affiliation: Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok 10210, Thailand.
authors.forename: Chantragan
authors.initials: C
authors.lastname: Srisomsap
mesh_terms: D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D018929:Cell Culture Techniques; D018281:Cholangiocarcinoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D017077:Culture Media, Conditioned; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D020543:Proteome / Q000235:genetics*; D053719:Tandem Mass Spectrometry; D059016:Tumor Microenvironment / Q000235:genetics*
chemical_list: D017077:Culture Media, Conditioned; D009363:Neoplasm Proteins; D020543:Proteome