title: Inflammation-related DNA damage and expression of CD133 and Oct3/4 in cholangiocarcinoma patients with poor prognosis.
abstract: Nitrative and oxidative DNA damage plays an important role in inflammation-related carcinogenesis. Chronic inflammation such as parasite infection and primary sclerosing cholangitis can be an etiological factor of cholangiocarcinoma. Using a proteomic approach and double-fluorescent staining, we identified high expression and colocalization of albumin and cytokeratin-19 in liver fluke-associated cholangiocarcinoma tissues, compared with normal livers from cholangiocarcinoma patients and cadaveric donors, respectively. Albumin was detected not only in cells of hyperplastic bile ducts and cholangiocarcinoma, but also in liver stem/progenitor cell origin, such as canal of Hering, ductules, and ductular reactions, suggesting the involvement of stem/progenitor cells in cholangiocarcinoma development. To clarify the involvement of liver stem/progenitor cells in cholangiocarcinoma, we examined several stem/progenitor cell markers (CD133, CD44, OV6, and Oct3/4) in cholangiocarcinoma tissues analyzed by immunohistochemical staining, and measured 8-oxodG levels by using HPLC-ECD as an inflammation-related DNA lesion. In addition, a stem/progenitor cell factor Bmi1, 8-nitroguanine (formed during nitrative DNA damage), DNA damage response (DDR) proteins (phosphorylated ATM and γ-H2AX), and manganese-SOD (Mn-SOD) were analyzed by immunohistochemistry. Stem/progenitor cell markers (CD133, OV6, CD44, and Oct3/4) were positively stained in 56, 38, 47, and 56% of 34 cholangiocarcinoma cases, respectively. Quantitative analysis of 8-oxodG revealed significantly increased levels in CD133- and/or Oct3/4-positive tumor tissues compared to negative tumor tissues, as well as 8-nitroguanine formation detected by immunohistochemistry. In the cases of CD44- and/or OV6-positive tissue, no significant difference was observed. Cholangiocarcinoma patients with CD133- and/or Oct3/4-positive tumor tissues showed significantly lower expression of Mn-SOD and higher DDR protein, γ-H2AX. Moreover, CD133- and/or Oct3/4-positive cholangiocarcinoma patients had significant associations with tumor histology types, tumor stage, and poor prognoses. Our results suggest that CD133 and Oct3/4 in cholangiocarcinoma are associated with increased formation of DNA lesions and the DDR protein, which may be involved in genetic instability and lead to cholangiocarcinoma development with aggressive clinical features.
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan; Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Faculty of Pharmaceutical Sciences, Suzuka University of Medical Science, Suzuka, Mie, 513-8670, Japan.
authors.forename: Raynoo
authors.initials: R
authors.lastname: Thanan
authors.affiliation: Department of Pathology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Chawalit
authors.initials: C
authors.lastname: Pairojkul
authors.affiliation: Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Somchai
authors.initials: S
authors.lastname: Pinlaor
authors.affiliation: Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Department of Surgery, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Narong
authors.initials: N
authors.lastname: Khuntikeo
authors.affiliation: Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Chaisiri
authors.initials: C
authors.lastname: Wongkham
authors.affiliation: Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Banchob
authors.initials: B
authors.lastname: Sripa
authors.affiliation: Faculty of Health Science, Suzuka, Mie, 510-0293, Japan.
authors.forename: Ning
authors.initials: N
authors.lastname: Ma
authors.affiliation: Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Kulthida
authors.initials: K
authors.lastname: Vaeteewoottacharn
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan; Faculty of Pharmaceutical Sciences, Suzuka University of Medical Science, Suzuka, Mie, 513-8670, Japan.
authors.forename: Ayako
authors.initials: A
authors.lastname: Furukawa
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan.
authors.forename: Hatasu
authors.initials: H
authors.lastname: Kobayashi
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Hiraku
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan.
authors.forename: Shinji
authors.initials: S
authors.lastname: Oikawa
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan; Faculty of Pharmaceutical Sciences, Suzuka University of Medical Science, Suzuka, Mie, 513-8670, Japan.
authors.forename: Shosuke
authors.initials: S
authors.lastname: Kawanishi
authors.affiliation: Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. Electronic address: puangrat@kku.ac.th.
authors.forename: Puangrat
authors.initials: P
authors.lastname: Yongvanit
authors.affiliation: Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507, Japan. Electronic address: mmurata@doc.medic.mie-u.ac.jp.
authors.forename: Mariko
authors.initials: M
authors.lastname: Murata
mesh_terms: D000080242:8-Hydroxy-2'-Deoxyguanosine; D000071916:AC133 Antigen; D000418:Albumins / Q000378:metabolism; D015703:Antigens, CD / Q000096:biosynthesis / Q000235:genetics*; D000943:Antigens, Differentiation / Q000096:biosynthesis; D001650:Bile Duct Neoplasms / Q000473:pathology*; D001653:Bile Ducts, Intrahepatic / Q000473:pathology; D002471:Cell Transformation, Neoplastic / Q000276:immunology; D018281:Cholangiocarcinoma / Q000473:pathology*; D015209:Cholangitis, Sclerosing; D004249:DNA Damage / Q000235:genetics* / Q000276:immunology; D004260:DNA Repair; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000032:analysis; D005260:Female; D042822:Genomic Instability / Q000235:genetics*; D006023:Glycoproteins / Q000096:biosynthesis / Q000235:genetics*; D006147:Guanine / Q000031:analogs & derivatives / Q000032:analysis / Q000096:biosynthesis; D006657:Histones / Q000096:biosynthesis; D006801:Humans; D018960:Hyaluronan Receptors / Q000096:biosynthesis; D007249:Inflammation / Q000276:immunology; D053539:Keratin-19 / Q000378:metabolism; D008099:Liver / Q000166:cytology; D008297:Male; D008875:Middle Aged; D050814:Octamer Transcription Factor-3 / Q000096:biosynthesis / Q000235:genetics*; D010084:Oxidation-Reduction; D010455:Peptides / Q000235:genetics*; D063150:Polycomb Repressive Complex 1 / Q000032:analysis; D011379:Prognosis; D013234:Stem Cells / Q000166:cytology; D013482:Superoxide Dismutase
chemical_list: C095838:8-nitroguanine; D000071916:AC133 Antigen; D000418:Albumins; D015703:Antigens, CD; D000943:Antigens, Differentiation; C069077:BMI1 protein, human; C497483:CD44 protein, human; D006023:Glycoproteins; C468783:H2AX protein, human; D006657:Histones; D018960:Hyaluronan Receptors; D053539:Keratin-19; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; C000606023:PROM1 protein, human; D010455:Peptides; C115011:oval cell marker OV-6; D006147:Guanine; D000080242:8-Hydroxy-2'-Deoxyguanosine; D013482:Superoxide Dismutase; D063150:Polycomb Repressive Complex 1; D003849:Deoxyguanosine
keywords: CD133; CD44; Cholangiocarcinoma; Clusters of differentiation 44; HPLC-ECD; High pressure liquid chromatography coupled with electrochemical detector; Inflammation; OV6; Oct3/4; Octamer-binding transcription factor 3 or 4; Oval marker 6; Oxidative stress; Stem/progenitor cell; clusters of differentiation 133