title: Characterization of the human gastric fluid proteome reveals distinct pH-dependent protein profiles: implications for biomarker studies.
abstract: Gastric fluid is a source of gastric cancer biomarkers. However, very little is known about the normal gastric fluid proteome and its biological variations. In this study, we performed a comprehensive analysis of the human gastric fluid proteome using samples obtained from individuals with benign gastric conditions. Gastric fluid proteins were prefractionated using ultracentrifuge filters (3 kDa cutoff) and analyzed by two-dimensional gel electrophoresis (2-DE) and multidimensional LC-MS/MS. Our 2-DE analysis of 170 gastric fluid samples revealed distinct protein profiles for acidic and neutral samples, highlighting pH effects on protein composition. By 2D LC-MS/MS analysis of pooled samples, we identified 284 and 347 proteins in acidic and neutral samples respectively (FDR ≤1%), of which 265 proteins (72.4%) overlapped. However, unlike neutral samples, most proteins in acidic samples were identified from peptides in the filtrate (i.e., <3 kDa). Consistent with this finding, immunoblot analysis of six potential gastric cancer biomarkers rarely detected full-length proteins in acidic samples. These findings have important implications for biomarker studies because a majority of gastric cancer patients have neutral gastric fluid compared to noncancer controls. Consequently, sample stratification, choice of proteomic approaches, and validation strategy can profoundly affect the interpretation of biomarker findings. These observations should help to refine gastric fluid biomarker studies.
authors.affiliation: Division of Medical Sciences, Humphrey Oei Institute of Cancer Research, National Cancer Centre , Singapore.
authors.forename: Siok Yuen
authors.initials: SY
authors.lastname: Kam
authors.forename: Thomas
authors.initials: T
authors.lastname: Hennessy
authors.forename: Seow Ching
authors.initials: SC
authors.lastname: Chua
authors.forename: Chee Sian
authors.initials: CS
authors.lastname: Gan
authors.forename: Robin
authors.initials: R
authors.lastname: Philp
authors.forename: Ka Ka
authors.initials: KK
authors.lastname: Hon
authors.forename: Liyun
authors.initials: L
authors.lastname: Lai
authors.forename: Weng Hoong
authors.initials: WH
authors.lastname: Chan
authors.forename: Hock Soo
authors.initials: HS
authors.lastname: Ong
authors.forename: Wai Keong
authors.initials: WK
authors.lastname: Wong
authors.forename: Kiat Hon
authors.initials: KH
authors.lastname: Lim
authors.forename: Khoon Lin
authors.initials: KL
authors.lastname: Ling
authors.forename: Hwee Sian
authors.initials: HS
authors.lastname: Tan
authors.forename: Mei Mei
authors.initials: MM
authors.lastname: Tan
authors.forename: Mengfatt
authors.initials: M
authors.lastname: Ho
authors.forename: Oi Lian
authors.initials: OL
authors.lastname: Kon
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D015180:Electrophoresis, Gel, Two-Dimensional; D004724:Endoscopy / Q000379:methods; D005260:Female; D005750:Gastric Juice / Q000378:metabolism*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting / Q000379:methods; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D008875:Middle Aged; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D014408:Biomarkers, Tumor; D020543:Proteome