Plasma proteomics of pancreatic cancer patients by multi-dimensional liquid chromatography and two-dimensional difference gel electrophoresis (2D-DIGE): up-regulation of leucine-rich alpha-2-glycoprotein in pancreatic cancer.
We investigated the aberrant expression of plasma proteins in patients with pancreatic cancer. High-abundance plasma proteins (albumin, transferrin, haptoglobin, alpha-1-antitrypsin, IgG and IgA) were depleted by use of an immuno-affinity column, and low-abundance ones were separated into five fractions by anion-exchange chromatography. The fractionated plasma proteins were subjected to 2D-DIGE with highly sensitive fluorescent dyes. The quantitative protein expression profiles obtained by 2D-DIGE were compared between two plasma protein mixtures: one from five non-cancer bearing healthy donors and the other from five patients with pancreatic cancer. Among 1200 protein spots, we found that 33 protein spots were differently expressed between the two mixtures; 27 of these were up-regulated and six were down-regulated in cancer. Mass spectrometry and database searching allowed the identification of the proteins corresponding to the gel spots. Up-regulation of leucine-rich alpha-2-glycoprotein (LRG), which has not previously been implicated in pancreatic cancer, was observed. Western blotting with an anti-LRG antibody validated the up-regulation of LRG in an independent series of plasma samples from healthy controls, patients with chronic pancreatitis, and patients with pancreatic cancer. Our results demonstrate the application of a combination of multi-dimensional liquid chromatography with 2D-DIGE for plasma proteomics and suggest the clinical utility of LRG plasma level measurement.
Kakisaka
Tatsuhiko
T
Proteome Bioinformatics Project, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan.
Kondo
Tadashi
T
Okano
Tetsuya
T
Fujii
Kiyonaga
K
Honda
Kazufumi
K
Endo
Mitsufumi
M
Tsuchida
Akihiko
A
Aoki
Tatsuya
T
Itoi
Takao
T
Moriyasu
Fuminori
F
Yamada
Tesshi
T
Kato
Harubumi
H
Nishimura
Toshihide
T
Todo
Satoru
S
Hirohashi
Setsuo
S
D000368:Aged; D001798:Blood Proteins / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005260:Female; D005456:Fluorescent Dyes; D006023:Glycoproteins / Q000097:blood*; D006801:Humans; D008297:Male; D008875:Middle Aged; D010190:Pancreatic Neoplasms / Q000097:blood*; D040901:Proteomics; D015854:Up-Regulation
D001798:Blood Proteins; D005456:Fluorescent Dyes; D006023:Glycoproteins; C015246:LRG1 protein, human