title: [Sorting of CD133(+) subset cells in human gastric cancer and the identification of their tumor initiating cell-like properties].
abstract: OBJECTIVE
  To sort CD133(+) subset cells in human gastric cancer (GC) and to identify their tumor initiating cell-like properties.
  
  
  METHODS
  The tissues of GC and normal tissues adjacent to GC were obtained from 50 patients. Samples were stained for CD133 by immunohistochemistry. Likewise, assessments of CD133 were undertaken by Western blot. Flow cytometry was used to determine the proportion of CD133(+) cells in four GC cell lines therein the KATO-III was sorted by magnetic activated cell sorting (MACS) method. The growing characteristics and the tumorigenic ability of CD133(+) cells were evaluated in vitro and in vivo. Meanwhile, the growth of single cells in suspension culture was observed and expression of stem cell-specific marker were determined using reverse transcription-polymerase chain reaction (RT-PCR).
  
  
  RESULTS
  The expression of CD133 was demonstrated on the cell membranes in the mucosa and submucosa of primary GC, which were higher than those in the normal gastric tissues adjacent to cancer (P<0.05). Four GC cell lines including KATO-III, SGC-7901, AGS and MKN-45 were found to contain (28 ± 2)%, (17 ± 2)%, (6 ± 2)%, and (4 ± 2)% of CD133(+) cells respectively. In addition, the purity of CD133(+) cells isolated from KATO-III by MACS was (91 ± 3)% and up to(95 ± 2)% after 1-week culture. CCK-8 detection showed that population doubling time of the CD133(+) cells was (21 ± 3)h, significantly shorter than that of the CD133(-) cells[(40 ± 8)h, P<0.05]. Notably, there was a remarkable difference of tumor formation rate between CD133(+) cells (100%), non-sorted cells (80%), and CD133(-) cells(0). The average mass and volume of tumor in group of CD133(+) cells was larger and heavier than those in non-sorted cells (P<0.05, P<0.05). Furthermore, the single cell proliferated well, formed the big sphere and semi-quantitative RT-PCR showed expression of stem cell markers such as Oct-4, Nanog, Sox-2, Musashi-1 and EGFR.
  
  
  CONCLUSIONS
  CD133 protein expression in primary lesions is higher than those in the normal gastric tissues. CD133(+) subset cells can be isolated, purified, and amplified in human GC, and possess some properties including the ability of self-renewal, proliferation, and higher tumorigenic ability in vivo and can express some stem cell markers.
authors.affiliation: Department of General Surgery, Shanghai Jiaotong University School of Medicine, Shanghai, China. Jiang_md@hotmail.com
authors.forename: Rui-qi
authors.initials: RQ
authors.lastname: Lu
authors.forename: Ju-gang
authors.initials: JG
authors.lastname: Wu
authors.forename: Guo-cai
authors.initials: GC
authors.lastname: Zhou
authors.forename: Hai-guang
authors.initials: HG
authors.lastname: Jiang
authors.forename: Ji-wei
authors.initials: JW
authors.lastname: Yu
authors.forename: Bo-jian
authors.initials: BJ
authors.lastname: Jiang
mesh_terms: D000071916:AC133 Antigen; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D045744:Cell Line, Tumor; D005260:Female; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008875:Middle Aged; D014411:Neoplastic Stem Cells / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism*; D013274:Stomach Neoplasms / Q000378:metabolism / Q000473:pathology*; D023041:Xenograft Model Antitumor Assays
chemical_list: D000071916:AC133 Antigen; D015703:Antigens, CD; D006023:Glycoproteins; C000606023:PROM1 protein, human; D010455:Peptides; C000606024:Prom1 protein, mouse