title: The specific linker phosphorylation of Smad2/3 indicates epithelial stem cells in stomach; particularly increasing in mucosae of Helicobacter-associated gastritis.
abstract: BACKGROUND
  The gastric corpus and antrum are believed to contain epithelial stem cells in the isthmus. However, the lack of useful markers has hindered studies of their origin. We explored whether Smad2/3, phosphorylated at specific linker threonine residues (pSmad2/3L-Thr), could serve as a marker for stem cells.
  
  
  METHODS
  Stomachs, small intestines, and colons from Helicobacter felis-infected and noninfected C57BL/6 mice were examined. Double immunofluorescent staining of pSmad2/3L-Thr with Ki67, cytokeratin 8, or doublecortin and calcium/calmodulin-dependent protein kinase-like-1 (DCAMKL1) was performed, and pSmad2/3L-Thr immunostaining-positive cells were counted. After immunofluorescent staining, we stained the same sections with hematoxylin-eosin and observed these cells under a light microscope.
  
  
  RESULTS
  In infected mice, pSmad2/3L-Thr immunostaining-positive cells were significantly increased in the corpus and antrum compared with those of noninfected mice (p < 0.0001). The number of Ki67 immunostaining-positive cells in the corpus and antrum of infected mice was also much greater than in the noninfected mice. Although pSmad2/3L-Thr immunostaining-positive cells were detected among the Ki67 cells, immunohistochemical co-localization of pSmad2/3L-Thr with Ki67 was never observed. pSmad2/3L-Thr immunostaining-positive cells showed immunohistochemical co-localization with cytokeratin 8, but some of them showed co-localization or adjacent localization with DCAMKL1 immunostaining-positive cells. Under a light microscope, pSmad2/3L-Thr immunostaining-positive cells indicated undifferentiated morphological features and were confirmed in the isthmus. In small intestines and colons, pSmad2/3L-Thr immunostaining-positive cells were detected in specific epithelial cells around crypt bases, where the respective putative stem cells are thought to exist.
  
  
  CONCLUSIONS
  We have identified the significant expression of pSmad2/3L-Thr in specific epithelial cells of the murine stomach and have suggested these cells to be epithelial stem cells.
authors.affiliation: The Third Department of Internal Medicine, Division of Gastroenterology and Hepatology, Kansai Medical University, 10-15 Fumizono-cho, Moriguchi, Osaka 570-8506, Japan. fukuitos@takii.kmu.ac.jp
authors.forename: Toshiro
authors.initials: T
authors.lastname: Fukui
authors.forename: Masanobu
authors.initials: M
authors.lastname: Kishimoto
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nakajima
authors.forename: Masao
authors.initials: M
authors.lastname: Yamashina
authors.forename: Shinji
authors.initials: S
authors.lastname: Nakayama
authors.forename: Takeo
authors.initials: T
authors.lastname: Kusuda
authors.forename: Yutaku
authors.initials: Y
authors.lastname: Sakaguchi
authors.forename: Katsunori
authors.initials: K
authors.lastname: Yoshida
authors.forename: Kazushige
authors.initials: K
authors.lastname: Uchida
authors.forename: Akiyoshi
authors.initials: A
authors.lastname: Nishio
authors.forename: Koichi
authors.initials: K
authors.lastname: Matsuzaki
authors.forename: Kazuichi
authors.initials: K
authors.lastname: Okazaki
mesh_terms: D000818:Animals; D003106:Colon / Q000378:metabolism / Q000382:microbiology; D004847:Epithelial Cells / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D005753:Gastric Mucosa / Q000378:metabolism / Q000382:microbiology / Q000473:pathology; D005756:Gastritis / Q000382:microbiology / Q000473:pathology; D016481:Helicobacter Infections / Q000382:microbiology / Q000473:pathology; D044922:Helicobacter felis / Q000302:isolation & purification; D007421:Intestine, Small / Q000378:metabolism / Q000382:microbiology; D019394:Ki-67 Antigen / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010766:Phosphorylation; D051899:Smad2 Protein / Q000378:metabolism*; D051900:Smad3 Protein / Q000378:metabolism*; D013234:Stem Cells / Q000378:metabolism*; D013270:Stomach / Q000382:microbiology
chemical_list: D019394:Ki-67 Antigen; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D051900:Smad3 Protein; C497791:Smad3 protein, mouse