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Identification of Stem Cells in the Epithelium of the Stomach Corpus and Antrum of Mice. (Gastroenterology, Sep 2016)

Matsuo J1, Kimura S2, Yamamura A2, Koh CP1, Hossain MZ1, Heng DL1, Kohu K1, Chih-Cheng Voon D3, Hiai H4, Unno M5, Yan So JB6, Zhu F7, Srivastava S1, Meng T8, Yeoh KG9, Osato M10, Ito Y11

1Cancer Science Institute of Singapore, National University of Singapore, 14 Medical Drive, Singapore 117599.
2Cancer Science Institute of Singapore, National University of Singapore, 14 Medical Drive, Singapore 117599; Department of Surgery, Graduate School of Medicine, Tohoku University, 1-1 Seiryo-machi, Aoba-ku, Miyagi, Japan 9808574.
3Cancer Science Institute of Singapore, National University of Singapore, 14 Medical Drive, Singapore 117599; Institute for Frontier Science Initiative and Division of Genetics, Cancer Research Institute, Kanazawa University, Kanazawa, Ishikawa, Japan.
4Kyoto Disease Model Institute, Kyoto Science and Technology Center, Rm 22, 14 Yoshidakawaramachi, Sakyo-ku, Kyoto 606-8305.
5Department of Surgery, Graduate School of Medicine, Tohoku University, 1-1 Seiryo-machi, Aoba-ku, Miyagi, Japan 9808574.
6Department of Surgery, Yong Loo Lin School of Medicine, National University of Singapore C/O NUHS Tower Block 1E Kent Ridge Road, Singapore 119228.
7Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, C/O NUHS Tower Block IE Kent Ridge Road, Singapore 119228.
8Department of Pathology, National University Health System, 1E Kent Ridge Road, Singapore 119228.
9Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, C/O NUHS Tower Block IE Kent Ridge Road, Singapore 119228; Department of Gastroenterology and Hepatology, National University Health System, 1E Kent Ridge Road, Singapore 119228.
10Cancer Science Institute of Singapore, National University of Singapore, 14 Medical Drive, Singapore 117599; Institute of Bioengineering and Nanotechnology, A*STAR, 31 Biopolis way, Singapore 138669; Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, 14 Medical Drive, Singapore 119228; International Research Center for Medical Sciences, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto 860-0811, Japan. Electronic address: csimo@nus.edu.sg.
11Cancer Science Institute of Singapore, National University of Singapore, 14 Medical Drive, Singapore 117599; Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, C/O NUHS Tower Block IE Kent Ridge Road, Singapore 119228. Electronic address: csiitoy@nus.edu.sg.

Abstract:

BACKGROUND & AIMS:

Little is known about mechanisms of gastric carcinogenesis, partly because it has been a challenge to identify characterize gastric stem cells. Runx genes regulate development and their products are transcription factors associated with cancer development. A Runx1 enhancer element, eR1 is a marker of hematopoietic stem cells. We studied expression from eR1 in stomach and the roles of gastric stem cells in gastric carcinogenesis in transgenic mice.

METHODS:

We used in situ hybridization and immunofluorescence analyses to study expression of Runx1 in gastric tissues from C57BL/6 (control) mice. We then created mice that expressed enhanced green fluorescent protein (EGFP) or CreERT2 under the control of eR1 (eR1-CreERT2;Rosa-LSL-tdTomato, eR1-CreERT2;Rosa-LSL-EYFP mice). Gastric tissues were collected and lineage-tracing experiments were performed. Gastric organoids were cultured from eR1-CreERT2(5-2);Rosa-LSL-tdTomato mice and immunofluorescence analyses were performed. We investigated the effects of expressing oncogenic mutations in stem cells under control of eR1 using eR1-CreERT2;LSL-KrasG12D/+ mice; gastric tissues were collected and analyzed by histology and immunofluorescence.

RESULTS:

Most proliferation occurred in the isthmus; 86% of proliferating cells were RUNX1 positive and 76% were MUC5AC positive. In eR1-EGFP mice, EGFP signals were mainly detected in the upper part of the gastric unit, and 83% of EGFP-positive cells were located in the isthmus/pit region. We found that eR1 marked undifferentiated stem cells in the isthmus and a smaller number of terminally differentiated chief cells at the base. eR1 also marked cells in the pyloric gland in the antrum. Lineage tracing experiments demonstrated that stem cells in the isthmus and antrum continuously gave rise to mature cells to maintain the gastric unit. eR1-positive cells in the isthmus and pyloric gland generated organoid cultures in vitro. In eR1-CreERT2;LSL-Kras G12D/+ mice, MUC5AC-positive cells rapidly differentiated from stem cells in the isthmus, resulting in distinct metaplastic lesions similar to that observed in human gastric atrophy.

CONCLUSIONS:

Using lineage tracing experiments in mice, we found that a Runx1 enhancer element, eR1, promotes its expression in the isthmus stem cells of stomach corpus as well; as pyloric gland in the antrum. We were able to use eR1 to express oncogenic mutations; in gastric stem cells, proving a new model for studies of gastric carcinogenesis.

PMID: 27670082