Gastroenterology

Gastroenterology

Volume 118, Issue 1, January 2000, Pages 60-69
Gastroenterology

Alimentary Tract
Overexpression of intestinal trefoil factor in human colon carcinoma cells reduces cellular growth in vitro and in vivo,☆☆

https://doi.org/10.1016/S0016-5085(00)70414-8Get rights and content

Abstract

Background & Aims: Intestinal trefoil factor (ITF) has a role in gastrointestinal mucosal integrity and the repair of damaged mucosa. However, little is known about its role in tumors. To analyze the role of ITF in colon carcinomas, overexpression of the ITF gene in colon carcinoma cells was used. Methods: Human colon carcinoma cell lines LoVo and SW837, expressing no endogenous ITF, and WiDr expressing a low level of ITF were stably transfected with an expression vector harboring human ITF complementary DNA. The effects of ITF overexpression on in vitro growth, morphology in collagen gel, response to epidermal growth factor (EGF), mitogen-activated protein kinase (MAPK) activity, and growth in nude mice were assessed. Results: Overexpression of ITF in LoVo and SW837 resulted in significantly reduced growth in vitro and in vivo. In collagen gels, the ITF-expressing LoVo clones formed smaller, more dispersed colonies. EGF-induced phosphorylation of MAPKs was modestly reduced in the ITF-expressing clones. The growth of WiDr was modestly suppressed only in vivo by ITF overexpression. Conclusions: Overexpression of ITF suppressed the growth of colon carcinoma cells. ITF may function as an inhibitory factor for the growth of colonic neoplasm.

GASTROENTEROLOGY 2000;118:60-69

Section snippets

Cell culture

All human colon carcinoma cell lines were maintained in a growth medium of a mixture of RPMI 1640 and Ham's F-12 supplemented with 10% heat-inactivated fetal bovine serum. LoVo was obtained from Riken Cell Bank (Tsukuba, Japan). SW837, WiDr, Colo205, and Caco-2 were from Dainihon Seiyaku (Osaka, Japan), and Colo320DM was from Japanese Cancer Research Resources Bank (Osaka, Japan). For 3-dimensional (3-D) collagen gel culture, 0.1% collagen type I gel (Koken, Tokyo, Japan) in the growth medium

Selection of ITF-expressing LoVo clones and effect of ITF overexpression on in vitro culture morphology and growth

An RT-PCR study revealed that 3 (SW837, Colo320DM, and LoVo) of 7 colon carcinoma cell lines examined did not express endogenous ITF (Figure 1A).

. (A) RT-PCR study for ITF mRNA of various colon carcinoma cell lines. (B) Representative results of RNA blot analysis for ITF mRNA of LoVo clones. pCIneo-ITF transfectants expressed ITF mRNA of molecular size around 0.75 kilobases (kb). Neither parent nor mock-transfected clones expressed ITF mRNA. ITF-29 expressed the most abundant ITF mRNA, and the

Discussion

The current view on the role of trefoil family peptides indicates that these peptides have an important role in promoting repair of damaged mucosa and differentiation of gastrointestinal epithelial cells. Although the precise mechanism by which trefoil peptides promote repair of damaged mucosa remains to be clarified, the migration-stimulatory activities of these peptides are thought to be important because the repair of ulcerated areas involves migration of epithelial cells over the ulcerated

Acknowledgements

The authors thank Drs. H. Maruyama, Nagoya City University, and Y. Nawa, Miyazaki Medical College, for generating anti-ITF antiserum; N. Iwakiri for skillful technical assistance; and Dr. K. Nabeshima for useful discussion and comment.

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    Address requests for reprints to: Masashi Koono, M.D., Second Department of Pathology, Miyazaki Medical College, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan. Fax: (81) 985-85-6003.

    ☆☆

    Supported in part by a grant from the Tokyo Biochemical Research Foundation, Japan, a grant from the Kurozumi Medical Foundation, and a grant-in-aid for research project from Miyazaki Medical College.

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