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. 2005 Jan;96(1):42-7.
doi: 10.1111/j.1349-7006.2005.00007.x.

EPHA2/EFNA1 expression in human gastric cancer

Ritsuko Nakamura  1 , Hideki Kataoka, Naomi Sato, Masao Kanamori, Megumi Ihara, Hisaki Igarashi, Sanjar Ravshanov, You-Jie Wang, Zhong-You Li, Takahiro Shimamura, Toshihiko Kobayashi, Hiroyuki Konno, Kazuya Shinmura, Masamitsu Tanaka, Haruhiko Sugimura
Affiliations

EPHA2/EFNA1 expression in human gastric cancer

Ritsuko Nakamura et al. Cancer Sci. 2005 Jan.

Abstract

The erythropoietin-producing hepatocellular (EPH)A2 receptor, tyrosine kinase, is overexpressed and phosphorylated in several types of human tumors and has been associated with malignant transformation. A recent report, however, indicated that stimulation of the EPHA2 receptor ligand, ephrinA1 (EFNA1), inhibits the growth of EPHA2-expressing breast cancer. The authors examined the expression of EPHA2 and EFNA1 using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) in four gastric cancer cell lines and 49 primary gastric cancer samples, as well as in normal gastric tissue. EPHA2 was more highly expressed in tumor tissue than in normal tissue in 27 cases (55%). EFNA1 was overexpressed in tumor tissue in 28 cases (57%). No significant correlation was detected between the expression levels and histologic features such as tumor size, age, vessel invasion, or lymph node involvement. However, EPHA2 overexpression was more prominent in macroscopic type 3 and 4 tumors than in type 1 or 2 advanced gastric cancer. The authors observed EPHA2 expression in three of the four gastric cancer cell lines (AGS, KATO3, and MKN74) that were examined. In one cell line, TMK1, EPHA2 expression was barely detectable using northern blotting, RT-PCR, and western blotting. In contrast, EFNA1 was detected in all cell lines. In the gastric cancer cell lines that endogenously expressed EPHA2, stimulation with ephrinA1-Fc led to decreased EPHA2 protein expression and increased EPHA2 phosphorylation. Finally, the growth of EPHA2-expressing cells was inhibited by repetitive stimulation with soluble ephrinA1-Fc. Taken together, these findings suggest that EPHA2 and EFNA1 expression may influence the behavior of human gastric cancer.

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Figures

Figure 1
Figure 1
Semi‐quantitative reverse transcription‐polymerase chain reaction data of two cases of primary gastric cancer and corresponding non‐tumor tissue, and gastric cancer cell lines. (A) Both ephrinA1 (EFNA1) and EPHA2 were overexpressed in these tumors (cases 1 and 2). ACTB is shown as an internal control. (B) EFNA1 was expressed in all gastric cell lines, particularly in TMK1. EPHA2 was detected in these four cell lines, but expression was weak in TMK1 compared with the other lines. N, non‐tumorous gastric tissue; T gastric cancer tissue.
Figure 2
Figure 2
Immunohistochemistry of EPHA2 in primary gastric cancer tissue. (A) EPHA2 was detected in cytoplasm and cell membrane of tumor cells. ×400. (B) The adjacent section after absorption with EPHA2 blocking peptide. ×400. (C) Western blotting using anti‐EPHA2 antibody to cell lysates of 293T transfected with a mock (–) and an EPHA2 encoding plasmid (+). (D) Normal (black arrow) and intestinal metaplasia (red arrow) portion. ×400. (E,F) Gastric adenomas with (E) and without (F) EPHA2 immunoreactivity. (G) Western blotting of the primary tissues to anti‐EPHA2 antibody. Case 1 and 2 are the same cases 1 and 2 shown in Fig. 1A. The lysates were from N (non‐tumorous gastric tissue) and T (gastric cancer tissue).
Figure 3
Figure 3
EphrinA1 (EFNA1) and EPHA2 expression by northern blotting. (A) EFNA1 is expressed in all gastric cancer cell lines, particularly TMK1. (B) EPHA2 is clearly detected in AGS, KATO3, and MKN74, but very weakly in TMK1. These data were quantitatively consistent with those obtained using semiquantitative reverse transcription‐polymerase chain reaction, shown in Figure 1. 293T is shown as a negative control.
Figure 4
Figure 4
Phosphorylation and degradation of EPHA2 stimulated by ephrinA1‐Fc in MKN74 cells and other gastric cell lines. (A,B) MKN74 cells were stimulated by ephrinA1‐Fc for the indicated periods (A) and at the noted concentrations (B). Total cell lysates were separated using sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and immunoblotted with anti‐EPHA2 antibody (upper panels). The cell lysates were immunoblotted with antiphosphotyrosine antibody after immunoprecipitation with anti EPHA2 and separated using SDS–PAGE (lower panels). (C) EPHA2 expression after EFNA1 stimulation in the other gastric cancer cell lines. AGS and KATO3 also showed an increase in EPHA2 phosphorylation and degradation of EPHA2 by ephrinA1‐Fc.
Figure 5
Figure 5
EphrinA1‐Fc inhibited growth of cells expressing EPHA2, but not cells that did not express EPHA2. (A) 1 × 105 of MKN74 or (B) TMK1 were seeded on a six‐well dish and, after 24 h, fed with cell medium containing ephrinA1‐Fc. The medium was changed every 24 h for 3 days, and the cell numbers were counted using a hemacytometer daily. EFNA1 stimulation inhibited MKN74 cell growth. The bars show means ± SE of cell numbers (three or more experiments).
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References

    1. Bohme B, VandenBos T, Cerretti DP, Park LS, Holtrich U, Rubsamen‐Waigmann H, Strebhardt K. Cell‐cell adhesion mediated by binding of membrane‐anchored ligand LERK‐ 2 to the EPH‐related receptor human embryonal kinase 2 promotes tyrosine kinase activity. J Biol Chem 1996; 271: 24747–52. - PubMed
    1. Mellitzer G, Xu Q, Wilkinson DG. Eph receptors and ephrins restrict cell intermingling and communication. Nature 1999; 400: 77–81. - PubMed
    1. George SE, Simokat K, Hardin J, Chisholm AD. The VAB‐1 Eph receptor tyrosine kinase functions in neural and epithelial morphogenesis in C. elegans. Cell 1998; 92: 633–43. - PubMed
    1. Adams RH, Wilkinson GA, Weiss C, Diella F, Gale NW, Deutsch U, Risau W, Klein R. Roles of ephrinB ligands and EphB receptors in cardiovascular development: demarcation of arterial/venous domains, vascular morphogenesis, and sprouting angiogenesis. Genes Dev 1999; 13: 295–306. - PMC - PubMed
    1. Pandey A, Shao H, Marks RM, Polverini PJ, Dixit VM. Role of B61, the ligand for the Eck receptor tyrosine kinase, in TNF‐ alpha‐induced angiogenesis. Science 1995; 268: 567–9. - PubMed

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