Biological characterization of three immortalized esophageal epithelial cell lines

Zeng,Fa‑Min; Xie,Yang‑Min; Liao,Lian‑Di; Li,Li‑Yan; Chen,Bo; Xie,Jian‑Jun; Xu,Li‑Yan; Li,En‑Min

doi:10.3892/mmr.2016.5813

Biological characterization of three immortalized esophageal epithelial cell lines

Authors:
- Fa‑Min Zeng
- Yang‑Min Xie
- Lian‑Di Liao
- Li‑Yan Li
- Bo Chen
- Jian‑Jun Xie
- Li‑Yan Xu
- En‑Min Li
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Affiliations: Key Laboratory of Molecular Biology for High Cancer Incidence in The Coastal Chaoshan Area, Medical College of Shantou University, Shantou, Guangdong 515041, P.R. China
Published online on: October 6, 2016 https://doi.org/10.3892/mmr.2016.5813
Pages: 4802-4810

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Abstract

The key molecular events that contribute to tumorigenesis are incompletely understood. The aim of the present study was to characterize and compare the biological phenotypes of three human telomerase reverse transcriptase (hTERT) and/or human papillomavirus 16 E6 and E7‑immortalized esophageal epithelial cell lines, NE2‑hTERT (NE2), NE3‑E6E7‑hTERT (NE3) and NEcA6‑E6E7‑hTERT (NEcA6). The present study used soft‑agar colony formation assays, tumorigenicity assays in nude mice, and cell proliferation, adhesion and migration assays to identify the biological characteristics of NE2, NE3 and NEcA6 cells. NE2 and NE3 cells exhibited characteristics of benign cells, such as the inability to grow in soft agar or form tumors in nude mice. By contrast, NEcA6 cells had undergone transformation, as demonstrated by the ability to grow in soft agar and form tumors in nude mice. In addition, NEcA6 cells exhibited increased migration and adhesion capabilities when compared with NE2 and NE3 cells. In order to identify mechanism(s) that may contribute to the altered biological phenotypes exhibited by these cells, the expression of three proteins involved in modulating cell migration [fascin, ezrin/radixin/moesin family proteins and phosphorylated‑focal adhesion kinase (Tyr 397)], as well as the expression status and subcellular localization of three key focal adhesions components (paxillin, talin and kindlin‑2) were examined. Paxillin, talin and kindlin‑2 were localized to adhesive sites that connect F‑actin with the extracellular matrix in transformed NEcA6 cells, but were distributed in a diffuse manner in NE2 and NE3 cells. Knockdown of kindlin‑2 in NE3 and NEcA6 cells decreased cell adhesion, however, NEcA6 cells demonstrated a greater sensitivity to knockdown of kindlin‑2. No significant differences were observed in the protein expression levels of fascin, exrin/radixin/moesin and p‑FAK in the three cell lines. In conclusion, these results demonstrate that these three focal adhesion components, particularly kindlin‑2, may contribute to the carcinogenesis of esophageal squamous cells.

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