Twist expression promotes migration and invasion in hepatocellular carcinoma.
BACKGROUND
Twist, a transcription factor of the basic helix-loop-helix class, is reported to regulate cancer metastasis. It is known to induce epithelial-mesenchymal transition (EMT). In this study, we evaluated the expression of twist and its effect on cell migration in hepatocellular carcinoma (HCC).


METHODS
We examined twist expression using immunohistochemistry in 20 tissue samples of hepatocellular carcinoma, and assessed twist expression in HCC cell lines by RT-PCR and Western blot analysis. Ectopic twist expression was created by introducing a twist construct in the twist-negative HCC cell lines. Endogenous twist expression was blocked by twist siRNA in the twist-positive HCC cell lines. We studied EMT related markers, E-cadherin, Vimentin, and N-cadherin by Western blot analysis. Cell proliferation was measured by MTT assay, and cell migration was measured by in vitro wound healing assay. We used immunofluorescent vinculin staining to visualize focal adhesion.


RESULTS
We detected strong and intermediate twist expression in 7 of 20 tumor samples, and no significant twist expression was found in the tumor-free resection margins. In addition, we detected twist expression in HLE, HLF, and SK-Hep1 cells, but not in PLC/RPF/5, HepG2, and Huh7 cells. Ectopic twist-expressing cells demonstrated enhanced cell motility, but twist expression did not affect cell proliferation. Twist expression induced epithelial-mesenchymal transition together with related morphologic changes. Focal adhesion contact was reduced significantly in ectopic twist-expressing cells. Twist-siRNA-treated HLE, HLF, and SK-Hep1 cells demonstrated a reduction in cell migration by 50, 40 and 18%, respectively.


CONCLUSIONS
Twist induces migratory effect on hepatocellular carcinoma by causing epithelial-mesenchymal transition.
Matsuo
Noriyuki
N
Department of Gastroenterology and Hepatology, Okayama University Graduate School of Medicine and Dentistry, 2-5-1 Shikata-cho, Okayama 700-8558, Japan. ayakouharujp@yahoo.co.jp
Shiraha
Hidenori
H
Fujikawa
Tatsuya
T
Takaoka
Nobuyuki
N
Ueda
Naoki
N
Tanaka
Shigetomi
S
Nishina
Shinichi
S
Nakanishi
Yutaka
Y
Uemura
Masayuki
M
Takaki
Akinobu
A
Nakamura
Shinichiro
S
Kobayashi
Yoshiyuki
Y
Nouso
Kazuhiro
K
Yagi
Takahito
T
Yamamoto
Kazuhide
K
D000368:Aged; D000369:Aged, 80 and over; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D002448:Cell Adhesion / Q000502:physiology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology*; D005260:Female; D020868:Gene Silencing; D006801:Humans; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009361:Neoplasm Invasiveness; D009687:Nuclear Proteins / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D051879:Twist-Related Protein 1 / Q000096:biosynthesis* / Q000235:genetics
D009687:Nuclear Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C103173:TWIST1 protein, human; D051879:Twist-Related Protein 1