Gastroenterology

Gastroenterology

Volume 128, Issue 1, January 2005, Pages 51-62
Gastroenterology

Basic-alimentary tract
Actinin-4 increases cell motility and promotes lymph node metastasis of colorectal cancer

https://doi.org/10.1053/j.gastro.2004.10.004Get rights and content

Background & Aims: Enhanced motility of cancer cells by remodeling of the actin cytoskeleton seems crucial in the process of cancer invasion and metastasis. We previously identified an actin-binding protein, actinin-4, as a new biomarker of cancer invasion and an indicator of prognosis for patients with breast cancer. However, its involvement in the mechanisms of cancer invasion and metastasis remains undetermined. The current study tested the role of actinin-4 in the motility and metastatic potential of colorectal cancer cells. Methods & Results: Quantitative immunofluorescence histochemistry showed that the expression level of the actinin-4 protein was increased in 73.1% (19/26) of the cases of colorectal cancer over the corresponding normal intestinal epithelium. The increased expression of actinin-4 was most significant in dedifferentiated cancer cells at the invasive front. A colorectal cancer cell clone capable of inducing actinin-4 using the tetracycline-regulatory system (designated DLD1 Tet-off ACTN-4) was established. Upon the induction of actinin-4, DLD1 Tet-off ACTN-4 cells spread filopodia and significantly increased their motility (P = .00027); actinin-4 protein was concentrated at the leading edges of these actin-rich podia. When injected into the mesocecum of severe combined immunodeficient mice, DLD1 Tet-off ACTN4 cells, but not the control cells, metastasized into regional mesenteric lymph nodes, resembling the behavior of clinical cancers. The expression of actinin-4 in focally dedifferentiated cancer cells at the invasive front was significantly correlated with the frequency of lymph node metastasis of colorectal cancer (P = .038). Conclusions: Actinin-4 actively increases cell motility and promotes lymph node metastasis of colorectal cancer.

Section snippets

Antibodies

Anti-hemagglutinin (HA) rat monoclonal antibody (clone 3F10) was purchased from Roche Diagnostics GmbH (Mannheim, Germany). Anti-actin mouse monoclonal antibody (clone AC-15) was purchased from Abcam Inc (Cambridge, England). Anti-E-cadherin mouse monoclonal HECD1 and anti-actinin-4 rabbit polyclonal (Ab-1) antibodies were produced in our laboratory, as described previously.16, 23 Another anti-actinin-4 polyclonal (Ab-2) antibody was newly raised against the synthetic peptide NQSYQYGPSSAGNGAGC

Increased expression of actinin-4 protein in colorectal cancer

The expression of actinin-4 protein was examined in 26 cases of colorectal carcinoma using immunofluorescence histochemistry (Figure 1). The expression level of actinin-4 was increased in the cancer cells when compared with normal intestinal epithelial cells in the same specimens (Figure 1A and B). We measured the fluorescence intensity along lines of the same length placed on 3 randomly selected normal colonic crypts and 3 cancerous glands in each case and found that the expression of

Discussion

Cell movement can be viewed as a dynamic multistep cycle of (1) extension of podia, (2) attachment of the leading edge of podia to the substrate, (3) forward retraction of the cell body, and (4) detachment of rear adhesions.30 Although the molecular mechanisms of podia formation are not perfectly understood, a certain set of actin-binding proteins seems to be involved, because filamentous actin is highly concentrated in the leading edge of podia.10, 11, 12, 13 In this study, we showed that DLD1

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    Supported by grants from the Ministry of Health, Labor and Welfare and by the Program for the Promotion of Fundamental Studies in Health Sciences of the Organization for Pharmaceutical Safety and Research of Japan. Y. H. is a recipient of the Research Resident Fellowship from the Foundation for the Promotion of Cancer Research.

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