Skip to main page content
U.S. flag

An official website of the United States government

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
Clinical Trial
. 2013 Aug 28;8(8):e72662.
doi: 10.1371/journal.pone.0072662. eCollection 2013.

miR-26a suppresses tumor growth and metastasis by targeting FGF9 in gastric cancer

Min Deng  1 , Hai-lin Tang, Xi-hong Lu, Mei-yuan Liu, Xiao-min Lu, Yi-xue Gu, Ji-fang Liu, Zhi-min He
Affiliations
Clinical Trial

miR-26a suppresses tumor growth and metastasis by targeting FGF9 in gastric cancer

Min Deng et al. PLoS One. .

Abstract

The role of miR-26a in cancer cells seemed controversial in previous studies. Until now, the role of miR-26a in gastric cancer remains undefined. In this study, we found that miR-26a was strongly downregulated in gastric cancer (GC) tissues and cell lines, and its expression levels were associated with lymph node metastasis and clinical stage, as well as overall survival and replase-free survival of GC. We also found that ectopic expression of miR-26a inhibited GC cell proliferation and GC metastasis in vitro and in vivo. We further identified a novel mechanism of miR-26a to suppress GC growth and metastasis. FGF9 was proved to be a direct target of miR-26a, using luciferase assay and western blot. FGF9 overexpression in miR-26a-expressing cells could rescue invasion and growth defects of miR-26a. In addition, miR-26a expression inversely correlated with FGF9 protein levels in GC. Taken together, our data suggest that miR-26a functions as a tumor suppressor in GC development and progression, and holds promise as a prognostic biomarker and potential therapeutic target for GC.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. miR-26a is frequently downregulated in human gastric cancer and correlates with GC prognosis.
(A) miR-26a was detected in 40 gastric cancer patients by qRT-PCR. Data are shown as log2 of the fold change in gastric cancer tissues (tumor) relative to adjacent normal tissues (normal). (B) Relative expression of miR-26a in 6 cell lines derived from gastric cancer and one nonmalignant gastric cell line (GES-1) was determined by qRT-PCR. Data are shown as log2 of the fold change in GC cell lines relative to GES-1. (C) miR-26a expression was analyzed in adjacent normal tissues (Normal) and GCs (Tumor) samples on the tissue microarrays by in situ hybridisation. Expression scores are shown as box plots. Representative images of miR-26a expression by in situ hybridization are shown. Original magnification: ×200. (D) Survival analysis of GC. OS and RFS curves for 126 GC patients with high or low miR-26a expression were constructed using the Kaplan-Meier method and evaluated using the log-rank test.
Figure 2
Figure 2. miR-26a inhibits cell migration, invasion, growth and soft agar colony formation, and induces cell apoptosis in GC.
(A, B) The wound healing assay (A) and invasion assay (B) of SGC-7901 and AGS cells infected with miR-26a or scramble lentivirus. The invasion assay was measured by way of Transwell assays with Matrigel. (C) The growth of SGC-7901 and AGS cells infected with miR-26a or scramble lentivirus was assayed. (D) Colony growth assays in soft agar were performed on SGC-7901 with overexpression of miR-26a or scramble. Representative images of the assays are shown (left panel). Original magnification: ×200. (E) Overexpression of miR-26a induces tumor cell apoptosis. SGC-7901 cells were infected with miR-26a or scramble lentivirus. The apoptotic cells were evaluated by Annexin V-FITC and propidium iodine staining and analyzed with FACS. All data are presented.as mean ± s.e.m from at least three separate experiments.
Figure 3
Figure 3. miR-26a inhibits GC cell growth and metastasis in vivo.
(A) Tumor growth in mouse xenograft models. SGC-7901 cells infected with miR-26a or scramble lentivirus were injected subcutaneously into nude mice. Tumor size was measured every 5 days. After 30 days, the mice were killed, necropsies were performed, and tumors were weighed. (B) Tumor metastasis in mouse xenograft models. SGC-7901 cells with overexpression of miR-26a or scramble were injected into the tail vein of nude mice. After 45 days, the mice were killed. micrometastases in lung per HE-stained section in individual mice were calculated. Each group had six mice. Original magnification, ×100; scale bar: 50 µm. All data are shown as mean±s.e.m.
Figure 4
Figure 4. miR-26a directly targets FGF9.
(A) The 3′-UTR element of FGF9 messenger RNA is partially complementary to miR-26a. miR-26a or scramble control and luciferase reporter containing either a wildtype or a mutant 3′-UTR were co-transfected into HEK-293T cells. And a Renilla luciferase expressing construct exerts as internal control. (B) Western blot analysis of FGF9 expression in SGC-7901 and AGS cells infected with miR-26a, and GES-1 transfected with miR-26a inhibitors (Anti-miR-26a). (C, D, E) FGF9 abrogates the suppressive roles of miR-26a in GC cell invasion and growth. SGC-7901 cells stably expressing miR-26a or scramble were transfect with or without FGF9 plasmids. Invasion assays(C), Apoptosis analysis (D), and Cell proliferation analysis (E) were performed with the above cells as described in Materials and Methods. Data are presented as mean±s.e.m from at least three independent experiments. (F) Spearman’s correlation scatter plot of the levels of miR-26a (determined by in situ hybridization) and FGF9 protein (determined by immunohistochemistry) in 126 GC specimens. Representative images of FGF9 expression by immunohistochemistry are shown (right panel). Original magnification: ×200.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Doench JG, Sharp PA (2004) Specificity of microRNA target selection in translational repression. Genes Dev 18: 504–511. - PMC - PubMed
    1. He L, Hannon GJ (2004) MicroRNAs: small RNAs with a big role in gene regulation. Nat Rev Genet 5: 522–531. - PubMed
    1. Yan LX, Huang XF, Shao Q, Huang MY, Deng L, et al. (2008) MicroRNA miR-21 overexpression in human breast cancer is associated with advanced clinical stage, lymph node metastasis and patient poor prognosis. RNA 14: 2348–2360. - PMC - PubMed
    1. Ma L, Young J, Prabhala H, Pan E, Mestdagh P, et al. (2010) miR-9, a MYC/MYCN-activated microRNA, regulates E-cadherin and cancer metastasis. Nat Cell Biol 12: 247–256. - PMC - PubMed
    1. Vosa U, Vooder T, Kolde R, Vilo J, Metspalu A, et al. (2013) Meta-analysis of microRNA expression in lung cancer. Int J Cancer 132: 2884–2893. - PubMed

Publication types

MeSH terms

Grants and funding

This work was supported by grants from the Nature Scientific Foundation of China (81101526, 31100936) and Guangzhou Medical College Doctor Start Foundation (2012C11). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.