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. 2019 Nov;18(5):5408-5416.
doi: 10.3892/ol.2019.10902. Epub 2019 Sep 20.

Talin1 knockdown prohibits the proliferation and migration of colorectal cancer cells via the EMT signaling pathway

Ling Ji  1   2 , Feizhao Jiang  2 , Xianping Cui  1 , Chengkun Qin  1
Affiliations

Talin1 knockdown prohibits the proliferation and migration of colorectal cancer cells via the EMT signaling pathway

Ling Ji et al. Oncol Lett. 2019 Nov.

Retraction in

Abstract

Colorectal cancer (CRC) is the third most commonly diagnosed cancer and the second highest cause of cancer-associated death worldwide. Talin1 activates integrins, which mediate cell adhesion, proliferation, tumorigenesis and metastasis. The aim of the present study was to determine talin1 expression levels in colorectal cancer (CRC) and investigate the role of talin1 in CRC proliferation and invasion in vitro and in vivo. Talin1 protein expression levels were detected in human CRC and adjacent normal tissues by immunohistochemistry. Talin1 short hairpin RNA and control vectors were designed and stably transfected into HCT116 CRC cells. Cell proliferation was determined by MTT assay. Cell migratory and invasive capabilities were detected by wound-healing and Matrigel invasion assays. The expression of proteins in the epithelial-to-mesenchymal transition signaling pathway was determined by western blotting and reverse transcription-quantitative PCR. The effect of talin1 on tumor growth was explored in vivo using BALB/c nude mice. Immunohistochemical analysis of CRC and adjacent normal tissue revealed that talin1 expression was upregulated in CRC. Talin1 knockdown significantly reduced the proliferation, migration and invasive ability of HCT116 cells compared with the control. Protein levels of phosphorylated STAT3 and vimentin were significantly lower in talin1-knockdown HCT116 cell lines compared with the control, whereas protein levels of E-cadherin were increased. Interleukin-6 mRNA levels were significantly increased in patients' blood samples compared with blood samples from healthy controls, as well as in CRC compared with adjacent normal tissue. In vivo experiments demonstrated that talin1 knockdown reduced CRC tumor growth and weight in nude mice. In conclusion, Talin1 knockdown may prevent the proliferation and migration of CRC cells by downregulating various factors involved in the epithelial-to-mesenchymal transition process, such as phosphorylated STAT3 and vimentin; therefore, talin1 may provide a novel therapeutic target for CRC.

Keywords: colorectal cancer; epithelial-to-mesenchymal transition; migration; proliferation; talin1.

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Figures

Figure 1.
Figure 1.
Talin1 expression is upregulated in colorectal cancer and it increases proliferation of HCT116 cells. (A) IHC demonstrating the distribution of talin1 in tumor (+++) and normal (+) tissues. Magnification, ×400. (B) Quantification of talin1 distribution in the IHC assays from at least 30 patients. (C) Western blotting results of talin1 protein expression in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control. (D) Talin1 mRNA levels in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control (n=5). (E) Talin1 protein levels in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control (n=3). (F) Knockdown of endogenous talin1 inhibited the proliferation of HCT116 cells. Data are expressed as the mean ± SEM. **P<0.01 and ***P<0.001 vs. control. IHC, immunohistochemistry; shRNA, short hairpin RNA.
Figure 2.
Figure 2.
Talin1 knockdown inhibits the migration and invasion of HCT116 cells. (A) Knockdown of endogenous talin1 inhibited the invasive ability of HCT116 cells. (B) Quantification results of the invasion assays. (C) Talin1 knockdown reduces migration of HCT116 cells. Magnification, ×400. (D) Quantification results of the wound-healing assays. Data are expressed as the mean ± SEM from at least three independent experiments. *P<0.05, **P<0.01 and ***P<0.001 vs. control. shRNA, short hairpin RNA.
Figure 3.
Figure 3.
Talin1 knockdown affects the expression of proteins associated with the EMT signaling pathway in HCT116 cells. (A) E-cadherin and vimentin protein levels were measured by western blotting in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control. (B) E-cadherin protein levels in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control (n=3). (C) Vimentin protein levels in in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control (n=3). (D) E-cadherin mRNA levels in tumor and normal tissues. (E) ZO-1 mRNA levels in tumor and normal tissues. (F) Occludin mRNA levels in tumor and normal tissues. (G) pSTAT3 and total STAT3 protein levels measured by western blotting. (H) pSTAT3 and total STAT3 protein levels in talin1-knockdown HCT116 cell lines and HCT116 cells transfected with the shRNA control (n=3). (I) IL-6 mRNA levels in blood samples from patients and healthy controls. (J) Serum IL-6 concentrations in blood samples from patients and healthy controls. (K) IL-6 mRNA levels in tumor and adjacent normal tissues. Data are presented as mean ± SEM. *P<0.05, **P<0.01 and ***P<0.001 vs. control. ZO-1, zonula occludens-1; pSTAT3, phosphorylated STAT3; IL-6, interleukin-6; shRNA, short hairpin RNA.
Figure 4.
Figure 4.
Talin1 knockdown suppressed tumor growth in a xenograft model. (A) Talin1 knockdown inhibited tumor growth. (B and C) Talin1 knockdown reduced (B) tumor weight and (C) tumor volume. (D) Talin1 knockdown did not affect mouse body weight. **P<0.01 and ***P<0.001 vs. control. shRNA, short hairpin RNA.
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