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. 2014 Jul 30;5(14):5570-80.
doi: 10.18632/onco_target.2141.

Small interfering RNA (siRNA)-mediated knockdown of macrophage migration inhibitory factor (MIF) suppressed cyclin D1 expression and hepatocellular carcinoma cell proliferation

Xiao-Hui Huang  1 , Wei-Hua Jian  2 , Zhao-Feng Wu  3 , Jie Zhao  1 , Hua Wang  1 , Wen Li  1 , Jin-Tang Xia  4
Affiliations

Small interfering RNA (siRNA)-mediated knockdown of macrophage migration inhibitory factor (MIF) suppressed cyclin D1 expression and hepatocellular carcinoma cell proliferation

Xiao-Hui Huang et al. Onco_target. .

Abstract

Macrophage migration inhibitory factor (MIF), a proinflammatory and immunoregulatory chemokine, plays important roles in cancer-related biological processes. However, few studies have focused on the clinical relevance of MIF and cyclin D1 expression in hepatocellular carcinoma cells (HCCs). In this study, MIF and cyclin D1 expression levels in HCC tissues and cell lines were significantly upregulated compared with adjacent normal tissues or a normal liver cell line. In HCC specimens, MIF expression positively correlated with cyclin D1 expression. Additionally, MIF and cyclin D1 expression positively correlated with tumor size. MIF knockdown inhibited the proliferation of PLC and HepG2 cells and promoted apoptosis. However, small interfering RNA (siRNA) against MIF did not influence the cell cycle in these cells. In an in vivo xenograft model, MIF knockdown reduced the tumor growth rate. The expression levels of Bcl-2, p-caspase-3, BIM and Bax were upregulated, while the expression levels of cyclin D1, p-Akt and p-ERK were downregulated in MIF-knockdown cells. These findings indicate that MIF siRNA reduces proliferation and increases apoptosis in HCC cells. MIF knockdown inhibits the expression of growth-related proteins and induces the expression of apoptosis-related proteins, supporting a role for MIF as a novel therapeutic _target for HCC.

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Conflict of interest statement

There are no conflicts of interest.

Figures

Figure 1
Figure 1. MIF and cyclin D1 are highly expressed in HCC
(A) Immunohistochemical analysis of MIF expression in HCC tissue (T) and paired adjacent non-cancerous liver tissue (N) (200X magnification). (B) Immunohistochemical analysis of cyclin D1 expression in HCC tissue (T) and paired, adjacent non-cancerous liver tissue (N) (200X magnification). (C) MIF protein expression in HCC tissue (T) and paired, adjacent non-cancerous liver tissue (N). (D) Cyclin D1 protein expression in HCC tissue (T) and paired adjacent non-cancerous liver tissue (N). (E) MIF and cyclin D1 expression in HCC and paired adjacent non-cancerous tissue by qRT-PCR. The data were normalized to β-actin levels, as a control. (F) Correlation of MIF and cyclinD1 expression in HCC tissues. **P<0.001 compared with adjacent noncancerous liver tissues.
Figure 2
Figure 2. MIF and cyclin D1 expression is upregulated in HCC cell lines
Western blot analysis of MIF and cyclin D1 expression in the human hepatocyte cell line LO2 and several HCC cell lines (BEL7402, PLC, HepG2, Huh-7 and Hep3B). β-Actin expression levels were used as internal controls.
Figure 3
Figure 3. Knockdown of MIF expression by MIF siRNA.PLC and HepG2 cells were transfected with different concentrations of MIF siRNA or control siRNA for 48 h.RNA and protein expression was analyzed by real-time PCR and Western blot analysis, respectively
(A) MIF and (B) cyclin D1 mRNA levels were downregulated by MIF siRNA. **P < 0.001 compared with controls. (C) Western blotting demonstrated that MIF protein and cyclin D1 protein levels were significantly decreased by MIF siRNA.
Figure 4
Figure 4. The effects of MIF knockdown on cell proliferation
(A) HepG2 cells and (B) PLC cells were transfected with MIF or control siRNA. Cell viability was assessed using MTT assays at five time points (0, 24, 48, 72, and 96 h). **P < 0.01. (C) Western blot analysis showed that knockdown of MIF expression reduced the expression of p-AKT and p-ERK in PLC and HepG2 cells.
Figure 5
Figure 5. The effects of MIF knockdown on cell apoptosis
(A) HepG2 cells and (B) PLC cells were transfected with MIF or control siRNA. Apoptosis was assessed using the Annexin V-FITC Apoptosis Detection Kit. **P < 0.01. (C) Western blot analysis showed that MIF knockdown increased the expression of Bcl-2, p-caspase-3, BIM and BAX in PLC and HepG2 cells. **P < 0.01.
Figure 6
Figure 6. The effects of MIF knockdown on PLC and HepG2 cell growth in nude mice
(A) Images of tumors extracted from MIF-knockdown and control groups (PLC and HepG2) after nude mice were euthanized 21 d post-tumor cell injection. (B) Growth curves for PLC tumors treated with MIF or control siRNA. (C) Growth curves for HepG2 tumors treated with MIF or control siRNA.
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References

    1. Thomas MB, Jaffe D, Choti MM, Belghiti J, Curley S, Fong Y, Gores G, Kerlan R, Merle P, O'Neil B, Poon R, Schwartz L, Tepper J, Yao F, Haller D, Mooney M, Venook A. Hepatocellular carcinoma: consensus recommendations of the National Cancer Institute Clinical Trials Planning Meeting. J Clin Oncol. 2010;28(25):3994–4005. - PMC - PubMed
    1. Chen L, Zhang Q, Chang W, Du Y, Zhang H, Cao G. Viral and host inflammation-related factors that can predict the prognosis of hepatocellular carcinoma. Eur J Cancer. 2012;48(13):1977–1987. - PubMed
    1. Adamali H, Armstrong ME, McLaughlin AM, Cooke G, McKone E, Costello CM, Gallagher CG, Leng L, Baugh JA, Fingerle-Rowson G, Bucala RJ, McLoughlin P, Donnelly SC. Macrophage migration inhibitory factor enzymatic activity, lung inflammation, and cystic fibrosis. Am J Respir Crit Care Med. 2012;186(2):162–169. - PMC - PubMed
    1. Arenberg D, Luckhardt TR, Carskadon S, Zhao L, Amin MA, Koch AE. Macrophage migration inhibitory factor promotes tumor growth in the context of lung injury and repair. Am J Respir Crit Care Med. 2010;182(8):1030–1037. - PMC - PubMed
    1. Wang D, Luo L, Chen W, Chen LZ, Zeng WT, Li W, Huang XH. Significance of the vascular endothelial growth factor and the macrophage migration inhibitory factor in the progression of hepatocellular carcinoma. Oncol Rep. 2014;31(3):1199–1204. - PubMed

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