Egr2 was down-regulated in HCC cell lines.

<p>(A) Relative expression of Egr2 in L02 and HCC cells was analyzed by qRT-PCR. (B) Relative expression of Egr2 in L02 and HCC cells was detected by immunohistochemistry. *P<0.05, **P<0.01.</p

The expression of miR-20a/LncRNA-AF113014/Egr2 in HCC.

<p>The expressions of Egr2, LncRNA-AF113014 and miR-20a in HCC tissues were analyzed with qPCR.</p

AF113014 inhibited proliferation of HCC cells <i>in vitro</i>.

<p>(A) Relative expressions of AF113014 in SMMC7721 and Huh-7 cells transfected with Ad-AF113014 or siAF113014. GAPDH was used as reference gene in real-time PCR. (B) Proliferations of SMMC7721 and Huh-7 cells transfected with Ad-AF113014 or siAF113014 were examined by MTS. (C) Proliferations of SMMC7721 and Huh-7 cells transfected with Ad-AF113014 or siAF113014 were examined by colony formation assay. *P<0.05, **P<0.01.</p

Egr2 Suppressed HCC cell proliferation <i>in vitro</i>.

<p>(A) Relative mRNA and protein expressions of Egr2 in Ad-Egr2 or siEgr2 transfected SMMC7721 cells. (B) Proliferations of SMMC7721 transfected with Ad-Egr2 or siEgr2 were testified by MTS. (C) Proliferations of SMMC7721 transfected with Ad-Egr2 or siEgr2 were examined by colony formation assay. *P<0.05, **P<0.01.</p

Egr2 was downstream target gene of AF113014.

<p>(A) The location of AF113014 in chromosome 10. (B, C) Relative mRNA and protein expressions of Egr2 in Ad-AF113014 or siAF113014 transfected SMMC7721 cells, respectively. GAPDH was used as an internal quantitative control. * P < 0.05, ** P<0.01.</p

Expressions of AF113014 were down-regulated in HCC cell lines.

<p>lncRNA-AF113014 expressions in L02 and HCC cell lines. *P<0.05 vs L02.</p

AF113014 up-regulated Egr2 expression by interacting with miR-20a.

<p>(A) Software predicted the binding sites between AF113014, miRNA-20a and Egr2-3'UTR. (B) Relative expressions of miRNA-20a in Ad-AF113014 or siAF113014 transfected SMMC7721 cells. U6 RNA was used as control. (C) Luciferase activity assay was performed to determine the relationship between AF113014, miRNA-20a and Egr2 in SMMC7721 cells.* P < 0.05, ** P<0.01. (D) qRT-PCR were performed to analyze the expressions of Egr2 after co-transfected with Ad-AF113014 and miRNA-20a, Ad-AF113014 and miRNA-20a-in, siAF113014 and miRNA-20a, siAF113014 and miRNA-20a-in, respectively. Ad-GFP+NC was used as control. (E) Western blot were performed to analyze the expressions of Egr2 after co-transfected with Ad-AF113014 and miRNA-20a, Ad-AF113014 and miRNA-20a-in, siAF113014 and miRNA-20a, siAF113014 and miRNA-20a-in, respectively. Ad-GFP+NC was used as control.</p

Proliferation founctions of AF113014 effected on miR-20a and Egr2 in HCC cells.

<p>(A) Proliferations of SMMC7721 cells were examined by MTS after co-transfected with Ad-AF113014, miR-20a, miR-20a-in, Ad-Egr2, si-Egr2 in groups. (B) Proliferations of SMMC7721 cells were examined by colony formation assay after co-transfected with Ad-AF113014, miR-20a, miR-20a-in, Ad-Egr2, si-Egr2 in groups. (C) Numeralization of (B).</p

AF113014 influenced tumor growth <i>in vivo</i>.

<p>(A) Representative tumors formed in Ad-GFP and Ad-AF113014 infected SMMC7721 cells, and the tumor growth curve of each group. * P < 0.05, ** P < 0.01. (B) Transplanted tumors sections were stained with Egr2 and Ki-67. (C) Western blot analysis of Egr2 expression in transplanted tumors. GAPDH was used as control.</p

Primer sequences used for PCR or plasmids construction and siRNA sequence.

<p>Primer sequences used for PCR or plasmids construction and siRNA sequence.</p