Journal: Hepatology (Baltimore, Md.)

Article Title: Oncogenic and tumor suppressive roles of polo-like kinases in human hepatocellular carcinoma.

doi: 10.1002/hep.23467

Figure Lengend Snippet: Fig. 1. Deregulation of the members of the PLK family and FOXM1 in human HCC. (A) Expression levels of PLK1-4 and FOXM1 in normal livers (NL, n 6), HCC (35 HCCB and 40 HCCP, respectively), and corresponding nontumorous surrounding livers (SLB and SLP, respectively) by real-time reverse-transcription PCR. Surrounding livers from the two prognostic subclasses did not show statistical differences (P 0.05). N-Target (NT) 2 Ct; Ct RNR18-Ct target gene. (B) Representative Western blots of lysates prepared from normal livers, SL, HCCB, and HCCP and immunoblotted with the indicated antibodies. (C) Western blotting optical densities were normalized to -actin values and expressed in arbitrary units. ■HCC and SL versus normal liver, at least P 0.01; †HCC subtypes versus corresponding surrounding liver, P 0.001; *HCCP versus HCCB for P 0.001 (Wilcoxon rank sum test).

Article Snippet: Human HCC cell lines (HepG2, HuH7, PLC, Hep3B, SNU-387, SNU-423, HLE, HuH6, SK-Hep1, and THLE-2), purchased from either the American Type Culture Collection or the Riken Cell Bank, were subjected to either small interfering RNA (siRNA) or demethylating treatments as reported in the Supporting Information.

Techniques: Expressing, Reverse Transcription, Western Blot

Journal: Hepatology (Baltimore, Md.)

Article Title: Oncogenic and tumor suppressive roles of polo-like kinases in human hepatocellular carcinoma.

doi: 10.1002/hep.23467

Figure Lengend Snippet: Fig. 3. Effect of siRNA-mediated silencing of PLK1-4 genes on cell viability of human HCC cell lines. Cell viability was assessed using gene-specific and control siRNAs after 48 hours of treatment. PLK1 was silenced in HepG2 and Hep3B cell lines (A), whereas PLK2-4 were suppressed in SNU-423 and HLE cells (B-D). Untreated cells were used for normalization. Each bar represents the mean standard deviation (n 9). ■siPLK1-treated cells versus transf C, at least P 0.001; †siPLK1-treated cells versus nonsense, P 0.001 (Student t test). C, untreated cells; ns, nonsense siRNA or control siRNA; siPLK1-4, siRNA against PLK1-4; transf C, transfection reagent control.

Article Snippet: Human HCC cell lines (HepG2, HuH7, PLC, Hep3B, SNU-387, SNU-423, HLE, HuH6, SK-Hep1, and THLE-2), purchased from either the American Type Culture Collection or the Riken Cell Bank, were subjected to either small interfering RNA (siRNA) or demethylating treatments as reported in the Supporting Information.

Techniques: Control, Standard Deviation, Transfection

Journal: Hepatology (Baltimore, Md.)

Article Title: Oncogenic and tumor suppressive roles of polo-like kinases in human hepatocellular carcinoma.

doi: 10.1002/hep.23467

Figure Lengend Snippet: Fig. 4. Effect of siRNA-mediated silencing of PLK1-4 genes on apoptosis in human HCC cell lines. Apoptosis was assessed using gene-specific and control siRNAs after 48 hours of treatment. PLK1 was silenced in HepG2 and Hep3B cell lines (A), whereas PLK2-4 were suppressed in SNU-423 and HLE cells (B-D). Untreated cells were used for normalization. Each bar represents the mean standard deviation (n 9). ■siPLK1-treated cells versus transf C, at least P 0.001; †siPLK1-treated cells versus ns, P 0.001 (Student t test). Abbreviations: C, untreated cells; ns, nonsense siRNA or control siRNA; siPLK1-4, siRNA against PLK1-4; transf C, transfection reagent control.

Article Snippet: Human HCC cell lines (HepG2, HuH7, PLC, Hep3B, SNU-387, SNU-423, HLE, HuH6, SK-Hep1, and THLE-2), purchased from either the American Type Culture Collection or the Riken Cell Bank, were subjected to either small interfering RNA (siRNA) or demethylating treatments as reported in the Supporting Information.

Techniques: Control, Standard Deviation, Transfection

Journal: Hepatology (Baltimore, Md.)

Article Title: Oncogenic and tumor suppressive roles of polo-like kinases in human hepatocellular carcinoma.

doi: 10.1002/hep.23467

Figure Lengend Snippet: Fig. 5. Effect of siRNA-mediated silencing of PLK1 on cell cycle and apoptotic cascades in human HCC cell lines. Western blot analysis revealed that inhibition of PLK1 expression results in a G2/M block (A) as well as increased PARP cleavage and a decline of the survivin levels (B), indicating apoptosis induction, when compared with control siRNA-treated cells. Abbreviations: C, untreated cells; ns, nonsense siRNA or control siRNA; PARP cl, cleaved PARP; PARP fl, full-length PARP; siPLK1, siRNA against PLK1; transf C, transfection reagent control.

Article Snippet: Human HCC cell lines (HepG2, HuH7, PLC, Hep3B, SNU-387, SNU-423, HLE, HuH6, SK-Hep1, and THLE-2), purchased from either the American Type Culture Collection or the Riken Cell Bank, were subjected to either small interfering RNA (siRNA) or demethylating treatments as reported in the Supporting Information.

Techniques: Western Blot, Inhibition, Expressing, Blocking Assay, Control, Transfection

Journal: Hepatology (Baltimore, Md.)

Article Title: Oncogenic and tumor suppressive roles of polo-like kinases in human hepatocellular carcinoma.

doi: 10.1002/hep.23467

Figure Lengend Snippet: Fig. 6. Mediators of apoptosis following PLK1 in human HCC cell lines either wild-type (HepG2) or deleted (Hep3B) for the p53 gene. (A) Induction of p53 and p73 genes and their putative target proteins, p21 and BAX, were detected by means of Western blotting in cells transfected with siRNA against PLK1 for 48 hours. An involvement of the mitochondrial apoptotic pathway was confirmed by way of western blotting analysis of (B) BAX and cytochrome C (Cytochr C) localization, and of (C) MCL1, BCL2, and cleaved caspase-3 protein levels (CASPASE cl). Abbreviations: C, untreated cells; ns, nonsense siRNA or control siRNA; siPLK1, siRNA against PLK1; transf C, transfection reagent. The arrows indicate the cleaved CASPASE3 products.

Article Snippet: Human HCC cell lines (HepG2, HuH7, PLC, Hep3B, SNU-387, SNU-423, HLE, HuH6, SK-Hep1, and THLE-2), purchased from either the American Type Culture Collection or the Riken Cell Bank, were subjected to either small interfering RNA (siRNA) or demethylating treatments as reported in the Supporting Information.

Techniques: Western Blot, Transfection, Control

Journal: Hepatology (Baltimore, Md.)

Article Title: Oncogenic and tumor suppressive roles of polo-like kinases in human hepatocellular carcinoma.

doi: 10.1002/hep.23467

Figure Lengend Snippet: Fig. 7. Effect of FOXM1 modulation on PLK1 protein levels in human HCC cell lines. (A) Overexpression of FOXM1 by transient transfection in SNU-182 cells up-regulates PLK1 levels. (B) Silencing of FOXM1 in Hep3B cells determines down-regulation of PLK1 levels. Equivalent results were obtained in HepG2 cells (data not shown). (C) Overexpression of wild-type or oncogenic mutant (Q61L) Ha-Ras leads to up-regulation of PLK1 through FOXM1. The SNU-182 cell line was transfected with either wild-type Ha-Ras or its mutated form (Q61L) alone, or associated with siRNA against FOXM1 (siFOXM1) or PLK1 (siPLK1). Representative Western blotting panels at 48 hours after transfection are shown. (D) Effect of transfection with wild-type Ha-Ras, its mutated form (H-Ras Q61L) alone, or associated with siRNA against FOXM1 (siFOXM1) or PLK1 (siPLK1), on in vitro growth of SNU-182 cells. Results at 48 hours after transfection are expressed as the mean standard deviation. Untreated cells (Control) were used for normalization. Experiments were performed at least three times in triplicate. *Wild-type Ha-Ras associated with siFOXM1 or siPLK1 versus Ha-Ras alone, P 0.001; ■mutated Ha-Ras Q61L associated with siFOXM1 or siPLK1 versus Ha-Ras Q61L alone, P 0.001 (Student t test).

Article Snippet: Human HCC cell lines (HepG2, HuH7, PLC, Hep3B, SNU-387, SNU-423, HLE, HuH6, SK-Hep1, and THLE-2), purchased from either the American Type Culture Collection or the Riken Cell Bank, were subjected to either small interfering RNA (siRNA) or demethylating treatments as reported in the Supporting Information.

Techniques: Over Expression, Transfection, Mutagenesis, Western Blot, In Vitro, Standard Deviation, Control