Correlation between miRNA-24 expression and TRIB3 mRNA and protein expression.

a<p>Spearman's rho.</p

Associations of <i>miRNA-24</i> expression levels with clinicopathological characteristics.

a<p>Due to missing values, numbers do not always add up to 94.</p>b<p><i>P</i> tested with Mann-Whitney U test or Kruskal-Wallis Test where appropriate.</p

Effect of siRNA mediated knockdown of TRIB3 on mRNA and protein levels.

<p>A. Fold induction of TRIB3 mRNA in MDA-MB-231 cells during normoxia and after 24 hours of hypoxia (1% oxygen) treated with negative control siRNA (SCR) or siRNA mediated TRIB3 knockdown (TRIB3-KD) (n = 8). B. Fold induction of TRIB3 protein intensity in MDA-MB-231 cells during normoxia or after 24 hours of hypoxia (1% oxygen) treated with negative control siRNA (SCR) or siRNA mediated TRIB3 knockdown (TRIB3-KD) (n = 3). Protein intensities were controlled for α-tubulin intensities on same blot. Fold induction are averages (+ SD) relative to levels in cells treated with control siRNA (SCR) during normoxia.</p

Effect of proteasome inhibition on TRIB3 protein levels.

<p>A. Western blot of MDA-MB-231 cells treated with MG132 and exposed to hypoxia (0.1% oxygen) immunostained for β-catenin (positive MG132 control), TRIB3 and α-tubulin (loading control). B. MDA-MB-231 cells treated with hydroxychloroquine (CQ) and exposed to hypoxia (1% oxygen) immunostained for LC3b (positive autophagy inhibition marker), TRIB3 and α-tubulin (loading control). C. MDA-MB-231 cells treated with cycloheximide (CHX) and exposed to hypoxia (1% oxygen) immunostained for HIF-1 (positive translation inhibition control), TRIB3 and α-tubulin (loading control). D. MCF-7 cells treated with cycloheximide (CHX) and exposed to hypoxia (1% oxygen) immunostained for HIF-1 (positive translation inhibition control), TRIB3 and α-tubulin (loading control).</p

TRIB3 mRNA translation during anoxia.

<p>A. Polysome profiles of MCF7 cells grown under aerobic of anoxic conditions during 24 hours. B. TRIB3 mRNA levels determined by Q-PCR in the total RNA fraction. C. TRIB3 mRNA distribution within the polysome. D. Change in translation efficiency of the TRIB3 mRNA after exposure to anoxia for 24 hrs. Experiments were performed in triplicates.</p

Breast cancer patient survival based on miRNA-24 expression levels.

<p>A. Disease free survival in patients stratified according to miRNA-24 expression below or above median. B. Overall survival in patients stratified according to miRNA-24 expression below or above median (0.9×10⁻³). C. Disease free survival in patients stratified according to tertiles miRNA-24 expression below 0.5×10⁻³, above 1.2×10⁻³ or in between. D. Overall survival in patients stratified according to tertiles miRNA-24 expression below 0.5×10⁻³, above 1.2×10⁻³ or in between.</p

miRNA 24 expression levels in groups with accordance and discordance between TRIB3 mRNA and protein.

<p>miRNA 24 expression levels in groups with accordance and discordance between TRIB3 mRNA and protein.</p

No cytotoxic effects of CAIX inhibition in SCCNij202.

<p>S4 does not affect proliferation within CAIX positive areas (A), apoptosis within CAIX positive areas (B) or the amount of necrosis (C). Abbreviations: BrdUrd, bromodeoxyuridine; CAIX, carbonic anhydrase IX; LI, labeling index; NS, not significant, 1× S4, 8 h, one i.p. injection S4, harvest after 8 hours; 1× S4, 24 h, one i.p. injection S4, harvest after 24 hours; 3× S4, one i.p. injection S4 a day for 3 days, harvest 8 hours after the last injection; 5× S4, one i.p. injection S4 a day for 5 days, harvest 8 hours after the last injection.</p

Intracellular pH and CAIX inhibition in SCCNij202.

<p>S4 does not change CHOP mRNA (A) and MMP9 mRNA (B). Abbreviations: CHOP, C/EBP homologous protein; MMP9, matrix metallopeptidase 9; NS, not significant; 1× S4, 8 h, one i.p. injection S4, harvest after 8 hours; 1× S4, 24 h, one i.p. injection S4, harvest after 24 hours; 3× S4, one i.p. injection S4 a day for 3 days, harvest 8 hours after the last injection; 5× S4, one i.p. injection S4 a day for 5 days, harvest 8 hours after the last injection.</p

CAIX ectodomain shedding after treatment with S4.

<p>S4 decreases CAIX ectodomain shedding in the serum, especially 8 hours after the first injection. Abbreviations: CAIX, carbonic anhydrase IX; 1× S4, 8 h, one i.p. injection S4, harvest after 8 hours; 1× S4, 24 h, one i.p. injection S4, harvest after 24 hours; 3× S4, one i.p. injection S4 a day for 3 days, harvest 8 hours after the last injection; 5× S4, one i.p. injection S4 a day for 5 days, harvest 8 hours after the last injection.</p