Cyr61 Levels in Various Breast Cancer Cell Lines – Baseline and in Response to IGF-1.

<p>(<b>A</b>) Baseline mRNA expression of Cyr61 in Wildtype MCF-7, SKBR3, BT474, and MDAMB231 as determined by RT-Q-PCR. (<b>B–D</b>) Cyr61 expression after IGF-1 treatment at baseline, 20 minutes, 4 hours, and 24 hours in (<b>B</b>) MCF-7 WT, (<b>C</b>) MCF-7 DN, and (<b>D</b>) MCF-7 AA cells. Cyr61 is increased by treatment of IGF-1, especially at 20 minutes. (<b>E</b>) Expression of Cyr61 mRNA in MCF-7 WT cells compared to MCF-7 AA (transfectant with constitutively active Akt), and MCF-7 DN (transfectant with dominant negative constitutively inactive Akt). (<b>F</b>) The results obtained in the MCF-7 cell line were confirmed by a second breast cancer cell line, SKBR3. All mRNA levels were adjusted for the housekeeping gene, 18S. P-values were calculated relative to MCF-7 WT in (A,E), to 0 Minutes in (B–D), and SKBR3 WT in (F). P<0.05 is statistically significant.</p

Confirmation of Cyr61 levels and Invasiveness in MCF-7 WT and MCF-7 CYA.

<p>(<b>A</b>) The mRNA expression of Cyr61 was assessed by RT-Q-PCR and (<b>B</b>) by immunofluorescent staining (FITC - green – Cyr61; DAPI – blue – nuclei). (<b>C</b>) Quantification of invasiveness from invasion chamber. (<b>D</b>) Red arrows demonstrate positive examples from invasion chamber. CYA cells are 9 times more invasive than the MCF-7 WT cells. All mRNA levels were adjusted for the housekeeping gene, 18S. P-values were calculated relative to MCF-7 and P<0.05 is statistically significant.</p

Proposed Schematic of IGF-1 mediated signaling in relation to Cyr61 expression and activity.

<p>Proposed Schematic of IGF-1 mediated signaling in relation to Cyr61 expression and activity.</p

Effects of IGF-1 and PD98059 Induction on Proliferation, Invasion, and Cyr61 Expression in MCF-7 WT and MCF-7 CYA.

<p>(<b>A</b>) The mRNA expression of Cyr61 was assessed by RT-Q-PCR. All mRNA levels were adjusted for the housekeeping gene, 18S. (<b>B</b>) Proliferation assay summary data after 48 hour IGF-1 (100 ng/mL) and PD98059 (30 µM) treatment in MCF-7 WT and MCF-7 CYA. Each condition was plated into 6 identical wells in a 96 well plate. All treatments were standardized relative to the untreated condition for each cell line. (<b>C</b>) Results from Invasion assay after 40 hour IGF-1 (100 ng/mL) and PD98059 (30 µM) treatment in MCF-7 WT and MCF-7 CYA. P-values were assessed relative to the untreated condition in each assay, with P<0.05 considered statistically significant.</p

Effects of IGF-1 and LY294002 Induction on Proliferation, Invasion, and Cyr61 Expression in MCF-7 WT and CYA.

<p>(<b>A</b>) The mRNA expression of Cyr61 was assessed by RT-Q-PCR. All mRNA levels were adjusted for the housekeeping gene, 18S. (<b>B</b>) Results from proliferation assay after 48 hour IGF-1 (100 ng/mL) and LY294002 (50 µM) treatment in MCF-7 WT and MCF-7 CYA. Each condition was plated into 6 identical wells in a 96 well plate. All treatments were standardized relative to the untreated condition for each cell line. (<b>C</b>) Results from Invasion assay after 40 hour IGF-1 (100 ng/mL) and LY294002 (50 µM) treatment in MCF-7 WT and MCF-7 CYA. P-values were assessed relative to the untreated condition in each assay, with P<0.05 considered statistically significant.</p

Inverse Relationship between IGF-1 induced Cyr61 upregulation and decreased E-cadherin and FOXO1 expression is confirmed in two cell lines.

<p>The mRNA expression of Cyr61, E-cadherin, and FOXO1 were assessed by RT-Q-PCR. All mRNA levels were adjusted for the housekeeping gene, 18S. (<b>A</b>) Expression of Cyr61, E-cadherin, and FOXO1 in response to IGF-1 induction (100 ng/mL) following serum starvation in MCF-7 WT cells; and (<b>B</b>) MDAMB231 cells. P-values were assessed relative to the untreated condition within each cell line, and P<0.05 considered statistically significant.</p

Correlation between DYRK2 expression and clinicopathological characteristics of CRC patients.

<p>Correlation between DYRK2 expression and clinicopathological characteristics of CRC patients.</p

mRNA expression of DYRK2 is lower in CRC compared to the normal colon/rectum.

<p>A. Gaedcke's colorectal carcinoma data set: paired normal colon vs paired CRC; B. Skrzypczak's colorectal carcinoma data set: Normal colon vs colorectal adenocarcinoma and colorectal carcinoma; C. Kaiser's colorectal carcinoma data set: normal colon vs different types of CRC; D. TCGA colorectal carcinoma data set: normal colon and rectum vs different type of CRC.</p

Decreased expression of DYRK2 in paraffin-embedded colorectal cancer tissues and advanced colorectal cancer.

<p>A, DYRK2 protein expressed in adjacent non-cancerous tissue (ANT) and colorectal cancer tissue (T) by IHC analysis. B, Quantitative analysis of DYRK2 expression in primary colorectal tumors and adjacent non-cancerous tissues (ANT) by IHC analysis (Wilcoxon signed rank test, n = 99, <i>P</i> < 0.001). C, Immuno-staining of DYRK2 in five pairs of representative colorectal tumor tissues (T) with adjacent non-cancerous tissues (ANT). D, Representative IHC analyses of DYRK2 expression at different clinical stages, two magnifications (100X and 200X).</p

Clinicopathological characteristics and DYRK2 expression of 181 patient samples of CRC.

<p>Clinicopathological characteristics and DYRK2 expression of 181 patient samples of CRC.</p