05) (Figure 4). Figure 4 Cell proliferation activity after transfection of CDK8-siRNA assessed by MTT assay. Curves of cell Selleck MGCD0103 growth after transfection for 24, 48 and 72 h by MTT assay. Results are given as means ± SD from three independent experiments. P < 0.05. Effect of CDK8-siRNA
transfection on the apoptosis LY2109761 molecular weight and cell cycle of HCT116 cells We performed experiments to evaluate the apoptosis and cell cycle of HCT116 cells by CDK8-siRNA. As shown in figure 5, following 48 h transfection, it was indicated that the rate of apoptosis in the CDK8-siRNA group (23.50 ± 1.20%) was significantly higher than that of the scrambled siRNA (4.87 ± 1.48%) and non-siRNA groups (4.77 ± 1.42%) (P < 0.01) (Figure 5A). On the other hand, the cell cycle analysis showed that G0/G1 phase of CDK8-siRNA transfected group in a ratio of 65.77 ± 1.17%, was significantly higher than that of scrambled siRNA (50.20 ± 2.43%) and non-siRNA group (54.33 ± 2.55%) (P < 0.01) (Figure 5B). Figure 5 Effect of CDK8-siRNA transfection on the apoptosis and cell cycle of HCT116 cells. 48 h after transfection, cell apoptosis (A) and cell cycle (B) were determined by flow cytometry. Quadrants D2-D4 represent necrotic/late apoptotic cells, viable cells, and early apoptotic cells, respectively.
Results are given as means ± SD from three independent experiments. CDK8 and β-catenin expression in fresh colon tumor and adjacent normal tissues To further confirm the expression of CDK8 and β-catenin in colon cancer, we detected the expression of CDK8 and β-catenin in fresh colon cancer tissues and adjacent normal tissues of the same patient. LY3023414 solubility dmso Real-time PCR was adopted to detect the mRNA levels. Results showed that mRNA expression levels of CDK8 and β-catenin in tumor tissues was significantly higher than in very adjacent normal tissues (P < 0.05) (Figure 6). In addition, the expression of CDK8 was correlated with the expression of β-catenin in both tumor tissues (r = 0.744, P < 0.01) and adjacent normal tissues (r = 0.650, P < 0.05). Figure 6 CDK8 and β-catenin mRNA expression in colon tumor and adjacent normal tissues detected
by real-time PCR. Fresh tumor and corresponding adjacent tissues from 12 patients were resected under sterile conditions and then snapfrozen in liquid nitrogen immediately. 200 mg tissue was taken out from liquid nitrogen and plused 1 ml Trizol when RNA was extaracted. Real-time PCR is performed for the expression levels of CDK8 (A) and β-catenin (B). Results are given as means ± SD from three independent experiments. P < 0.05. Shown by IHC, the positive cells were expressed as brown particles distributed in the cytoplasm of tumor cells (Figure 7). Of the 47 colon tumors, the positive rate of CDK8 and β-catenin was 76.6% (36/47) and 95.7% (45/47). However, in adjacent normal tissues, the positive rate was 21.3% (10/47) and 88.9% (40/47), respectively.