Effect of clofenotane on epithelialmesenchymal transition in colorectal cancer DLD1 cells / 中国药理学与毒理学杂志

ABSTRACT

OBJECTIVE To explore the effect of clofenotane (DDT) on epithelial-mesenchymal transition (EMT) and the relevant molecular mechanism in human colorectal cancer cells. METHODS Human colorectal cancer cells DLD1 were treated with DDT 0.01, 0.1, 1.0, 10.0 and 100.0 nmol·L-1 for 48 h. Then, the morphology of DLD1 cells was observed. mRNA levels of E-cadherin, N-cadherin, vimentin and Snail1 were detected by real-time PCR. Protein expression of STAT3 signaling pathway of proteins STAT3 and p-STAT3 was detected by Western blotting. STAT3 inhibitor WP1006 (5μmol · L-1) was added to determine its impact on DDT-induced alternation of STAT3/Snail1 signaling and EMT-related molecules. Protein expression of STAT3 and p-STAT3 was detected by Western blotting and mRNA levels of E-cadherin, N-cadherin, Vimentin and Snail1 were detected by real-time PCR. RESULTS DLD1 cell morphology was changed after exposure to DDT 0.01-100.0 nmol · L- 1. Meanwhile, real-time PCR showed that the mRNA level of E-cadherin was significantly decreased compared with normal cell control (P<0.01), which was 42.4±2.8%of that in the normal control group. The mRNA levels of N-cadherin, Vimentin and Snail1 were significantly increased (P<0.01), which were 1.91±0.1, 1.5±0.2 and 1.5±0.1 times that of the normal control group. DDT 0.1, 1.0 and 10.0 nmol · L-1 exposure induced up-regulation of STAT3 and p-STAT3 protein levels (P<0.01), which were 2.1 and 1.8 times that of the normal control group. The addition of STAT3 inhibitor WP1066 (5 μmol · L-1) prevented STAT3 from phosphorylation as well as the up-regulation of Snail1(P<0.01), which was (56.3 ± 0.9)% that of the DDT 1.0 nmol · L-1 treat?ment group. Compared with DDT treatment alone, the mRNA levels of EMT-related molecules were remarkably reversed by WP1066 (5 μmol · L- 1) co-treatment, increasing E-cadherin but decreasing N-cadherin and vimentin in DLD1 cells(P<0.01), which were 50.2±2.9%and 61.6±6.1%of those in the DDT 1.0 nmol · L- 1 treatment group, respectively. CONCLUSION DDT alters the expressions of EMT-related molecules including E-cadherin, N-cadherin and vimentin via STAT3/Snail1 signaling, thus promoting the EMT process in human colorectal cancer cells. This progress may be closely related to DDT-induced colorectal cancer development.