ABSTRACT
Objective: To investigate the effect of cell division cycle 42 (Cdc 42) gene on the formation of filopodia suppressed by non-steroidal anti-inflammatory drugs (NSAIDs), and to explore the probable signal transduction pathway involving Cdc 42 in the inhibition effects on migration and invasion abilities of breast cancer cells induced by NSAIDs. Methods: MCF-7 cells were divided into four groups: blank control group, NS-398 (100 μmol/L)-treated group, cyclooxygenase-2 (Cox-2) small interfering RNA (siRNA)-transfected group and the negative siRNA-transfected group. The expressions of Cox-2 and Cdc42 mRNAs were detected by real-time fluorogentic quantitative PCR(RFQ-PCR), and the expressions of Cox-2 and Cdc42 proteins were measured by Western blotting. Actin-tracker Green fluorescent probe was used to examined the morphology of filopodias in MCF-7 cells. The invasion ability of MCF-7 cells was detected by using the Martrigel-coated Transwell. Results: The filopodias in MCF-7 cells disappeared in the NS-398-treated group, and the invasion ability of MCF-7 cells was also significantly decreased in this group compared with that in the blank control group (P0.05), and the difference was also not seen in the expression of Cdc42 mRNA or protein between these two groups (P>0.05). The active-Cdc42 level was significantly decreased in the NS-398-treated group compared with that in the blank control group (P<0.05). The filopodias in the Cox-2 siRNA-transfected group disappeared, and the invasion ability of MCF-7 cells transfected with Cox-2 siRNA was significantly decreased compared with that in the negative siRNA-transfected group (P<0.05). The active-Cdc42 level was also significantly decreased in MCF-7 cells transfected with Cox-2 siRNA compared with those transfected with negative siRNA (P<0.05). Conclusion: NSAIDs can suppress the invasion ability of breast cancer cells. This effect may be achieved by inhibiting the activity of Cox-2, decreasing the expression level of active-Cdc42, and suppressing the formation of filopodias.
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the changes of Cdc42 expression under estrogen stimulation, and to explore the signaling pathway of intracellular material transportation caused by estrogen.</p><p><b>METHODS</b>MTT was used to test the drug sensitivity of cells. Real-time PCR was used to evaluate the expression of Cdc42 mRNA. The amount of ADM accumulated in MCF-7 cells was detected by flow cytometry. The protein levels of active-Cdc42 and Total-Cdc42 were measured by Western blot.</p><p><b>RESULTS</b>IC(50) of ADM in MCF-7 cells was increased from (0.098 ± 0.011) µg/ml to (0.134 ± 0.130) µg/ml (P < 0.05) after estrogen stimulation. The amount of ADM accumulated in MCF-7 cells was reduced from 7.253 ± 0.310 to 3.233 ± 0.313 (P < 0.05). All of Cdc42 mRNA, active-Cdc42 protein and total-Cdc42 protein were increased (P < 0.05). After the treatment with siRNA, the IC(50) of ADM in siRNA group was decreased to (0.057 ± 0.017) µg/ml (P < 0.05) compared with that in the control group. The amount of accumulated ADM was significantly increased in the siRNA group, and all the expression levels of Cdc42 mRNA, active-Cdc42 protein and total-Cdc42 protein were decreased in the siRNA group (P < 0.05).</p><p><b>CONCLUSIONS</b>Estrogen enhances the drug resistance in breast cancer cells. The mechanism of this effect may be via the enhancing Cdc42 expression and decreasing the accumulation of chemotherapeutic drugs in the cancer cells.</p>