Effects of microRNA-136 on proliferation of lung adenocarcinoma A549 cells

Abstract

Objective To investigate the effects of microRNA (miRNA, miR)-136 on A549 cells proliferation. Methods (1) The expression of miR-136 in A549 cells and 16HBE cells was detected by reverse transcriptase-polymerase chain reaction (RT-PCR). 40 nmol/L miR-136 inhibitor was transfected into A549 cells, and the effect of inhibition was detected by RT-PCR. (2) Cell proliferation was tested by methyl thiazol tetrazolium (MTT) assay and soft agar assay. (3) The effect of 20 nmol/L and 40 nmol/L miR-136 inhibitor on extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway was determined by Western blotting. Results (1) Compared to 16HBE cells (0.35±0.12), the relative expression of miR-136 was significantly higher in A549 cells (1.39±0.33, t=13.165, P=0.002). The expression level of miR-136 was significantly decreased in A549 cells after transfection with anti-miR-136 oligonucleotide (0.21±0.08) compared to control group (1.32±0.29, t=15.016, P=0.000). (2) The MTT assay revealed that the absorbance of anti-miR-136 oligonucleotide transfected A549 cells was decreased to 1.25±0.04 (t=7.258, P=0.013) on the 5th day compared to the control group. The soft agar assay showed that the colony number of anti-miR-136 oligonucleotide transfected cells (7.55±2.48) was significantly reduced compared to the control group (21.2±6.15, t=9.127, P=0.008). (3) Western blotting showed that anti-miR-136 oligonucleotide resulted in a significant reduction of ERK1/2 phosphorylation in a dose-dependent manner. Compared to the control group, transfection of anti-miR-136 oligonucleotide on the concentration of 20 nmol/L and 40 nmol/L led to 50% and 90% decrease of ERK1/2 phosphorylation respectively in A549 cells (χ2=8.339, P=0.009; χ2=11.023, P=0.000). Conclusion Down-regulation of miR-136 decreases A549 cells proliferation by inhibiting ERK1/2 pathway. Key words: Lung neoplasms; MicroRNA-136; Extracellular signal-regulated kinase 1/2; Cell proliferation