Research Paper Volume 11, Issue 23 pp 11329—11346

Interplay of MKP-1 and Nrf2 drives tumor growth and drug resistance in non-small cell lung cancer

Figure 1. MKP-1 regulates the proliferation and drug resistance of A549 NSCLC cells. Two stable MKP-1-knockdown cell lines, siMKP1-C1 and siMKP1-C2, were generated after stable transfection with the pGFP-V-RS-MKP-1 plasmid into A549 cells. siCon expressing empty pGFP-V-RS vector was used as a negative control. (A) MKP-1 mRNA levels in siCon, siMKP1-C1, and siMKP1-C2 cells as determined by Taqman RT-PCR. The 18S rRNA level was used as an internal control and the value for siCon cells was set at 100%. Values are mean ± SD, n = 3. (B) Immunoblots of whole-cell lysates probed with anti-MKP-1, anti-Nrf2 or anti-actin. The relative levels of MKP-1 and Nrf2 normalized to actin are shown above each lane. Blots are representative three separate experiments. (C) Knockdown of MKP-1 reduces cell migration. Scratch assay images of siMKP1-C1 (b and e), siMKP1-C2 (c and f) and siCon cells (a and d) acquired at 0 (a-c) and 24 h (d-f). Red lines define the areas lacking cells. Statistics are shown in (g). Values are mean ± SD, n = 3. (D) MKP-1 promotes motility of NSCLC cells. Images of transwell migration assays of siMKP1-C1 (b), siMKP1-C2 (c), and siCon cells (a). (d) Statistics for three experiments. The number of siCon cells was set at 100%. Values are mean ± SD, n = 3. (E) Knockdown of MKP-1 decreases cell proliferation. Cells were cultured for 24, 48, or 72 h and the numbers determined by MTS assays. The value for the same cells at 0 h was set at 1. Values are mean ± SD, n = 3. (F) Knockdown of MKP-1 increases sensitivity to cisplatin in NSCLC cells. siMKP1-C1, siMKP1-C2 and siCon cells were exposed to cisplatin (0–25 μg/ml) for 48 h. The cell viability was determined by MTT method. The value of DMSO treatment was set at 1. Values are means ± SD, n = 3. *p <0.05, **p <0.01.