miR-146a indirectly regulates CCL2 expression. (a) Renilla luciferase activity was measured in A549 cells transfected with synthetic miRNAs and the pLightSwitch_CCL2 3ʹ UTR construct. CCL2 3ʹ UTR luciferase activity was normalized to activity from the pLightSwitch_GAPDH 3ʹ UTR construct in A549 cells exposed to the same miRNA condition. Luciferase activity was further normalized to total protein concentrations of each sample. Luciferase activity was significantly reduced in cells transfected with synthetic miR-146a. (*) p < 0.02, n = 4. (b) The CCL2 3ʹ UTR luciferase assay was repeated as in (a), but additional samples were generated with the synthetic miRNAs and pLightSwitch_CCL2 3ʹ UTR construct co-transfected with a HuR expression vector, pcDNA3.1(+)-HuR-FLAG. Overexpressing HuR could rescue the luciferase activity that was inhibited by synthetic miR-146a, but levels were not as high as luciferase activity in cells transfected with both pcDNA3.1(+)-HuR-FLAG and the negative control miRNA. (*) p < 0.05, n = 3; (**) p < 0.03, n = 3.