Effect of transfection of miRNA antisense molecules on TNF-αsynthesis by RA FLS.

<p><b>A, B.</b> MiR-125b and miR-939 levels were determined by quantitative RT-PCR in RA FLS and THP-1 cells stimulated with LPS for 3 h and 6 h. RNAU6 was used as endogenous control for data normalization. The control (C) corresponded to untreated cells. RA FLS were transfected with either miR-125b or miR-939 antisense molecules or in combination or with the Clear-miR™ negative control (control). LPS or medium (C) activation of transfected cells was performed 24 h post-transfection, for 3 h and 6 h. Non transfected RA FLS were used as negative controls (NT). <b>C, D.</b> TNF-α and IL-6 release were determined by ELISA in culture supernatants harvested 6 h after stimulation with LPS or medium (C). <b>E.</b> Intracellular TNF-α expression was determined in transfected FLS and activated with LPS for 24 h. Data are expressed as the mean of triplicate samples +/− SD of three independent experiments for each patient.