Figure 3
From: miR-744-5p contributes to ocular inflammation in patients with primary Sjogrens Syndrome
Poly(I:C) mediated inflammation is reduced in human conjunctival cells treated with a miR-744-5p antagomir ex vivo. (A) PCR analysis of cytokeratin-18 (CK18) and cytokeratin-19 (CK19) in primary human conjunctival epithelial cells (HConEC) from Innoprot. Results are representative of 3 samples, with 18 s used as a control. The full gel image is included in the supplementary data. (B–E) Human conjunctival epithelial cells (Innoprot) were transfected with a FITC-labeled miR-744-5p antagomir for 72 hours. (B–D) Transfection of miR-744-5p antagomir was visualised using an inverted bright field microscope (Leica, DMIL) x20 magnification. (E) Uptake FITC-labelled miR-744-5p antagomir assessed by flow cytometry after 72 h of culture (filled histogram) relative to negative antagomir-treated cells (unfilled histogram). Data are representative of three independent experiments. (F–H) Human conjunctival epithelial cells (Innoprot) were treated with a miR-744-5p antagomir for 72 hours. Cells were lither left untreated or cultured with polyinosinic:polycytidylic acid (Poly I:C) during the final 24 hours of culture. (F) Expression of Pellino3 in miR-744-5p antagomir transfected human conjunctival epithelial cells (Innoprot) after 72 hours was determined by real-time PCR. Values are the mean ± SD of 6 samples, *P ≤ 0.03. (G,H) Levels of CXCL10 and Rantes in human conjunctival epithelial cells (Innoprot) as determined by ELISA. Values are the mean ± SD of 3 samples, *P ≤ 0.03, **P ≤ 0.005. UT = untransfected cells, Neg A = negative antagomir and 744A = miR-744-5p antagomir.
