CD4⁺NKG2D⁺ T cells autoimmune diseases.

<p>This figure depicts the ratio of CD4⁺NKG2D⁺ T cells within the total peripheral CD4 pool in control, RA, SLE and SS individuals. The variability in SS patients is due to 2 individuals whom represent an unusually high ratio (see main text for explanation). None of the differences is significant as assessed by the Wilcoxon text: p = 0.1658 for controls vs. RA; p = 0.9547 for controls vs. SLE and p = 0.5012 for controls vs. SS.</p

Expression analysis of <i>MICA</i> and <i>MICB</i> by real-time quantitative PCR in HeLa cells.

<p><i>MICA</i> (A) and <i>MICB</i> (B) expression profile as analyzed by RT-qPCR. ▪ Samples 15 minutes after heat shock, ♦ Samples 60 minutes after heat shock, ▴ untreated controls.</p

Characteristics of patients and healthy controls.

*<p>DAS (Disease Activity Score) 28 for RA, SLEDAI (Systemic Lupus Erythematosus Disease Activity Index) for SLE and mean AVS (AVS: Analogic Visual Scale) of subjective sicca syndrome, asthenia and pain for SS.</p>**<p>Abbreviations: NSAID: Non-Steroidal Anti-Inflammatory Drugs; SMARD: Symptom Modifying Anti-Rheumatic Drugs; DCART: Disease Controlling Anti-Rheumatic Therapy; ANA: antinuclear antibody (indirect immunofluorescence on Hep2 cells), ENA: extractable nuclear antigens; dsDNA: double-stranded DNA, RF: Rheumatoid Factor, CCP: cyclic citrullinated peptide.</p

Expression analysis of <i>MICA</i> and <i>MICB</i> by real-time quantitative PCR in Sjögren's syndrome accessory salivary glands.

<p>Relative expression level of <i>MICA</i> (A) and <i>MICB</i> (B) in Sjögren's syndrome patients as compared to healthy controls. The differences between patients and controls are not significant as assessed by Mann-Whitney <i>U</i> test (α = 0.27 for <i>MICA</i> and α = 0.53 for <i>MICB</i>).</p

MICA expression in Rheumatoid Arthritis synovitis and Sjögren syndrome accessory salivary glands.

<p>Immunohistochemical analysis of (A) (B) RA synovitis and an (C) (D) SS accessory salivary gland stained with an anti-MICA monoclonal antibody. MIC expression is clearly of epithelial/fibroblastic nature in both tissues/organs.</p

Flow cytometric analysis of the CD4⁺NKG2D⁺ population.

<p>This example, taken from one of the two SS patients (Mrs XET) harbouring a particularly high ratio of CD4⁺NKG2D⁺ T cells (36.2%) illustrates the methodology used in order to enumerate these cells in all individuals (controls and patients) included in this work during a routine 50 000 events analysis.</p

Northern blot analysis of <i>MICA</i> and <i>MICB</i> expression in various tumors.

<p>The same disposition as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0000518#pone-0000518-g001" target="_blank">Figure 1</a> applies, but this time the blots contain tumor and control (adjacent disease-free tissue) lanes.</p

Immunohistochemistry of RA joint infiltrate.

<p>(A) Color stain by Hematoxylin-eosin staining. Immunohistochemical analysis using (B) CD4 (C) CD8 and (D) NKG2D antibodies. The infiltrate is clearly of lymphocytic origin with a majority of CD4 T cells.</p

Effect of LPS on TNF-αmRNA expression in RA FLS and THP-1 cells.

<p><b>A, B.</b> TNF-α mRNA expression was determined by RT-PCR in RA FLS (<b>A</b>) and THP-1 cells (<b>B</b>) stimulated with LPS (1 µg/ml) for 2 h, 4 h and 6 h. Control cells were incubated for 4 h with medium (C). <b>C, D.</b> RA FLS and THP-1 cells were stimulated for 3 h with LPS and then incubated for another 1, 2, 3 and 4 h with actinomycin D (5 µg/ml). Control cells were incubated for 3 h with medium. TNF-α mRNA expression was determined by RT-PCR. The results are representative of three different experiments for each patient.</p

Effect of transfection of miR-346 antagomirs on TNF-α mRNA stability and release in RA FLS.

<p><b>A.</b> FLS were transfected with miR-346 antisense molecules or with the Clear-miR™ negative control (control), activated with LPS 24 h post-transfection for 3 h and incubated for another 1, 2, 3 and 4 h with actinomycin D. Control cells were incubated for 3 h with medium (C). NT: non transfected cells. <b>B, C</b>: TNF-α expression was detected using cellular ELISA or western blotting with anti-TNF-α antibodies, in FLS transfected with miR-346 antisense molecules or with the Clear-miR™ negative control (control) or in non transfected FLS (NT). 24 h post-transfection, FLS were either incubated in medium (C) or activated with LPS for 6 h. The results are representative of three different experiments for each patient. <b>D.</b> TNF-α release was determined by ELISA in culture supernatants after stimulation with LPS or medium (C). Data are expressed as the mean of triplicate samples +/− SD of three independent experiments for each patient. p<0.01.</p