TNF Intracellular staining in CD14 positive monocytes from healthy control and RA patients.

<p>A) Thawed A) Cryopreserved PBMC from control and RA patients were stimulated for 4hrs with LPS (1 µg/ml), heat-aggregated Ig [HAG] (100 µg/ml) or the absence of stimulation (NS). Representative dot plots/histograms are shown for all treatments in a HC individual and for HAG stimulation in a RA patient (bottom panel). Intracellular staining was performed using anti-human TNF-PE or isotype control-PE antibodies and MFI units of expression measured. B) Intracellular staining for TNF in response to LPS (left panel) and HAG (right panel) in healthy control (HC, n = 8) and RA (n = 15) subjects. C) Correlation between HAG-stimulated TNF intracellular staining and % CD16 positive monocytes in RA patients. The percentage of CD14⁺⁺ monocytes expressing CD16 (x-axis) and the MFI of anti-TNF-PE expression (y-axis) were determined by flow cytometry. Open diamonds represent early RA patient and shaded diamonds represent long-standing RA patients. Correlation coefficient spearman's rho = 0·813, p<0.001.</p

Early RA patient baseline demographics and associations with response to steroid/methotrexate therapy.

<p>A cohort of early RA patients (n = 73) were recruited for investigation. Baseline characteristics of the cohort is summarised in column [Entire cohort]. Patients with EULAR response data at week 14 post-initiation of therapy (n = 42) were furthermore split into non-responders [NR] or moderate/good responders [MOD/GOOD] and baseline characteristics in each subgroup analysed separately. Statistically significant differences between the two EULAR response subgroups [NR vs. MOD/GOOD] were examined. (‡) fishers statistical test (φ) mann-whitney U test.</p

FcγRIIIa/CD16 expression on RA CD14⁺⁺ monocytes pre- and post- methotrexate therapy.

<p>A) The percentage of CD16 positive CD14⁺⁺ monocytes was measured at baseline (y-axis) and correlated with patient responses to methotrexate treatment. Patients were classified at week 14 post-initiation of therapy as non-responders (NR, n = 12), moderate (MOD, n = 12) or good (GOOD, n = 15) according to EULAR criteria and compared using Mann Whitney U test. Dots represent outliers. (B) The percentage reduction in DAS28-ESR between 0 and 14 weeks post-initiation of therapy (x-axis) and the % of CD14⁺⁺ cells expressing CD16 at baseline (y-axis). Negative levels indicate increased DAS28ESR between baseline and week 14. Statistics; Spearman's correlation. C) The percentage of CD16 positive CD14⁺⁺ monocytes was measured at baseline (wk0), 2 and 14 weeks after initiation of steroid/methotrexate treatment in early DMARD-naïve RA patients. Samples at 2 weeks mainly reflect influence of steroid treatment whereas week 14 samples will mainly reflect the effect of methotrexate therapy. Wilcoxon signed ranks test used to compare paired samples pre- and post- therapy.</p