Characteristics of the patients and controls enrolled in the study.

<p>Characteristics of the patients and controls enrolled in the study.</p

pSS patients with increased B cell hyperactivity can be identified using hierarchical clustering of serum sncRNA expression levels.

<p>Nine sncRNAs were selected based on their differential expression in the discovery array and subsequent technical replication. These sncRNAs were measured using single-assay RT-qPCR in samples from both cohorts (n = 75). Unsupervised hierarchical clustering was performed on the expression of the nine selected sncRNAs in all 37 pSS patients. Grey fields depict unavailable data points (A). Clinical parameters and frequency of positivity for anti-La (SSB) autoantibodies were compared between the three clusters (B). The patients in each cluster were compared using Kruskal-Wallis H test with post-hoc Dunn’s test of multiple comparisons and Fisher’s exact test. For dot plots, medians ± IQR are shown.</p

IL-7 and TLR7 synergistically increases CD4 T cell proliferation in T and B co-cultures.

<p>Representative FACS stainings for KI67⁺ CD4 T cells from unstimulated, IL-7, and TLR7/IL-7 stimulated T/B cell co-cultures in the absence or presence of monocytes/macrophages are shown (n = 5). IL-7 significantly stimulates proliferation of CD4 T cells, which is synergistically increased when combined with TLR7 stimulation (<b>A, B</b>). IL-7-induced CD4 T cell proliferation is enhanced in the presence of monocytes/macrophages, but no additive effect is observed with a combination of IL-7 and TLR7 stimulation (<b>C, D</b>).*p<0.05 and **p<0.001 indicate statistical significance compared to medium values.</p

IL-7 synergistically increases proliferation of TLR7-stimulated B cells in co-culture with CD4 T cells, which is enhanced by monocytes/macrophages.

<p>Isolated B cells co-cultured 1∶1 (5.10⁵ each) together with CD4 T cells for 6 days show an increased lymphocytic proliferation upon TLR7 or IL-7 stimulation, which is additively increased upon combined stimulation with IL-7/TLR7 (n = 8) (A). A similar, but overall enhanced effect for the total proliferation is seen when monocytes/macrophages are added to the culture (5.10⁴ B). Representative FACS stainings for KI67⁺ B cells from an unstimulated, TLR7, IL-7, and TLR7/IL-7 stimulated CD4 T/B cell co-culture -/+ monocytes/macrophages are shown as well as the average data (n = 5) (C, D). TLR7 induces a significant increase in the percentage of KI67⁺ B cells. IL-7 stimulation induces a small, but statistically significant increase in Ki67⁺ B cells. When TLR7 and IL-7 are added together a synergistic increase in proliferation is observed (C). Overall the effects are enhanced by addition of monocytes/macrophages to the T/B cell co-cultures (D). * and ** indicate a statistical significant differences of p<0.05 and p<0.01, respectively, as compared to medium values or between treatments.</p

Correlations between serum sncRNA levels and disease parameters in pSS patients.

<p>Correlations between serum sncRNA levels and disease parameters in pSS patients.</p

RT-qPCR data of all nine sncRNAs included in the validation phase.

<p>Serum sncRNAs were measured using single Taqman qRT-PCR in the validation cohort (n = 45). ΔCt per sample was calculated using the expression of an exogenous spiked-in Arabidopsis thaliana miRNA to correct for technical variation. The relative expression of each sample was calculated as fold change (FC) in comparison with the ΔCt mean of the HC group in the respective cohort. Medians ± IQR are shown.</p

Results from discovery and validation cohort analyses.

<p>Results from discovery and validation cohort analyses.</p