Fig. 6: In vitro experiments corroborate that specific IFNα-related features are observed in the NR model. | Nature Communications

Fig. 6: In vitro experiments corroborate that specific IFNα-related features are observed in the NR model.

From: Type 1 interferon signature and allograft inflammatory factor-1 contribute to refractoriness to TNF inhibition in ankylosing spondylitis

Fig. 6

a In vitro bulk RNA-seq experimental design. CD14+ monocytes were isolated from human PBMCs and treated with combinations of cytokines and adalimumab (ADA) to create six conditions before harvest and library construction. ‘I’ denotes IFNα and ‘T’ denotes TNFα. b Heatmap of DEGs identified through k-means clustering across six conditions. Modules are named and characterized on the left. Statistical confirmation was performed using the MAST algorithm (Seurat implementation), with significance determined by Bonferroni-adjusted P < 0.05 and a log2FC > 0.1. c Heatmap showing the relative comparison of modules scored against 11 CD14+ monocyte clusters from patients with AS. Columns are ordered based on the Euclidean distance. d Bar plots showing the pathway analysis and TF analysis for module 4, with the top three P value terms highlighted. Terms with an adjusted P < 0.05 are in green; non-significant terms are in gray. Combined scores were calculated by multiplying the Fisher exact test log(P value) by the z-score of the deviation from the expected rank. (BH adjustment, two-sided) e Venn diagram of ADA-upregulated genes in the R and NR models. Roman numeral I denotes R model-specific genes, II denotes genes upregulated in both models, and III denotes NR model-specific genes, with counts provided. Bar graphs show the counts and proportion of genes upregulated by IFNα added to TNFα or to control within each category.f Line graph of deconvoluted proportions (n = 3) from in vitro bulk RNA-seq data using patient data as the reference. Key clusters from patient data are in bold. Error bars represent the standard error of the mean (SEM). g Box plot comparing the cosine similarity of deconvoluted data for the key clusters in bold from the I + T condition between R_Pre and NR_Pre samples (R n = 12, NR n = 10) and analyzed using the Wilcoxon rank sum exact test. Boxplot definitions as in Fig. 2d. Source data are provided as a Source Data file.

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