Fig 1: Intradermal recombinant IL-1α triggers both heat and mechanical hypersensitivity in the absence of capsaicin (a). Response latency (s) to radiant heat measured before (0) and 15 and 30 min after 20 μg/ml injection of 5 μg/ml recombinant mouse IL-1α (m-IL-1α) in sterile PBS + 10% FBS (n = 6). Injected and un-injected paws (blue). Mean ± SE response latencies to heat 15 min following injection to: m-IL-1α: 9.66 ± 1.26 s and un-injected 16.06 ± 1.84 s. (b) Percentage change from baseline for Individual mice in response to m-IL-1a (15 min) − 31.03 ± 6.36% and un-injected paws 10.54 ± 10.06%. Paw injection| time interaction analysis of variance measured by two-way ANOVA and Tukey's HSD correction for multiple comparisons p = 0.0018. m-IL-1α injected|un-injected p = 0.0075 (15 min) and m-IL-1α injected|un-injected (30 min) p = 0.0235. (c) Mean ± SE paw withdrawal thresholds to mechanical stimuli before (0) and 15 and 30 min after 20 μL injection of 5 μg/ml recombinant mouse IL-1α (m-IL-1α) in sterile PBS + 10% FBS (n = 6 mice). Mean ± SE for m-IL-1α: 3.58 ± 0.18 g and un-injected = 5.32 ± 0.52 g. (d) Average percent change from baseline for each animal at indicated time points. Mean ± SE values at 15 min were m-IL-1a = − 27.25 ± 2.22% and un-injected = 3.78 ± 9.42%. Paw injection| time interaction analysis of variance measured by two-way ANOVA and Tukey's HSD correction for multiple comparisons p = 0.0063. At 15 min m-IL-1α injected|un-injected p = 0.0205; 30 min m-IL-1α injected|un-injected p = 0.0068.
Fig 2: Dysplastic Basal Cells Induce an Inflammatory Phenotype in Pulmonary Fibroblasts in vitro via Diffusible Factors.(A) Graphic summarizing the generation of conditioned media from ectopic, post-influenza KRT5+ basal cells used to treat fibroblasts. (B) Key inflammatory fibroblast marker genes Saa3 and Ccl2 expression was assayed by RT-qPCR for control and conditioned media treated fibroblasts on plastic, 16kPa, and 0.5kPa plating conditions. Plotted as fold change relative to Plastic Control. (C) PCA Plot showing the 16 samples used for bulk RNA-seq analysis. (D) Volcano plot illustrating significantly differentially expressed genes between control and conditioned media treated fibroblasts cultured on 0.5kPa plates with selected cytokines and chemokines labeled. P value < 0.01, Fold Change > 4 ,184 upregulated 68 downregulated. We note upregulation of Il1a in the fibroblasts themselves upon conditioned media treatment. (E) Heatmap depicting the relative expression of selected inflammatory, fibrotic, and proliferative fibroblast marker genes between the samples depicted in (C). (F) Functional enrichment analysis of the top 92 significantly upregulated genes (P value < 0.01, Fold Change > 4 ) shared between 0.5kPa and Plastic conditioned media treated fibroblasts via Metascape. (B) Data shows mean fold change ± SD and are pooled from 3 independent experiments (n=3 mice). P values were calculated using unpaired t-tests: **P < 0.01, ***P < 0.001, ****P < 0.0001.
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