interleukin 1 beta il 1β  (R&D Systems)

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: (A) Schematic of the primary mouse astrocyte cultures. (B) Representative immunofluorescence images of cultured astrocytes stained for GFAP, showing characteristic astrocyte morphology. (C) Overview of RNA-seq and capped small (cs)RNA-seq data generation from astrocytes. The schematic shows the typical distribution of csRNA-seq and RNA-seq at various genomic locations, which allows the identification of Transcriptional Start Site (TSSs) using HOMER2. (D) Volcano plot of RNA-seq differential expression in astrocytes treated with vehicle (Veh) or IL-1B (10 ng/mL, 1 h). (E) Pathway enrichment analysis of IL-1B-induced differentially expressed genes. (F) RT-qPCR validation of selected IL-1B-responsive genes in astrocytes

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: Immunofluorescence, Cell Culture, Staining, RNA Sequencing, Quantitative Proteomics, Quantitative RT-PCR, Biomarker Discovery

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: Genome browser views showing RNA-seq signal at the Cxcl1, Ccl20, Csf2 , and Il6 loci in astrocytes treated with vehicle or IL-1B. These representative examples illustrate the robust transcriptional induction of canonical inflammatory genes following IL-1B stimulation. Gene models and genomic coordinates are shown in mm10 .

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: RNA Sequencing

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: (A) Pairwise correlation analysis of csRNA-seq replicates from untreated and IL-1B-treated astrocytes. (B) Distribution of strand-specific csRNA-seq reads around GENCODE-annotated TSSs, showing strong enrichment at annotated transcriptional start sites. (C) Genomic annotation of astrocytes TSRs, partitioned across promoter, intronic, intergenic, and other genomic features. (D) Average chromatin profiles at promoter-distal TSRs, showing ATAC-seq and H3K27ac enrichment around transcribed regulatory elements. (E) Average csRNA-seq signal centered on promoter-distal ATAC-seq peaks, comparing transcribed and non-transcribed accessible regions (No Tx: 225,150 peaks, Tx: 14,836 peaks). (F) Average mCH profiles at promoter-proximal TSRs, promoter-distal transcribed TSRs, and non-transcribed distal accessible regions in astrocytes (data from ). (G) Distribution of NFIA and TEAD4 ChIP-seq peaks found overlapping TSRs and ATAC-seq peaks in astrocytes. (H)ChIP-seq read density for NFIA and TEAD4 centered on csRNA-seq defined TSRs, showing transcription factor binding immediately upstream of the primary TSS.

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: ChIP-sequencing, Binding Assay

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: (A)Volcano plot of differentially regulated csRNA-seq signal at astrocyte TSRs following IL-1B stimulation, identifying induced and repressed regulatory elements. (B) Genome browser tracks depicting induction of promoter and distal regulatory enhancer transcription by csRNA-seq at the Ccl2 locus. (C) GREAT functional enrichment analysis of IL-1B induced TSRs. (D) Spatial clustering of IL-1B induced TSRs, depicting the density of TSRs in each category adjacent to Induced TSRs. (E) De novo motif enrichment analysis of IL-1B induced TSRs by HOMER. (F) Average csRNA-seq signal centered on promoter-distal NF-κB/p65 peaks, showing increased bidirectional transcription at p65-bound distal elements after IL-1B stimulation, consistent with eRNA induction. (G) Spatial density of NF-κB, AP1, and IRF motifs relative to TSRs (TF binding sites per bp per TSS), showing upstream enrichment of these motifs at IL-1B-induced TSRs compared with unchanged or repressed TSRs.

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: Functional Assay, Binding Assay

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: (A) Genome browser example of an IL-1B-induced locus (Cxcl10) in astrocytes, showing increased transcription initiation after stimulation. (B) Scatter plot comparing IL-1B-induced Log2 csRNA-seq changes at the promoter vs. RNA-seq changes across genes, highlighting genes regulated primarily at initiation (along x-axis) versus those showing stronger changes at the mRNA level (along y-axis). (C) Genome browser tracks at the Junb locus showing increased gene expression and RNAPII elongation in the gene body with limited change in promoter initiation and RNAPII promoter levels, consistent with regulation being mediated primarily at the level of transcription elongation rather than increased initiation. (D) RNAPII ChIP-seq levels at the promoters of IL-1B induced genes stratified by genes with minimal versus strong increases in csRNA-seq initiation activity. (E) Scatter plot comparing changes in overall TSR levels (Log2 Fold change, NT vs. IL-1B) versus their WIP score significance (the −Log10 p-value), identifying TSRs with altered initiation patterns independent of changes in total transcriptional output. (F) Representative examples of TSRs exhibiting a strong change in initiation pattern (top, WIP score −1.61, Lg10 p-value = 9.54e-05) versus a strong change in overall initiation levels with minimal change in initiation shape(WIP score −0.13, Log10 p-value = 0.84). (G) Scatter plot of TF motif enrichment in TSRs with significant changes in overall activity versus changes in TSS positions, highlighting differential associations of NF-κB, TEAD, and NFI motifs with these TSRs classes. (H) Venn diagram showing the overlap of ChIP-seq peaks for NFIA and TEAD4 before and after IL-1B stimulation in astrocytes. (I) Motif enrichment analysis of condition-specific (either Veh or IL-1B) TEAD4- and NFIA-bound regions, showing IL-1B-specific enrichment for inflammatory TF motifs, including NF-κB, IRF (IRF8), and AP1 (i.e. Fra1).

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: RNA Sequencing, Gene Expression, ChIP-sequencing, Activity Assay

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: (A) Venn diagram showing overlap between IL-1B-induced genes in astrocytes and KLA-induced genes in bone marrow-derived macrophages (BMDMs). (B) Venn diagram showing overlap between induced TSRs in astrocytes and BMDMs, revealing largely distinct stimulus-responsive enhancer landscapes despite partial overlap in induced genes. (C) Genome browser view of Tnfaip3 locus illustrating that astrocytes and BMDMs induce the same gene but use different enhancers upstream a shared promoter. (D) Top: Venn diagram showing overlap of NF-κB binding sites between activated astrocytes and BMDMs. Bottom: Fraction of astrocyte-specific, shared, and BMDM-specific p65 peaks associated with TSRs induced in astrocytes only, in both cell types, and in BMDM only. (E) Heatmap of de novo motif enrichment at cell type-specific induced TSRs, showing enrichment of lineage-associated motifs for each cell type. (F) Fraction of induced TSR classes bound by lineage-associated TFs, showing preferential association of astrocyte-induced TSRs with NFI and TEAD4 and macrophage-induced TSRs with PU.1 and CEBPα.

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: Derivative Assay, Binding Assay

Journal: bioRxiv

Article Title: Transcription initiation profiling defines the regulatory logic of astrocyte gene regulation

doi: 10.64898/2026.05.03.722406

Figure Lengend Snippet: (A) Average eRNA signal centered on astrocyte-specific, shared, and BMDM-specific NF-κBp65 peaks, showing cell type-matched induction of regulatory transcription at p65 bound sites. (B) Fraction of astrocyte-specific, shared, and BMDM-specific p65 peaks overlapping accessible chromatin regions in astrocytes or BMDMs, indicating that NF-κB recruitment occurs preferentially at cell-type-specific open chromatin regions. (C) Violin plots showing increase in ChIP-seq signal for p65, NFIA, TEAD4 at astrocyte IL-1B-induced TSRs and for p65, PU.1, and CEBPβ in macrophage KLA-induced TSRs.

Article Snippet: Two days before collection, the astrocytes were plated at a density of 500,000 cells per well on 4-well chamber slides to perform immunocytochemistry or a density of 5.5 million cells per 15 cm dish to be treated with 10ng/mL of recombinant human interleukin 1 beta (IL-1B) (Invivogen, Cat#rcyec-h) for 1hr before sample collection.

Techniques: ChIP-sequencing

Journal: Journal of Orthopaedic Translation

Article Title: Intermittent intra-articular delivery of FGF8b enhances cartilage homeostasis and attenuates osteoarthritis progression

doi: 10.1016/j.jot.2025.101037

Figure Lengend Snippet: FGF8b prevents human cartilage from IL-1β induced degeneration. A. Safranin O staining revealed increased proteoglycan content in the extracellular matrix of human cartilage sections after FGF8b treatment for Ex vivo OA model. B-D. IHC staining for Acan, Col X, and Mmp13 was performed on human articular cartilage explants treated with IL-1β and FGF8b. Scale bar: 100 μm, n = 3. E-G. Statistical analysis of relative expression of chondro-homeostasis-associated genes in cartilage explants. H. Human primary chondrocytes were treated with varying concentrations of FGF8b for 24 h, followed by western blotting analysis. I-J. The signal intensities of Sox9 and Runx2 were quantitatively analyzed. K. Human primary chondrocytes were treated with IL-1β (10 ng/ml) to induce an OA-like chondrocyte model, along with FGF8b (25 ng/ml) for 24 h. Western blotting was performed to assess the effects of FGF8b on chondrocyte homeostasis-related markers. L-M. Quantitative analysis of Mmp13 and Adamts5 gray value. n = 3. ns: not significant, ∗P < 0.05, ∗∗P < 0.01, and ∗∗∗P < 0.001.

Article Snippet: Explants were then treated for 4 days with 10 ng/ml interleukin-1β (IL-1β) (201-LB-005, R&D Systems) and 100 ng/ml FGF8b (100-25, PeproTech) in this serum-free medium.

Techniques: Staining, Ex Vivo, Immunohistochemistry, Expressing, Western Blot

Journal: Journal of Orthopaedic Translation

Article Title: Intermittent intra-articular delivery of FGF8b enhances cartilage homeostasis and attenuates osteoarthritis progression

doi: 10.1016/j.jot.2025.101037

Figure Lengend Snippet: FGF8b exerts a chondroprotective effect via the FGFR3-PI3K-AKT signaling pathway. A. Surface plasmon resonance analysis shows the high-affinity binding of FGF8b to FGFR3. B-C. FGF8b treatment of SW1353 chondrocytes for 15 min increased the expression of phosphorylated FGFR3. D-E. The knockdown efficiency of FGFR3 following transfection with FGFR3 siRNA in SW1353 cells was assessed by Western blotting. F-H. The chondroprotective effect of FGF8b for 24 h was partially attenuated after FGFR3 knockdown. I. KEGG enrichment analysis of transcriptome sequencing data suggested an upregulation of the PI3K-AKT signaling pathway following FGF8b treatment for osteoarthritis. J-L. The FGFR3-PI3K-AKT signaling pathway was downregulated in chondrocytes treated with IL-1β for 15 min, whereas FGF8b treatment activated the FGFR3-PI3K-AKT signaling pathway. M. The activation of the PI3K-AKT signaling pathway by FGF8b is dependent on FGFR3. N-P. The chondroprotective effect of FGF8b was partially abolished upon blockade of the PI3K-AKT signaling pathway using the LY294002 inhibitor under inflammatory conditions. n = 3, ns: not significant, ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001 and ∗∗∗∗P < 0.0001.

Article Snippet: Explants were then treated for 4 days with 10 ng/ml interleukin-1β (IL-1β) (201-LB-005, R&D Systems) and 100 ng/ml FGF8b (100-25, PeproTech) in this serum-free medium.

Techniques: SPR Assay, Binding Assay, Expressing, Knockdown, Transfection, Western Blot, Sequencing, Activation Assay