il 1α recombinant protein  (Cell Signaling Technology Inc)

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: On GD15.5, pregnant mice [‘Vehicle’ group (n = 50) and ‘Broad Spectrum Chemokine Inhibitor’ (BSCI) group (n = 50)] were injected intravenously with the first dose of BSCI (10 mg/kg/day) or vehicle (saline). In 24 hrs, on GD16.5, pregnant mice received a second injection of BSCI or vehicle. At the same time, half of the vehicle group (n = 25) and half of the BSCI group (n = 25) received ultrasound-guided intra-amniotic infusions of IL-1α (IL-1α; 400 ng/amniotic sac). The second half of the vehicle group (n = 25) and half of the BSCI group (n = 25) received an infusion of sterile saline. The IL-1α injections were administered into each amniotic sac (number of fetuses was 5-9). (A) Long-term effect of BSCI. Pregnant mice were observed until delivery to record the PTB rate. All BSCI-treated animals from the IL-1α and saline groups that did not deliver preterm were given daily injections of BSCI. Mice from all four study groups (n=6/group) that carried the pregnancy to term were killed after delivery on GD19.5. The number of live pups per litter, birth weights, and placental weights were recorded. (B) Short-term effect of BSCI. In a replicate group of animals, pregnant BSCI-treated and vehicle-treated mice were sacrificed 2,8 and 24 hr post-IL-1α induction. On GD16.5, two hrs after IL-1α injection (t = 2 hr), the first six animals from each groups were killed, and maternal and fetal tissues were collected for analysis. After 8 hrs (t = 8 hr), six animals from each group were killed and maternal and fetal tissues were collected for analysis. The remaining animals (n=6/group) were killed during PTB, or exactly 24 hrs after the administration of IL-1α (t = 24 hr), and tissues were collected for biochemical and immunohistochemical evaluation.

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Injection, Saline, Sterility, Immunohistochemical staining

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: (A) To induce in utero sterile inflammation, the alarmin IL-1α (400 ng/amniotic sac) was intra-amniotically administered to C57BL/6 dams under ultrasound guidance on gestational day (GD) 16.5. Dams were monitored until delivery, and neonatal survival and weight were recorded. (B) Gestational lengths are shown for each treatment group in time or GD. The dotted line indicates the threshold of term deliveries after GD 18. (C) Rates of preterm birth among dams injected with saline (n = 6), IL-1α (n = 8), BSCI (n = 5), and IL-1α + BSCI (n = 10) are shown as bar plots. (D) The fetal weights at delivery were calculated as an average of 3 pups per dam. Representative images show a fetus from each group. The dotted line indicates the threshold of viable pups. Scale bar represents 1 cm. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: In Utero, Sterility, Injection, Saline

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: Pro-inflammatory (IL-1α, IL-6, IL-12p70, TNF-α, CSF2) and anti-inflammatory (IL-10) cytokines; protein expression was detected by multiplex magnetic bead assay following local IL-1α administration and treatment with BSCI. Shown are vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected BSCI-treated samples (striped grey bars), n =5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference between groups is indicated by *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Expressing, Multiplex Assay, Injection

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: (A) Pro-inflammatory (Il6, Il1β, Tnfα, Csf2/Gm-scf) and anti-inflammatory (Il10) cytokines; (B) Chemokines Ccl2 (Mcp1), Ccl4 (Mip1β), Cxcl1 (KC/Groα), and Cxcl2 (Mip2α) transcripts were detected by Real-Time RT-PCR in mouse liver. Shown are samples from vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected + BSCI-treated pregnant animals (striped grey bars), n = 5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by * (P < 0.05), ** (P < 0.01) and *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Quantitative RT-PCR, Injection

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: (A) Pro-inflammatory (Il6, Il1β, Tnfα, Csf2/Gmscf) and anti-inflammatory (Il10) cytokines; (B) Chemokines (Ccl2/Mcp1, Ccl4/Mip1β, Cxcl1/KC/Groα and Cxcl2/Mip2α); (C) Pro-labor genes Nfkb1, Ptgs2/Cox2, Akr1c18/20αHSD, and Gja1/Cx43 transcripts were detected by Real-Time RT-PCR. (D) Pro-inflammatory cytokines (CSF2, CSF3, IL-6) and chemokines (CCL2, CCL3, CCL4, CXCL1, Eotaxin) proteins were detected by multiplex magnetic bead assay (BioRad). Shown are vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected BSCI-treated samples (striped grey bars), n = 5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by * (P < 0.05), ** (P < 0.01), and *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Quantitative RT-PCR, Multiplex Assay, Injection

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: (A) Pro-inflammatory (Il6, Il1β, Tnfα, Csf2) and anti-inflammatory (Il10) cytokines; (B) Chemokines (Ccl2/Mcp1, Ccl4/Mip1β, Cxcl1/KC/Groα and Cxcl2/Mip2α) mRNA levels were detected by Real-Time RT-PCR. (C) Pro-inflammatory (CSF2, CSF3, IL-6, IL-12(p40), IL-1β, TNF-α), anti-inflammatory (IL-10) cytokines; and (D) chemokines (CCL2, CCL3, CCL4, CCL5, CXCL1, Eotaxin) protein levels were detected by multiplex magnetic bead assay. Shown are vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected BSCI-treated samples (striped grey bars), n=5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by * (P < 0.05), ** (P < 0.01), and *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Quantitative RT-PCR, Multiplex Assay, Injection

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: Pro-inflammatory (IL-6, TNF-α, CSF2) and anti-inflammatory (IL-10) cytokines; protein expression was detected by multiplex magnetic bead assay following local IL-1α administration and treatment with BSCI. Shown are vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected BSCI-treated samples (striped grey bars), n = 5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by * (P < 0.05).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Expressing, Multiplex Assay, Injection

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: (A) Pro-inflammatory (Il6, Il1β, Tnfα, Csf2/Gmscf) and anti-inflammatory (Il10) cytokines; (B) Chemokines (Ccl2 (Mcp1), Ccl4 (Mip1β), Cxcl1 (KC or Groα), and Cxcl2 (Mip2α)) mRNA expression was detected by Real-Time RT-PCR. Shown are vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected BSCI-treated samples (striped grey bars), n = 5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by * (P < 0.05), ** (P < 0.01), and *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Expressing, Quantitative RT-PCR, Injection

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: (A) A UCSC bedgraph of transcript reads mapped to the mm10 genome demonstrating an increase in expression of the chemokine Cxcl2 in response to IL-1α, and repression of Cxcl2 expression with BSCI pre-treatment at 24 hrs (n=3). (B) Principal component analyses demonstrating similarity in transcript profiles (RNASeq) between saline vehicle control (V/V), BSCI/V, IL-1α/BSCI, and the independent clustering of IL-1α/V. (C) Bar graph showing comparison of differentially expressed genes identified (abs. fold change ≥ 2; padj < 0.05; Y-axis) in V/BSCI, IL-1α/V, and IL-1α/BSCI compared to V/V control. The number of differentially expressed genes is on the Y-axis and listed above each bar, while differentially expressed genes common between conditions are shown below the X-axis. (D) An example of a UCSC bedgraph track generated from ATAC-seq data mapped to the mm10 genome demonstrating an increase in accessibility around the chemokine Cxcl2 in response to IL-1α, and repression of this increase in accessibility by BSCI (n=3). Bars under UCSC tracks identify where ATAC-seq peaks were called using macs2. (E) Principal component analyses demonstrating similarity in chromatin accessibility (ATACseq) between V/V, V/BSCI, IL-1α/BSCI, and the independent clustering of IL-1α/V. (F) Volcano plot showing regions of chromatin gaining and losing accessibility between IL-1α/Vehicle and V/V and (G) BSCI/IL-1α and V/V. Y-axis is -log 10 (padj) and X-axis log 2 (foldchange); dashed lines represent significance thresholds (abs. fold change ≥ 2; padj < 0.05). Samples in grey did not meet criteria for significance.

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Expressing, RNA sequencing, Saline, Control, Comparison, Generated

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: Volcano plots showing differentially expressed proteins identified by mass spectrometry, with each dot representing one protein. In both panels, red dots indicate significantly increased (upregulated) proteins and blue dots indicate significantly decreased (downregulated) proteins. (A) Comparison of IL-1α/vehicle (IL-1α/V) vs vehicle/vehicle (V/V) tissue, with selected downregulated proteins annotated. (B) Comparison of IL-1α/BSCI vs IL-1α/vehicle (IL-1α/V) tissue, with selected upregulated proteins annotated. Gene Ontology (GO) enrichment analyses of differentially expressed proteins: (C) IL-1α/vehicle (IL-1α/V) vs vehicle (V/V) and (D) IL-1α/BSCI vs IL-1α/vehicle. In both, enriched biological processes are shown by fold enrichment, with point size reflecting enrichment magnitude and color representing statistical significance (−log10[FDR]). (E) Bar graphs showing raw mass spectrometry intensity values of selected extracellular matrix (ECM) proteins across V/V, IL-1α/V, and IL-1α/BSCI groups, with particular emphasis on fibrillar collagen-associated proteins (including COL3A1, COL1A1, and COL1A2). Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by * (P < 0.05), ** (P < 0.01), and *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Mass Spectrometry, Comparison

Journal: bioRxiv

Article Title: A Broad-Spectrum Chemokine Inhibitor Prevents Preterm Labor in Mice by Supressing Inflammation Induced by Intra-Amniotic Injection of Interleukin-1 alpha

doi: 10.64898/2026.03.04.709657

Figure Lengend Snippet: Macrophages were identified using anti-F4/80 antibody (yellow), using anti-CD86 for pro-inflammatory M1 macrophages (red), and using anti-CD206 for homeostatic M2 macrophages (green). Bright field (BF, top row) images were used to mask the borders between the myometrium and the decidua. (A) Shown are the representative immunofluorescence images with negative control inlets (images taken with respective IgG controls for each antibody). (B) Qupath software was used to quantify the number of macrophages in the myometrium and decidua. The bar graphs show quantification of vehicle (white bars), BSCI-treated (striped bars), IL-1α-injected (grey bars), and IL-1α-injected BSCI-treated samples (striped grey bars), n = 5–6/group. Two-way ANOVA was utilized, followed by a Bonferroni post-test. Results were expressed as mean ± SEM. Significant difference is indicated by ** (P < 0.01) and *** (P < 0.001).

Article Snippet: According to previous optimized conditions by Motomura et al. , ultrasound-guided intra-amniotic injection of IL-1α (Cat #200-LA/CF, R&D Systems, Inc., Minneapolis, MN, USA) at concentrations of 400 ng per 25 μl of sterile 1× PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Immunofluorescence, Negative Control, Software, Injection