Mouse IL-1 beta/IL-1F2 Biotinylated Antibody

Detection of Mouse IL-1 beta/IL-1F2 by Western Blot

Adenosine mediates increase in pro-IL-1 beta via a HIF-1 alpha-dependent pathway. (a) Consensus NF kappaB and HRE binding sites in the IL-1 beta promoter. (b) LPS primed PECs obtained from A2AR-cKO and controls were stimulated with or without NECA or CGS21680. (c) LPS primed PECs obtained from HIF-1 alpha-cKO and controls were stimulated with or without NECA at time points as indicated. Cells were harvested and RNA isolated after each treatment and gene expression of Il1b and Hif1 alpha quantified by real-time PCR. (d) THP-1 cells were transfected with HRE- promoter luciferase construct and beta-galactosidase plasmid in the presence or absence of CREB dominant negative plasmid (CREB delta), and then primed with LPS/PMA followed by NECA. (e) THP-1 cells were transfected with human IL-1 beta promoter luciferase construct and beta-galactosidase plasmid in the presence or absence of CREB delta, and then primed with LPS/PMA followed by CGS21680 or NECA. Luciferase activities were measured and normalized to beta-galactosidase activity and normalized with controls. Data are mean± SD of triplicate cultures and are representative of three independent experiments. (f) CD14-MD2-TLR4- HEK 293 cells were transfected with NF kappaB promoter luciferase construct and Renilla luciferase (Rluc) control reporter vector, and then treated with CGS21680, NECA or ZM241395 in the presence of LPS, Luciferase activities were measured and normalized to Rluc activity and the normalized value with controls as indicated. Data are mean± SD of triplicate cultures and are representative of three independent experiments for b-f. (g) LPS primed PECs obtained from HIF-1 alpha-cKO and controls were treated with or without NECA at different time points as indicated followed pulsing with ATP. Cell supernatants were collected in 5 hrs after ATP pulsing. IL-1 beta was measured in cell supernatant by ELISA. Data are expressed as the mean ± SD from three independent experiments. (h) LPS primed PECs obtained from HIF-1 alpha-cKO and control mice were treated with or without NECA at different time points as indicated followed by pulsing with ATP. Cell lysates were collected after ATP pulsing. HIF-1 alpha, pro-caspase-1, Caspaspe-1 and pro-IL-1 beta was measured in cell supernatant by western blot. Immunoblots shown are representative results from at least three independent experiments. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24352507), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse IL-1 beta/IL-1F2 by Western Blot

Liver injury and fibrosis is dependent on A2A receptor signaling in macrophages(a)A2AR-cKO and control mice were injected intraperitoneally with LPS (1 mg/kg) and D-galactosamine (500 mg kg−1) for 6 hrs followed by liver tissue and serum collection for H&E staining and ALT assay. (b) Liver RNA samples were collected and Il1b gene assayed by real-time PCR using specific primers. (c) Liver tissue lysates were assayed for pro-caspase-1, cleaved caspase-1 (p10), and beta-actin protein level by immunoblot analysis using specific antibodies. Data are expressed as the mean ± SD from 10–11 mice from each group for a-d. (d) Serum was collected for measurement of IL-1 beta. (e)A2AR-cKO and control mice were injected intraperitoneally with single dose of TAA followed by liver tissue collection as indicated for H&E staining. (f) Liver RNA samples were collected and Il1b gene was assayed by real-time PCR. (g) Liver tissue was also obtained at day 7 after TAA injection and stained for H&E and Sirius red for fibrosis. (h) Sera were collected and the serum ALT assay was performed (Data are expressed as the mean ± SD from 5 mice in each group). * p <0.05 determined by Student’s t-test. Scale bars correspond to 500μm. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24352507), licensed under a CC-BY license. Not internally tested by R&D Systems.