was detected by the Bridge-It cAMP designer cAMP assay (Mediomics, St. Louis, MO). Cholangiocytes (10,000/well) were incubated with TLCA, OA, C1 and C2 (all, 25 µM), SBI-115 (100 and 200 µM) and pasireotide (20 µM, MedChemexpress, Monmouth Junction, NJ) for 15–30 min. Doses of drugs were chosen based on published data9 (link), 18 (link) or dose ranging (SBI-115).
Sbi 115
Manufactured by MedChemExpress
Sourced in United States
SBI-115 is a research reagent manufactured by MedChemExpress. It is a small molecule compound. This product is intended for laboratory research use only and not for any clinical or diagnostic applications.
Automatically generated - may contain errors
Lab products found in correlation
Gly β mca, by MedChemExpress (2 mentions)
Gw4064, by MedChemExpress (1 mentions)
Vancomycin, by Merck Group (1 mentions)
P iκb, by Affinity Biosciences (1 mentions)
Occludin, by Affinity Biosciences (1 mentions)
Claudin 3, by Affinity Biosciences (1 mentions)
Il 1β, by Cell Signaling Technology (1 mentions)
β actin, by Affinity Biosciences (1 mentions)
Myeloperoxidase mpo assay kit, by Nanjing Jiancheng (1 mentions)
Anti phycoerythrin pe microbeads, by Miltenyi Biotec (1 mentions)
Ls column, by Miltenyi Biotec (1 mentions)
Ldh cytotoxicity assay kit, by Beyotime (1 mentions)
Mouse il 10 quantikine elisa kit, by R&D Systems (1 mentions)
Camp parameter assay kit, by R&D Systems (1 mentions)
Forskolin, by Selleck Chemicals (1 mentions)
5 protocols using sbi 115
1
Cholangiocyte cAMP Response to Bile Acids
2
Evaluating SLA-I Expression in IPEC-J2 Cells
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When IPEC-J2 cells reached approximately 70% confluence, the medium was changed to 10/20 μM LCA (83443, Sigma, USA) solution, 2 μM GW4064 (HY-50108, MedChemExpress, USA), 5 μM CCDC (SD2384, Beyotime, China), 2 μM SBI-115 (HY-111534, MedChemExpress, USA), or 2 μM Gly-β-MCA (HY-114392, MedChemExpress, USA) (the culture medium for the blank group was replaced with basal DMEM) and incubated for 12 h. Each sample was adjusted to 105 cells after incubation, and a portion was treated with SLA-I (XK3759109, Invitrogen, USA) antibody for 20 min while shielded from light. This fraction was then analyzed using a flow cytometry device. The expression of SLA-I in the remaining cells was then determined by ELISA after the other cells were ultrasonically lysed.
3
Inflammatory Signaling Pathway Regulation
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Deoxycholic acid (DCA), cholic acid (CA), and vancomycin were bought from Sigma Aldrich (St. Louis, MO, USA). SBI-115, MDL-12330A, and H89 were purchased from MedChemExpress (MCE, USA). The primary antibodies, including phosphorylation-p65 (p-p65, #AF2006; RRID: AB_2834435), p-65 (#AF5006; RRID: AB_2834847), p-IκB (#AF2002; RRID: AB_2834433), IκB (#AF5002; RRID: AB_2834792), Occludin (#DF7504; RRID: AB_2841004), ZO-1 (#AF5145; RRID: AB_2837631), Claudin-3 (#AF0129; RRID: AB_2833313) and β-actin (#AF7018; RRID: AB_2839420) were obtained from Affinity Biosciences (OH, USA). NLRP3 (#15101), ASC (#67824), and IL-1β (#12242) were bought from Cell Signaling Technology (CST, Boston, USA). Goat anti-rabbit or Rabbit anti-mouse secondary antibodies were bought from ImmunoWay Biotechnology Company. Mouse TNF-α (Cat #430915) and IL-1β (Cat #432615) ELISA assay kits were obtained from Biolegend (CA, USA). Myeloperoxidase (MPO) assay kit was bought from Nanjing Jiancheng Bioengineering Institute (Nanjing, China).
4
PEDV-induced Cytotoxicity Assay
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On day 8 after piglets were challenged with PEDV, jejunal monocytes were stained using PE-labeled anti-pig CD8 (559584, BD, USA), and cells were subsequently collected as effector cells by positive screening in LS Columns (130-122-729, Miltenyi Biotec, Germany) using Anti-Phycoerythrin (PE) MicroBeads (130-048-801, Miltenyi Biotec, Germany). IPEC-J2 cells were grown to 70% confluence, the medium was discarded, and 10 μM LCA, 2 μM SBI-115 (HY-111534, MedChemExpress, USA), or 2 μM Gly-β-MCA (HY-114392, MedChemExpress, USA) was added. IPEC-J2 cells were subsequently infected with PEDV (103.5 PFU ml−1) for 8 h. After coculture of the effector and target cells for 4 h, the supernatant was collected, and the cytotoxicity of each group was assayed using the LDH Cytotoxicity Assay Kit (C0017, Beyotime, China) according to the manufacturer’s instructions.
5
Activation of TGR5 Signaling Pathway
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TGR5 antagonist SBI 115 was purchased from MedChemExpress (Monmouth Junction, NJ, USA) and forskolin (adenylate cyclase activator to increase cAMP) was purchased from Selleck Chemicals (Houston, TX, USA). All antibodies used are listed in Table S1 and specific primer sequences are listed in Supporting Information Table S2 . cAMP and IL-10 were detected with the cAMP Parameter Assay Kit and Mouse IL-10 Quantikine ELISA Kit (R&D Systems, Minneapolis, MN, USA).
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