Fig. 7: SCM-198 induces Ca2+ influx controlled by the Y274 residue of AdipoR2.
a Time-course dynamics of Flow-4 AM fluorescence in AML12 hepatocytes, treated with DMSO or SCM-198 following H2O2 exposure. b Quantification of intracellular Ca2+ content in AML12 hepatocytes treated with DMSO or SCM-198 (0.1, 0.5, 1 µM) for 24 h after H2O2 exposure (n = 4 experimental repeats). c Dynamics of Flow-4 AM fluorescence in AML12 hepatocytes, treated with DMSO or SCM-198 following H2O2 exposure, with or without BATPA. d Ca2+ quantitative assay for AML12 hepatocytes treated with SCM-198 after H2O2 exposure, with or without AdipoR2 knockdown (n = 3 experimental repeats). e Molecular docking analysis of R275TM5-D117N-terminus interaction of ADIPOR2 (PDB ID: 6KS1), viewed from the intracellular side. The binding of Y274TM5 by SCM-198 increases the hydrophobicity of R275TM5, potentially leading to the opening of the internal cavity, allowing ion inflow. R275TM5, D117N-terminus, H348 TM7, and R335TM6/ICL3 are shown as sticks. f Dynamics of Flow-4 AM fluorescence in Adipor2-/- mouse primary hepatocytes with overexpression of AdipoR2WTor AdipoR2Y274A, treated with SCM-198 after H2O2 exposure. g, h Quantification of intracellular Ca2+ content (g) and NO concentration (h) in Adipor2-/- mouse primary hepatocytes with overexpression of Adipo2WTor AdipoR2Y274A, treated with SCM-198 after H2O2 exposure (n = 5 experimental repeats). i A model of actions of SCM-198. There exists an AdipoR2-CaM-CaMKII-NOS3 axis in liver cells. TAA/APAP or H2O2 injuries reduce phosphorylation levels of NOS3 and CaMKII. The binding of SCM-198 to AdipoR2 through R335 and Y274 causes Ca2+ influx and increases phosphorylation levels of NOS3 and CaMKII, leading to quick upregulation of NO synthesis. Created in BioRender. Lin, G. (2024) https://BioRender.com/n69o964. Data in (b, d, g, h) are presented as mean ± SEM. The p-values were determined using SPSS v20 for t test, two-tail analysis. ns: not significant. Source data are provided as a Source Data file.
