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Image Search Results
Journal: Nature Communications
Article Title: G z ESTY as an optimized cell-based assay for initial steps in GPCR deorphanization
doi: 10.1038/s41467-025-59850-8
Figure Lengend Snippet: A Real-time analysis of ligand-mediated activation of a battery of G i/o/z -PCRs (red) and G s -PCR (green) in cells co-transfected with indicated receptors and the 3-in-one G z ESTY plasmid in the presence of 50 µM IBMX. Cells transfected only with the 3-in-one plasmid served as control (white). The following agonists were applied at 4 min: 10 µM dopamine (D2R, D1R), 100 µM clonidine (ADRA2A, ADRA2B, ADRA2C), 10 µM GABA (GABABR), 10 µM DAMGO (MOR), 1 µM SNC-80 (DOR), 10 µM salvinorin A (KOR), 10 µM human galanin (1–30) (GAL1R), 100 µM serotonin (5-HT1B), 10 µM human neuropeptide Y (13–36) (NPYR1), 10 µM lysophosphatidic acid (LPAR2), 10 µM 2-arachidonoyl glycerol (CB1R, CB2R), 10 µM N-formyl-met-leu-phe (FPR1), 1 mM isobutyric acid (FFAR3), 10 µM somatostatin 14 (SSTR1), 10 µM neuropeptide FF (NPFFR1), 10 µM SEW2871 (S1PR1), 10 µM histamine (HRH3), 10 µM quinpirole (D3R), 10 µM TFLLR (PAR1), 10 µM SLIGKV (PAR2), 10 µM MK-6892 (HCA2R), 1 µM teriparatide PTH (PTH1R), 10 µM AB-MECA (ADORA2B), 10 µM NDP-α-MSH (MC4R), and 10 µM CGRP (CLR). Data are shown as means ± SEM; N = 5 independent transfections. B GPCRs that couple to G s and/or G i/o/z and can potentially be detected by GZESTY are highlighted and include 213 out of 249 total ligand-activated GPCRs (86%) (adapted from GPRCdb.org). C Quantification of agonist-induced activity for 24 G i/o/z -coupled GPCRs. On the right, the response to agonist of the last seven GPCRs is also reported with a different scale. Data are shown as means ± SEM of the fold change obtained; N = 5 independent transfections.
Article Snippet: The following chemicals were purchased: clonidine (Tocris), dopamine (Tocris), GABA (Tocris), serotonin (Tocris), 1-oleoyl lysophosphatidic acid (Tocris), DAMGO (MedChemExpress), TFLLR (MedChemExpress), human PAR-2 (1-6, SLIGKV) (MedChemExpress), human galanin (1–30) (MedChemExpress), IBMX (MedChemExpress), SEW2871 (MedChemExpress), somatostatin-14 (Cpc Scientific), human neuropeptide Y (13-36) (Cpc Scientific), neuropeptide FF (Thermo Scientific Chemicals), MK-6892 (MedChemExpress), 2-arachidonoyl glycerol (Cayman Chemicals), N-Formyl-Met-Leu-Phe (R&D systems), SNC80 (Adipogen), isobutyric acid (TCI chemicals), salvinorin A (ChromaDex Inc.), morphine (Mallinckrodt Chemical Company), human β-endorphin (Sigma-Aldrich), teriparatide (MedChemExpress), NDP-α-MSH (Phoenix Pharmaceuticals), AB-MECA (MedChemExpress), quinpirole (Tocris),
Techniques: Activation Assay, Battery, Transfection, Plasmid Preparation, Control, Activity Assay
Journal: Molecular Metabolism
Article Title: Adipose mTORC2 is essential for sensory innervation in white adipose tissue and whole-body energy homeostasis
doi: 10.1016/j.molmet.2022.101580
Figure Lengend Snippet: Sensory but not sympathetic innervation is altered upon loss of adipose mTORC2 . (A) 2D representatives of a 3D reconstruction of inguinal WAT (iWAT) four weeks after tamoxifen treatment immunostained with tyrosine hydroxylase (TH; yellow). (A1-2) Low magnification projection of sympathetic neuronal network in control and iAdRiKO mice (N = 4; 5). Scale bar = 500 μm. (A3-4) High magnification projection of sympathetic neurons in iWAT parenchyma of control mice and iAdRiKO (N = 19; 10). Scale bar = 100 μm. (B) Immunoblot analysis of iWAT from control and iAdRiKO mice four weeks after tamoxifen treatment. Hormone-sensitive lipase (HSL). (n = 6; 6). (C) 2D representatives of a 3D reconstruction of iWAT four weeks after tamoxifen treatment immunostained with calcitonin gene-related peptide (CGRP; magenta). (C1-2) Low magnification projection of sensory neuronal network in control and iAdRiKO mice (N = 12; 19). Scale bar = 500 μm. (C3-4) Low magnification cross section of sensory neuronal network in control and iAdRiKO mice (N = 12; 19). Nerve bundle (1), innervation along blood vessel (2), tissue autofluorescence (green). Scale bar = 500 μm. (C5-6) High magnification projection of sensory neurons in iWAT parenchyma of control mice and iAdRiKO (N = 16; 11). Scale bar = 100 μm. (C7-8) High magnification cross section of neurons in control and iAdRiKO mice (N = 16; 11). Innervation along blood vessel (2), parenchymal innervation (3), tissue autofluorescence (green). Scale bar = 100 μm. (D) Quantification of the total neurite length of CGRP-positive neurons in iWAT parenchyma of control mice and iAdRiKO four weeks after tamoxifen treatment (N = 6).
Article Snippet: CGRP levels in lysates were determined using mouse calcitonin gene-related peptide (CGRP) ELISA kit (CSB-EQ027706MO, Cusabio) according to the manufacturer's instructions.
Techniques: Control, Western Blot
Journal: Molecular Metabolism
Article Title: Adipose mTORC2 is essential for sensory innervation in white adipose tissue and whole-body energy homeostasis
doi: 10.1016/j.molmet.2022.101580
Figure Lengend Snippet: Loss of adipose mTORC2 reduces sensory neurons in the proximity of adipocytes . (A) Low magnification projection of inguinal WAT (iWAT) four weeks after tamoxifen treatment co-immunostained with tyrosine hydroxylase (TH, yellow) and calcitonin gene-related peptide (CGRP, magenta) (N = 15; 18). Scale bar = 500 μm. (B) High magnification projection of iWAT four weeks after tamoxifen treatment co-immunostained with TH (yellow) and CGRP (magenta) (N = 21; 22). Scale bar = 100 μm. (C) Subsequent sections (P1–P6; 6.48/6.28 μm interval, respectively) of control and iAdRiKO iWAT immunostained with TH (yellow) and CGRP (magenta) illustrating single nerve fibers innervating the periphery. Region of interest: Nerve ending and potential synapses (arrows). Tissue autofluorescence = green. Scale bar = 100 μm.
Article Snippet: CGRP levels in lysates were determined using mouse calcitonin gene-related peptide (CGRP) ELISA kit (CSB-EQ027706MO, Cusabio) according to the manufacturer's instructions.
Techniques: Control
Journal: Molecular Metabolism
Article Title: Adipose mTORC2 is essential for sensory innervation in white adipose tissue and whole-body energy homeostasis
doi: 10.1016/j.molmet.2022.101580
Figure Lengend Snippet: Adipose mTORC2 maintains sensory neurons . (A–B) High magnification projection (A) and quantification of the total neurite length (B) of CGRP-positive neurons in iWAT parenchyma of control mice and iAdRiKO three days after tamoxifen treatment (N = 6). Scale bar = 100 μm. (C–D) High magnification projection (C) and quantification of the total neurite length (D) of CGRP-positive neurons in iWAT parenchyma of control mice and iAdRiKO two weeks after tamoxifen treatment (N = 6). Scale bar = 100 μm; Student's t-test, ∗∗∗p < 0.001.
Article Snippet: CGRP levels in lysates were determined using mouse calcitonin gene-related peptide (CGRP) ELISA kit (CSB-EQ027706MO, Cusabio) according to the manufacturer's instructions.
Techniques: Control
Journal: Molecular Metabolism
Article Title: Adipose mTORC2 is essential for sensory innervation in white adipose tissue and whole-body energy homeostasis
doi: 10.1016/j.molmet.2022.101580
Figure Lengend Snippet: GAP43 expression is downregulated in CGRP-positive neurons upon loss of adipose mTORC2 . (A) Immunoblot analysis of inguinal WAT (iWAT) tissue from control and iAdRiKO mice two weeks after tamoxifen treatment. (n = 6; 6). (B) Immunoblot analysis of iWAT tissue from control and iAdRiKO mice four weeks after tamoxifen treatment. (n = 6; 6). (C) Immunoblot analysis of surgically denervated iWAT depot (denervation) compared to iWAT depot from sham-operated mice (sham). Neurofilament heavy polypeptide (NFH). (n = 5; 5). (D) Representative image of a large nerve bundle in iWAT of control mice immunostained with growth-associated protein 43 (GAP43)-pS41 and calcitonin gene-related peptide (CGRP). (N = 11; 9). (E) Representative image of a large nerve bundle in iWAT of control mice immunostained with GAP43-pS41 and tyrosine hydroxylase (TH). (N = 19; 11).
Article Snippet: CGRP levels in lysates were determined using mouse calcitonin gene-related peptide (CGRP) ELISA kit (CSB-EQ027706MO, Cusabio) according to the manufacturer's instructions.
Techniques: Expressing, Western Blot, Control
Journal: Journal of Clinical and Translational Hepatology
Article Title: RAMP1 Protects Hepatocytes against Ischemia-reperfusion Injury by Inhibiting the ERK/YAP Pathway
doi: 10.14218/JCTH.2023.00339
Figure Lengend Snippet: (A) Protein levels of RAMP1 in Control and H/R groups with and without CGRP agonist. (B) Relative cell activity was determined by CCK-8 assay of RAMP1 in the control and H/R groups using a CGRP agonist. (C) CCK-8 assay was performed using CGRP agonists, YAP inhibitors, and agonists in H/R. (D) CCK-8 assay was performed using a CGRP agonist combined with an ERK inhibitor or agonist in H/R. (E) CCK-8 experiments were performed using ERK inhibitors in combination with YAP inhibitors or agonists in H/R. (F) CCK-8 assays were performed using ERK agonists combined with a YAP inhibitor or agonist in H/R. (G) Statistical analysis of flow cytometry to detect the proportion of apoptosis in H/R with a CGRP agonist, YAP inhibitor and agonist, and ERK inhibitor and agonist. (H) Statistical data in H/R, ERK inhibitor combined with YAP inhibitor or agonist were used to detect the proportion of apoptosis by flow cytometry. (I) Statistical data in H/R using ERK agonists combined with YAP inhibitors or agonists to detect the proportion of apoptosis. CGRP agonist: Caltonin gene-related peptide (CGRP) II, rat TFA, ERK agonist (C16-PAF); ERK inhibitor (SCH772984); YAP agonist (PY-60); and YAP inhibitor (Verteporfin). The cells were treated with apoptosis pathway inhibitors and subjected to 6 h of reperfusion after hypoxia. All data are presented as the mean ± SD. * p <0.05, ** p <0.01, *** p <0.001 using Student’s two-tailed t-test. CCK-8, cell counting Kit-8; CGRP agonist, caltonin gene-related peptide agonist; HIRI, hepatic ischemia-reperfusion injury.
Article Snippet: YAP phosphorylation inhibitor: Truli (E1061, Selleck, 0.2 nm) and PY-60 (HY-141644, MCE; 1.6 µm; 10 mg/kg for mice in vivo , i.p. ); ERK phosphorylation inhibitor: Temuterkib (HY-101494, MCE; 5 nm) and SCH772984 (HY-50846, MCE; 300 nm, 5 mg/kg for mice in vivo , i.p. ); ERK agonist: C16-PAF(HY-108635, MCE, 1 µm , ); STAT3 inhibitor : Sttatic (HY-13818, MCE; IC50:10 µm); p-AKT inhibitor: MK2206 (HY-10358, MCE; 65 nm); Caspase-3 inhibitor: Z-VAD (HY-16658B, MCE; 10 µm ; 10 mg/kg for mice in vivo , i.p. ); CGRP agonist: Calcitonin Gene Related Peptide (CGRP) II,
Techniques: Control, Activity Assay, CCK-8 Assay, Flow Cytometry, Two Tailed Test, Cell Counting