grk5 inhibitor  (MedChemExpress)

Journal: International Journal of Obesity (2005)

Article Title: GRK5 is required for adipocyte differentiation through ERK activation

doi: 10.1038/s41366-025-01712-w

Figure Lengend Snippet: A Eight-week-old male C57BL/6J mice were fed chow or a high fat diet (Envigo #TD 88137, 42% from fat, 0.2% total cholesterol) for 16 weeks and their body weight was measured ( n = 5/diet group). B Mice were then fasted for 24 h and their epididymal (Epi) visceral (Vis) white adipose tissue (WAT) and brown adipose tissue (BAT) RNA was extracted and reverse-transcribed into cDNA for real-time PCR quantification of Grk5 normalized to 18 s (endogenous control). C Six-week-old male C57BL/6J mice were fed chow or a high fat diet (Research Diets Inc #D12492, 60% from fat) for 12 weeks and their body composition such as fat mass was quantified by EcoMRI ( n = 10/diet group). D Adipocyte fraction and stromal vascular (SV) cell fraction were isolated from the Epi Vis WAT of overnight fasted mice. Both fractions’ RNA was extracted and reverse-transcribed into cDNA for real-time PCR quantification of Grk5 normalized to 18 s (endogenous control). All results are mean ± SEM, presented as the fold change compared to chow-fed mouse group and analyzed using a two-tailed Student’s unpaired t test ( A – C ), or the fold change compared to chow SV fractions and analyzed using a one-way ANOVA with Sidak multiple comparisons ( D ).

Article Snippet: A pyridine-based bicyclic compound of small molecule GRK5 inhibitor, GRK5-IN-2, was purchased from MedChemExpress (HY-136561).

Techniques: Reverse Transcription, Real-time Polymerase Chain Reaction, Control, Isolation, Two Tailed Test

Journal: International Journal of Obesity (2005)

Article Title: GRK5 is required for adipocyte differentiation through ERK activation

doi: 10.1038/s41366-025-01712-w

Figure Lengend Snippet: A Cellular proteins of undifferentiated wildtype (WT) control and GRK5 knockout (KO) 3T3-L1 preadipocytes ( n = 4/genotype) were harvested and subjected to Western blot using anti-GRK5, anti-GRK2, and anti-α-tubulin antibodies. B After 2 days of growth, proliferation was assessed in undifferentiated WT and GRK5 KO 3T3-L1 preadipocytes ( n = 6/genotype). The percentage of EdU-positive cells (pink) was calculated by merging EdU (red) and Hoechst 3342 (blue) staining. C WT and GRK5 KO 3T3-L1 cells ( n = 3/genotype) were proliferated for 2 days (Day 0) and then differentiated into adipocytes for 9 days. Day 0, 3, 6, and 9 cells were lipid-extracted to measure triacylglycerol (TAG) mass by a colorimetric assay. Daily Cytation images at 10x magnification were taken during 9 days of adipocyte differentiation. All results are mean ± SEM and analyzed using a two-tailed Student’s unpaired t test ( B ) and a two-way ANOVA with Sidak multiple comparisons ( C ).

Article Snippet: A pyridine-based bicyclic compound of small molecule GRK5 inhibitor, GRK5-IN-2, was purchased from MedChemExpress (HY-136561).

Techniques: Control, Knock-Out, Western Blot, Staining, Colorimetric Assay, Two Tailed Test

Journal: International Journal of Obesity (2005)

Article Title: GRK5 is required for adipocyte differentiation through ERK activation

doi: 10.1038/s41366-025-01712-w

Figure Lengend Snippet: Cellular RNA was extracted from wildtype (WT) control and GRK5 knockout (KO) 3T3-L1 cells ( n = 3/genotype) and reverse-transcribed into cDNA for real-time PCR quantification of Acc1, Pparγ , Fasn , Cd36, Fabp4 , Dgat2 , and Lipin1 normalized to 18 s (endogenous control). Cellular proteins of undifferentiated (Day 0) and differentiated (Day 9) WT control and GRK5 KO 3T3-L1 cells ( n = 2/genotype) were harvested and subjected to Western blot using anti-GRK5, anti-PPARγ, anti-CD36, and anti-α-tubulin antibodies. All results are mean ± SEM and presented as the fold change compared to WT at Day 0 and analyzed using a two-way ANOVA with Sidak multiple comparisons.

Article Snippet: A pyridine-based bicyclic compound of small molecule GRK5 inhibitor, GRK5-IN-2, was purchased from MedChemExpress (HY-136561).

Techniques: Control, Knock-Out, Reverse Transcription, Real-time Polymerase Chain Reaction, Western Blot

Journal: International Journal of Obesity (2005)

Article Title: GRK5 is required for adipocyte differentiation through ERK activation

doi: 10.1038/s41366-025-01712-w

Figure Lengend Snippet: A Pathway analysis of RNA sequencing data using Limma. B Wildtype (WT) control and GRK5 knockout (KO) 3T3-L1 preadipocytes ( n = 2/genotype) were differentiated for 2 days and treated with 1 μg/ml of insulin for 5, 10, and 15 min. Then, the cellular proteins were harvested and subjected to Western blot using anti-GRK5, anti-phosphorylated (p)-ERK, anti-ERK, and anti-GAPDH antibodies.

Article Snippet: A pyridine-based bicyclic compound of small molecule GRK5 inhibitor, GRK5-IN-2, was purchased from MedChemExpress (HY-136561).

Techniques: RNA Sequencing, Control, Knock-Out, Western Blot

Journal: International Journal of Obesity (2005)

Article Title: GRK5 is required for adipocyte differentiation through ERK activation

doi: 10.1038/s41366-025-01712-w

Figure Lengend Snippet: A Alignment of the GRK5 (green) and GRK2 (cyan) crystal structures. There is an alpha helix (shown in red) present near the binding pocket in GRK5, not present in GRK2. B Relative docking positions of GRK5-IN-2 in GRK5 and GRK2. C The dose-response curves of GRK5-IN-2 and staurosporine were determined by a GRK5 kinase system and a luminescent ADP detection assay. D Wildtype 3T3-L1 preadipocytes were differentiated and concurrently treated without (DMSO vehicle) or with GRK5-IN-2 ( n = 3/dose) for 7 days. Cells were lipid-extracted to measure triacylglycerol (TAG) mass by an enzymatic colorimetric assay. E Day 3 differentiated wildtype 3T3-L1 cell cultures were pretreated without (DMSO vehicle) or with GRK5-IN-2 (40 μM) for 30 min and then treated with 1 μg/ml of insulin plus 0.5 μCi/ml of [1,2- 14 C]-acetic acid for 60 and 120 min ( n = 3/time point). Cells were lipid-extracted, and TAG, free cholesterol (FC), cholesteryl ester (CE), and phospholipid (PL) were separated using thin layer chromatography. [ 14 C]-TAG, [ 14 C]-FC, [ 14 C]-CE, and [ 14 C]-PL were quantified by liquid scintillation counting. All results are mean ± SEM and analyzed using a two-way ANOVA with Sidak multiple comparisons.

Article Snippet: A pyridine-based bicyclic compound of small molecule GRK5 inhibitor, GRK5-IN-2, was purchased from MedChemExpress (HY-136561).

Techniques: Binding Assay, Detection Assay, Enzymatic Colorimetric Assay, Thin Layer Chromatography