Journal: Journal of the Endocrine Society

Article Title: Insulin receptor trafficking and interactions in muscle cells

doi: 10.1210/jendso/bvag020

Figure Lengend Snippet: Biochemical and live-cell imaging characterization of the effects of insulin on INSR. (A) Internalized to total INSR ratio was quantified using surface biotinylation in undifferentiated C2C12 myoblasts. Cells were incubated in PBS (no insulin) or serum-free DMEM containing 0, 0.2, 2, or 20 nM insulin for 15 minutes. The ratios are normalized to the 0 nM group of each gel ( P > .05 when not specified). (B) Western blot showing SNAP-tagged INSR expressed from lentiviral vector in comparison to wild-type INSR in C2C12 myoblasts. (C, D) Representative images of INSR-A-SNAP-labeled using an Alexa Fluor 488 cell nonpermeable dye in 0, 0.2, or 20 nM insulin conditions in live undifferentiated C2C12 myoblasts from (C) 0 to 15 minutes or (D) after 20 minutes using a spinning disk confocal microscope. Time-lapse images of INSR vesicle interactions starting from the selected subregions (white squares) of snapshots are shown in the insets ( n = 3 cells).

Article Snippet: Cell nonpermeable SNAP-Surface Alexa Fluor 488 substrate (Cat. #S9129S, New England Biolabs) was dissolved and diluted to 5 μM concentration in serum-free medium containing 0.5% bovine serum albumin according to the manufacturer's instruction.

Techniques: Live Cell Imaging, Incubation, Western Blot, Plasmid Preparation, Comparison, Labeling, Microscopy

Journal: iScience

Article Title: FDA-approved PDE4 inhibitors alleviate the dominant toxicity of ALS-FTD-associated CHCHD10 S59L in Drosophila and human cells

doi: 10.1016/j.isci.2026.114879

Figure Lengend Snippet: Forskolin and Roflumilast alleviate mitophagy and muscle damage in Drosophila HeLa YFP−Parkin cells were co-transfected with FLAG-tagged CHCHD10 S59L and mCherry-LC3 in the DMSO, Forskolin, and Roflumilast-containing medium. (A) Representative images of CHCHD10 S59L expressing mitochondria cultured in DMSO, Forskolin (20 μM), and Roflumilast (1 μM) containing media. Cells were immunostained with anti-FLAG (green, CHCHD10 S59L ) and anti-TOM20 (white, mitochondria) antibodies. Arrows indicate LC3 puncta in the CHCHD10 S59L expressing mitochondria. Scale bars, 20uM. (B) Graphical quantification of the percentage of HeLa YFP−parkin shows LC3-mcherry puncta forming in the CHCHD10 S59L expressing mitochondria. Data are shown as mean ± SD (one-way ANOVA and post hoc Dunnett’s test, two-sided, comparison with DMSO). (C) Representative micrographs of indirect flight muscles from 10-day-old adult flies (MHC-Gal4, UAS-S81L) fed with Forskolin (50 μM) and Roflumilast (40 μM), immunostained with streptavidin-Alexa Fluor 488 and phalloidin-Alexa Fluor 594. DMSO was used as a vehicle. Scale bars, 10uM. (D) Graphical representation of ATP levels in thoraxes from 10-day-old flies using four independent replicates for males and females, respectively. Data are shown as mean ± SD (one-way ANOVA and post hoc Dunnett’s test, two-sided, ∗∗∗ p < 0.001, compared to DMSO).

Article Snippet: Sections were incubated with streptavidin-Alexa Fluor 488 and phalloidin–Alexa Fluor 594 (Invitrogen) overnight at 4°C for mitochondrial and muscle fiber staining respectively and mounted with Prolong Diamond Antifade Reagent with DAPI.

Techniques: Transfection, Expressing, Cell Culture, Comparison, Muscles