Journal: International Journal of Molecular Sciences

Article Title: miR-193b-3p Promotes Proliferation of Goat Skeletal Muscle Satellite Cells through Activating IGF2BP1

doi: 10.3390/ijms232415760

Figure Lengend Snippet: miR-193b-3p induces IGF2BP through a seed-sites match. ( A ) miR-193b-3p promotes IGF2BP1 transcripts. ( B ) miR-193b-3p elevates IGF2BP1 protein. Western blotting (WB) assay was performed to detect IGF2BP1 protein. GAPDH works as a loading control. ( C ) The expression profile of IGF2BP1 in goat tissues and cells. ( D ) miR-193b-3p targets 3′ UTR of IGF2BP1. The potential seed match between miR-193b-3p within 3′ UTR regions of IGF2BP1 is predicted. The blue line marks the complementary nucleotide. Mfe indicates free energy between IGF2BP1-wt and miR-193b-3p. ( E ) The addition of miR-193b-3p activates luciferase activity of wild-type (R-Luc-IGF2BP1-wt) in MuSCs. ( F ) miR-193b-3p activates luciferase activity of R-Luc-IGF2BP1-wt in HeLa and mouse C2C12 myoblast. Data are shown as mean ± MSE. An unpaired two-tailed t -test is used to evaluate the means difference. *** p < 0.001, ** p < 0.01, * p < 0.05, and § p < 0.1.

Article Snippet: For the luciferase reporter assays, goat MuSCs (~1 × 10 4 cells per well), mouse C2C12 myoblasts (~1 × 10 5 cells per well, Thermo Fisher Scientific, Waltham, MA, USA), and HeLa cells (~2 × 10 5 cells per well, ATCC, Manassas, VA, USA) were cultured in GM on 24-well plates, and cotransfected with vectors (640 ng/well for each vector) using Lipofectamine 2000 reagent when cells reached 80–90% confluence.

Techniques: Western Blot, Control, Expressing, Luciferase, Activity Assay, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: miR-193b-3p Promotes Proliferation of Goat Skeletal Muscle Satellite Cells through Activating IGF2BP1

doi: 10.3390/ijms232415760

Figure Lengend Snippet: miR-193b-3p negatively regulates mouse IGF2BP1 and suppresses C2C12 proliferation. ( A , B ) miR-193b-3p decreases transcripts of IGF2BP1 ( A ) but barely affects the abundance of its neighboring genes in C2C12 ( B ). ( C ) miR-193b-3p deduces C2C12 proliferation. Left panel, Representative immunofluorescence images of EdU staining cells in miR-193b-3p-disturbed C2C12 (mi-Ctrl and mi-193b-3p, in-Ctrl and in-193b-3p, 50 nM). Scale bar = 100 μm. Right panel, Fold change of EdU + cells (ratio of EdU + myoblasts to all) are evaluated using randomly selected fields and normalized to control. Each treatment is repeated ten times. ( D ) Cell cycle is affected by miR-193b-3p mimic. Flow cytometric assay is performed for cells treated with mi-193b-3p or mi-Ctrl ( n = 3). Data are shown as mean ± MSE. ** p < 0.01, * p < 0.05, and § p < 0.1.

Article Snippet: For the luciferase reporter assays, goat MuSCs (~1 × 10 4 cells per well), mouse C2C12 myoblasts (~1 × 10 5 cells per well, Thermo Fisher Scientific, Waltham, MA, USA), and HeLa cells (~2 × 10 5 cells per well, ATCC, Manassas, VA, USA) were cultured in GM on 24-well plates, and cotransfected with vectors (640 ng/well for each vector) using Lipofectamine 2000 reagent when cells reached 80–90% confluence.

Techniques: Immunofluorescence, Staining, Control, Flow Cytometry