Fig. 4
From: MSC-sEVs exacerbate senescence by transferring bisecting GlcNAcylated GPNMB
The bisecting GlcNAc modification on GPNMB. (A) Expression of GPNMB with bisecting GlcNAc in young and D-Gal m-MSCs by IP assay. (B) Intact glycoproteomic analysis by combination of PHA-E enrichment and LC-MS/MS analysis (schematic). (C) Volcano of differentially expressed glycopeptides in young and D-Gal induced m-MSCs. (D) Representative MS/MS spectrum of peptide NDRN#LSDEIFLR of GPNMB with bisecting GlcNAc in m-MSCs. (E, F) Wild-type and mutant GPNMB at Asn 249 were overexpressed in MSC-T, and bisecting GlcNAc on GPNMB in MSC-T/OEG and MSC-T/OEG-MU cells was assayed by IP and western blotting. (G, H) MSC-T/OEM3 and MSC-T/OEG-MU cells were treated with Chlo or MG132 for indicated times, and GPNMB expression was evaluated by western blotting. (I) qRT-PCR results of expression of senescence markers in MSC-T and MSC-T/OEG-MU cells. (J) Western blotting analysis and quantification of senescence markers in MSC-T and MSC-T/OEG-MU cells. (K) Apoptosis of MSC-T, MSC-T/OEG and MSC-T/OEG-MU cells