Project description:All cell lines were purchased from ATCC (LGC Standards S.r.l., Milan, Italy), except T-47D and MDA-MB-468 that were obtained from the National Cancer Institute Developmental Therapeutics Program (Frederick, MD).
Project description:Fukutin-related protein (FKRP) is a glycosyltransferase involved in glycosylation of alpha-dystroglycan (a-DG). Mutations in FKRP are associated with muscular dystrophies (MD) ranging from limb-girdle LGMDR9 to Walker-Warburg syndrome (WWS), a severe type of congenital MD. Although hypoglycosylation of a-DG is the main hallmark of this group of diseases, a full understanding of the underlying pathophysiology is still missing. Here, we investigated molecular mechanisms impaired by FKRP mutations in pluripotent stem (PS) cell-derived myotubes. FKRP deficient myotubes show transcriptome alterations in genes involved in extracellular matrix receptor interactions, calcium signaling, PI3K-Akt pathway, and lysosomal function. Accordingly, using a panel of patient-specific LGMDR9 and WWS induced PS cell-derived myotubes, we found a significant reduction in the autophagy-lysosome pathway for both disease phenotypes. Additionally, we show that WWS myotubes display decreased ERK1/2 activity and increased apoptosis, which were restored in gene edited myotubes. Our results suggest the autophagy-lysosome pathway and apoptosis may contribute to the FKRP-associated MD pathogenesis.
Project description:All cell lines were purchased from ATCC (LGC Standards S.r.l., Milan, Italy), except T-47D and MDA-MB-468 that were obtained from the National Cancer Institute Developmental Therapeutics Program (Frederick, MD). Six untreated breast cancer cell lines grown according to the growth protocol reported below
Project description:Sample multiplexing using isobaric tagging is a powerful strategy for proteome-wide protein quantification. One major caveat of isobaric tagging is ratio compression that results from the isolation, fragmentation, and quantification of co-eluting, near-isobaric peptides, a phenomenon typically referred to as “ion interference.” A robust platform to ensure quality control, optimize parameters, and enable comparisons across samples is essential as new instrumentation and analytical methods evolve. Here, we introduce TKO-iQC, an integrated platform consisting of the Triple Knock-Out (TKO) yeast digest standard and an automated web-based database search and protein profile visualization application. We highlight two new TKO standards based on the TMTpro reagent (TKOpro9 and TKOpro16), as well as an updated TKO Viewing Tool, TVT2.0. TKO-iQC greatly facilitates the comparison of instrument performance with a straightforward and streamlined workflow.
Project description:The Z-disc is a protein-rich structure critically important for myofibril development and integrity. In order to monitor the quantiative changes in C2C12 myoblast during myogenesis, a quantitative dimethyl-labelling approach was performed with d0 myoblasts, d5 myotubes and electrical puls stimulated d5 myotubes.
Project description:objection: The immune inflammatory disorders rheumatoid arthritis (RA), psoriatic arthritis (PsA) and psoriasis (Ps) share common pathologic features and show responsiveness to anti-tumor necrosis factor (TNF) agents yet they are phenotypically distinct. The aim of this study was to examine if anti-TNF therapy is associated with divergent gene expression profiles in circulating cells and target tissues of patients with these diseases Method: Peripheral blood CD14+ and CD14- cells were isolated from 9 RA, 12 PsA and 10 Ps patients before and after infliximab (IFX) treatment. Between April 2007 and June 2009, 31 patients with active RA, PsA and Ps who were naïve to anti-TNF agents, were recruited from the Faculty Rheumatology Clinics at the University of Rochester Medical Center after informed, written consent was obtained in a protocol approved by the Research Subjects Review Board at the University of Rochester Medical Center. Of the 31 subjects, 9 had active RA and 12 had PsA despite treatment with Disease Modifying Anti-Rheumatic Drugs (DMARDs). Also, 10 patients with extensive Ps (>5% BSA) documented by a dermatologist, were enrolled and they were examined by a rheumatologist to exclude the presence of inflammatory arthritis. Nineteen healthy controls were also recruited.