Dataset Information

TMTpro: Design and initial evaluation of a novel Proline-based isobaric 16-plex Tandem Mass Tag reagent set

ABSTRACT: The design and synthesis of a novel proline-reporter-based isobaric Tandem Mass Tag 16 tag set (TMTpro) was carried out and the data uploaded here is a comparison of the performance of the new TMTpro tags with the current commercially available dimethylpiperidine-reporter-based TMT10/11 reagents. Data from 2 experiments are provided.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Rattus Norvegicus (rat) Homo Sapiens (human) Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: Michael Bremang

LAB HEAD: Ian Pike

PROVIDER: PXD014750 | Pride | 2019-11-25

REPOSITORIES: Pride

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TMTpro: Design, Synthesis, and Initial Evaluation of a Proline-Based Isobaric 16-Plex Tandem Mass Tag Reagent Set.

Thompson Andrew A Wölmer Nikolai N Koncarevic Sasa S Selzer Stefan S Böhm Gitte G Legner Harald H Schmid Peter P Kienle Stefan S Penning Petra P Höhle Claudia C Berfelde Antje A Martinez-Pinna Roxana R Farztdinov Vadim V Jung Stephan S Kuhn Karsten K Pike Ian I

Analytical chemistry 20191203 24

The design and synthesis of a proline-based reporter isobaric Tandem Mass Tag structure (TMTpro) is presented. An analysis is made of the performance of the new TMTpro tags in comparison with the current commercially available dimethylpiperidine-reporter-based TMT10/11 reagents. The new reporter structure provides a set of 16 tags for use with resolution of 6.3 mDa mass differences in high resolution mass spectrometers and a set of 9 reagents with 1 Da spacing between reporter ions for single da ...[more]

PMID: 31738517

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Project description:Chronic calcineurin inhibitor (CNI) nephrotoxicity is a major drawback in current immunosuppressive regimens. Pathology includes vascular and tubulointerstitial alterations. Although still commonly in use, cyclosporine A (CsA) is increasingly exchanged for tacrolimus (Tac) for a more favorable clinical outcome. Data on their differential pathogenetic effects in the kidney are, however, still scarce. We asked how CsA and Tac may affect adverse renal responses differentially. Methods- Wistar rats were divided into groups receiving cyclosporineÊ AÊ (n=4, target trough plasma level 3 mg/ml) and the respective vehicle (n=4, saline), or tacrolimus (Tac, FK506, n= 6, target trough plasma level 3.5 ng/ml) and the respective vehicle (n=6, DMSO/PEG500), via subcutaneously implanted osmotic minipumps for 28 days. Clinical parameters from plasma and urine samples were controlled. Animals were prepared for high-end morphological analysis, elective immunostaining, and high-throughput technology. Large scale electron microscopy (EM), confocal, stimulated emission-depletion (STED) and 3D-structured illumination (SIM) microscopy were used for pathology. Protein biochemistry, transcriptome, proteome, and phosphoproteome technologies were used to identify gene expressional differences. Results - In rats, CsA and Tac produced common as well as distinct alterations in glomeruli and tubulointerstitium. Both drugs caused cortical fibrotic foci with sclerotic glomeruli and damaged tubules. With CsA, glomerular damage was milder than with Tac. Proximal tubules showed widespread dysmorphic lysosomes with peripheral LAMP1 signal as well as autophagic and mitophagic vacuoles along with high apoptosis rate and diminished albumin uptake; megalin-dependent endocytosis was causally involved in the changes, and lysosomal exocytosis was sharply stimulated at the luminal cell pole. KIM1 signal was moderate. With Tac, these changes were far less pronounced, but the incidence of focal tubular necrosis along with intense KIM-1 signal was high. High throughput analysis showed differential changes between CsA and Tac with almost no overlap between the respective spectra. CsA caused upregulation of components of the unfolded protein response and apoptosis, whereas Tac resulted in RAS-associated stimulation and vascular deterioration. Conclusions- CNI nephrotoxicity presented with widely differing pathogenetic characteristics upon chronical CsA vs. Tac treatments. Beyond its known vascular effects, CsA markedly affected proximal tubular integrity, whereas Tac was primarily affecting vascular/glomerular components. Both medications produced comparable degrees of nephron loss and fibrosis. Results may serve to better adjust immunosuppressive treatment combinations in transplant patients.

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