Using proteomics, we investigated the temporal expression profiles of proteins in rat sciatic nerve in course of aging by 2D-DIGE and mass spectrometry. The differential proteomic expression was compared with that of the surrounding muscle tissue. The proteomic investigation was performed by DIGE and mass spectrometry on sciatic nerves and muscles of 4 rats 8 and 24 month-old, respectively. Ground frozen nerves and muscles were solubilized, centrifuged and quantitated. Protein labelling with cyanine dyes (Cy3 and Cy5) was performed according to the manufacturer's instructions (GE Healthcare). These protocols suggest an internal standard be run on all gels within an experiment. The internal standard is created by pooling an aliquot of all biological samples in the experiment in order to warrant that every protein from all samples will be represented. Each sample was labelled with Cy5 dye and internal standard with Cy3 dye (400 pmol dye/50 ug sample). After labelling, protein samples were mixed with an equal volume of 2X sample buffer (Urea 8M, DTT 130 mM, CHAPS 4% and IPG buffer pH 3-10 2%).Gels were scanned directly between low fluorescence plates using a Typhoon 9200 laser scanner (GE Healthcare) according to the manufacturer's recommendations (532 and 633 nm laser; emission filters of 520 and 670 nm).Determination of protein spot abundance was performed using DeCyder software (version 5.0, GE Healthcare).A standard 2D map of sciatic nerve was defined and compared to a standard map of rat skeletal muscle. Qualitative comparison shows a low degree of similarity in proteome expression between nerve and muscle tissues. Nevertheless, we can identify some common proteins. Proteins differentially expressed in sciatic nerve and in muscle tissue in course of aging were characterized by MALDI TOF and ESI MS/MS.Quantitative differential expression provides up and down regulated proteins in common between the two tissues. Differences in protein expression identified during aging in sciatic nerve were retained also in muscle samples indicating the muscle could be a possible target for neurodegenerative studies.
|Titolo:||Comparative proteomic profile of sciatic nerve and muscle tissue in old and young rats|
|Data di pubblicazione:||lug-2006|
|Enti collegati al convegno:||Italian Proteomic Association|
|Citazione:||Comparative proteomic profile of sciatic nerve and muscle tissue in old and young rats / D. Capitanio, M. Vasso, M. Moriggi, A. Viganò, S. De Palma, M. Ripamonti, V. Magnaghi, C. Gelfi, D. Capitanio. ((Intervento presentato al 1. convegno ItPA congress tenutosi a Pisa nel 2006.|
|Appare nelle tipologie:||14 - Intervento a convegno non pubblicato|