Artikel
LMD-assisted Comparison of Muscle Proteome in Patients with Filaminopathy and matched Controls
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Autoren
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Verena Theis
- Medical Proteome Center ZKF I, Functional Proteomics, Bochum, Germany
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Alexandra Maerkens
- Medical Proteome Center ZKF I, Functional Proteomics, Bochum, Germany
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Rudolf Andre Kley
- Bergmannsheil University Hospital, Neurology, Bochum, Germany
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Matthias Vorgerd
- Bergmannsheil University Hospital, Neurology, Bochum, Germany
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Katrin Marcus
- Medical Proteome Center ZKF I, Functional Proteomics, Bochum, Germany
| Veröffentlicht: | 11. September 2012 |
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Gliederung
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Background: Filaminopathy is a subtype of myofibrillar myopathies (MFM) caused by filamin C mutations. Filaminopathy is characterized by focal myofibrillar destruction and protein aggregation in muscle fibers. The aim of our study was to identify regulated proteins in muscle fibers of filaminopathy patients that have not (yet) shown protein aggregation to get insights in early pathogenesis of filaminopathy. To achieve this goal, we used two different proteomic approaches: a combination of mass spectrometry and spectral index calculation and 2D Difference Gel Electrophoresis (DIGE) using saturation labeling.
Method: Defined areas of muscle tissue without protein aggregates were collected from muscle biopsies of 5 filaminopathy patients and 5 healthy controls by using laser microdissection. The controls were matched to the patients by age, origin of muscle biopsy and gender. For spectral index calculation the samples were digested with trypsin and analyzed by nanoflow LC electrospray MS/MS. Proteins with a ratio >1.8 (patient samples compared to controls) were accepted as regulated proteins in muscle proteome. For 2D DIGE the muscle tissue was collected in saturation label buffer and then analyzed with 2D DIGE. The data were analyzed by DeCyder and potential differential proteins were identified by mass spectrometry.
Results: A total of 355 different proteins were identified with spectral index calculation. 11 of these proteins were up-regulated including obscurin, calsequestrin-2 and musculoskeletal embryonic nuclear protein 1. 16 proteins were down-regulated, e.g myozenin 1. Filamin C was identified in each sample but seems not to be regulated in early stages of filaminopathy. The 2D DIGE detected 16 differential protein spots.
Conclusion: Our approach combining spectral index calculation and 2D DIGE gives new insights in the muscle proteome of filaminopathy patients. The identification of regulated proteins may help to understand the early pathogenesis of this MFM subtype and to establish new biomarkers.
