MALDI Imaging-Massenspektrometrie bei hämophiler Arthropathie

Aim: Arthropathy as a result of repeated joint bleeding is a severe complication in patients with hemophilia. In the evaluation of synovial tissue specimens, histology alone is non-specific and there is a considerable morphological overlap with other joint diseases. There are several biomarkers found to be involved in the disease process such as iron, ferritin and fibrinogen molecules that are abundant in high concentration in blood [1, 2]. We applied MALDI IMS as a new technique to show the distribution of various proteins, at the peptide level, that have been suggested to play a key role in the pathogenesis of the disease. Methods: Synovial tissue of 6 patients with hemophilic arthropathy were sectioned and stained with iron staining (Berlin-blue). A serial section of each sample was subjected to in situ trypsin digestion at 37 °C for 1.5 h in a humidity chamber after deparaffination and antigen retrieval. Trypsin solution (0.1µg/µl) was deposited on the tissue sections by vibrational spraying (ImagePrep, Bruker Daltonik GmbH). Matrix solution (7 mg/ml in 50/50 acetonitrile /0.5% TFA) was deposited onto digested sections using the same devise. MALDI imaging data were acquired at a spatial resolution of 200 µm using a Bruker Autoflex Speed TOF/TOF system and visualized with FlexImaging software. Spectral analysis was performed with FlexAnalysis 3.3 and ClinProTools 3.3 software programs (Bruker Daltonik GmbH). Results: We detected several peaks predictive in mass to charge ratio with ferritin light (m/z 1608) and heavy chain (m/z 1345), alpha- (m/z 1071) and beta- (m/z 1274) hemoglobin subunits, truncated coagulation factor VIII peptide (m/z 1502, 1176), beta- and gamma fibrinogen peptides (m/z 980, 1032, 1117 and 1683), and annexin A2 (m/z 1111, 1268, 1460, 2164). Additionally, the distribution of these proteins in synovial tissue sections was demonstrated. Conclusion: MALDI IMS detected and mapped specific proteins in the synovial membrane of patients with hemophilic arthropathy that are probable drivers in the pathogenesis of the disease. References: 1. Roosendaal G, van den Berg HM, Lafeber FP, Bijlsma J. [Pathology of synovitis and hemophilic arthropathy]. Orthopade 1999; 28: 323-328. 2. Valentino LA, Hakobyan N, Enockson C. Blood-induced joint disease: the confluence of dysregulated oncogenes, inflammatory signals, and angiogenic cues. Semin Hematol 2008; 45: S50-57.