Mass Spectrometric-Based Proteomic Analysis of Cutaneous Occlusive Vascular Diseases in Formalin-Fixed Tissues
Wonwoo Shon, Michael J Camilleri, Lori A Erickson, Ahmet Dogan, Thomas J Flotte. Mayo Clinic, Rochester, MN
Background: Cutaneous occlusive vascular disease results from various etiologies. These intravasculuar thrombi are indistinguishable by light microscopy alone. As the management relies on treatment of the underlying disease, accurate classification of this disorder is of paramount importance. In this study, we evaluated various types of cutaneous occlusive vascular disease by laser microdissection and mass spectrometry based proteomic analysis.
Design: All available materials from 16 skin biopsies with intravascular thrombosis (7 calciphylaxis, 5 cryoglobulinemia, 1 leukocytoclastic vasculitis, 1 disseminated intravascular coagulation, 1 stasis ulcer, and 1 antiphospholipid syndrome) were retrieved. In each case, intravascular thrombi were identified under fluorescent light and microdissected. These fragments were digested into tryptic peptide and analyzed by nano-flow liquid chromatography electrospray tandem mass spectrometry (LC-MS/MS). The results were assigned peptide and protein probability scores, and they were displayed by the use of Scaffold program. Clinical information was obtained from our medical record system.
Results: Microscopically, all skin biopsies contained intravascular thrombi within the dermis and/or subcutaneous tissues. Secondary changes such as surface ulceration, dermal necrosis, RBC extravasation, and/or calcification (both intra and extravascular) were present in many of the biopsies. Proteomic analysis by LC-MS/MS was able to accurately identify specific proteins in all cases tested. Fibronogens were found in 9/16 cases (56%). In 4 of 5 cryoglobulinemia cases, the thrombi contained immunoglobulins (2 IgM and κ, 1 IgG and λ, 1 IgM, IgG, and κ) and the results were able to correlate with the bone marrow immunophenotypic data in patients with hematopoietic disorder. Interestingly, IgA, IgG, κ and λ were also detected in the intravascular thrombi of a patient with antiphospholipid syndrome.
Conclusions: LC-MS/MS based proteomic analysis of cutaneous intravascular thrombosis selectively identified immunoglobulin depositions in patients with cryoglobulinemia (both monoclonal and mixed). Detection of immunoglobulin constituents of the thrombi enhances our ability to further type cryoglobulinemia accurately independent of clinical information. Future study of additional cutaneous intravascular thrombosis in patients with other systemic diseases, should further clarify potential diagnostic roles for this novel application for cutaneous occlusive vascular diseases in formalin-fixed tissues.
Wednesday, March 21, 2012 1:00 PM
Poster Session VI # 131, Wednesday Afternoon