| 1. |
- Hamsten, Carl, et al.
(författare)
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Elevated levels of FN1 and CCL2 in bronchoalveolar lavage fluid from sarcoidosis patients
- 2016
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Ingår i: Respiratory Research. - : BioMed Central. - 1465-9921 .- 1465-993X. ; 17
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Tidskriftsartikel (refereegranskat)abstract
- Background: Sarcoidosis is a granulomatous systemic inflammatory disease in which more than 90 % of all patients develop pulmonary manifestations. Several gene associations have previously been described, but established and clinically useful biomarkers are still absent. This study aimed to find proteins in bronchoalveolar lavage (BAL) fluid that can be associated with the disease. Methods: We developed and performed profiling of 94 selected proteins in BAL fluid and serum samples obtained from newly diagnosed and non-treated patients with sarcoidosis. Using multiplexed immunoassays, a total of 317 BAL and 217 serum samples were analyzed, including asthmatic patients and healthy individuals as controls. Results: Our analyses revealed increased levels of eight proteins in sarcoidosis patients compared to controls. Out of these, fibronectin (FN1) and C-C motif chemokine 2 (CCL2) revealed the strongest associations. In addition, cadherin 5 (CDH5) was found to correlate positively with lymphocyte cell numbers in BAL fluid. Conclusions: Applying a high throughput proteomics screening technique, we found proteins of potential clinical relevance in the context of sarcoidosis.
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| 2. |
- Häggmark, Anna, et al.
(författare)
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Proteomic Profiling Reveals Autoimmune Targets in Sarcoidosis
- 2015
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Ingår i: American Journal of Respiratory and Critical Care Medicine. - 1073-449X .- 1535-4970. ; 191:5, s. 574-583
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Tidskriftsartikel (refereegranskat)abstract
- Rationale: There is a need to further characterize the antibody repertoire in relation to sarcoidosis and potentially related autoantigens. Objectives: We investigated bronchoalveolar lavage (BAL) and serum samples from patients with sarcoidosis and healthy and diseased control subjects to discover sarcoidosis-associated autoantigens. Methods: Antigen microarrays built on 3,072 protein fragments were used to screen for IgG reactivity in 73 BAL samples from subjects with sarcoidosis, subjects with asthma, and healthy subjects. A set of 131 targets were selected for subsequent verification on suspension bead arrays using 272 additional BAL samples and 141 paired sera. Reactivity to four antigens was furthermore analyzed in 22 unprocessed BAL samples from patients with fibrosis and 269 plasma samples from patients diagnosed with myositis. Measurements and Main Results: Reactivity toward zinc finger protein 688 and mitochondrial ribosomal protein L43 were discovered with higher frequencies in patients with sarcoidosis, for mitochondrial ribosomal protein L43 especially in patients with non-Lofgren syndrome. Increased reactivity toward nuclear receptor coactivator 2 was also observed in patients with non-Lofgren syndrome as compared with patients with Lofgren syndrome. The antigen representing adenosine diphosphate-ribosylation factor GTPase activating protein 1 revealed high reactivity frequency in all sample groups but with significantly higher level of IgG reactivities in patients with sarcoidosis. Conclusions: Autoantigen reactivity was present in most BAL and serum samples analyzed, and the results revealed high interindividual heterogeneity, with most of the reactivities observed in single individuals only. Four proteins are here proposed as sarcoidosis-associated autoimmune targets and of interest for further validation in independent cohorts.
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| 3. |
- Wiklundh, Emil
(författare)
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Immunopeptidomics and autoantigens of interstitial lung diseases
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- We investigate Sarcoidosis and Idiopathic Pulmonary Fibrosis, IPF, to find markers for early fibrosis development. The aetiologies for both diseases are unknown and there is no specific treatment, moreover, there is a lack of diagnostic biomarkers for both diagnose and disease activity. Overall, the diseases have a considerable effect on patients’ physical health and quality of life. To identify patients at risk of rapid development of fibrosis, it is vital to improve patient care. We hypothesize that identification of specific antigens can help in the exposition of the pathogenesis of sarcoidosis as well as of IPF, and in the long perspective this could lead to identification of therapeutic targets. In project I we investigated the presence of proteins in BAL and serum from sarcoidosis patients and controls to discover disease associated proteins. In total eight proteins had increased levels in sarcoidosis patients, two were the most prominent (Fibronectin 1, FN1 and C-C motif chemokine 2, CCL2) that displayed the greatest differences between cohorts. Furthermore, the protein cadherin 5 (CDH5) had a positive association to lymphocyte count in BAL, interesting since T-lymphocytes are the major cell type in sarcoidosis. Potentially this could provide a way of monitoring lymphocyte presence in the lung through blood sample. In project II the large source of antigens from the Human Protein Atlas Project, a large library of protein fragments and antibodies that represent virtually all proteins in the body, was used to screen the immunoglobulin G specificity towards 3072 selected antigens in serum and BAL samples from patients with either of the sarcoidosis subcategories (Löfgren's syndrome (LS), non-Löfgren's sarcoidosis (nLS)), and asthma as well as healthy controls. A selected set of antigens went on to be analyzed in mSBA analysis. Sarcoidosis patients demonstrated an elevated reactivity frequencies toward Zinc finger protein 688 (ZNF688) and mitochondrial ribosomal protein L43 (MRPL43), particularly MRLP43 displayed a higher frequency in patients with non-Löfgren syndrome. Even though the protein fragment representing adenosine diphosphate-ribosylation factor GTPase activating protein 1 (ARFGAP1) showed high reactivity frequency in all sample groups, it was still significantly elevated in patients with sarcoidosis compared to the other cohorts. In project III we used a mass spectrometry-based method to analyze and characterize the Fc regions of human IgGs in paired serum and BAL fluid. Antibodies were isolated using a fast and reliable approach via MelonGel extraction. The isolated IgGs were digested by trypsin and separated by nLC (nano liquid chromatography) as in conventional proteomics, but peptide fragmentation was performed by both high resolution HCD MS/MS (Higher-energy C-trap dissociation) and high-resolution ETD MS/MS (Electron-transfer dissociation). We identified a candidate marker IgG4, which corresponded well to inflammatory activity in chronic lung diseases while also correlating between BAL and serum (R2=0.95), thus being readily available for sample collection. The IgG galactosylation marker could prove to be useful in clinical settings by monitoring chronic pulmonary inflammation status. Based on the results of project II we concluded that the Fc galactosylation status of IgG4 could potentially be used as a serum marker for severity in pulmonary inflammation. In project IV project, we used the mSBA approach for both sarcoidosis and IPF samples in order to evaluate similarities and differences between fibrosis associated diseases. Autoantigens were found in a majority of patient samples, including healthy controls, although that cohort displayed a lower frequency of autoreactivity and also comparatively lower titers. Altogether, autoreactivity was higher in IPF and in nLS patients compared to all other groups; both groups included a higher fibrosis rate than the other diseases potentially linking a general presence of autoreactive antibodies to fibrosis development. Reactivity toward collagen 5A1 (COL5A1) was discovered with a statistical significant higher frequency in patients with IPF compared to all other groups apart from the nLS group. The protein epitope of COL5A1 is proposed as an autoimmune target and/or marker of fibrosis, and is of interest for further investigation. In this thesis we have profiled the repertoire of proteins and autoantibodies in serum and BAL from patients with sarcoidosis, in addition we also investigated and compared the presence of autoreactive antibodies in patients with sarcoidosis and IPF, as well as control subjects and various comparable diseases. We have also proposed the ratio between agalactosylated and galactosylated forms of the Fc region in IgG4 as a marker for severe chronic lung disease. These discoveries open up for more studies to characterize, and test functionality, of these autoantibodies and proteins in the setting of sarcoidosis, IPF, or inflammatory and/or pulmonary diseases.
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