| 1. |
- Holmquist, Emelie
(författare)
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Novel complement regulatory mechanisms in disease
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The complement system is one of the most important defence mechanisms against bacteria and pathogens. It acts within the framework of both innate and adaptive immunity. In addition to direct elimination of pathogens, complement also supports waste removal (dying cells, immune complexes, and misfolded proteins) and guides effector mechanisms of the adaptive immune response. Key events in complement activation are the formation of the C3 and C5 convertase enzyme complexes, the release of chemoattractants, the opsonization with C3b, and cell lysis due to the assembly of the membrane-attack complex (MAC). The proteolytic cascade of the complement system must be tightly regulated, since both excessive and insufficient activation significantly contributes to the pathology of many diseases. To protect its own cells and tissues, our body expresses several soluble and membrane-bound complement regulators. Here, we study the function of three of these regulators. Sushi Domain-Containing Protein 4 (SUSD4) is a poorly studied human protein. We show that SUSD4 is a novel complement inhibitor that interferes with the formation of the C3 converts. Additionally, we found that SUSD4 is expressed by breast cancer cells, and that this expression is correlated with a better patient prognosis. Factor I is a well-known complement inhibitor involved in the degradation of the activation products C4b and C3b. We detected factor I expression in breast cancer, and determined that breast cancer cells are able to produce the proteolytically active form of this protein. Analysis of patient data revealed that factor I expression correlates with poor survival rates. Cartilage Oligomeric Matrix Protein (COMP) is a large protein involved in the organisation of collagen in the extracellular matrix. We have previously shown that it can both activate and inhibit the complement system. Here, we determined that COMP expression is unregulated in breast cancer tissue where it contributes to a more aggressive form of cancer. COMP expression correlates with poor prognosis and faster recurrence of the disease. Tumors expressing COMP, grown in vivo, were significantly larger and more invasive as compared to control tumors. Interestingly, COMP affected the metabolism and protein processing machinery of the cancer cells, helping them adapt better to a harsh environment. In summary, this thesis describes novel, disease-related, functions of three complement regulators.
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| 2. |
- Okroj, Marcin, et al.
(författare)
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Functional analyses of complement convertases using c3 and c5-depleted sera.
- 2012
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 7:10
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Tidskriftsartikel (refereegranskat)abstract
- C3 and C5 convertases are central stages of the complement cascade since they converge the different initiation pathways, augment complement activation by an amplification loop and lead to a common terminal pathway resulting in the formation of the membrane attack complex. Several complement inhibitors attenuate convertase formation and/or accelerate dissociation of convertase complexes. Functional assays used to study these processes are often performed using purified complement components, from which enzymatic complexes are reconstituted on the surface of erythrocytes or artificial matrices. This strategy enables identification of individual interactions between convertase components and putative regulators but carries an inherent risk of detecting non-physiological interactions that would not occur in a milieu of whole serum. Here we describe a novel, alternative method based on C3 or C5-depleted sera, which support activation of the complement cascade up to the desired stages of convertases. This approach allows fast and simple assessment of the influence of putative regulators on convertase formation and stability. As an example of practical utility of the assay, we performed studies on thioredoxin-1 in order to clarify the mechanism of its influence on complement convertases.
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| 3. |
- Okroj, Marcin, et al.
(författare)
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Heavy chains of inter alpha inhibitor (IαI) inhibit the human complement system at early stages of the cascade .
- 2012
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Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 287:24, s. 20100-20110
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Tidskriftsartikel (refereegranskat)abstract
- Inter alpha inhibitor (IαI) is an abundant serum protein consisting of three polypeptides: two heavy chains (HC1 and HC2) and bikunin, a broad-specificity Kunitz-type proteinase inhibitor. The complex is covalently held together by chondroitin sulphate but during inflammation IαI may interact with TNF-stimulated gene 6 protein (TSG-6), which supports transesterification of heavy chains to hyaluronan. Recently, IαI was shown to inhibit mouse complement in vivo and to protect from complement-mediated lung injury but the mechanism of such activity was not elucidated. Using human serum depleted from IαI, we found that IαI is not an essential human complement inhibitor as reported for mice and that such serum has unaltered hemolytic activity. However, purified human IαI inhibited classical, lectin and alternative complement pathways in vitro when added in excess to human serum. The inhibitory activity was dependent on heavy chains but not bikunin and detected at the level of initiating molecules (MBL, properdin) in the lectin/ alternative pathways or C4b in the classical pathway. Furthermore, IαI affected formation and assembly of C1 complex and prevented assembly of the classical pathway C3-convertase. Presence and putative interactions with TSG-6 did not affect the ability of IαI to inhibit complement thus implicating IαI as a potentially important complement inhibitor once enriched onto hyaluronan moieties in the course of local inflammatory processes. In support of this, we found a correlation between IαI/HC-containing proteins and hemolytic activity of synovial fluid from patients suffering from rheumatoid arthritis.
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| 4. |
- Okroj, Marcin, et al.
(författare)
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Local expression of complement factor I in breast cancer cells correlates with poor survival and recurrence.
- 2015
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Ingår i: Cancer Immunology and Immunotherapy. - : Springer Science and Business Media LLC. - 1432-0851 .- 0340-7004. ; 64:4, s. 467-478
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Tidskriftsartikel (refereegranskat)abstract
- Tumor cells often evade killing by the complement system by overexpressing membrane-bound complement inhibitors. However, production of soluble complement inhibitors in cells other than hepatocytes was rarely reported. We screened several breast cancer cell lines for expression of soluble complement inhibitor, complement factor I (FI). We also analyzed local production of FI in tissue microarrays with tumors from 130 breast cancer patients by in situ hybridization and immunohistochemistry. We found expression of FI in breast adenocarcinoma cell line MDA-MB-468 and confirmed its functional activity. Expression of FI at mRNA and protein levels was also confirmed in tumor cells and tumor stroma, both in fibroblasts and infiltrating immune cells. Multivariate Cox regression analyses revealed that high expression of FI protein in tumor cells was correlated with significantly shorter cancer-specific survival (HR 2.8; 95 % CI 1.0-7.5; p = 0.048) and recurrence-free survival (HR 3.4; 95 % CI 1.5-7.4; p = 0.002). High FI expression was positively correlated with tumor size (p < 0.001), and Nottingham histological grade (p = 0.015) and associated with estrogen and progesterone receptor status (p = 0.03 and p = 0.009, respectively). Our data show that FI is expressed in breast cancer and is associated with unfavorable clinical outcome.
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| 5. |
- Zöller, Bengt, et al.
(författare)
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Familial Segregation of Venous Thromboembolism in Sweden: A Nationwide Family Study of Heritability and Complex Segregation Analysis
- 2021
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Ingår i: Journal of the American Heart Association. - 2047-9980. ; 10:24
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundThis is the first nationwide segregation analysis that aimed to determine whether familial venous thromboembolism (VTE) is attributable to inheritance and/or shared environment, and the possible mode of inheritance.Methods and ResultsThe Swedish Multi‐Generation Register was linked to the Swedish patient register for the period 1964 to 2015. Three generational families of Swedish‐born individuals were identified. Heritability was examined using Falconer regression. Complex segregation analysis was conducted using the Statistical Analysis for Genetic Epidemiology software (version 6.4, 64‐bit Linux). Among the 4 301 174 relatives from 450 558 pedigrees, 177 865 (52% women) individuals were affected with VTE. VTE occurred in 2 or more affected relatives in 61 217 (13.6%) of the pedigrees. Heritability showed age and sex dependence with higher heritability for men and young individuals. In 18 933 pedigrees, VTE occurred only in the first generation and was not inherited. Segregation analysis was performed in the remaining 42 284 pedigrees with inherited VTE and included 939 192 individuals. Prevalence constraints were imposed in the models to allow for the selection of the pedigrees analyzed. The sporadic nongenetic model could be discarded. The major‐type‐only model, with a correlation structure compatible with some polygenic effects, was the preferred model. Among the Mendelian models, the mixed codominant (plus polygenic) model was preferred.ConclusionsThis nationwide segregation analysis of VTE supports a genetic cause of the familial aggregation of VTE. Heritability was higher for men and younger individuals, suggesting a Carter effect, in agreement with a multifactorial threshold inheritance.
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