| 1. |
- Vaziri-Sani, Fariba, et al.
(författare)
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Phenotypic expression of factor H mutations in patients with atypical hemolytic uremic syndrome
- 2006
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Ingår i: Kidney International. - : Nature Publishing Group. - 0085-2538 .- 1523-1755. ; 69:6, s. 981-988
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Tidskriftsartikel (refereegranskat)abstract
- We investigated the phenotypic expression of factor H mutations in two patients with atypical hemolytic uremic syndrome (HUS). Factor H in serum was assayed by rocket immunoelectrophoresis, immunoblotting, and double immunodiffusion and in tissue by immunohistochemistry. Functional activity was analyzed by hemolysis of sheep erythrocytes and binding to endothelial cells. A homozygous mutation in complement control protein (CCP) domain 10 of factor H was identified in an adult man who first developed membranoproliferative glomerulonephritis and later HUS. C3 levels were very low. The patient had undetectable factor H levels in serum and a weak factor H 150 kDa band. Double immunodiffusion showed partial antigenic identity with factor H in normal serum owing to the presence of factor H-like protein 1. Strong specific labeling for factor H was detected in glomerular endothelium, mesangium and in glomerular and tubular epithelium as well as in bone marrow cells. A heterozygous mutation in CCP 20 of factor H was found in a girl with HUS. C3 levels were moderately decreased at onset. Factor H levels were normal and a normal 150 kDa band was present. Double immunodiffusion showed antigenic identity with normal factor H. Factor H labeling was minimal in the renal cortex. Factor H dysfunction was demonstrated by increased sheep erythrocyte hemolysis and decreased binding to endothelial cells. In summary, two different factor H mutations associated with HUS were examined: in one, factor H accumulated in cells, and in the other, membrane binding was reduced.
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| 2. |
- Bergenfelz, Anders, et al.
(författare)
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Complications to thyroid surgery : results as reported in a database from a multicenter audit comprising 3,660 patients
- 2008
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Ingår i: Langenbeck's archives of surgery (Print). - : Springer. - 1435-2443 .- 1435-2451. ; 393:5, s. 667-673
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND AND AIM: During recent years, more radical surgery for thyroid disease, i.e., total instead of subtotal resection, has been evident. Results following this strategy on national levels are scarce.MATERIALS AND METHODS: From 2004 to 2006, 26 Scandinavian Departments registered 3,660 thyroid operations in a database. Risk factors for complications were analyzed with multiple logistic regression.RESULTS: After thyroidectomy, re-bleeding occurred in 2.1% and was associated with older age (OR 1.04; p < 0.0001) and male gender (OR 1.90; p = 0.014). Postoperative infection occurred in 1.6% and associated with lymph node operation (OR 8.18; p < 0.0001). Postoperative unilateral paresis of the recurrent laryngeal nerve was diagnosed 3.9% and bilateral paresis in 0.2%. Unilateral paresis was associated with older age, intrathoracic goiter, thyreotoxicosis, and if routine laryngoscopy was practiced (OR 1.92; p = 0.0002). After 6 months, the incidence of nerve paresis was 0.97%. After bilateral thyroid surgery (n = 1,648), hypocalcaemia treated with vitamin D analogue occurred in 9.9% of the patients at the first follow-up and in 4.4% after 6 months.CONCLUSION: Complications to thyroid surgery are not uncommon. The high frequency of hypocalcaemia treated with vitamin D after 6 months is a cause of concern.
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| 3. |
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| 4. |
- Karpman, D, et al.
(författare)
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Platelet activation in hemolytic uremic syndrome
- 2006
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Ingår i: Seminars in Thrombosis and Hemostasis. - : Georg Thieme Verlag. - 0094-6176 .- 1098-9064. ; 32:2, s. 128-145
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Tidskriftsartikel (refereegranskat)abstract
- Platelet consumption in platelet-fibrin aggregates leading to thrombocytopenia and small vessel obstruction are major features of the hemolytic uremic syndrome (HUS). Although thrombocytopenia has been correlated to poor prognosis, the mechanisms by which thrombocytopenia develops in HUS have not been completely elucidated. However, plausible explanations have been platelet contact with thrombogenic surfaces and/or direct contact with an aggregating agent. This article summarizes several mechanisms of platelet activation, interactions with leukocytes, chemokine release, complement activation, and antimicrobial defense. Specific mechanisms are outlined by which platelets may be activated, leading to thrombocytopenia during HUS. In diarrhea-associated HUS Shiga toxin has been shown to injure the endothelium, thus exposing the subendothelium, releasing tissue factor, and rendering the vessel wall prothrombotic. Shiga toxin also binds to and activates platelets. The toxin may activate endothelial cells and platelets simultaneously. In atypical HUS the alternative complement pathway is activated because of mutations in complement regulatory proteins. Mutated factor H does not bind to endothelium and platelets efficiently, enabling complement activation on these cells. In thrombotic thrombocytopenic purpura, intravascular platelet clotting Occurs due to dysfunction of the von Willebrand factor (VWF)-cleaving protease ADAMTS13. Thrombi are formed by binding of platelets to ultralarge VWF multimers.
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| 5. |
- Aradottir, Sigridur Sunna, et al.
(författare)
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Complement dysregulation associated with a genetic variant in factor H-related protein 5 in atypical hemolytic uremic syndrome
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Ingår i: Pediatric Nephrology. - 1432-198X. ; 39:4, s. 1105-1111
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Atypical hemolytic uremic syndrome (aHUS) can be associated with mutations, deletions, or hybrid genes in factor H-related (FHR) proteins.METHODS: A child with aHUS was investigated. Genetics was assessed by Sanger and next generation sequencing. Serum FHR5 was evaluated by immunoblotting, ELISA, and by induction of rabbit red blood cell hemolysis in the presence/absence of recombinant human rFHR5. Mutagenesis was performed in HEK cells.RESULTS: A heterozygous genetic variant in factor H-related protein 5 (CFHR5), M514R, was found in the child, who also had a homozygous deletion of CFHR3/CFHR1, and antibodies to factor H, as well as low levels of C3. Patient serum exhibited low levels of FHR5. In the presence of rabbit red blood cells, patient serum induced hemolysis which decreased when rFHR5 was added at physiological concentrations. Similar results were obtained using serum from the father, bearing the CFHR5 variant without factor H antibodies. Patient FHR5 formed normal dimers. The CFHR5 M514R variant was expressed in HEK cells and minimal secretion was detected whereas the protein level was elevated in cell lysates.CONCLUSIONS: Decreased secretion of the product of the mutant allele could explain the low FHR5 levels in patient serum. Reduced hemolysis when rFHR5 was added to serum suggests a regulatory role regarding complement activation on red blood cells. As such, low levels of FHR5, as demonstrated in the patient, may contribute to complement activation. A higher resolution version of the Graphical abstract is available as Supplementary information.
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| 6. |
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| 7. |
- Aradottir, Sigridur Sunna, et al.
(författare)
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Factor D Inhibition Blocks Complement Activation Induced by Mutant Factor B Associated With Atypical Hemolytic Uremic Syndrome and Membranoproliferative Glomerulonephritis
- 2021
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Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- Complement factor B (FB) mutant variants are associated with excessive complement activation in kidney diseases such as atypical hemolytic uremic syndrome (aHUS), C3 glomerulopathy and membranoproliferative glomerulonephritis (MPGN). Patients with aHUS are currently treated with eculizumab while there is no specific treatment for other complement-mediated renal diseases. In this study the phenotype of three FB missense variants, detected in patients with aHUS (D371G and E601K) and MPGN (I242L), was investigated. Patient sera with the D371G and I242L mutations induced hemolysis of sheep erythrocytes. Mutagenesis was performed to study the effect of factor D (FD) inhibition on C3 convertase-induced FB cleavage, complement-mediated hemolysis, and the release of soluble C5b-9 from glomerular endothelial cells. The FD inhibitor danicopan abrogated C3 convertase-associated FB cleavage to the Bb fragment in patient serum, and of the FB constructs, D371G, E601K, I242L, the gain-of-function mutation D279G, and the wild-type construct, in FB-depleted serum. Furthermore, the FD-inhibitor blocked hemolysis induced by the D371G and D279G gain-of-function mutants. In FB-depleted serum the D371G and D279G mutants induced release of C5b-9 from glomerular endothelial cells that was reduced by the FD-inhibitor. These results suggest that FD inhibition can effectively block complement overactivation induced by FB gain-of-function mutations.
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| 8. |
- Arvidsson, Ida, et al.
(författare)
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Apyrase decreases phage induction and Shiga toxin release from E. coli O157:H7 and has a protective effect during infection
- 2022
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Ingår i: Gut microbes. - : Informa UK Limited. - 1949-0976 .- 1949-0984. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Shiga toxin (Stx)-producing enterohemorrhagic Escherichia coli (EHEC) cause gastrointestinal infection and, in severe cases, hemolytic uremic syndrome which may lead to death. There is, to-date, no therapy for this infection. Stx induces ATP release from host cells and ATP signaling mediates its cytotoxic effects. Apyrase cleaves and neutralizes ATP and its effect on Stx and EHEC infection was therefore investigated. Apyrase decreased bacterial RecA and dose-dependently decreased toxin release from E. coli O157:H7 in vitro, demonstrated by reduced phage DNA and protein levels. The effect was investigated in a mouse model of E. coli O157:H7 infection. BALB/c mice infected with Stx2-producing E. coli O157:H7 were treated with apyrase intraperitoneally, on days 0 and 2 post-infection, and monitored for 11 days. Apyrase-treated mice developed disease two days later than untreated mice. Untreated infected mice lost significantly more weight than those treated with apyrase. Apyrase-treated mice exhibited less colonic goblet cell depletion and apoptotic cells, as well as lower fecal ATP and Stx2, compared to untreated mice. Apyrase also decreased platelet aggregation induced by co-incubation of human platelet-rich-plasma with Stx2 and E. coli O157 lipopolysaccharide in the presence of collagen. Thus, apyrase had multiple protective effects, reducing RecA levels, stx2 and toxin release from EHEC, reducing fecal Stx2 and protecting mouse intestinal cells, as well as decreasing platelet activation, and could thereby delay the development of disease.
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| 9. |
- Arvidsson, Ida, et al.
(författare)
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Early terminal complement blockade and C6 deficiency are protective in enterohemorrhagic Escherichia coli-infected mice
- 2016
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Ingår i: Journal of Immunology. - : The American Association of Immunologists. - 0022-1767 .- 1550-6606. ; 197:4, s. 1276-1286
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Tidskriftsartikel (refereegranskat)abstract
- Complement activation occurs during enterohemorrhagic Escherichia coli (EHEC) infection and may exacerbate renal manifestations. In this study, we show glomerular C5b-9 deposits in the renal biopsy of a child with EHEC-associated hemolytic uremic syndrome. The role of the terminal complement complex, and its blockade as a therapeutic modality, was investigated in a mouse model of E. coli O157:H7 infection. BALB/c mice were treated with monoclonal anti-C5 i.p. on day 3 or 6 after intragastric inoculation and monitored for clinical signs of disease and weight loss for 14 d. All infected untreated mice (15 of 15) or those treated with an irrelevant Ab (8 of 8) developed severe illness. In contrast, only few infected mice treated with anti-C5 on day 3 developed symptoms (three of eight, p < 0.01 compared with mice treated with the irrelevant Ab on day 3) whereas most mice treated with anti-C5 on day 6 developed symptoms (six of eight). C6-deficient C57BL/6 mice were also inoculated with E. coli O157:H7 and only 1 of 14 developed disease, whereas 10 of 16 wild-type mice developed weight loss and severe disease (p < 0.01). Complement activation via the terminal pathway is thus involved in the development of disease in murine EHEC infection. Early blockade of the terminal complement pathway, before the development of symptoms, was largely protective, whereas late blockade was not. Likewise, lack of C6, and thereby deficient terminal complement complex, was protective in murine E. coli O157:H7 infection.
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| 10. |
- Arvidsson, Ida, et al.
(författare)
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Shiga toxin-induced complement-mediated hemolysis and release of complement-coated red blood cell-derived microvesicles in hemolytic uremic syndrome.
- 2015
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Ingår i: Journal of Immunology. - : The American Association of Immunologists. - 1550-6606 .- 0022-1767. ; 194:5, s. 2309-2318
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Tidskriftsartikel (refereegranskat)abstract
- Shiga toxin (Stx)-producing Escherichia coli (STEC) cause hemolytic uremic syndrome (HUS). This study investigated whether Stx2 induces hemolysis and whether complement is involved in the hemolytic process. RBCs and/or RBC-derived microvesicles from patients with STEC-HUS (n = 25) were investigated for the presence of C3 and C9 by flow cytometry. Patients exhibited increased C3 deposition on RBCs compared with controls (p < 0.001), as well as high levels of C3- and C9-bearing RBC-derived microvesicles during the acute phase, which decreased after recovery. Stx2 bound to P1 (k) and P2 (k) phenotype RBCs, expressing high levels of the P(k) Ag (globotriaosylceramide), the known Stx receptor. Stx2 induced the release of hemoglobin and lactate dehydrogenase in whole blood, indicating hemolysis. Stx2-induced hemolysis was not demonstrated in the absence of plasma and was inhibited by heat inactivation, as well as by the terminal complement pathway Ab eculizumab, the purinergic P2 receptor antagonist suramin, and EDTA. In the presence of whole blood or plasma/serum, Stx2 induced the release of RBC-derived microvesicles coated with C5b-9, a process that was inhibited by EDTA, in the absence of factor B, and by purinergic P2 receptor antagonists. Thus, complement-coated RBC-derived microvesicles are elevated in HUS patients and induced in vitro by incubation of RBCs with Stx2, which also induced hemolysis. The role of complement in Stx2-mediated hemolysis was demonstrated by its occurrence only in the presence of plasma and its abrogation by heat inactivation, EDTA, and eculizumab. Complement activation on RBCs could play a role in the hemolytic process occurring during STEC-HUS.
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| 11. |
- Békássy, Zivile D., et al.
(författare)
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Aliskiren inhibits renin-mediated complement activation
- 2018
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Ingår i: Kidney International. - : Elsevier BV. - 0085-2538. ; 94:4, s. 689-700
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Tidskriftsartikel (refereegranskat)abstract
- Certain kidney diseases are associated with complement activation although a renal triggering factor has not been identified. Here we demonstrated that renin, a kidney-specific enzyme, cleaves C3 into C3b and C3a, in a manner identical to the C3 convertase. Cleavage was specifically blocked by the renin inhibitor aliskiren. Renin-mediated C3 cleavage and its inhibition by aliskiren also occurred in serum. Generation of C3 cleavage products was demonstrated by immunoblotting, detecting the cleavage product C3b, by N-terminal sequencing of the cleavage product, and by ELISA for C3a release. Functional assays showed mast cell chemotaxis towards the cleavage product C3a and release of factor Ba when the cleavage product C3b was combined with factor B and factor D. The renin-mediated C3 cleavage product bound to factor B. In the presence of aliskiren this did not occur, and less C3 deposited on renin-producing cells. The effect of aliskiren was studied in three patients with dense deposit disease and this demonstrated decreased systemic and renal complement activation (increased C3, decreased C3a and C5a, decreased renal C3 and C5b-9 deposition and/or decreased glomerular basement membrane thickness) over a follow-up period of four to seven years. Thus, renin can trigger complement activation, an effect inhibited by aliskiren. Since renin concentrations are higher in renal tissue than systemically, this may explain the renal propensity of complement-mediated disease in the presence of complement mutations or auto-antibodies.
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| 12. |
- Békassy, Zivile, et al.
(författare)
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Eculizumab in an anephric patient with atypical haemolytic uraemic syndrome and advanced vascular lesions.
- 2013
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Ingår i: Nephrology Dialysis Transplantation. - : Oxford University Press (OUP). - 1460-2385 .- 0931-0509. ; 28:11, s. 2899-2907
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Tidskriftsartikel (refereegranskat)abstract
- Atypical haemolytic uraemic syndrome (aHUS) is associated with dysfunction of the alternative pathway of complement. Disease activity subsides as renal failure progresses but recurs upon renal transplantation, indicating that viable renal tissue contributes to disease activity. We present evidence of cerebrovascular occlusive disease indicating that vascular injury may occur in the absence of kidneys.
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| 13. |
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| 14. |
- Békassy, Zivile, et al.
(författare)
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Intestinal damage in enterohemorrhagic Escherichia coli infection.
- 2011
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Ingår i: Pediatric Nephrology. - : Springer Science and Business Media LLC. - 1432-198X .- 0931-041X. ; oct, s. 2059-2071
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Tidskriftsartikel (refereegranskat)abstract
- Enterohemorrhagic Escherichia coli (EHEC) infection leads to marked intestinal injury. Sigmoid colon obtained from two children during EHEC infection exhibited abundant TUNEL-positive cells. To define which bacterial virulence factors contribute to intestinal injury the presence of Shiga toxin-2 (Stx2), intimin and the type III secretion system were correlated with symptoms and intestinal damage. C3H/HeN mice were inoculated with Stx2-producing (86-24) and non-producing (87-23) E. coli O157:H7 strains and 86-24 mutants lacking eae, encoding intimin (strain UMD619) or escN regulating the expression of type III secretion effectors (strain CVD451). Severe symptoms developed in mice inoculated with 86-24 and 87-23. Few mice inoculated with the mutant strains developed severe symptoms. Strain 86-24 exhibited higher fecal bacterial counts, followed by 87-23, whereas strains UMD619 and CVD451 showed minimal fecal counts. More TUNEL-positive cells were found in proximal and distal colons of mice inoculated with strain 86-24 compared with strains 87-23 and CVD451 (p
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| 15. |
- Brackman, Damien, et al.
(författare)
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Thrombotic microangiopathy mimicking membranoproliferative glomerulonephritis
- 2011
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Ingår i: Nephrology Dialysis Transplantation. - : Oxford University Press (OUP). - 1460-2385 .- 0931-0509. ; 26:10, s. 3399-3403
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Tidskriftsartikel (refereegranskat)abstract
- A 4-year-old boy presented with proteinuria and developed progressive renal failure over 6 years. In the patient's family, five individuals were affected with atypical haemolytic uraemic syndrome (aHUS) but not the patient. Renal biopsies (n = 3) showed glomerular basement membrane thickening with double contours, endothelial swelling and deposits of C3 and C1q. Electron microscopy revealed mesangial and subendothelial electron-dense deposits. Complement mutations in membrane cofactor protein (Y155D) and C3 (R713W and G1094R) were detected in all affected family members. The patient also had transient autoantibodies to factor H. The findings suggest that aHUS and glomerulopathy resembling membranoproliferative glomerulonephritis may have a common molecular background.
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| 16. |
- Friberg Hed, Niklas, et al.
(författare)
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Arginase release in hemolytic uremic syndrome affects the vasculature
- 2023
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Hemolysis is a cardinal feature of hemolytic uremic syndrome (HUS) and during hemolysis excess arginase 1 is released from erythrocytes. Increased arginase activity leads to reduced arginine, as it is converted to urea and ornithine, and thereby reduced nitric oxide bioavailability, with secondary vascular injury. In this study we investigated arginase release in HUS patients and laboratory models. Two separate cohorts of patients (n=47 in total) with HUS associated with Shiga toxin-producing enterohemorrhagic E. coli (EHEC) and pediatric controls (n=35) were investigated. HUS patients had excessively high arginase 1 levels and activity (conversion to urea and ornithine) in plasma/serum during the acute phase, compared to remission and controls. Arginase 1 levels correlated with lactate dehydrogenase activity, indicating hemolysis, as well as the need for dialysis treatment. Patients also exhibited high levels of plasma alpha-1-microglobulin, a heme scavenger. Two mouse models were used, mice were challenged intragastrically with E. coli O157:H7 or injected intraperitoneally with Shiga toxin 2. Both models exhibited significantly elevated plasma arginase 1 levels and activity. Arginase 1 levels correlated with lactate dehydrogenase activity and alpha-1-microglobulin in the plasma of EHEC-infected mice. In vitro perfusion of Shiga toxin 2- and E. coli O157-lipopolysaccharide-stimulated human blood cells combined with ADAMTS13-deficient plasma over glomerular endothelial cells induced hemolysis and the release of bioactive arginase 1. The high levels of arginase released during HUS could thereby contribute to microvascular injury during HUS.
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| 17. |
- Friberg, Niklas, et al.
(författare)
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Red blood cell-derived arginase release in hemolytic uremic syndrome
- 2024
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Ingår i: Journal of Translational Medicine. - 1479-5876. ; 22:1
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundHemolysis is a cardinal feature of hemolytic uremic syndrome (HUS) and during hemolysis excess arginase 1 is released from red blood cells. Increased arginase activity leads to reduced L-arginine, as it is converted to urea and L-ornithine, and thereby reduced nitric oxide bioavailability, with secondary vascular injury. The objective of this study was to investigate arginase release in HUS patients and laboratory models and correlate arginase levels to hemolysis and kidney injury.MethodsTwo separate cohorts of patients (n = 47 in total) with HUS associated with Shiga toxin-producing enterohemorrhagic E. coli (EHEC) and pediatric controls (n = 35) were investigated. Two mouse models were used, in which mice were either challenged intragastrically with E. coli O157:H7 or injected intraperitoneally with Shiga toxin 2. An in vitro model of thrombotic microangiopathy was developed in which Shiga toxin 2- and E. coli O157 lipopolysaccharide-stimulated human blood cells combined with ADAMTS13-deficient plasma were perfused over glomerular endothelial cells. Two group statistical comparisons were performed using the Mann–Whitney test, multiple groups were compared using the Kruskal–Wallis test followed by Dunn’s procedure, the Wilcoxon signed rank test was used for paired data, or linear regression for continuous variables.ResultsHUS patients had excessively high plasma arginase 1 levels and activity (conversion of L-arginine to urea and L-ornithine) during the acute phase, compared to remission and controls. Arginase 1 levels correlated with lactate dehydrogenase activity, indicating hemolysis, as well as the need for dialysis treatment. Patients also exhibited high levels of plasma alpha-1-microglobulin, a heme scavenger. Both mouse models exhibited significantly elevated plasma arginase 1 levels and activity. Plasma arginase 1 levels correlated with lactate dehydrogenase activity, alpha-1-microglobulin and urea levels, the latter indicative of kidney dysfunction. In the in vitro model of thrombotic microangiopathy, bioactive arginase 1 was released and levels correlated to the degree of hemolysis.ConclusionsElevated red blood cell-derived arginase was demonstrated in HUS patients and in relevant in vivo and in vitro models. The excessively high arginase levels correlated to the degree of hemolysis and kidney dysfunction. Thus, arginase inhibition should be investigated in HUS.
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| 18. |
- Johansson, Karl E., et al.
(författare)
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Shiga toxin signals via ATP and its effect is blocked by purinergic receptor antagonism
- 2019
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 9:1
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Tidskriftsartikel (refereegranskat)abstract
- Shiga toxin (Stx) is the main virulence factor of enterohemorrhagic Escherichia coli (EHEC), that cause gastrointestinal infection leading to hemolytic uremic syndrome. The aim of this study was to investigate if Stx signals via ATP and if blockade of purinergic receptors could be protective. Stx induced ATP release from HeLa cells and in a mouse model. Toxin induced rapid calcium influx into HeLa cells, as well as platelets, and a P2X1 receptor antagonist, NF449, abolished this effect. Likewise, the P2X antagonist suramin blocked calcium influx in Hela cells. NF449 did not affect toxin intracellular retrograde transport, however, cells pre-treated with NF449 exhibited significantly higher viability after exposure to Stx for 24 hours, compared to untreated cells. NF449 protected HeLa cells from protein synthesis inhibition and from Stx-induced apoptosis, assayed by caspase 3/7 activity. The latter effect was confirmed by P2X1 receptor silencing. Stx induced the release of toxin-positive HeLa cell- and platelet-derived microvesicles, detected by flow cytometry, an effect significantly reduced by NF449 or suramin. Suramin decreased microvesicle levels in mice injected with Stx or inoculated with Stx-producing EHEC. Taken together, we describe a novel mechanism of Stx-mediated cellular injury associated with ATP signaling and inhibited by P2X receptor blockade.
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| 19. |
- Johansson, Karl, et al.
(författare)
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Shiga Toxin-Bearing Microvesicles Exert a Cytotoxic Effect on Recipient Cells Only When the Cells Express the Toxin Receptor
- 2020
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Ingår i: Frontiers in cellular and infection microbiology. - : Frontiers Media SA. - 2235-2988. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Shiga toxin is the main virulence factor of non-invasive enterohemorrhagic Escherichia coli strains capable of causing hemolytic uremic syndrome. Our group has previously shown that the toxin can reach the kidney within microvesicles where it is taken up by renal cells and the vesicles release their cargo intracellularly, leading to toxin-mediated inhibition of protein synthesis and cell death. The aim of this study was to examine if recipient cells must express the globotriaosylceramide (Gb3) toxin receptor for this to occur, or if Gb3-negative cells are also susceptible after uptake of Gb3-positive and toxin-positive microvesicles. To this end we generated Gb3-positive A4GALT–transfected CHO cells, and a vector control lacking Gb3 (CHO-control cells), and decreased Gb3 synthesis in native HeLa cells by exposing them to the glycosylceramide synthase inhibitor PPMP. We used these cells, and human intestinal DLD-1 cells lacking Gb3, and exposed them to Shiga toxin 2-bearing Gb3-positive microvesicles derived from human blood cells. Results showed that only recipient cells that possessed endogenous Gb3 (CHO-Gb3 transfected and native HeLa cells) exhibited cellular injury, reduced cell metabolism and protein synthesis, after uptake of toxin-positive microvesicles. In Gb3-positive cells the toxin introduced via vesicles followed the retrograde pathway and was inhibited by the retrograde transport blocker Retro-2.1. CHO-control cells, HeLa cells treated with PPMP and DLD-1 cells remained unaffected by toxin-positive microvesicles. We conclude that Shiga toxin-containing microvesicles can be taken up by Gb3-negative cells but the recipient cell must express endogenous Gb3 for the cell to be susceptible to the toxin.
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| 20. |
- Karpman, Diana, et al.
(författare)
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Complement Interactions with Blood Cells, Endothelial Cells and Microvesicles in Thrombotic and Inflammatory Conditions.
- 2015
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Ingår i: Advances in Experimental Medicine and Biology. - Cham : Springer International Publishing. - 0065-2598. - 9783319186023 ; 865, s. 19-42
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Konferensbidrag (refereegranskat)abstract
- The complement system is activated in the vasculature during thrombotic and inflammatory conditions. Activation may be associated with chronic inflammation on the endothelial surface leading to complement deposition. Complement mutations allow uninhibited complement activation to occur on platelets, neutrophils, monocytes, and aggregates thereof, as well as on red blood cells and endothelial cells. Furthermore, complement activation on the cells leads to the shedding of cell derived-microvesicles that may express complement and tissue factor thus promoting inflammation and thrombosis. Complement deposition on red blood cells triggers hemolysis and the release of red blood cell-derived microvesicles that are prothrombotic. Microvesicles are small membrane vesicles ranging from 0.1 to 1 μm, shed by cells during activation, injury and/or apoptosis that express components of the parent cell. Microvesicles are released during inflammatory and vascular conditions. The repertoire of inflammatory markers on endothelial cell-derived microvesicles shed during inflammation is large and includes complement. These circulating microvesicles may reflect the ongoing inflammatory process but may also contribute to its propagation. This overview will describe complement activation on blood and endothelial cells and the release of microvesicles from these cells during hemolytic uremic syndrome, thrombotic thrombocytopenic purpura and vasculitis, clinical conditions associated with enhanced thrombosis and inflammation.
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| 21. |
- Karpman, Diana, et al.
(författare)
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Platelet activation in hemolytic uremic syndrome.
- 2006
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Ingår i: Seminars in Thrombosis and Hemostasis. - : Georg Thieme Verlag KG. - 1098-9064 .- 0094-6176. ; 32:2, s. 128-145
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Forskningsöversikt (refereegranskat)
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| 22. |
- Lanke, Elsa, et al.
(författare)
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Characterization of a novel mutation in the von Willebrand factor propeptide in a distinct subtype of recessive von Willebrand disease.
- 2008
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Ingår i: Thrombosis and Haemostasis. - 0340-6245. ; 100:2, s. 211-216
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Tidskriftsartikel (refereegranskat)abstract
- von Willebrand factor (VWF) is a plasma protein that consists of a series of multimers of which the high-molecular-weight VWF multimers are the most potent in platelet adhesion and aggregation. The propeptide of the VWF (VWFpp) is known to be essential in the process of multimer assembly. Genetic studies were performed in a patient with a phenotype of von Willebrand disease (VWD) characterized by very low plasma factor VIII and VWF levels and a VWF consisting of only a dimeric band and total absence of all multimers in plasma. The patient was found to be homozygous for the novel C570S mutation, caused by a 1709G>C transition in exon 14 of the VWF gene coding for the propeptide. Three asymptomatic relatives were found to be heterozygous. In-vitro mutagenesis and expression in COS-7 cells confirmed the detrimental effect of the mutation on VWF multimerization. Our findings show that the C570S mutation in the VWFpp abolishes multimerization of VWF. The mutation probably disrupts the normal configuration of the VWFpp, which is essential for correct orientation of the protomers and ultimately multimerization. The mutant amino acid is located in a region that is highly conserved across several species which underlines its critical role. This variant constitutes a distinct subtype of recessive 2A VWD with the exclusive presence of the dimeric form of VWF in plasma.
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| 23. |
- Lanke, Elsa, et al.
(författare)
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N1421K mutation in the glycoprotein Ib binding domain impairs ristocetin- and botrocetin-mediated binding of von Willebrand factor to platelets.
- 2008
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Ingår i: European Journal of Haematology. - : Wiley. - 1600-0609 .- 0902-4441. ; 81, s. 384-390
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Tidskriftsartikel (refereegranskat)abstract
- von Willebrand disease (VWD) is a common inheritable bleeding disorder caused by deficiency of von Willebrand Factor (VWF), which is involved in platelet adhesion and aggregation. We report a family consisting of three patients with VWD characterized by an apparently normal multimeric pattern, moderately decreased plasma factor VIII (FVIII) and VWF levels, and disproportionately low plasma VWF:RCo levels. The patients were found to be heterozygous for the novel N1421K mutation, caused by a 4263C>G transversion in exon 28 of the VWF gene coding for the A1 domain. Botrocetin- and ristocetin-mediated binding of plasma VWF to GPIb were reduced in the patients. In vitro mutagenesis and expression in COS-7 cells confirmed the impairment of the mutant in botrocetin- and ristocetin-mediated VWF binding to GPIb. VWF collagen binding capacity was unaffected in plasma from the heterozygous individuals as well as in medium from transfected COS-7 cells. Our findings indicate that the N1421K substitution in the VWF affects the GPIb binding site or a recognition element by a conformational change of the A1 domain.
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| 24. |
- Lopatko Fagerström, Ingrid, et al.
(författare)
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Blockade of the kallikrein-kinin system reduces endothelial complement activation in vascular inflammation
- 2019
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Ingår i: EBioMedicine. - : Elsevier BV. - 2352-3964. ; 47, s. 319-328
-
Tidskriftsartikel (refereegranskat)abstract
- Background: The complement and kallikrein-kinin systems (KKS) are activated during vascular inflammation. The aim of this study was to investigate if blockade of the KKS can affect complement activation on the endothelium during inflammation. Methods: Complement deposition on endothelial microvesicles was assayed in vasculitis patient plasma samples and controls. Plasma was perfused over glomerular endothelial cells and complement deposition assayed by flow cytometry. The effect of the kinin system was assessed using kinin receptor antagonists and C1-inhibitor. The in vivo effect was assessed in kidney sections from mice with nephrotoxic serum-induced glomerulonephritis treated with a kinin receptor antagonist. Findings: Vasculitis patient plasma had significantly more C3- and C9-positive endothelial microvesicles than controls. Perfusion of patient acute-phase plasma samples over glomerular endothelial cells induced the release of significantly more complement-positive microvesicles, in comparison to remission or control plasma. Complement activation on endothelial microvesicles was reduced by kinin B1- and B2-receptor antagonists or by C1-inhibitor (the main inhibitor of the classical pathway and the KKS). Likewise, perfusion of glomerular endothelial cells with C1-inhibitor-depleted plasma induced the release of complement-positive microvesicles, which was significantly reduced by kinin-receptor antagonists or C1-inhibitor. Mice with nephrotoxic serum-induced glomerulonephritis exhibited significantly reduced glomerular C3 deposition when treated with a B1-receptor antagonist. Interpretation: Excessive complement deposition on the endothelium will promote endothelial injury and the release of endothelial microvesicles. This study demonstrates that blockade of the KKS can reduce complement activation and thereby the inflammatory response on the endothelium. Funding: Full details are provided in the Acknowledgements/Funding section.
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| 25. |
- Manea Hedström, Minola, et al.
(författare)
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ADAMTS13 phenotype in plasma from normal individuals and patients with thrombotic thrombocytopenic purpura.
- 2007
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Ingår i: European Journal of Pediatrics. - : Springer Science and Business Media LLC. - 1432-1076 .- 0340-6199. ; 166:3, s. 249-257
-
Tidskriftsartikel (refereegranskat)abstract
- The activity of ADAMTS 13, the von Willebrand factor cleaving protease, is deficient in patients with thrombotic thrombocytopenic purpura (TTP). In the present study, the phenotype of ADAMTS13 in TTP and in normal plasma was demonstrated by immunoblotting. Normal plasma (n=20) revealed a single band at 190 kD under reducing conditions using a polyclonal antibody, and a single band at 150 kD under non-reducing conditions using a monoclonal antibody. ADAMTS 13 was not detected in the plasma from patients with congenital TTP (n=5) by either antibody, whereas patients with acquired TTP (n=2) presented the normal phenotype. Following immunoadsorption of immunoglobulins, the ADAMTS 13 band was removed from the plasma of the patients with acquired TTP, but not from that of normal individuals. This indicates that ADAMTS13 is complexed with immunoglobulin in these patients. The lack of ADAMTS13 expression in the plasma from patients with hereditary TTP may indicate defective synthesis, impaired cellular secretion, or enhanced degradation in the circulation. This study differentiated between normal and TTP plasma, as well as between congenital and acquired TTP. This method may, therefore, be used as a complement in the diagnosis of TTP.
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| 26. |
- Manea Hedström, Minola, et al.
(författare)
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Podocytes express ADAMTS13 in normal renal cortex and in patients with thrombotic thrombocytopenic purpura
- 2007
-
Ingår i: British Journal of Haematology. - : Wiley. - 0007-1048 .- 1365-2141. ; 138:5, s. 651-662
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Tidskriftsartikel (refereegranskat)abstract
- Congenital thrombotic thrombocytopenic purpura (TTP) is associated with ADAMTS13 mutations. The major site of ADAMTS13 synthesis is the liver. Expression in other tissues, and in TTP, has not been shown. In this study, ADAMTS13 protein expression was investigated in normal kidney and in renal tissue from two TTP patients, with a compound heterozygous mutation (P353L and P457L) and a homozygous mutation (4143insA). Real-time polymerase chain reaction demonstrated ADAMTS13 mRNA in normal kidney. ADAMTS13 was detected in the glomeruli and tubuli of normal and TTP kidney using anti-ADAMTS13 antibodies. In the glomeruli, expression was localised to podocytes (as demonstrated by counterstaining with two podocyte markers) and endothelium. Similar distribution was detected in the TTP kidneys. Electron microscopy detected ADAMTS13 in podocytes, endothelium and glomerular basement membrane. Cultured human podocytes expressed ADAMTS13 mRNA and protein, and podocyte lysate exhibited von Willebrand factor-cleaving activity. Mutation expression studies of the P353L and P457L mutations showed partially impaired secretion and lower activity of the secreted mutants. Impaired secretion has previously been shown for the 4143insA mutation. Podocyte-derived ADAMTS13 may offer local protection in the high-shear microcirculation of the glomerulus. The mutations in the two TTP patients studied enabled protein expression in the podocytes but affected protease secretion.
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| 27. |
- Mossberg, Maria, et al.
(författare)
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Cl-Inhibitor Decreases the Release of Vasculitis-Like Chemotactic Endothelial Microvesicles
- 2017
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Ingår i: Journal of the American Society of Nephrology. - : AMER SOC NEPHROLOGY. - 1046-6673 .- 1533-3450. ; 28:8, s. 2472-2481
-
Tidskriftsartikel (refereegranskat)abstract
- The kinin system is activated during vasculitis and may contribute to chronic inflammation. C1-inhibitor is the main inhibitor of the kinin system. In this study, we investigated the presence of the kinin B1 receptor on endothelial microvesicles and its contribution to the inflammatory process. Compared with controls (n=15), patients with acute vasculitis (n=12) had markedly higher levels of circulating endothelial micro vesicles, identified by flow cytometry analysis, and significantly more microvesicles that were positive for the kinin B1 receptor (Pamp;lt;0.001). Compared with microvesicles from wild-type cells, B1 receptor-positive microvesicles derived from transfected human embryonic kidney cells induced a significant neutrophil chemotactic effect, and a B1 receptor antagonist blocked this effect. Likewise, patient plasma induced neutrophil chemotaxis, an effect decreased by reduction of microvesicle levels and by blocking the B1 receptor. We used a perfusion system to study the effect of patient plasma (n=6) and control plasma (n=6) on the release of microvesicles from glomerular endothelial cells. Patient samples induced the release of significantly more B1 receptor-positive endothelial microvesicles than control samples, an effect abrogated by reduction of the microvesicles in the perfused samples. Perfusion of C1-inhibitor depleted plasma over glomerular endothelial cells promoted excessive release of B1 receptor-positive endothelial microvesicles compared with normal plasma, an effect significantly decreased by addition of C1-inhibitor or B1 receptor-antagonist. Thus, B1 receptor-positive endothelial microvesicles may contribute to chronic inflammation by inducing neutrophil chemotaxis, and the reduction of these microvesicles by C1-inhibitor should be explored as a potential treatment for neutrophil-induced inflammation.
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| 28. |
- Nilsson, Sara, et al.
(författare)
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A mutation in factor I that is associated with atypical hemolytic uremic syndrome does not affect the function of factor I in complement regulation.
- 2007
-
Ingår i: Molecular Immunology. - : Elsevier BV. - 1872-9142 .- 0161-5890. ; 44:8, s. 1835-1844
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Tidskriftsartikel (refereegranskat)abstract
- Factor I (FI) is the major complement inhibitor that degrades Ob and C4b in the presence of cofactors such as factor H (FH) and membrane cofactor protein (MCP). Recently, mutations and polymorphisms in complement regulator molecules FH and MCP but also in FI have been associated with atypical hemolytic uremic syndrome (aHUS). HUS is a disorder characterized by hemolytic anemia, thrombocytopenia and acute renal failure. In this study, we report three unrelated patients with an identical heterozygous mutation, G261D, in the FI heavy chain who developed severe aHUS at different time points in their lives. Two of the patients also have polymorphisms in FH previously associated with risk of developing aHUS. Testing in particular one patient and control serum samples we did not observe major differences in complement hemolytic activity, FI plasma levels or the capability to degrade C4b or Ob. A recombinant protein was produced in order to analyze the functional consequences of the mutation. Mutant FI had a slightly different migration pattern during electrophoresis under reducing conditions. An alteration due to alternative splicing or glycosylation was ruled out, thus the altered migration may be due to proximity of the mutation to a cysteine residue. The recombinant mutant FI degraded Ob and C4b in a manner comparable to wild-type protein. In conclusion, despite the association between the heterozygous mutation in FI and aHUS we did not observe any abnormalities in the function of FI regarding complement regulation.
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| 29. |
- Nilsson, Sara C., et al.
(författare)
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A mutation in factor I that is strongly associated with atypical hemolytic uremic syndrome does not affect the function of factor I in complement regulation
- 2007
-
Ingår i: Molecular Immunology. - 0161-5890 .- 1872-9142. ; 44:1-3, s. 221-221
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Factor I (FI) is the major complement inhibitor that degrades C3b and C4b in the presence of cofactors C4b binding protein (C4BP), factor H (FH), membrane cofactor protein (MCP) or complement receptor 1 (CR1). Recently, mutations and polymorphisms in complement regulator molecules FH and MCP but also in FI have been associated with atypical hemolytic uremic syndrome (aHUS). HUS is a disorder characterized by hemolytic anemia, thrombocytopenia and acute renal failure. In this study we report three unrelated patients with an identical heterozygous mutation, G261D, in FI heavy chain who developed severe aHUS at different time points in their lives. Two patients also have polymorphisms in FH previously associated with risk of developing aHUS. Testing in particular one patient and control serum samples we did not observe major differences in complement hemolytic activity, FI plasma levels or the capability to degrade C4b or C3b. A recombinant protein was produced in order to analyze the functional consequences of the mutation. Mutant FI had a slightly different migration pattern during electrophoresis under reducing conditions. An alteration due to alternative splicing or glycosylation was ruled out, thus the altered migration may be due to proximity of the mutation to a cysteine residue. The recombinant mutant FI degraded C3b and C4b in a manner comparable to wild type protein. In conclusion, despite the strong association between the heterozygous mutation in FI and aHUS we did not observe any abnormalities in the function of FI regarding complement regulation.
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| 30. |
- Rydberg, Viktor, et al.
(författare)
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Genetic investigation of Nordic patients with complement-mediated kidney diseases
- 2023
-
Ingår i: Frontiers in Immunology. - 1664-3224. ; 14
-
Tidskriftsartikel (refereegranskat)abstract
- BackgroundComplement activation in atypical hemolytic uremic syndrome (aHUS), C3 glomerulonephropathy (C3G) and immune complex-mediated membranoproliferative glomerulonephritis (IC-MPGN) may be associated with rare genetic variants. Here we describe gene variants in the Swedish and Norwegian populations.MethodsPatients with these diagnoses (N=141) were referred for genetic screening. Sanger or next-generation sequencing were performed to identify genetic variants in 16 genes associated with these conditions. Nonsynonymous genetic variants are described when they have a minor allele frequency of ResultsIn patients with aHUS (n=94, one also had IC-MPGN) 68 different genetic variants or deletions were identified in 60 patients, of which 18 were novel. Thirty-two patients had more than one genetic variant. In patients with C3G (n=40) 29 genetic variants, deletions or duplications were identified in 15 patients, of which 9 were novel. Eight patients had more than one variant. In patients with IC-MPGN (n=7) five genetic variants were identified in five patients. Factor H variants were the most frequent in aHUS and C3 variants in C3G. Seventeen variants occurred in more than one condition.ConclusionGenetic screening of patients with aHUS, C3G and IC-MPGN is of paramount importance for diagnostics and treatment. In this study, we describe genetic assessment of Nordic patients in which 26 novel variants were found.
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| 31. |
- Sartz, Lisa, et al.
(författare)
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A novel C3 mutation causing increased formation of the C3 convertase in familial atypical hemolytic uremic syndrome.
- 2012
-
Ingår i: Journal of Immunology. - : The American Association of Immunologists. - 0022-1767 .- 1550-6606. ; 188:4, s. 2030-2037
-
Forskningsöversikt (refereegranskat)abstract
- Atypical hemolytic uremic syndrome has been associated with dysregulation of the alternative complement pathway. In this study, a novel heterozygous C3 mutation was identified in a factor B-binding region in exon 41, V1636A (4973 T > C). The mutation was found in three family members affected with late-onset atypical hemolytic uremic syndrome and symptoms of glomerulonephritis. All three patients exhibited increased complement activation detected by decreased C3 levels and glomerular C3 deposits. Platelets from two of the patients had C3 and C9 deposits on the cell surface. Patient sera exhibited more C3 cleavage and higher levels of C3a. The C3 mutation resulted in increased C3 binding to factor B and increased net formation of the C3 convertase, even after decay induced by decay-accelerating factor and factor H, as assayed by surface plasmon resonance. Patient sera incubated with washed human platelets induced more C3 and C9 deposition on the cell surface in comparison with normal sera. More C3a was released into serum over time when washed platelets were exposed to patient sera. Results regarding C3 and C9 deposition on washed platelets were confirmed using purified patient C3 in C3-depleted serum. The results indicated enhanced convertase formation leading to increased complement activation on cell surfaces. Previously described C3 mutations showed loss of function with regard to C3 binding to complement regulators. To our knowledge, this study presents the first known C3 mutation inducing increased formation of the C3 convertase, thus explaining enhanced activation of the alternative pathway of complement.
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| 32. |
- Schmitt, Roland, et al.
(författare)
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IgA nephropathy associated with a novel N-terminal mutation in factor H.
- 2011
-
Ingår i: European Journal of Pediatrics. - : Springer Science and Business Media LLC. - 1432-1076 .- 0340-6199. ; 170, s. 107-110
-
Tidskriftsartikel (refereegranskat)abstract
- Most patients with IgA nephropathy exhibit complement deposition in the glomerular mesangium. Certain cases of IgA nephropathy have been associated with reduced levels of complement factor H. A recent study could not demonstrate mutations at the C-terminal of factor H. We describe a novel heterozygous mutation in factor H, position A48S (nucleotide position 142 G > T, alanine > serine), detected in exon 2 of a 14-year-old girl with IgA nephropathy. The patient exhibited reduced levels of C3 and factor H, the latter suggesting that the mutation affected factor H secretion. The patient developed initial signs and symptoms of glomerulonephritis at the age of 9 years but presented again at the age of 14 years with weight gain, renal failure, nephrotic-range proteinuria and malignant hypertension. Blood tests suggested the development of microangiopathic hemolytic anemia (MAHA) but the renal biopsy was mostly indicative of chronic changes associated with IgA nephropathy as well as vascular changes associated with malignant hypertension. Immunofluorescence exhibited deposits of IgA, C3, and IgM. Screening of the factor H gene revealed, in addition to the mutation, three heterozygous hemolytic uremic syndrome -associated risk polymorphisms (-257 c/t, 2089 a/g, and 2881 g/t) which may have increased the patient's susceptibility to the occurrence of MAHA triggered by malignant hypertension. The combined clinical picture of IgA nephropathy and MAHA may have been partly related to the alterations in factor H.
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| 33. |
- Schmitt, Roland, et al.
(författare)
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The Combined Role of Galactose-Deficient IgA1 and Streptococcal IgA-Binding M Protein in Inducing IL-6 and C3 Secretion from Human Mesangial Cells: Implications for IgA Nephropathy.
- 2014
-
Ingår i: Journal of Immunology. - : The American Association of Immunologists. - 1550-6606 .- 0022-1767. ; 193:1, s. 317-326
-
Tidskriftsartikel (refereegranskat)abstract
- IgA nephropathy (IgAN) is characterized by mesangial cell proliferation and extracellular matrix expansion associated with immune deposits consisting of galactose-deficient polymeric IgA1 and C3. We have previously shown that IgA-binding regions of streptococcal M proteins colocalize with IgA in mesangial immune deposits in patients with IgAN. In the present study, the IgA-binding M4 protein from group A Streptococcus was found to bind to galactose-deficient polymeric IgA1 with higher affinity than to other forms of IgA1, as shown by surface plasmon resonance and solid-phase immunoassay. The M4 protein was demonstrated to bind to mesangial cells not via the IgA-binding region but rather via the C-terminal region, as demonstrated by flow cytometry. IgA1 enhanced binding of M4 to mesangial cells, but not vice versa. Costimulation of human mesangial cells with M4 and galactose-deficient polymeric IgA1 resulted in a significant increase in IL-6 secretion compared with each stimulant alone. Galactose-deficient polymeric IgA1 alone, but not M4, induced C3 secretion from the cells, and costimulation enhanced this effect. Additionally, costimulation enhanced mesangial cell proliferation compared with each stimulant alone. These results indicate that IgA-binding M4 protein binds preferentially to galactose-deficient polymeric IgA1 and that these proteins together induce excessive proinflammatory responses and proliferation of human mesangial cells. Thus, tissue deposition of streptococcal IgA-binding M proteins may contribute to the pathogenesis of IgAN.
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| 34. |
- Ståhl, Anne-lie, et al.
(författare)
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A Novel Mechanism of Bacterial Toxin Transfer within Host Blood Cell-Derived Microvesicles.
- 2015
-
Ingår i: PLoS Pathogens. - : Public Library of Science (PLoS). - 1553-7366 .- 1553-7374. ; 11:2
-
Tidskriftsartikel (refereegranskat)abstract
- Shiga toxin (Stx) is the main virulence factor of enterohemorrhagic Escherichia coli, which are non-invasive strains that can lead to hemolytic uremic syndrome (HUS), associated with renal failure and death. Although bacteremia does not occur, bacterial virulence factors gain access to the circulation and are thereafter presumed to cause target organ damage. Stx was previously shown to circulate bound to blood cells but the mechanism by which it would potentially transfer to target organ cells has not been elucidated. Here we show that blood cell-derived microvesicles, shed during HUS, contain Stx and are found within patient renal cortical cells. The finding was reproduced in mice infected with Stx-producing Escherichia coli exhibiting Stx-containing blood cell-derived microvesicles in the circulation that reached the kidney where they were transferred into glomerular and peritubular capillary endothelial cells and further through their basement membranes followed by podocytes and tubular epithelial cells, respectively. In vitro studies demonstrated that blood cell-derived microvesicles containing Stx undergo endocytosis in glomerular endothelial cells leading to cell death secondary to inhibited protein synthesis. This study demonstrates a novel virulence mechanism whereby bacterial toxin is transferred within host blood cell-derived microvesicles in which it may evade the host immune system.
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| 35. |
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| 36. |
- Ståhl, Anne-Lie, et al.
(författare)
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Factor H dysfunction in patients with atypical hemolytic uremic syndrome contributes to complement deposition on platelets and their activation.
- 2008
-
Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 111, s. 5307-5315
-
Tidskriftsartikel (refereegranskat)abstract
- Atypical hemolytic uremic syndrome (aHUS) may be associated with mutations at the C-terminal of factor H (FH). FH binds to platelets via the C-terminal as previously shown using a construct consisting of short consensus repeats (SCRs) 15-20. Four FH mutations, in SCR15 (C870R) and SCR20 (V1168E, E1198K, E1198Stop) in aHUS patients, were studied regarding their ability to allow complement activation on platelet surfaces. Purified FH-E1198Stop mutant exhibited reduced binding to normal washed platelets compared to normal FH, detected by flow cytometry. Washed platelets taken from the four aHUS patients during remission exhibited C3 and C9 deposition, as well as CD40-ligand (CD40L) expression indicating platelet activation. Combining patient serum/plasma with normal washed platelets led to C3 and C9 deposition, CD40L and CD62P expression, aggregate formation and generation of tissue factor-expressing microparticles. Complement deposition and platelet activation were reduced when normal FH was pre-incubated with platelets and were minimal when using normal serum. The purified FH-E1198Stop mutant added to FH-deficient plasma (complemented with C3) allowed considerable C3 deposition on washed platelets, in comparison to normal FH. In summary, mutated FH enables complement activation on the surface of platelets and their activation, which may contribute to the development of thrombocytopenia in aHUS.
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| 37. |
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| 38. |
- Tati, Ramesh, et al.
(författare)
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Complement Activation Associated with ADAMTS13 Deficiency in Human and Murine Thrombotic Microangiopathy.
- 2013
-
Ingår i: Journal of Immunology. - : The American Association of Immunologists. - 1550-6606 .- 0022-1767. ; 191:5, s. 2184-2193
-
Tidskriftsartikel (refereegranskat)abstract
- This study addressed the contribution of ADAMTS13 deficiency to complement activation in thrombotic thrombocytopenic purpura (TTP). Renal tissue and blood samples were available from 12 TTP patients. C3 and C5b-9 deposition were demonstrated in the renal cortex of two TTP patients, by immunofluorescence and immunohistochemistry, respectively. C3 was also demonstrated in the glomeruli of Shiga toxin-2-treated Adamts13(-/-) mice (n = 6 of 7), but less in mice that were not Shiga toxin-2 treated (n = 1 of 8, p < 0.05) or wild-type mice (n = 0 of 7). TTP patient plasma (n = 9) contained significantly higher levels of complement-coated endothelial microparticles than control plasma (n = 13), as detected by flow cytometry. Exposure of histamine-stimulated primary glomerular endothelial cells to platelet-rich plasma from patients, or patient platelet-poor plasma combined with normal platelets, in a perfusion system, under shear, induced C3 deposition on von Willebrand factor-platelet strings (on both von Willebrand factor and platelets) and on endothelial cells. Complement activation occurred via the alternative pathway. No C3 was detected when cells were exposed to TTP plasma that was preincubated with EDTA or heat-inactivated, or to control plasma. In the perfusion system, patient plasma induced more release of C3- and C9-coated endothelial microparticles compared with control plasma. The results indicate that the microvascular process induced by ADAMTS13 deficiency triggers complement activation on platelets and the endothelium, which may contribute to formation of thrombotic microangiopathy.
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| 39. |
- Tati, Ramesh, et al.
(författare)
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Neutrophil Protease Cleavage of Von Willebrand Factor in Glomeruli - An Anti-thrombotic Mechanism in the Kidney
- 2017
-
Ingår i: EBioMedicine. - : Elsevier BV. - 2352-3964. ; 16, s. 302-311
-
Forskningsöversikt (refereegranskat)abstract
- Adequate cleavage of von Willebrand factor (VWF) prevents formation of thrombi. ADAMTS13 is the main VWF-cleaving protease and its deficiency results in development of thrombotic microangiopathy. Besides ADAMTS13 other proteases may also possess VWF-cleaving activity, but their physiological importance in preventing thrombus formation is unknown. This study investigated if, and which, proteases could cleave VWF in the glomerulus. The content of the glomerular basement membrane (GBM) was studied as a reflection of processes occurring in the subendothelial glomerular space. VWF was incubated with human GBMs and VWF cleavage was assessed by multimer structure analysis, immunoblotting and mass spectrometry. VWF was cleaved into the smallest multimers by the GBM, which contained ADAMTS13 as well as neutrophil proteases, elastase, proteinase 3 (PR3), cathepsin-G and matrix-metalloproteinase 9. The most potent components of the GBM capable of VWF cleavage were in the serine protease or metalloprotease category, but not ADAMTS13. Neutralization of neutrophil serine proteases inhibited GBM-mediated VWF-cleaving activity, demonstrating a marked contribution of elastase and/or PR3. VWF-platelet strings formed on the surface of primary glomerular endothelial cells, in a perfusion system, were cleaved by both elastase and the GBM, a process blocked by elastase inhibitor. Ultramorphological studies of the human kidney demonstrated neutrophils releasing elastase into the GBM. Neutrophil proteases may contribute to VWF cleavage within the subendothelium, adjacent to the GBM, and thus regulate thrombus size. This anti-thrombotic mechanism would protect the normal kidney during inflammation and could also explain why most patients with ADAMTS13 deficiency do not develop severe kidney failure.
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| 40. |
- Tati, Ramesh, et al.
(författare)
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Phenotypic Expression of ADAMTS13 in Glomerular Endothelial Cells
- 2011
-
Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:6
-
Tidskriftsartikel (refereegranskat)abstract
- Background: ADAMTS13 is the physiological von Willebrand factor (VWF)-cleaving protease. The aim of this study was to examine ADAMTS13 expression in kidneys from ADAMTS13 wild-type (Adamts13(+/+)) and deficient (Adamts13(-/-)) mice and to investigate the expression pattern and bioactivity in human glomerular endothelial cells. Methodology/Principal Findings: Immunohistochemistry was performed on kidney sections from ADAMTS13 wild-type and ADAMTS13-deficient mice. Phenotypic differences were examined by ultramorphology. ADAMTS13 expression in human glomerular endothelial cells and dermal microvascular endothelial cells was investigated by real-time PCR, flow cytometry, immunofluorescence and immunoblotting. VWF cleavage was demonstrated by multimer structure analysis and immunoblotting. ADAMTS13 was demonstrated in glomerular endothelial cells in Adamts13(+/+) mice but no staining was visible in tissue from Adamts13(-/-) mice. Thickening of glomerular capillaries with platelet deposition on the vessel wall was detected in Adamts13(-/-) mice. ADAMTS13 mRNA and protein were detected in both human endothelial cells and the protease was secreted. ADAMTS13 activity was demonstrated in glomerular endothelial cells as cleavage of VWF. Conclusions/Significance: Glomerular endothelial cells express and secrete ADAMTS13. The proteolytic activity could have a protective effect preventing deposition of platelets along capillary lumina under the conditions of high shear stress present in glomerular capillaries.
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| 41. |
- Tontanahal, Ashmita, et al.
(författare)
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IgG Binds Escherichia coli Serine Protease EspP and Protects Mice From E. coli O157:H7 Infection
- 2022
-
Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 13
-
Tidskriftsartikel (refereegranskat)abstract
- Shiga toxin-producing Escherichia coli O157:H7 is a virulent strain causing severe gastrointestinal infection, hemolytic uremic syndrome and death. To date there are no specific therapies to reduce progression of disease. Here we investigated the effect of pooled immunoglobulins (IgG) on the course of disease in a mouse model of intragastric E. coli O157:H7 inoculation. Intraperitoneal administration of murine IgG on day 3, or both on day 3 and 6, post-inoculation improved survival and decreased intestinal and renal pathology. When given on both day 3 and 6 post-inoculation IgG treatment also improved kidney function in infected mice. Murine and human commercially available IgG preparations bound to proteins in culture filtrates from E. coli O157:H7. Bound proteins were extracted from membranes and peptide sequences were identified by mass spectrometry. The findings showed that murine and human IgG bound to E. coli extracellular serine protease P (EspP) in the culture filtrate, via the IgG Fc domain. These results were confirmed using purified recombinant EspP and comparing culture filtrates from the wild-type E. coli O157:H7 strain to a deletion mutant lacking espP. Culture filtrates from wild-type E. coli O157:H7 exhibited enzymatic activity, specifically associated with the presence of EspP and demonstrated as pepsin cleavage, which was reduced in the presence of murine and human IgG. EspP is a virulence factor previously shown to promote colonic cell injury and the uptake of Shiga toxin by intestinal cells. The results presented here suggest that IgG binds to EspP, blocks its enzymatic activity, and protects the host from E. coli O157:H7 infection, even when given post-inoculation.
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| 42. |
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