Molecular Studies and an ex vivo Complement Assay on Endothelium Highlight the Genetic Complexity of Atypical Hemolytic Uremic Syndrome: The Case of a Pedigree With a Null CD46 Variant

Piras, Rossella; Iatropoulos, Paraskevas; Bresin, Elena; Todeschini, Marta; Gastoldi, Sara; Valoti, Elisabetta; Alberti, Marta; Mele, Caterina; Galbusera, Miriam; Cuccarolo, Paola; Benigni, Ariela; Remuzzi, Giuseppe; Noris, Marina

doi:10.3389/fmed.2020.579418

Cited by 9 publications

(11 citation statements)

References 47 publications

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“…Rare variants in complement genes or FHAA confirmed the predisposition in about half the patients with massive ex vivo C5b9 formation on the resting endothelium, resembling primary aHUS. 3 Our observation that patients with neither genetic variants nor FHAA may present with massive ex vivo C5b9 formation points to a circulating factor that affects complement regulation, 18,35 either related to common variants in complement genes (i.e., minor allele frequency $0.1%) with in vitro studies showing functional consequences or a yet unidentified factor. TMA recurrence was common in patients with pathogenic variants, corroborating previous studies.…”

Section: Discussionmentioning

confidence: 99%

Functional and Genetic Landscape of Complement Dysregulation Along the Spectrum of Thrombotic Microangiopathy and its Potential Implications on Clinical Outcomes

Timmermans

Damoiseaux

Wérion

et al. 2021

Kidney International Reports

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Introduction The syndromes of thrombotic microangiopathy (TMA) are diverse and represent severe endothelial damage caused by various mechanisms. The complement system plays a major role in a subset of patients with TMA, and its recognition is of clinical importance because it guides choice and duration of treatment. Methods We studied a well-defined cohort of patients with TMA and hypothesized that assessment of serum-induced ex vivo C5b9 formation on the endothelium and screening for rare variants in complement genes can better categorize TMA. Results Massive ex vivo C5b9 formation was found in all patients with primary atypical hemolytic uremic syndrome ( n / N = 11/11) and in 59% of patients with TMA and coexisting conditions ( n / N = 30/51). Massive ex vivo C5b9 formation was associated with rare genetic variants (45% [ n / N = 20/44] vs. 0% [ n / N = 0/21] patients with normal ex vivo C5b9 formation; P < 0.001). Massive ex vivo C5b9 formation was associated with favorable renal response to therapeutic complement inhibition in patients with TMA and coexisting conditions (86% [ n / N = 12/14] vs. 31% [ n / N = 5/16] of untreated patients; P < 0.001), indicating complement-mediated TMA rather than secondary disease. Among treated patients, the odds ratio for 1-year kidney survival was 12.0 (95% confidence interval 1.2-115.4). TMA recurrence was linked to rare genetic variants in all cases. Patients with normal ex vivo C5b9 formation had an acute, nonrelapsing form of TMA. Conclusions Ex vivo C5b9 formation and genetic testing appears to categorize TMAs into different groups because it identifies complement as a driving factor of disease, with potential therapeutic and prognostic implications.

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Section: Discussionmentioning

confidence: 99%

Functional and Genetic Landscape of Complement Dysregulation Along the Spectrum of Thrombotic Microangiopathy and its Potential Implications on Clinical Outcomes

Timmermans

Damoiseaux

Wérion

et al. 2021

Kidney International Reports

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“…These tests consist of the quantification of complement activation products (C3 activation fragments and C5b-9) deposits on EC by immunofluorescence (IF) measured by confocal microscopy or flow cytometry (fluorescence-activated cell sorting, FACS) after incubation with a serum sample of interest. Different protocols have been proposed to study complement activation on the EC surface in several pathological conditions, including aHUS ( 30 , 78 – 88 ), TMA of other etiologies ( 89 , 90 ), HELLP syndrome and pre-eclampsia ( 91 ), C3G ( 14 , 92 ), lupus nephritis (LN) ( 93 , 94 ), APS ( 95 , 96 ), SCD ( 29 ), hemolytic anemia ( 97 ) and hyperhemolytic transfusion reaction without hemoglobinopathy ( 98 ).…”

Section: Study Of Complement Deposition On Cultured Endothelial Cellsmentioning

confidence: 99%

Ex Vivo Test for Measuring Complement Attack on Endothelial Cells: From Research to Bedside

et al. 2022

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As part of the innate immune system, the complement system plays a key role in defense against pathogens and in host cell homeostasis. This enzymatic cascade is rapidly triggered in the presence of activating surfaces. Physiologically, it is tightly regulated on host cells to avoid uncontrolled activation and self-damage. In cases of abnormal complement dysregulation/overactivation, the endothelium is one of the primary targets. Complement has gained momentum as a research interest in the last decade because its dysregulation has been implicated in the pathophysiology of many human diseases. Thus, it appears to be a promising candidate for therapeutic intervention. However, detecting abnormal complement activation is challenging. In many pathological conditions, complement activation occurs locally in tissues. Standard routine exploration of the plasma concentration of the complement components shows values in the normal range. The available tests to demonstrate such dysregulation with diagnostic, prognostic, and therapeutic implications are limited. There is a real need to develop tools to demonstrate the implications of complement in diseases and to explore the complex interplay between complement activation and regulation on human cells. The analysis of complement deposits on cultured endothelial cells incubated with pathologic human serum holds promise as a reference assay. This ex vivo assay most closely resembles the physiological context. It has been used to explore complement activation from sera of patients with atypical hemolytic uremic syndrome, malignant hypertension, elevated liver enzymes low platelet syndrome, sickle cell disease, pre-eclampsia, and others. In some cases, it is used to adjust the therapeutic regimen with a complement-blocking drug. Nevertheless, an international standard is lacking, and the mechanism by which complement is activated in this assay is not fully understood. Moreover, primary cell culture remains difficult to perform, which probably explains why no standardized or commercialized assay has been proposed. Here, we review the diseases for which endothelial assays have been applied. We also compare this test with others currently available to explore complement overactivation. Finally, we discuss the unanswered questions and challenges to overcome for validating the assays as a tool in routine clinical practice.

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“…Thus, a majority of mutations result in haploinsufficiency. However, a smaller portion of individuals are homozygous [ 39 , 41 , 42 ]. Additionally, a specific MCP SNP block in the promoter region, termed the MCPggaac risk haplotype, may be associated with decreased transcriptional activity.…”

Section: Deficiency Statesmentioning

confidence: 99%

“…The IVS2 + 2T > G variant was the most prevalent mutation (and a ‘hot spot’) in a cohort of aHUS-afflicted Indian children [ 46 ]. Further, it was also the most prevalent mutation (13/485) in an international aHUS cohort analyzed by Piras et al [ 41 ]. These studies highlight the potential variable outcome of intronic mutations and the importance of their rigorous analyses.…”

Section: Deficiency Statesmentioning

confidence: 99%

Membrane cofactor protein (MCP; CD46): deficiency states and pathogen connections

Liszewski

Atkinson

2021

Current Opinion in Immunology

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Molecular Studies and an ex vivo Complement Assay on Endothelium Highlight the Genetic Complexity of Atypical Hemolytic Uremic Syndrome: The Case of a Pedigree With a Null CD46 Variant

Cited by 9 publications

References 47 publications

Functional and Genetic Landscape of Complement Dysregulation Along the Spectrum of Thrombotic Microangiopathy and its Potential Implications on Clinical Outcomes

Functional and Genetic Landscape of Complement Dysregulation Along the Spectrum of Thrombotic Microangiopathy and its Potential Implications on Clinical Outcomes

Ex Vivo Test for Measuring Complement Attack on Endothelial Cells: From Research to Bedside

Membrane cofactor protein (MCP; CD46): deficiency states and pathogen connections

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