134 results found
Ahmad A, Mandwie M, O'Sullivan KM, et al., 2022, Conversion of the liver into a biofactory for DNaseI using adeno-associated virus vector gene transfer reduces neutrophil extracellular traps in a model of Systemic Lupus Erythematosus., Hum Gene Ther
Adeno-associated virus (AAV) vectors are proving to be clinically transformative tools in the treatment of monogenic genetic disease. Rapid ongoing development of this technology promises to not only increase the number of monogenic disorders amenable to this approach, but also to bring diseases with complex multigenic and non-genetic aetiologies within therapeutic reach. Here we explore the broader paradigm of converting the liver into a biofactory for systemic output of therapeutic molecules using AAV-mediated delivery of DNaseI as an exemplar. DNaseI can clear neutrophil extracellular traps (NETs), which are nuclear-protein structures possessing anti-microbial action that are also involved in the pathophysiology of clinically troubling immune-mediated diseases. However, a translational challenge is short half-life of the enzyme in vivo (<5 hours). The current study demonstrates that AAV-mediated liver-targeted gene transfer stably induces serum DNaseI activity to >190-fold above physiological levels. In lupus-prone mice (NZBWF1) activity was maintained for longer than 6 months, the latest time point tested, and resulted in a clear functional effect with reduced renal presence of neutrophils, NETs, IgG and complement C3. However, treatment in this complex disease model did not extend life-span, improve serological endpoints or preserve renal function indicating there are elements of pathophysiology not accessible to DNaseI in the NZBWF1 model. We conclude that a translational solution to the challenge of short half-life of DNaseI is AAV-mediated gene delivery and that this may be efficacious in treating disease where NETs are a dominant pathological mechanism.
Gilmore A, Wilson H, Cairns T, et al., 2022, Immune gene expression and functional networks in distinct lupus nephritis classes, Lupus Science & Medicine, Vol: 9, ISSN: 2053-8790
Objective: To explore the utility of the NanoString platform in elucidating kidney immune transcripts for class III, IV and V lupus nephritis (LN) using a retrospective cohort of formalin-fixed paraffin-embedded (FFPE) kidney biopsy tissue.Methods: Immune gene transcript analysis was performed using the NanoString nCounter platform on RNA from LN (n=55), thin basement membrane disease (TBM, n=14) and membranous nephropathy (MN, n=9) FFPE kidney biopsy tissue. LN samples consisted of single class III (n=11), IV (n=23) and V (n=21) biopsies with no mixed lesions. Differential gene expression was performed with NanoString nSolver, with visualisations of volcano plots and heatmaps generated in R. Significant transcripts were interrogated to identify functional networks using STRING and Gene ontogeny terms. Results: In comparison to TBM, we identified 52 significantly differentially expressed genes common to all three LN classes. Pathway analysis showed enrichment for type I interferon (IFN) signalling, complement and MHC II pathways, with most showing the highest expression in class IV LN. Our class IV LN biopsies also showed significant upregulation of NF-κB signalling and immunological enrichment in comparison to class V LN biopsies. Transcripts from the type I IFN pathway distinguished class V LN from MN. Conclusion: Our whole kidney section transcriptomic analysis provided insights into the molecular profile of class III, IV and V LN. The data highlighted important pathways common to all three classes and pathways enriched in our class IV LN biopsies. The ability to reveal molecular pathways in LN using FFPE whole biopsy sections could have clinical utility in treatment selection for LN.
Wong EKS, Marchbank KJ, Lomax-Browne H, et al., 2021, C3 Glomerulopathy and Related Disorders in Children: Etiology-Phenotype Correlation and Outcomes, CLINICAL JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY, Vol: 16, Pages: 1639-1651, ISSN: 1555-9041
Dotz V, Visconti A, Lomax-Browne HJ, et al., 2021, O- and N-glycosylation of serum immunoglobulin A is associated with IgA nephropathy and glomerular function., Journal of the American Society of Nephrology, Vol: 32, Pages: 1-12, ISSN: 1046-6673
BACKGROUND: IgA nephropathy (IgAN) is the most common primary glomerular disease worldwide and is a leading cause of renal failure. The disease mechanisms are not completely understood, but a higher abundance of galactose-deficient IgA is recognized to play a crucial role in IgAN pathogenesis. Although both types of human IgA (IgA1 and IgA2) have several N-glycans as post-translational modification, only IgA1 features extensive hinge-region O-glycosylation. IgA1 galactose deficiency on the O-glycans is commonly detected by a lectin-based method. To date, limited detail is known about IgA O- and N-glycosylation in IgAN. METHODS: To gain insights into the complex O- and N-glycosylation of serum IgA1 and IgA2 in IgAN, we used liquid chromatography-mass spectrometry (LC-MS) for the analysis of tryptic glycopeptides of serum IgA from 83 patients with IgAN and 244 age- and sex-matched healthy controls. RESULTS: Multiple structural features of N-glycosylation of IgA1 and IgA2 were associated with IgAN and glomerular function in our cross-sectional study. These features included differences in galactosylation, sialylation, bisection, fucosylation, and N-glycan complexity. Moreover, IgA1 O-glycan sialylation was associated with both the disease and glomerular function. Finally, glycopeptides were a better predictor of IgAN and glomerular function than galactose-deficient IgA1 levels measured by lectin-based ELISA. CONCLUSIONS: Our high-resolution data suggest that IgA O- and N-glycopeptides are promising targets for future investigations on the pathophysiology of IgAN and as potential noninvasive biomarkers for disease prediction and deteriorating kidney function.
Medjeral-Thomas NR, Cook HT, Pickering MC, 2021, Complement activation in IgA nephropathy, Springer Seminars in Immunopathology, Vol: 43, Pages: 679-690, ISSN: 1863-2297
IgA nephropathy pathogenesis is incompletely understood, and this limits the development of disease-specific biomarkers and effective therapies. Evidence of complement activity in IgA nephropathy is well established. However, a growing body of research indicates complement activity is an important contributor to IgA nephropathy pathology. In particular, multiple associations have been identified between complement alternative, lectin and terminal pathway proteins and IgA nephropathy severity. Recently, we have also gained insight into possible mechanisms that could link glomerular IgA deposition, complement activity, glomerular inflammation and disease severity. Ongoing clinical trials of therapeutic complement inhibitors will provide insight into the importance of complement activity to IgA nephropathy pathogenesis. Further research into mechanisms of complement activity is essential to improving our understanding and management of patients with IgA nephropathy.
Daugan MV, Revel M, Thouenon R, et al., 2021, Intracellular Factor H Drives Tumor Progression Independently of the Complement Cascade, CANCER IMMUNOLOGY RESEARCH, Vol: 9, Pages: 909-925, ISSN: 2326-6066
Gyapon-Quast F, Goicoechea de Jorge E, Malik T, et al., 2021, Defining the glycosaminoglycan interactions of complement factor H-related protein 5, Journal of Immunology, Vol: 207, Pages: 534-541, ISSN: 0022-1767
Complement activation is an important mediator of kidney injury in glomerulonephritis. Complement factor H (FH) and FH-related protein 5 (FHR-5) influence complement activation in C3 glomerulopathy and IgA nephropathy by differentially regulating glomerular complement. FH is a negative regulator of complement C3 activation. Conversely, FHR-5 in vitro promotes C3 activation either directly or by competing with FH for binding to complement C3b. The FH-C3b interaction is enhanced by surface glycosaminoglycans (GAGs) and the FH-GAG interaction is well-characterized. In contrast, the contributions of carbohydrates to the interaction of FHR-5 and C3b are unknown. Using plate-based and microarray technologies we demonstrate that FHR-5 interacts with sulfated GAGs and that this interaction is influenced by the pattern and degree of GAG sulfation. The FHR-5-GAG interaction that we identified has functional relevance as we could show that the ability of FHR-5 to prevent binding of FH to surface C3b is enhanced by surface kidney heparan sulfate. Our findings are important in understanding the molecular basis of the binding of FHR-5 to glomerular complement and the role of FHR-5 in complement-mediated glomerular disease.
Ahmad A, Mandwie M, Dreismann AK, et al., 2021, Adeno-associated virus vector gene delivery elevates Factor I levels and down-regulates the complement alternative pathway in vivo., Human Gene Therapy, Vol: 32, Pages: 1370-1381, ISSN: 1043-0342
The complement system is a key component of innate immunity but impaired regulation influences disease susceptibility, including age-related macular degeneration (AMD) and some kidney diseases. Whilst complete complement inhibition has been used successfully to treat acute kidney disease, key unresolved challenges include strategies to modulate rather than completely inhibit the system and to deliver therapy potentially over decades. Elevating concentrations of complement regulator factor I (CFI) restricts complement activation in vitro and this approach was extended in the current study to modulate complement activation in vivo. Sustained increases in CFI levels were achieved using an adeno-associated virus (AAV) vector to target the liver, inducing a 4- to 5-fold increase in circulating CFI levels. This led to decreased activity of the alternative pathway as demonstrated by a reduction in the rate of iC3b deposition and more rapid formation of C3 degradation products. In addition, vector application in a mouse model of systemic lupus erythematosus (NZBWF1), where tissue injury is in part complement dependent, resulted in reduced complement C3 and IgG renal deposition. Collectively, these data demonstrate that sustained elevation of CFI reduces complement activation in vivo providing proof-of-principle support for the therapeutic application of AAV gene delivery to modulate complement activation.
Medjeral-Thomas N, Troldborg A, Hansen A, et al., 2021, Protease inhibitor plasma concentrations associate with COVID-19 infection, Oxford Open Immunology, Vol: 2, ISSN: 2633-6960
Protease inhibitors influence a range of innate immunity and inflammatory pathways. We quantified plasma concentrations of key anti-inflammatory protease inhibitors in chronic haemodialysis patients with COVID-19. The samples were collected early in the disease course to determine whether plasma protease inhibitor levels associated with the presence and severity of COVID-19. We used antibody-based immunoassays to measure plasma concentrations of C1 esterase inhibitor (C1-INH), alpha2-macroglobulin (α2M), antithrombin, and inter-alpha-inhibitor heavy chain 4 (ITIH4) in 100 serial samples from 27 haemodialysis patients with COVID-19. ITIH4 was tested in two assays, one measuring intact ITIH4 and another also detecting any fragmented ITIH4 (total ITIH4). Control cohorts were 32 haemodialysis patients without COVID-19 and 32 healthy controls. We compared protease inhibitor concentration based on current and future COVID-19 severity and with CRP. Results were adjusted for repeated measures and multiple comparisons. Analysis of all available samples demonstrated lower plasma C1-INH and α2M and higher total ITIH4 in COVID-19 compared to dialysis controls. These differences were also seen in the first sample collected after COVID-19 diagnosis, a median of four days from diagnostic swab. Plasma ITIH4 levels were higher in severe than non-severe COVID-19. Serum CRP correlated positively with plasma levels of antithrombin, intact ITIH4, and total ITIH4. In conclusion, plasma protease inhibitor concentrations are altered in COVID-19.
Kerr H, Herbert AP, Makou E, et al., 2021, Murine factor H Co-produced in yeast with protein disulfide isomerase ameliorated C3 dysregulation in factor H-deficient mice, Frontiers in Immunology, Vol: 12, Pages: 1-17, ISSN: 1664-3224
Recombinant human factor H (hFH) has potential for treating diseases linked to aberrant complement regulation including C3 glomerulopathy (C3G) and dry age-related macular degeneration. Murine FH (mFH), produced in the same host, is useful for pre-clinical investigations in mouse models of disease. An abundance of FH in plasma suggests high doses, and hence microbial production, will be needed. Previously, Pichia pastoris produced useful but modest quantities of hFH. Herein, a similar strategy yielded miniscule quantities of mFH. Since FH has 40 disulfide bonds, we created a P. pastoris strain containing a methanol-inducible codon-modified gene for protein-disulfide isomerase (PDI) and transformed this with codon-modified DNA encoding mFH under the same promoter. What had been barely detectable yields of mFH became multiple 10s of mg/L. Our PDI-overexpressing strain also boosted hFH overproduction, by about tenfold. These enhancements exceeded PDI-related production gains reported for other proteins, all of which contain fewer disulfide-stabilized domains. We optimized fermentation conditions, purified recombinant mFH, enzymatically trimmed down its (non-human) N-glycans, characterised its functions in vitro and administered it to mice. In FH-knockout mice, our de-glycosylated recombinant mFH had a shorter half-life and induced more anti-mFH antibodies than mouse serum-derived, natively glycosylated, mFH. Even sequential daily injections of recombinant mFH failed to restore wild-type levels of FH and C3 in mouse plasma beyond 24 hours after the first injection. Nevertheless, mFH functionality appeared to persist in the glomerular basement membrane because C3-fragment deposition here, a hallmark of C3G, remained significantly reduced throughout and beyond the ten-day dosing regimen.
Medjeral-Thomas NR, Pickering MC, Cook HT, 2021, Complement and kidney disease, new insights., Curr Opin Nephrol Hypertens, Vol: 30, Pages: 310-316
PURPOSE OF REVIEW: In this review, we discuss recent studies showing the importance of the complement pathway in kidney disease. RECENT FINDINGS: Recent findings in C3 glomerulopathy (C3G) include: acute postinfectious glomerulonephritis is characterised by the presence of antifactor B antibodies; human leukocyte antigen type, but not rare complement gene variation, is associated with primary immunoglobulin-associated membranoproliferative GN and C3G. Immunohistochemistry in C3G shows that factor H related protein 5 (FHR5) is the most prevalent complement protein and correlates with kidney function. A multicentre study supported the use of mycophenolate mofetil (MMF) in C3G even after a propensity matching analysis. In immunoglobulin A nephropathy (IgAN) several studies have emphasised the importance of complement. Imbalances of circulating FH and FHR1 and FHR5, which interfere with the regulatory functions of FH, associate with IgAN. Immunohistochemistry has shown associations between glomerular FHR5 deposition and C3 activation; glomerular FHR5 associated with clinical markers of IgAN severity. Data also suggest the lectin complement pathway contributes to IgAN severity. We also discuss complement activation in thrombotic microangiopathy and other kidney diseases. SUMMARY: Complement activity can be detected in a wide range of kidney diseases and this provides pathogenic insight and potential for therapy with the ongoing development of several drugs directed at complement activation.
Medjeral-Thomas N, Troldborg A, Hansen A, et al., 2021, Plasma lectin pathway complement proteins in patients with COVID-19 and renal disease, Frontiers in Immunology, Vol: 12, ISSN: 1664-3224
We do not understand why non-white ethnicity and chronic kidney disease increase susceptibility to COVID-19. The lectin pathway of complement activation is a key contributor to innate immunity and inflammation. Concentrations of plasma lectin pathway proteins influence pathway activity and vary with ethnicity. We measured circulating lectin proteins in a multi-ethnic cohort of chronic kidney disease patients with and without COVID19 infection to determine if lectin pathway activation was contributing to COVID19 severity.We measured 11 lectin proteins in serial samples from a cohort of 33 patients with chronic kidney impairment and COVID19. Controls were single plasma samples from 32 patients on dialysis and 32 healthy individuals. We demonstrated multiple associations between recognition molecules and associated proteases of the lectin pathway and COVID-19, including COVID-19 severity. Some of these associations were unique to patients of Asian and White ethnicity. Our novel findings demonstrate that COVID19 infection alters the concentration of plasma lectin proteins and some of these changes were linked to ethnicity. This suggests a role for the lectin pathway in the host response to COVID-19 and suggest that variability within this pathway may contribute to ethnicity-associated differences in susceptibility to severe COVID-19.
Botto M, Buang N, Tapeng L, et al., 2021, Type I interferons affect the metabolic fitness of CD8+ T cells from patients with systemic lupus erythematosus, Nature Communications, Vol: 12, Pages: 1-15, ISSN: 2041-1723
The majority of patients with systemic lupus erythematosus (SLE) have high expression of type I IFN-stimulated genes. Mitochondrial abnormalities have also been reported, but the contribution of type I IFN exposure to these changes is unknown. Here, we show downregulation of mitochondria-derived genes and mitochondria-associated metabolic pathways in IFN-High patients from transcriptomic analysis of CD4+ and CD8+ T cells. CD8+ T cells from these patients have enlarged mitochondria and lower spare respiratory capacity associated with increased cell death upon rechallenge with TCR stimulation. These mitochondrial abnormalities can be phenocopied by exposing CD8+ T cells from healthy volunteers to type I IFN and TCR stimulation. Mechanistically these ‘SLE-like’ conditions increase CD8+ T cell NAD+ consumption resulting in impaired mitochondrial respiration and reduced cell viability, both of which can be rectified by NAD+ supplementation. Our data suggest that type I IFN exposure contributes to SLE pathogenesis by promoting CD8+ T cell death via metabolic rewiring.
Malik TH, Gitterman DP, Lavin DP, et al., 2021, Gain-of-function factor H–related 5 protein impairs glomerular complement regulation resulting in kidney damage, Proceedings of the National Academy of Sciences, Vol: 118, Pages: 1-9, ISSN: 0027-8424
Genetic variation within the factor H–related (FHR) genes is associated with the complement-mediated kidney disease, C3 glomerulopathy (C3G). There is no definitive treatment for C3G, and a significant proportion of patients develop end-stage renal disease. The prototypical example is CFHR5 nephropathy, through which an internal duplication within a single CFHR5 gene generates a mutant FHR5 protein (FHR5mut) that leads to accumulation of complement C3 within glomeruli. To elucidate how abnormal FHR proteins cause C3G, we modeled CFHR5 nephropathy in mice. Animals lacking the murine factor H (FH) and FHR proteins, but coexpressing human FH and FHR5mut (hFH-FHR5mut), developed glomerular C3 deposition, whereas mice coexpressing human FH with the normal FHR5 protein (hFH-FHR5) did not. Like in patients, the FHR5mut had a dominant gain-of-function effect, and when administered in hFH-FHR5 mice, it triggered C3 deposition. Importantly, adeno-associated virus vector-delivered homodimeric mini-FH, a molecule with superior surface C3 binding compared to FH, reduced glomerular C3 deposition in the presence of the FHR5mut. Our data demonstrate that FHR5mut causes C3G by disrupting the homeostatic regulation of complement within the kidney and is directly pathogenic in C3G. These results support the use of FH-derived molecules with enhanced C3 binding for treating C3G associated with abnormal FHR proteins. They also suggest that targeting FHR5 represents a way to treat complement-mediated kidney injury.
Gilmore AC, Zhang Y, Cook HT, et al., 2021, Complement activity is regulated in C3 glomerulopathy by IgG-factor H fusion proteins with and without properdin targeting domains, Kidney International, Vol: 99, Pages: 396-404, ISSN: 0085-2538
C3 glomerulopathy is characterized by accumulation of complement C3 within glomeruli. Causes include, but are not limited to, abnormalities in factor H, the major negative regulator of the complement alternative pathway. Factor H-deficient (Cfh-/-) mice develop C3 glomerulopathy together with a reduction in plasma C3 levels. Using this model, we assessed the efficacy of two fusion proteins containing the factor H alternative pathway regulatory domains (FH1-5) linked to either a non-targeting mouse immunoglobulin (IgG-FH1-5) or to an anti-mouse properdin antibody (Anti-P-FH1-5). Both proteins increased plasma C3 and reduced glomerular C3 deposition to an equivalent extent, suggesting that properdin-targeting was not required for FH1-5 to alter C3 activation in either plasma or glomeruli. Following IgG-FH1-5 administration, plasma C3 levels temporally correlated with changes in factor B levels whereas plasma C5 levels correlated with changes in plasma properdin levels. Notably, the increases in plasma C5 and properdin levels persisted for longer than the increases in C3 and factor B. In Cfh-/- mice IgG-FH1-5 reduced kidney injury during accelerated serum nephrotoxic nephritis. Thus, our data demonstrate that IgG-FH1-5 restored circulating alternative pathway activity and reduced glomerular C3 deposition in Cfh-/- mice and that plasma properdin levels are a sensitive marker of C5 convertase activity in factor H deficiency. The immunoglobulin conjugated FH1-5 protein, through its comparatively long plasma half-life, may be a potential therapy for C3 glomerulopathy.
Rovin BH, Caster DJ, Cattran DC, et al., 2021, Management and treatment of glomerular diseases (part 2): Conclusions from a kidney disease: Improving global outcomes (kdigo) controversies conference, Nephrology (Saint-Petersburg), Vol: 25, Pages: 96-119, ISSN: 1561-6274
In November 2017, the Kidney Disease: Improving Global Outcomes (KDIGO) initiative brought a diverse panel of experts in glomerular diseases together to discuss the 2012 KDIGO glomerulonephritis guideline in the context of new developments and insights that had occurred over the years since its publication. During this KDIGO Controversies Conference on Glomerular Diseases, the group examined data on disease pathogenesis, biomarkers, and treatments to identify areas of consensus and areas of controversy. This report summarizes the discussions on primary podocytopathies, lupus nephritis, anti-neutrophil cytoplasmic antibody-associated nephritis, complement mediated kidney diseases, and monoclonal gammopathies of renal significance.
Prendecki M, Clarke C, McKinnon T, et al., 2021, SARS-CoV-2 antibody point-of-care testing in dialysis and kidney transplant patients with COVID-19., Kidney Medicine, Vol: 3, Pages: 54-59.e1, ISSN: 2590-0595
Rationale & Objective: A number of serologic tests for immunoglobulin G (IgG) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are now commercially available, including multiple lateral flow immunoassays (LFIAs), which have the advantage of being inexpensive and easy to use, without the reliance on laboratory facilities. However, data on the development of humoral immunity to SARS-CoV-2 in patients with kidney disease is limited, and the utility of an LFIA to test for antibodies in these patients has not been assessed. Study Design: Observational study. Setting & Participants: 60 patients (40 hemodialysis and 20 kidney transplant recipients) with SARS-CoV-2 infection confirmed by viral reverse transcriptase-polymerase chain reaction (RT-PCR) testing and 88 historic negative-control samples (collected before September 2019). Test: A commercially available LFIA to test for SARS-CoV-2 IgG in patients with infection confirmed by viral RT-PCR testing. Outcomes: Sensitivity and specificity of the LFIA to detect SARS-CoV-2 IgG in dialysis patients and transplant recipients. Results: 56/58 (96.6%) patients (38/39 hemodialysis and 18/19 transplant recipients) tested positive for SARS-CoV-2 IgG. 5/7 (71.4%) patients who were negative on preliminary testing had detectable IgG when retested more than 21 days postdiagnosis. Median times to first and second tests after diagnosis were 17 (interquartile range, 15-20) and 35 (interquartile range, 30-39) days, respectively. Calculation of test characteristics gave sensitivity of 96.6% (95% CI, 88.3%-99.4%) and specificity of 97.7% (95% CI, 92.0-99.6%). Limitations: Possible exposure to other beta-coronaviruses that may cross-react with the antigen used in the LFIA cannot be excluded. Conclusions: Symptomatic dialysis patients and transplant recipients commonly develop an immune response against SARS-CoV-2 infection that can be detected using an LFIA. Used diligently, an LFIA could be used to help scre
Gisby J, Clarke C, Medjeral-Thomas N, et al., 2020, Longitudinal proteomic profiling of high-risk patients with COVID-19 reveals markers of severity and predictors of fatal disease, eLife, Vol: 10, Pages: 1-30, ISSN: 2050-084X
End-stage kidney disease (ESKD) patients are at high risk of severe COVID-19. We performed dense serial blood sampling in hospitalised and non-hospitalised ESKD patients with COVID-19 (n=256 samples from 55 patients) and used Olink immunoassays to measure 436 circulating proteins. Comparison to 51 non-infected ESKD patients revealed 221 proteins differentially expressed in COVID-19, of which 69.7% replicated in an independent cohort of 46 COVID-19 patients. 203 proteins were associated with clinical severity scores, including IL6, markers of monocyte recruitment (e.g. CCL2, CCL7), neutrophil activation (e.g proteinase-3) and epithelial injury (e.g. KRT19). Random Forests machine learning identified predictors of current or future severity such as KRT19, PARP1, PADI2, CCL7, and IL1RL1 (ST2). Survival analysis with joint models revealed 69 predictors of death including IL22RA1, CCL28, and the neutrophil-derived chemotaxin AZU1 (Azurocidin). Finally, longitudinal modelling with linear mixed models uncovered 32 proteins that display different temporal profiles in severe versus non-severe disease, including integrins and adhesion molecules. Our findings point to aberrant innate immune activation and leucocyte-endothelial interactions as central to the pathology of severe COVID-19. The data from this unique cohort of high-risk individuals provide a valuable resource for identifying drug targets in COVID-19.
Prendecki M, Clarke C, Medjeral-Thomas N, et al., 2020, Temporal changes in complement activation in haemodialysis patients with COVID-19 as a predictor of disease progression, Clinical Kidney Journal, Vol: 13, Pages: 889-896, ISSN: 2048-8505
Background: Complement activation may play a pathogenic role in patients with severe coronavirus disease 2019 (COVID-19) by contributing to tissue inflammation and microvascular thrombosis. Methods: Serial samples were collected from patients receiving maintenance haemodialysis (HD). Thirty-nine patients had confirmed COVID-19 and 10 patients had no evidence of COVID-19. Plasma C5a and C3a levels were measured using enzyme-linked immunosorbent assay. Results: We identified elevated levels of plasma C3a and C5a in HD patients with severe COVID-19 compared with controls. Serial sampling identified that C5a levels were elevated prior to clinical deterioration in patients who developed severe disease. C3a more closely mirrored both clinical and biochemical disease severity. Conclusions: Our findings suggest that activation of complement plays a role in the pathogenesis of COVID-19, leading to endothelial injury and lung damage. C5a may be an earlier biomarker of disease severity than conventional parameters such as C-reactive protein and this warrants further investigation in dedicated biomarker studies. Our data support the testing of complement inhibition as a therapeutic strategy for patients with severe COVID-19.
Lou H, Wojciak-Stothard B, Ruseva MM, et al., 2020, Autoantibody-dependent amplification of inflammation in SLE, Cell Death and Disease, Vol: 11, ISSN: 2041-4889
Anti-double stranded DNA antibodies (anti-dsDNA) are a hallmark of SLE but their role in disease pathogenesis is not fully resolved. Anti-dsDNA in serum are highly heterogeneous therefore in this study, we aimed to dissect the functional specificities of anti-dsDNA using a panel of human monoclonal antibodies (humAbs) generated from patients with active lupus nephritis. A total of 46 ANA reactive humAbs were isolated and divided into four broad classes based on their reactivity to histones, DNA and Crithidia. Functional analysis indicated that one subclass of antibodies bound strongly to decondensed DNA areas in neutrophil extracellular traps (NETs) and protected NETs from nuclease digestion, similar to the sera from active SLE patients. In addition, these anti-dsDNA antibodies could stimulate type I interferon responses in mononuclear phagocytic cells, or NF-kB activity in endothelial cells, by uptake of NETs-anti-NETs immune complexes and subsequently trigging inflammatory responses in an Fc-gamma receptor (Fcg-R)-dependant manner. Together our data suggest that only a subset of anti-dsDNA antibodies is capable to amplify inflammatory responses by deposit in the nephritic kidney in vivo, protecting NETs digestion as well as uptake of NETs immune complexes into Fcg-R-expressing cells in vitro.
Clarke C, Prendecki M, Dhutia A, et al., 2020, High prevalence of asymptomatic COVID-19 infection in hemodialysis patients detected using serologic screening, Journal of the American Society of Nephrology, Vol: 31, Pages: 1969-1975, ISSN: 1046-6673
BACKGROUND: Strategies to minimize the risk of transmission and acquisition of COVID-19 infection in patients with ESKD receiving in-center hemodialysis have been rapidly implemented across the globe. Despite these interventions, confirmed COVID-19 infection rates have been high in the United Kingdom. Prevalence of asymptomatic disease in an adult hemodialysis population has not been reported. Also, to our knowledge, the development of humoral response to SARS-CoV-2 has not been previously reported in this population. Although serologic testing does not provide information on the infectivity of patients, seroprevalence studies may enable investigation of exposure within dialysis units and hence, assessment of current screening strategies. METHODS: To investigate the seroprevalence of SARS-CoV-2 antibodies in a hemodialysis population, we used the Abbott IgG assay with the Architect system to test serum samples from 356 patients receiving in-center hemodialysis for SARS-CoV-2 antibodies. RESULTS: Of 356 patients, 121 had been symptomatic when screened before a dialysis session and received an RT-PCR test; 79 (22.2% of the total study population) tested positive for COVID-19. Serologic testing of all 356 patients found 129 (36.2%) who tested positive for SARS-CoV-2 antibodies. Only two patients with PCR-confirmed infection did not seroconvert. Of the 129 patients with SARS-CoV-2 antibodies, 52 (40.3%) had asymptomatic disease or undetected disease by PCR testing alone. CONCLUSIONS: We found a high seroprevalence of SARS-CoV-2 antibodies in patients receiving in-center hemodialysis. Serologic evidence of previous infection in asymptomatic or PCR-negative patients suggests that current diagnostic screening strategies may be limited in their ability to detect acute infection.
Merle NS, Leon J, Poillerat V, et al., 2020, Circulating FH protects kidneys from tubular injury during systemic hemolysis, Frontiers in Immunology, Vol: 11, Pages: 1-13, ISSN: 1664-3224
Intravascular hemolysis of any cause can induce acute kidney injury (AKI). Hemolysis-derived product heme activates the innate immune complement system and contributes to renal damage. Therefore, we explored the role of the master complement regulator Factor H (FH) in the kidney's resistance to hemolysis-mediated AKI. Acute systemic hemolysis was induced in mice lacking liver expression of FH (hepatoFH−/−, ~20% residual FH) and in WT controls, by phenylhydrazine injection. The impaired complement regulation in hepatoFH−/− mice resulted in a delayed but aggravated phenotype of hemolysis-related kidney injuries. Plasma urea as well as markers for tubular (NGAL, Kim-1) and vascular aggression peaked at day 1 in WT mice and normalized at day 2, while they increased more in hepatoFH−/− compared to the WT and still persisted at day 4. These were accompanied by exacerbated tubular dilatation and the appearance of tubular casts in the kidneys of hemolytic hepatoFH−/− mice. Complement activation in hemolytic mice occurred in the circulation and C3b/iC3b was deposited in glomeruli in both strains. Both genotypes presented with positive staining of FH in the glomeruli, but hepatoFH−/− mice had reduced staining in the tubular compartment. Despite the clear phenotype of tubular injury, no complement activation was detected in the tubulointerstitium of the phenylhydrazin-injected mice irrespective of the genotype. Nevertheless, phenylhydrazin triggered overexpression of C5aR1 in tubules, predominantly in hepatoFH−/− mice. Moreover, C5b-9 was deposited only in the glomeruli of the hemolytic hepatoFH−/− mice. Therefore, we hypothesize that C5a, generated in the glomeruli, could be filtered into the tubulointerstitium to activate C5aR1 expressed by tubular cells injured by hemolysis-derived products and will aggravate the tissue injury. Plasma-derived FH is critical for the tubular protection, since
Laskowski J, Renner B, Pickering MC, et al., 2020, Complement factor H-deficient mice develop spontaneous hepatic tumors, JOURNAL OF CLINICAL INVESTIGATION, Vol: 130, Pages: 4039-4054, ISSN: 0021-9738
Nayagam JS, McGrath S, Montasser M, et al., 2020, Successful simultaneous liver-kidney transplantation for renal failure associated with hereditary complement C3 deficiency., American Journal of Transplantation, Vol: 20, Pages: 2260-2263, ISSN: 1600-6135
Hereditary complement C3 deficiency is associated with recurrent bacterial infections and proliferative glomerulonephritis. We describe a case of an adult with complete deficiency of complement C3 due to homozygous mutations in C3 gene: c.1811delT (Val604Glyfs*2), recurrent bacterial infections, crescentic glomerulonephritis and end-stage renal failure. Following isolated kidney transplantation he would remain C3 deficient with a similar, or increased, risk of infections and glomerulonephritis. As C3 is predominantly synthesised in the liver, with a small proportion of C3 monocyte-derived and kidney-derived, he proceeded to simultaneous liver-kidney transplantation. The procedure has been successful with restoration of his circulating C3 levels, normal liver and kidney function at 26 months of follow up. Simultaneous liver-kidney transplant is a viable option to be considered in this rare setting.
Kasanmoentalib ES, Seron MV, Engelen-Lee JY, et al., 2019, Complement factor H contributes to mortality in humans and mice with bacterial meningitis, Journal of Neuroinflammation, Vol: 16, Pages: 1-14, ISSN: 1742-2094
BackgroundThe complement system is a vital component of the inflammatory response occurring during bacterial meningitis. Blocking the complement system was shown to improve the outcome of experimental pneumococcal meningitis. Complement factor H (FH) is a complement regulatory protein inhibiting alternative pathway activation but is also exploited by the pneumococcus to prevent complement activation on its surface conferring serum resistance.MethodsIn a nationwide prospective cohort study of 1009 episodes with community-acquired bacterial meningitis, we analyzed whether genetic variations in CFH influenced FH cerebrospinal fluid levels and/or disease severity. Subsequently, we analyzed the role of FH in our pneumococcal meningitis mouse model using FH knock-out (Cfh−/−) mice and wild-type (wt) mice. Finally, we tested whether adjuvant treatment with human FH (hFH) improved outcome in a randomized investigator blinded trial in a pneumococcal meningitis mouse model.ResultsWe found the major allele (G) of single nucleotide polymorphism in CFH (rs6677604) to be associated with low FH cerebrospinal fluid concentration and increased mortality. In patients and mice with bacterial meningitis, FH concentrations were elevated during disease and Cfh−/− mice with pneumococcal meningitis had increased mortality compared to wild-type mice due to C3 depletion. Adjuvant treatment of wild-type mice with purified human FH led to complement inhibition but also increased bacterial outgrowth which resulted in similar disease outcomes.ConclusionLow FH levels contribute to mortality in pneumococcal meningitis but adjuvant treatment with FH at a clinically relevant time point is not beneficial.
IgA nephropathy (IgAN) is common and often progresses to end stage renal disease. IgAN encompasses a wide range of histology and clinical features. IgAN pathogenesis is incompletely understood; the current multi-hit hypothesis of IgAN pathogenesis does not explain the range of glomerular inflammation and renal injury associated with mesangial IgA deposition. Although associations between IgAN and glomerular and circulating markers of complement activation are established, the mechanism of complement activation and contribution to glomerular inflammation and injury are not defined. Recent identification of specific complement pathways and proteins in severe IgAN cases had advanced our understanding of complement in IgAN pathogenesis. In particular, a growing body of evidence implicates the complement factor H related proteins 1 and 5 and lectin pathway as pathogenic in a subset of patients with severe disease. These data suggest complement deregulation and activity may be dominant drivers of renal injury in IgAN. Thereby, markers of complement activation may identify IgAN patients likely to progress to significant renal impairment and complement inhibition may emerge as an effective method of preventing and reducing glomerular injury in IgAN.
Vyas FS, Yang Y, Pappworth IY, et al., 2019, A MOUSE HOMODIMERIC MINI-FH CONSTRUCT PROVIDES LONG TERM PROTECTION FROM KIDNEY DISEASE IN A MOUSE MODEL OF C3G, 17th European Meeting on Complement in Human Disease (EMCHD), Publisher: PERGAMON-ELSEVIER SCIENCE LTD, Pages: 452-453, ISSN: 0161-5890
Medjeral-Thomas NR, Moffitt H, Lomax-Browne HJ, et al., 2019, Glomerular complement factor H–related protein 5 (FHR5) is highly Prevalent in C3 glomerulopathy and associated with renal impairment, Kidney International Reports, Vol: 4, Pages: 1387-1400, ISSN: 2468-0249
IntroductionTherapeutic agents that target complement are increasingly available for glomerular diseases. However, the mechanisms linking glomerular complement deposition with inflammation and damage are incompletely understood. Complement factor H–related protein 5 (FHR5) interacts with complement C3 and is considered to promote activation. Circulating and glomerular FHR5 associates with IgA nephropathy and abnormal FHR5 associates with familial C3 glomerulopathy (C3G). We characterized glomerular FHR5 staining in C3G and assessed its relationships with histological features of glomerular injury and clinical outcome.MethodsWe developed FHR5 staining protocols for formalin-fixed paraffin-embedded (FFPE) renal tissue and applied them to surplus biopsy sections from a C3G cohort.ResultsGlomerular FHR5 was highly prevalent in native and transplant C3G and correlated with glomerular C3 and C5b-9 staining. Glomerular FHR5 staining correlated negatively with estimated glomerular filtration rate (eGFR) (P = 0.04, difference of medians 19.7 ml/min per 1.73 m2; 95% confidence interval [CI] 1.1–43.0) and positively with a membranoproliferative glomerulonephritis pattern at diagnostic biopsy (odds ratio 18; 95% CI 1.6–201; P = 0.049). Glomerular FHR5 staining intensity positively correlated with glomerular complement C3b/iC3b/C3c (Pearson’s correlation coefficient [R] = 0.59; P = 0.0008), C3dg (R = 0.47; P = 0.02) and C5b9 (R = 0.44, P = 0.02).ConclusionsGlomerular FHR5 is highly prevalent in C3G, interacts with glomerular C3, and is associated with markers of disease severity. Glomerular FHR5 likely exacerbates complement-mediated glomerular damage in C3G and its interaction with glomerular complement might be exploited to target complement therapeutic agents.
Kiss MG, Ozsvar-Kozma M, Porsch F, et al., 2019, Complement factor H modulates splenic B cell development and limits autoantibody production, Frontiers in Immunology, Vol: 10, Pages: 1-12, ISSN: 1664-3224
Complement factor H (CFH) has a pivotal role in regulating alternative complement activation through its ability to inhibit the cleavage of the central complement component C3, which links innate and humoral immunity. However, insights into the role of CFH in B cell biology are limited. Here, we demonstrate that deficiency of CFH in mice leads to altered splenic B cell development characterized by the accumulation of marginal zone (MZ) B cells. Furthermore, B cells in Cfh−/− mice exhibit enhanced B cell receptor (BCR) signaling as evaluated by increased levels of phosphorylated Bruton's tyrosine kinase (pBTK) and phosphorylated spleen tyrosine kinase (pSYK). We show that enhanced BCR activation is associated with uncontrolled C3 consumption in the spleen and elevated complement receptor 2 (CR2, also known as CD21) levels on the surface of mature splenic B cells. Moreover, aged Cfh−/− mice developed splenomegaly with distorted spleen architecture and spontaneous B cell-dependent autoimmunity characterized by germinal center hyperactivity and a marked increase in anti-double stranded DNA (dsDNA) antibodies. Taken together, our data indicate that CFH, through its function as a complement repressor, acts as a negative regulator of BCR signaling and limits autoimmunity.
Foschi V, Bortolotti D, Doyle AF, et al., 2019, Analysis of HLA-G expression in renal tissue in lupus nephritis: a pilot study, Lupus, Vol: 28, Pages: 1091-1100, ISSN: 1477-0962
BACKGROUND: The study aimed to investigate whether HLA-G antigen is expressed in the kidneys of patients affected by lupus nephritis (LN) and whether its detection in renal biopsies could be adopted as a marker of treatment response and prognosis. METHODS: Thirty renal biopsies from patients with LN were selected and analyzed through immunohistochemistry. Laboratory and clinical data were retrospectively collected at baseline, 6 and 12 months and at the latest clinical appointment. A number of patients (63.3%) were treated with rituximab (RTX) +/- methylprednisolone in the induction phase. The expression of HLA-G in glomeruli, tubules and infiltrating cells was examined and compared between lupus patients who achieved either complete or partial renal response and those who did not respond to treatment. RESULTS: HLA-G staining was observed in the glomeruli of 20 of 30 samples from patients with LN. The expression of the antigen was detected in podocytes, along glomerular capillary walls, on parietal glomerular epithelial cells and within the juxtaglomerular apparatus. Seventy per cent of patients whose glomeruli expressed HLA-G achieved partial or complete response at 6 months and 75% at the latest available follow up compared with 30% and 40%, respectively, of those who did not show any expression. The pattern of staining in tubules and infiltrating cells was highly variable precluding any clinical correlation. CONCLUSION: This study demonstrates that HLA-G is expressed in renal tissue in LN. Our retrospective data suggest that its expression could correlate with response to treatment.
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