Patients with hypertension-associated thrombotic microangiopathy may present with complement abnormalities
2017; Elsevier BV; Volume: 91; Issue: 6 Linguagem: Inglês
10.1016/j.kint.2016.12.009
ISSN1523-1755
AutoresSjoerd A.M.E.G. Timmermans, Myrurgia A. Abdul‐Hamid, Joris Vanderlocht, Jan Damoiseaux, Chris Reutelingsperger, Pieter van Paassen, Frederiek de Heer, Mariëlle Krekels, Frank Stifft, Gaico H. Verseput, Siska Boorsma, Willem S. de Grave, J J Huitema, J.J.J.M. Wirtz, N. ter Braak, L.A.M. Frenken, S. Gaertner, M. Christiaans, T P Fung, M. Gelens, Jeroen P. Kooman, Karel M.L. Leunissen, Elisabeth J R Litjens, J van der Net, Frank M. van der Sande, E. van Duijnhoven,
Tópico(s)Renin-Angiotensin System Studies
ResumoThrombotic microangiopathy (TMA) is a pattern of endothelial damage that can be found in association with diverse clinical conditions such as malignant hypertension. Although the pathophysiological mechanisms differ, accumulating evidence links complement dysregulation to various TMA syndromes and in particular the atypical hemolytic uremic syndrome. Here, we evaluated the role of complement in nine consecutive patients with biopsy-proven renal TMA attributed to severe hypertension. Profound hematologic symptoms of TMA were uncommon. In six out of nine patients, we found mutations C3 in three, CFI in one, CD46 in one, and/or CFH in two patients either with or without the risk CFH-H3 haplotype in four patients. Elevated levels of the soluble C5b-9 and renal deposits of C3c and C5b-9 along the vasculature and/or glomerular capillary wall, confirmed complement activation in vivo. In contrast to patients without genetic defects, patients with complement defects invariably progressed to end-stage renal disease, and disease recurrence after kidney transplantation seems common. Thus, a subset of patients with hypertension-associated TMA falls within the spectrum of complement-mediated TMA, the prognosis of which is poor. Hence, testing for genetic complement abnormalities is warranted in patients with severe hypertension and TMA on renal biopsy to adopt suitable treatment options and prophylactic measures. Thrombotic microangiopathy (TMA) is a pattern of endothelial damage that can be found in association with diverse clinical conditions such as malignant hypertension. Although the pathophysiological mechanisms differ, accumulating evidence links complement dysregulation to various TMA syndromes and in particular the atypical hemolytic uremic syndrome. Here, we evaluated the role of complement in nine consecutive patients with biopsy-proven renal TMA attributed to severe hypertension. Profound hematologic symptoms of TMA were uncommon. In six out of nine patients, we found mutations C3 in three, CFI in one, CD46 in one, and/or CFH in two patients either with or without the risk CFH-H3 haplotype in four patients. Elevated levels of the soluble C5b-9 and renal deposits of C3c and C5b-9 along the vasculature and/or glomerular capillary wall, confirmed complement activation in vivo. In contrast to patients without genetic defects, patients with complement defects invariably progressed to end-stage renal disease, and disease recurrence after kidney transplantation seems common. Thus, a subset of patients with hypertension-associated TMA falls within the spectrum of complement-mediated TMA, the prognosis of which is poor. Hence, testing for genetic complement abnormalities is warranted in patients with severe hypertension and TMA on renal biopsy to adopt suitable treatment options and prophylactic measures. Severe hypertension may induce thrombotic microangiopathy (TMA) within the renal vasculature associated with fibrinoid necrosis of arterioles and the glomerular capillary tufts. The exact mechanism remains to be established, but TMA may occur when autoregulation fails to counteract the hypertension-induced shear stress. In those patients with hypertension as the primary pathologic process, aggressive management of blood pressure is effective in resolving acute features of TMA and at least partially restoring renal function.1Zhang B. Xing C. Yu X. et al.Renal thrombotic microangiopathies induced by severe hypertension.Hypertens Res. 2008; 31: 479-483Crossref PubMed Scopus (43) Google Scholar However, numerous other processes may be relevant, particularly in those patients who are not responding to standard treatment and are becoming dialysis-dependent. During the last decade, the alternative pathway (AP) of complement activation has been linked to TMA and in particular to the atypical hemolytic uremic syndrome (aHUS): a rare syndrome of microangiopathic hemolytic anemia, thrombocytopenia, and renal insufficiency. AP is a continuously active immune surveillance and effector system operating in circulation and on the cell surface, which is tightly regulated to prevent damage to the self. In aHUS, AP dysregulation can occur at the endothelial surface, leading to the formation of the terminal complement complex (i.e., C5b-9) and subsequent endothelial cell damage.2de Jorge E.G. Macor P. Paixao-Cavalcante D. et al.The development of atypical hemolytic uremic syndrome depends on complement C5.J Am Soc Nephrol. 2011; 22: 137-145Crossref PubMed Scopus (88) Google Scholar, 3Noris M. Galbusera M. Gastoldi S. et al.Dynamics of complement activation in aHUS and how to monitor eculizumab therapy.Blood. 2014; 124: 1715-1726Crossref PubMed Scopus (223) Google Scholar AP dysregulation can be due to mutations in genes that either regulate or activate AP and/or autoantibodies that inhibit complement-regulatory proteins.4Noris M. Caprioli J. Bresin E. et al.Relative role of genetic complement abnormalities in sporadic and familial aHUS and their impact on clinical phenotype.Clin J Am Soc Nephrol. 2010; 5: 1844-1859Crossref PubMed Scopus (698) Google Scholar, 5Maga T.K. Nishimura C.J. Weaver A.E. et al.Mutations in alternative pathway complement proteins in American patients with atypical hemolytic uremic syndrome.Hum Mutat. 2010; 31: E1445-E1460Crossref PubMed Scopus (226) Google Scholar The penetrance of aHUS is incomplete, indicating that a second hit such as hypertension is required for disease manifestations.6Sullivan M. Rybicki L.A. Winter A. et al.Age-related penetrance of hereditary atypical hemolytic uremic syndrome.Ann Hum Genet. 2011; 75: 639-647Crossref PubMed Scopus (29) Google Scholar The prognosis is extremely poor,4Noris M. Caprioli J. Bresin E. et al.Relative role of genetic complement abnormalities in sporadic and familial aHUS and their impact on clinical phenotype.Clin J Am Soc Nephrol. 2010; 5: 1844-1859Crossref PubMed Scopus (698) Google Scholar, 7Bresin E. Daina E. Noris M. et al.for the International Registry of Recurrent and Familial HUS/TTPOutcome of renal transplantation in patients with non-Shiga toxin-associated hemolytic uremic syndrome: prognostic significance of genetic background.Clin J Am Soc Nephrol. 2006; 1: 88-99Crossref PubMed Scopus (185) Google Scholar but blockade of the terminal complement pathway has dramatically improved the clinical outcome.8Legendre C.M. Licht C. Muus P. et al.Terminal complement inhibitor eculizumab in atypical hemolytic-uremic syndrome.N Engl J Med. 2013; 368: 2169-2181Crossref PubMed Scopus (999) Google Scholar, 9Licht C. Greenbaum L.A. Muus P. et al.Efficacy and safety of eculizumab in atypical hemolytic uremic syndrome from 2-year extensions of phase 2 studies.Kidney Int. 2015; 87: 1061-1073Abstract Full Text Full Text PDF PubMed Scopus (275) Google Scholar, 10Fakhouri F. Hourmant M. Campistol J.M. et al.Terminal complement inhibitor eculizumab in adult patients with atypical hemolytic uremic syndrome: a single-arm, open-label trial.Am J Kidney Dis. 2016; 68: 84-93Abstract Full Text Full Text PDF PubMed Scopus (185) Google Scholar In clinical practice, it is often a diagnostic challenge to differentiate hypertension-associated TMA from complement-mediated disease. This is particularly the case in patients presenting without profound hemolysis and/or thrombocytopenia. If one refrains from a complete diagnostic work-up, including a comprehensive search for complement abnormalities, a subset of patients may progress to end-stage renal disease without receiving optimal treatment. Moreover, the correct diagnosis is of utmost importance to adopt suitable prophylactic measures prior to kidney transplantation. Here, we hypothesized that AP dysregulation is an often unrecognized, but treatable cause of hypertension-associated TMA. To test this hypothesis, we in retrospect thoroughly analyzed AP in 9 patients with severe hypertension diagnosed with TMA on renal biopsy. The study included 8 cases without profound hematologic signs of TMA. Furthermore, to explore whether indeed the prognosis of these patients is poor, we evaluated the long-term renal outcome, including the disease course after transplantation. Fourteen consecutive patients who fulfilled the inclusion criteria of hypertension-associated TMA were included from January 2005 onward; 5 patients were excluded because of secondary TMA (antiphospholipid syndrome, n = 1; scleroderma renal crisis, n = 1), immune-complex glomerulonephritis (n = 2), or the lack of DNA material (n = 1). Hence, 9 patients were included. All 9 patients were evaluated at the Maastricht University Medical Centre, and 6 of them were referred from an outside institution. The baseline characteristics have been depicted in Table 1. At the time of presentation, a clinical diagnosis of malignant nephrosclerosis was clinically inferred. In all patients, mild-to-moderate hypertensive retinopathy was found, and papilledema was observed in 1 case (no. 8). Indeed, 7 patients had a known medical history of hypertension, including 2 patients with documented episodes of preeclampsia (nos. 1, 7) and/or malignant hypertension (no. 7). Proteinuria and hematuria were found in patients not presenting with anuria. Renal biopsies revealed characteristic lesions of TMA including endothelial cell swelling, reduplication of the glomerular basement membrane, wrinkling of the glomerular capillary wall, and/or mesangiolysis. Fibrin thrombi were localized in the glomeruli of 6 and in the vasculature of 5 tissue samples. Prominent intimal fibrosis, myxoid intimal alterations, and/or fibrinoid necrosis of the renal arteries were also found, reflecting preexisting severe hypertension. Assays of a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13, or ADAMTS13, at presentation were normal in 5 patients tested (nos. 1–5) and immunological tests were uniformly negative. Furthermore, iatrogenic causes, infections, and pregnancy were ruled out. However, aHUS was not suspected because profound hematologic signs of TMA were lacking in all but 1 patient (no. 3); moreover, none of the patients had a family history consistent with familial aHUS. Thus, a diagnosis of hypertension-associated TMA was established.Table 1Baseline clinical features and laboratory evaluationPatient No.Age (yr)SexBP (mm Hg)SCr (μmol/l)uProt (g/d)uRBCESRDHb (mmol/l)LDH (U/l)MAHAaCases with hemolytic anemia and schistocytes on peripheral blood smear were defined as MAHA.Platelets (×109/l)bLower limit of normal = 130 ×109/l.138.4F184/1401730NANAY5.11800Y224240.3M205/11411952.3YY5.71104Y158337.7M200/1205863.9YY5.32125Y100432.0F180/1201138NANAY5.91486Y142565.0M195/1051621.5YN7.9271N98641.1F180/1203340.7YY7.5291N285728.5F224/12210651.6YY5.1298N228827.9M240/1506731.6YY7.9165N133944.0F220/1206490.4YY8.2339N340BP, blood pressure; ESRD, end-stage renal disease; F, female; Hb, hemoglobin; LDH, lactate dehydrogenase; M, male; MAHA, microangiopathic hemolytic anemia; N, no; NA, not applicable; SCr, serum creatinine; uProt, proteinuria; uRBC, hematuria; Y, yes.a Cases with hemolytic anemia and schistocytes on peripheral blood smear were defined as MAHA.b Lower limit of normal = 130 ×109/l. Open table in a new tab BP, blood pressure; ESRD, end-stage renal disease; F, female; Hb, hemoglobin; LDH, lactate dehydrogenase; M, male; MAHA, microangiopathic hemolytic anemia; N, no; NA, not applicable; SCr, serum creatinine; uProt, proteinuria; uRBC, hematuria; Y, yes. DNA samples were tested; genetic AP abnormalities were identified in 6 (67%) of 9 patients, most of which were found in heterozygosity (Table 2). The mutated genes included C3, CFI, CFH, and CD46. The C3 and CFI mutations included the missense variants c.481C>T (R161W) and c.452A>G (N151S), respectively. The CFH mutations included c.2558G>A (C853R) and c.2850G>T (Q950H), the latter of which is a rare variant of unknown significance present also in the normal population. The CD46 mutation included the 6 base-pair deletion c.811_816delGAGACT (ΔD237/S238). Four patients carried the -332C>T and c.2808G>T single nucleotide polymorphisms that tag the CFH-H3 haplotype.11Caprioli J. Castelletti F. Bucchioni S. et al.for the International Registry of Recurrent and Familial HUS/TTPComplement factor H mutations and gene polymorphisms in haemolytic uraemic syndrome: the C-257T, the A2089G and the G2881T polymorphisms are strongly associated with the disease.Hum Mol Genet. 2003; 12: 3385-3395Crossref PubMed Scopus (270) Google Scholar, 12Bresin E. Rurali E. Caprioli J. et al.for the European Working Party on Complement Genetics in Renal DiseasesCombined complement gene mutations in atypical hemolytic uremic syndrome influence clinical phenotype.J Am Soc Nephrol. 2013; 24: 475-486Crossref PubMed Scopus (261) Google Scholar Also, the homozygous genomic deletion of CFHR1 and CFHR3 was identified in 1 patient, and circulating factor H autoantibodies were not found.Table 2Complement abnormalitiesPatient No.Mutation(s)CFH-H311Caprioli J. Castelletti F. Bucchioni S. et al.for the International Registry of Recurrent and Familial HUS/TTPComplement factor H mutations and gene polymorphisms in haemolytic uraemic syndrome: the C-257T, the A2089G and the G2881T polymorphisms are strongly associated with the disease.Hum Mol Genet. 2003; 12: 3385-3395Crossref PubMed Scopus (270) Google ScholarFHAACP (%)aNormal >75%.AP (%)bNormal >40%.sC5b-9 (ng/ml)cNormal 75%.b Normal >40%.c Normal <337 ng/ml.d Genetic abnormality was found in homozygosity. Open table in a new tab AP, functional activity of the alternative pathway; CP, functional activity of the classical pathway; FHAA, factor H autoantibodies; N, no; ND, not determined; Y, yes. At the time of renal biopsy, plasma sC5b-9 levels were measured by enzyme-linked immunosorbent assays (ELISAs) and although functional studies of AP and classical pathway were unremarkable, increased sC5b-9 levels were found in all patients. The upper limit of normal was set at 337 ng/ml (n = 20 healthy control subjects, mean ± 2 SD), whereas sC5b-9 levels ranged from 440 to 4,200 ng/ml in our cohort (Table 2). Renal biopsies were stained for complement components. Tissue specimens of 8 patients were available. Deposits of C3c and C5b-9 were found in patients with complement abnormalities along the vasculature and/or glomerular capillary wall (Figure 1, Supplementary Table S1), confirming complement activation.2de Jorge E.G. Macor P. Paixao-Cavalcante D. et al.The development of atypical hemolytic uremic syndrome depends on complement C5.J Am Soc Nephrol. 2011; 22: 137-145Crossref PubMed Scopus (88) Google Scholar Also, deposits of C4d, a biomarker for complement activation via the classical and/or lectin pathway, colocalized with C3c and C5b-9. Six of 8 tissue specimens revealed entrapment of aspecific IgM along the vasculature and/or glomerular capillary wall, whereas staining for IgG and IgA was negative. Follow-up ranged from 0.2 to 9.3 years (Figure 2). Eight of 9 patients progressed to end-stage renal disease despite aggressive management of hypertension and normalization of blood pressure. It is noteworthy that eculizumab was started in patient 6 when the CFI mutation was found after 3 months of dialysis. Renal function recovered (estimated glomerular filtration rate: 38 ml/min) within a 12-month treatment period. Subsequently, 6 allografts (3 living-unrelated, 1 living-related, 2 cadaveric) were transplanted in 4 patients, all of whom received tacrolimus. TMA recurrence manifested in 4 grafts (Figure 2), either with (no. 3) or without (nos. 1, 2) microangiopathic hemolytic anemia and thrombocytopenia. Blood pressure was tightly regulated, and the presence of donor-specific alloantibodies and infections as endothelium-damaging events were ruled out. Thus, we linked TMA recurrence to AP dysregulation;13Reynolds J.C. Agodoa L.Y. Yuan C.M. Abbott K.C. Thrombotic microangiopathy after renal transplantation in the United States.Am J Kidney Dis. 2003; 42: 1058-1068Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar moreover, the transplant biopsies confirmed complement activation in vivo (Figure 1, Supplementary Table S1). Recurrent disease was associated either with C3 (nos. 1, 3) or CD46/CFH (no. 2) mutations. TMA recurrence however, is uncommon among patients with mutated CD46. CD46 is a transmembrane protein widely expressed in the kidney, and consequently, the expression of CD46 in the allograft is driven by endothelial cells from the donor. Therefore, we screened the living-related donor (no. 2’s mother) and traced the CD46-ΔD237/S238 mutation. Plasma exchange was initiated in all patients with established recurrent disease; however, graft loss occurred in 3 of 4 disease episodes. At the time of transplantation, the genotype of patient 4 was known and up-front eculizumab was started, which prevented disease recurrence. We examined AP in 9 patients who have been diagnosed with hypertension-associated TMA, and although profound hematologic signs of TMA were uncommon, we identified a high prevalence (67%) of genetic defects associated with impaired complement regulation. Elevated plasma levels of sC5b-9, as well as renal deposits of C3c and C5b-9 at the endothelial surface, confirmed complement activation. Furthermore, both the renal survival and outcome after transplantation were poor. Therefore, we conclude that our patients fall within the spectrum of complement-mediated TMA, which is supported by the favorable response to complement blockade in 2 patients. Most patients who present with severe hypertension and advanced renal failure are not biopsied, because a diagnosis of malignant nephrosclerosis is clinically inferred. Here, we demonstrate that in these patients TMA can develop on the background of genetic defects in AP regulation even though profound hematologic abnormalities as seen in aHUS appeared uncommon. AP activation is a physiologic process that is tightly controlled by complement regulatory proteins. Factor I and cofactor molecules such as CD46 (i.e., membrane cofactor protein) and factor H are required to cleave C3b into inactive metabolites. CFI-N151S,14Bienaime F. Dragon-Durey M.A. Regnier C.H. et al.Mutations in components of complement influence the outcome of factor I-associated atypical hemolytic uremic syndrome.Kidney Int. 2010; 77: 339-349Abstract Full Text Full Text PDF PubMed Scopus (137) Google Scholar CD46-ΔD237/S238,15Richards A. Kemp E.J. Liszewski M.K. et al.Mutations in human complement regulator, membrane cofactor protein (CD46), predispose to development of familial hemolytic uremic syndrome.Proc Natl Acad Sci U S A. 2003; 100: 12966-12971Crossref PubMed Scopus (342) Google Scholar and CFH-C853R16Hakobyan S. Tortajada A. Harris C.L. et al.Variant-specific quantification of factor H in plasma identifies null alleles associated with atypical hemolytic uremic syndrome.Kidney Int. 2010; 78: 782-788Abstract Full Text Full Text PDF PubMed Scopus (40) Google Scholar have been associated with a reduced expression of the respective proteins, affecting AP regulation at the endothelial surface. The functional consequence of the CFH-Q950H variant of unknown significance remains speculative, however.17Mohlin F.C. Nilsson S.C. Levart T.K. et al.Functional characterization of two novel non-synonymous alterations in CD46 and a Q950H change in factor H found in atypical hemolytic uremic syndrome patients.Mol Immunol. 2015; 65: 367-376Crossref PubMed Scopus (21) Google Scholar C3-R161W has been associated with a hyperactive C3 convertase, enhancing complement activation on endothelial cells.18Roumenina L.T. Frimat M. Miller E.C. et al.A prevalent C3 mutation in aHUS patients causes a direct C3 convertase gain of function.Blood. 2012; 119: 4182-4191Crossref PubMed Scopus (109) Google Scholar Also, the risk CFH-H3 haplotype was found,11Caprioli J. Castelletti F. Bucchioni S. et al.for the International Registry of Recurrent and Familial HUS/TTPComplement factor H mutations and gene polymorphisms in haemolytic uraemic syndrome: the C-257T, the A2089G and the G2881T polymorphisms are strongly associated with the disease.Hum Mol Genet. 2003; 12: 3385-3395Crossref PubMed Scopus (270) Google Scholar lowering the threshold for TMA onset.12Bresin E. Rurali E. Caprioli J. et al.for the European Working Party on Complement Genetics in Renal DiseasesCombined complement gene mutations in atypical hemolytic uremic syndrome influence clinical phenotype.J Am Soc Nephrol. 2013; 24: 475-486Crossref PubMed Scopus (261) Google Scholar AP dysregulation and the formation of C5b-9 can occur on activated but not on resting endothelial cells,3Noris M. Galbusera M. Gastoldi S. et al.Dynamics of complement activation in aHUS and how to monitor eculizumab therapy.Blood. 2014; 124: 1715-1726Crossref PubMed Scopus (223) Google Scholar underlining the importance of a complement-amplifying condition such as hypertension-induced shear stress.19Yin W. Ghebrehiwet B. Weksler B. Peerschke E.I. Regulated complement deposition on the surface of human endothelial cells: effect of tobacco smoke and shear stress.Thromb Res. 2008; 122: 221-228Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar In our cohort, severe hypertension occurred in patients with evidence of preexisting hypertension, and we therefore propose that hypertension triggered complement activation, leading to overactivation of the AP and the development of TMA. However, no demonstrable genetic mutations were found in 3 of 9 patients, who may have abnormalities in unscreened regions of the complement-associated genes or genes that have not yet been linked to TMA. Recent data demonstrated that plasma levels of sC5b-9 are elevated in the acute phase of complement-mediated TMA.3Noris M. Galbusera M. Gastoldi S. et al.Dynamics of complement activation in aHUS and how to monitor eculizumab therapy.Blood. 2014; 124: 1715-1726Crossref PubMed Scopus (223) Google Scholar, 20Cataland S.R. Holers V.M. Geyer S. et al.Biomarkers of terminal complement activation confirm the diagnosis of aHUS and differentiate aHUS from TTP.Blood. 2014; 123: 3733-3738Crossref PubMed Scopus (112) Google Scholar, 21Volokhina E.B. Westra D. van der Velden T.J. et al.Complement activation patterns in atypical haemolytic uraemic syndrome during acute phase and in remission.Clin Exp Immunol. 2015; 181: 306-313Crossref PubMed Scopus (33) Google Scholar In line with these data, increased sC5b-9 levels were found in all our patients, indicating activation of the terminal complement pathway. Renal biopsies were also stained for complement components to analyze complement activation at the endothelial surface. C3c and C5b-9 deposits were found, confirming complement activation of the early and terminal complement pathways, respectively. Furthermore, we validate the observations by Chua et al.22Chua J.S. Baelde H.J. Zandbergen M. et al.Complement factor C4d is a common denominator in thrombotic microangiopathy.J Am Soc Nephrol. 2015; 26: 2239-2247Crossref PubMed Scopus (76) Google Scholar that C4d deposits are a common denominator in TMA (data not shown); although C4d is a biomarker for complement activation via the classical and/or lectin pathway, the significance of both pathways remains elusive. Hypertension-induced shear stress, however, might have been responsible for complement activation via the classical pathway.19Yin W. Ghebrehiwet B. Weksler B. Peerschke E.I. Regulated complement deposition on the surface of human endothelial cells: effect of tobacco smoke and shear stress.Thromb Res. 2008; 122: 221-228Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar As compared to hypertension-associated TMA,1Zhang B. Xing C. Yu X. et al.Renal thrombotic microangiopathies induced by severe hypertension.Hypertens Res. 2008; 31: 479-483Crossref PubMed Scopus (43) Google Scholar the renal prognosis of aHUS is extremely poor with high rates of end-stage renal disease and disease recurrence after transplantation.4Noris M. Caprioli J. Bresin E. et al.Relative role of genetic complement abnormalities in sporadic and familial aHUS and their impact on clinical phenotype.Clin J Am Soc Nephrol. 2010; 5: 1844-1859Crossref PubMed Scopus (698) Google Scholar, 23Le Quintrec M. Zuber J. Moulin B. et al.Complement genes strongly predict recurrence and graft outcome in adult renal transplant recipients with atypical hemolytic and uremic syndrome.Am J Transplant. 2013; 13: 663-675Crossref PubMed Scopus (189) Google Scholar We therefore questioned to what extent the disease course of our patients with hypertension-associated TMA has been affected by AP dysregulation. In our cohort, patients with genetically confirmed complement abnormalities invariably progressed to end-stage renal disease despite aggressive management of blood pressure, whereas TMA recurrence after transplantation was common and linked to underlying complement abnormalities. Disease recurrence was associated with a CD46 mutation in patient 2, which is usually associated with a favorable graft survival and low recurrence rates.24Noris M. Remuzzi G. Managing and preventing atypical hemolytic uremic syndrome recurrence after kidney transplantation.Curr Opin Nephrol Hypertens. 2013; 22: 704-712Crossref PubMed Scopus (51) Google Scholar Interestingly, the same CD46 mutation was traced in the patient’s mother who donated the kidney, explaining the discrepant disease course after transplantation. TMA, however, did not occur in the donor, providing evidence that additional factors, such as the CFH-H3 haplotype,12Bresin E. Rurali E. Caprioli J. et al.for the European Working Party on Complement Genetics in Renal DiseasesCombined complement gene mutations in atypical hemolytic uremic syndrome influence clinical phenotype.J Am Soc Nephrol. 2013; 24: 475-486Crossref PubMed Scopus (261) Google Scholar are important for the penetrance of disease. In those patients with recurrent disease, graft survival was poor despite plasma exchange. Taken together, our genetic and clinicopathologic findings are consistent with those observed in aHUS, indicating that our patients fall within the spectrum of complement-mediated TMA. The lack of profound hematologic signs of TMA is remarkable, although exceptionally reported in aHUS,25Sallee M. Ismail K. Fakhouri F. et al.Thrombocytopenia is not mandatory to diagnose haemolytic and uremic syndrome.BMC Nephrol. 2013; 14: 3Crossref PubMed Scopus (29) Google Scholar reflecting a more gradual disease course, the progression of which can be affected by triggers such as pregnancy.26Fakhouri F. Roumenina L. Provot F. et al.Pregnancy-associated hemolytic uremic syndrome revisited in the era of complement gene mutations.J Am Soc Nephrol. 2010; 21: 859-867Crossref PubMed Scopus (312) Google Scholar More modest complement activation and dysregulation have also been linked to TMA-like syndromes such as preeclampsia, which may have been the triggering event in 2 of our patients (nos. 1, 7). This suggests that ongoing damage to the endothelial cells within the renal vasculature can occur irrespective of hematologic signs of TMA. Therefore, novel techniques are needed to better determine the level of both the activation and regulation of the AP at the endothelial surface. Landmark trials have demonstrated the dramatic effects of the complement blocker eculizumab in aHUS.8Legendre C.M. Licht C. Muus P. et al.Terminal complement inhibitor eculizumab in atypical hemolytic-uremic syndrome.N Engl J Med. 2013; 368: 2169-2181Crossref PubMed Scopus (999) Google Scholar, 9Licht C. Greenbaum L.A. Muus P. et al.Efficacy and safety of eculizumab in atypical hemolytic uremic syndrome from 2-year extensions of phase 2 studies.Kidney Int. 2015; 87: 1061-1073Abstract Full Text Full Text PDF PubMed Scopus (275) Google Scholar, 10Fakhouri F. Hourmant M. Campistol J.M. et al.Terminal complement inhibitor eculizumab in adult patients with atypical hemolytic uremic syndrome: a single-arm, open-label trial.Am J Kidney Dis. 2016; 68: 84-93Abstract Full Text Full Text PDF PubMed Scopus (185) Google Scholar In our study, eculizumab was initiated in 2 patients. The renal function of patient 6 dramatically improved and dialysis treatment could be stopped within a year. Furthermore, up-front eculizumab treatment prevented disease recurrence in patient 4 who carried a high-risk CFH mutation.23Le Quintrec M. Zuber J. Moulin B. et al.Complement genes strongly predict recurrence and graft outcome in adult renal transplant recipients with atypical hemolytic and uremic syndrome.Am J Transplant. 2013; 13: 663-675Crossref PubMed Scopus (189) Google Scholar The favorable response to complement blockade confirms the pivotal role of AP dysregulation. Future studies, however, are needed to further examine the timing and duration of treatment with complement-inhibiting agents. In conclusion, we believe that screening for abnormalities of the AP, both in kidney biopsies and by genetic testing, is mandatory in all patients with TMA on renal biopsy attributed to severe hypertension. Our finding that AP dysregulation is the key causative factor of renal failure in these patients has a major impact on treatment and prognosis. Consecutive patients with biopsy-proven renal TMA who presented with severe hypertension were recruited from January 2005 onward. Severe hypertension was defined as blood pressure levels of ≥180 mm Hg systolic and/or 120 mm Hg diastolic and evidence of impending or progressive target organ dysfunction secondary to hypertension.27Chobanian A.V. Bakris G.L. Black H.R. et al.Seventh report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure.Hypertension. 2003; 42: 1206-1252Crossref PubMed Scopus (10557) Google Scholar, 28van den Born B.J. van der Hoeven N.V. Groot E. et al.Association between thrombotic microangiopathy and reduced ADAMTS13 activity in malignant hypertension.Hypertension. 2008; 51: 862-866Crossref PubMed Scopus (70) Google Scholar Patients were screened for secondary causes of TMA such as autoimmune diseases, drugs, infections, and pregnancy. ADAMTS13 assays were performed in patients with systemic hemolysis and/or thrombocytopenia to rule out thrombotic thrombocytopenic purpura. Patients without a definite clinical diagnosis were diagnosed as hypertension-associated TMA; these patients were included. Patients with immune-complex glomerulonephritis were excluded. Clinical and laboratory data were documented at the time of renal biopsy and during follow-up. The information was specified in our Limburg Renal Registry and the patient’s medical records.29van Paassen P. van Breda Vriesman P.J. van Rie H. Tervaert J.W. Signs and symptoms of thin basement membrane nephropathy: a prospective regional study on primary glomerular disease—The Limburg Renal Registry.Kidney Int. 2004; 66: 909-913Abstract Full Text Full Text PDF PubMed Scopus (105) Google Scholar The study was approved by the local ethics committee of the Maastricht University Medical Centre and is in accordance with the Declaration of Helsinki. Coding regions of CFH (NM_000186), CFI (NM_000204.3), CD46 (NM_002389.3), CFB (NM_001710.5), and C3 (NM_000064.2) were amplified and screened for mutations and polymorphisms using DNA sequencing.30Westra D. Volokhina E. van der Heijden E. et al.Genetic disorders in complement (regulating) genes in patients with atypical haemolytic uraemic syndrome (aHUS).Nephrol Dial Transplant. 2010; 25: 2195-2202Crossref PubMed Scopus (75) Google Scholar, 31Volokhina E. Westra D. Xue X. et al.Novel C3 mutation p.Lys65Gln in aHUS affects complement factor H binding.Pediatr Nephrol. 2012; 27: 1519-1524Crossref PubMed Scopus (35) Google Scholar Rearrangements in the CFH-CFHR1-5 genomic region were analyzed by multiplex-ligation probe amplification.32Maga T.K. Meyer N.C. Belsha C. et al.A novel deletion in the RCA gene cluster causes atypical hemolytic uremic syndrome.Nephrol Dial Transplant. 2011; 26: 739-741Crossref PubMed Scopus (36) Google Scholar In selected cases, the presence of circulating factor H autoantibodies was assessed by ELISA.33Dragon-Durey M.A. Loirat C. Cloarec S. et al.Anti-factor H autoantibodies associated with atypical hemolytic uremic syndrome.J Am Soc Nephrol. 2005; 16: 555-563Crossref PubMed Scopus (404) Google Scholar At the time of renal biopsy, serum and plasma samples were obtained, processed, and immediately stored at −80°C to prevent in vitro complement activation.34Yang S. McGookey M. Wang Y. et al.Effect of blood sampling, processing, and storage on the measurement of complement activation biomarkers.Am J Clin Pathol. 2015; 143: 558-565Crossref PubMed Scopus (54) Google Scholar AP and classical pathway functional assays (AP50 and CH50, respectively; Eurodiagnostica, Malmö, Sweden) were completed.35Seelen M.A. Roos A. Wieslander J. et al.Functional analysis of the classical, alternative, and MBL pathways of the complement system: standardization and validation of a simple ELISA.J Immunol Methods. 2005; 296: 187-198Crossref PubMed Scopus (232) Google Scholar Furthermore, plasma sC5b-9 levels were determined using a capture ELISA (BD Biosciences, San Diego, CA) according to the manufacturer’s instructions. Renal biopsies were processed for light-, immunofluorescence-, and electron microscopy as described.29van Paassen P. van Breda Vriesman P.J. van Rie H. Tervaert J.W. Signs and symptoms of thin basement membrane nephropathy: a prospective regional study on primary glomerular disease—The Limburg Renal Registry.Kidney Int. 2004; 66: 909-913Abstract Full Text Full Text PDF PubMed Scopus (105) Google Scholar Also, 2-μm frozen sections were analyzed for deposition of C4d and C5b-9 using mouse anti-C4d mAb (Quidel, San Diego, CA) and mouse anti-C5b-9 mAb (Quidel) as primary antibodies; fluorescein isothiocyanate–labeled antimouse Ab (Dako, Glostrup, Denmark) was used as secondary antibody. The deposits were scored on a scale from 0 to 3. PvP has received an unrestricted educational research grant from Alexion Pharmaceuticals Inc., which has not had any influence on the results or interpretations in this article. All the other authors declared no competing interests. We gratefully thank N. Bijnens, E. Geelkens, H. van Rie, and R. Theunissen (Laboratory of Clinical Immunology, Maastricht University Medical Center, Maastricht, the Netherlands). We also thank all participating nephrologists of the Limburg Renal Registry: F. de Heer, M. Krekels, F. Stifft, G. Verseput (Orbis Medical Center, Sittard, the Netherlands); S. Boorsma, W. Grave, J. Huitema, J. Wirtz (St. Laurentius Hospital, Roermond, the Netherlands); N. ter Braak, L. Frenken, and S. Gaertner (Atrium Medical Center, Heerlen, the Netherlands); and M. Christiaans, T. Fung, M. Gelens, J. Kooman, K. Leunissen, E. Litjens, J. van der Net, F. van der Sande, and E. van Duijnhoven (Department of Internal Medicine, Division of Nephrology, Maastricht University Medical Center, Maastricht, the Netherlands). Download .docx (.01 MB) Help with docx files Table S1Renal C3c and C5b-9 deposits. Severe hypertension with renal thrombotic microangiopathy: what happened to the usual suspect?Kidney InternationalVol. 91Issue 6PreviewPatients with atypical hemolytic uremic syndrome (aHUS) and malignant hypertension can both present with concomitant hypertension and thrombotic microangiopathy (TMA), rendering policy decisions complex. Timmermans et al. report that patients with severe hypertension and renal TMA might have unrecognized aHUS with underlying complement abnormalities. Based on this, they assert that all patients presenting with severe hypertension and renal TMA should be evaluated for aHUS. It remains uncertain whether this holds equally true for patients with malignant hypertension and renal TMA. Full-Text PDF Open ArchiveThe perfect stormKidney InternationalVol. 92Issue 1PreviewTimmermans et al. describe 9 patients with malignant hypertension, renal thrombotic microangiopathy (TMA), and complement alternative pathway (AP) dysregulation.1 To their list of complement gene defects, we would add a pathogenic heterozygous factor H–related protein 3 variant (CFHR3-I280KfsX6)2 in a 30-year-old male twin presenting with a picture identical to their study population: native renal biopsy showing acute TMA (Figure 1); microangiopathic hemolytic anemia (as in 4 of 9 patients in Timmermans et al. Full-Text PDF Open Archive
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