Csuka, Dorottya

Drożdż, Dorota

Miklaszewska, Monika

Pállinger, Éva

Takács, Beáta

Zachwieja, Katarzyna

Repository of the Academy's Library

862 M. Miklaszewska et al.PRACE  KAZUISTYCZNEHemolytic uremic syndrome with Mycoplasma pneumoniae infection and membrane cofactor protein mutation – case reportZespół hemolityczno-mocznicowy w przebiegu zakażenia Mycoplasma pneumoniae oraz mutacją błonowego kofaktora białkowego - opis przypadku 1Department of Pediatric Nephrology, Jagiellonian University Medical College,Cracow, Poland; Head: Prof. Dorota Drożdż MD, PhD.2Semmelweis University, Department of Genetics, Cell- and Immunobiology, Budapest, Hungary Head: Prof. Edit Buzás MD, DSc3Semmelweis University, 3rd Department of Medicine, Research Laboratory and Füst György Complement Diagnostic Laboratory, Budapest, HungaryHead:Prof. Zoltán Prohászka MD, DScAdditional key words:atypical hemolytic uremic syndromemembrane cofactor proteinMycoplasma pneumoniae infectionDodatkowe słowa kluczowe:atypowy zespół hemolityczno-mocznicowybłonowy kofaktor białkowyzakażenie Mycoplasma pneumoniae    Address for correspondence:Dr. Monika MiklaszewskaWielicka St. 26530-663 Kraków, PolandE-mail address: mmiklasz@wp.plMonika MIKLASZEWSKA1Katarzyna ZACHWIEJA1Dorota DROŻDŻ1Éva PÁLLINGER2Beáta TAKÁCS3Ágnes SZILÁGYI3Dorottya CSUKA3Zoltán PROHÁSZKA3 Background: Thrombotic microan-giopathies (TMA) are rare life-threate-ning diseases of various etiologies, making the identification of the spe-cific forms and appropriate treatment difficult. The aim of this work is to present the history of a patient with atypical hemolytic uremic syndrome (aHUS) that developed in the context of Mycoplasma pneumoniae infec-tion.  Case presentation: A 5 - year old, Caucasian, previously healthy girl presented with symptoms of HUS, without preceding diarrhoea and with ongoing upper respiratory tract infection. ADAMTS13 deficiency and presence of Shiga-like toxin producing E. coli (STEC) was excluded, and the diagnosis of aHUS verified. She requ-ired peritoneal dialysis for 4 days and fresh frozen plasma (FFP) treatment was started with good clinical respon-se. Serological investigation for Myco-plasma pneumoniae was positive (IgM) leading to the initiation of clarithromy-cin therapy. The complement profile (classical pathway activity, C3 and C4 serum levels were slightly decreased, no signs of alternative pathway dysre-gulation) was indicative for classical pathway activation and consumption. The genetic screening revealed a novel non-synonymous variation in the CD46 (MCP) gene in heterozygous form that causes a proline to leucine change at codon 155 of the MCP (P155L). The CD46 P155L variation was associa-ted in the samples of the patient and family members with decreased MCP protein expression on the surface of granulocytes. In addition to the P155L mutation, multiple frequent aHUS risk variations were also identified. Conclusion: The diagnosis of aHUS is challenging and is based mainly on the exclusion of ADAMTS13 deficient thrombotic thrombocytopenic purpu-ra (TTP) and typical HUS caused by STEC. Our patient had single-episodic HUS in the context of upper-airway infection, and finally a functionally relevant CD46 (MCP) mutation was identified. The complexity of aHUS, Wstęp: Mikroangiopatie zakrz-epowe  są to rzadkie, zagrażające życiu choroby o zróżnicowanej etiologii, co sprawia, że szczegółowa ich diag-nostyka oraz ustalenie optymalnego postępowania terapeutycznego są niezwykle trudne.  Celem pracy jest przedstawienie przypadku dziewczynki z atypowym zespołem hemolityczno – mocznicowym (aZHM), do którego rozwoju doszło w trakcie zakażenia Mycoplasma pneumoniae.  Opis przy-padku: 5 – letnia, dotychczas zdrowa dziewczynka, rasy kaukaskiej została przyjęta do Kliniki Nefrologii Dziecięcej z objawami ZHM, bez poprzedzającej biegunki, z objawami trwającej in-fekcji górnych dróg oddechowych. Po wykluczeniu niedoboru ADAMTS13 oraz zakażenia STEC (Shiga-like toxin producing E. coli)  - postawiono rozpoz-nanie aZHM. Dziewczynka przez 4 doby wymagała leczenia dializą otrzewnową, ponadto zastosowano wlewy świeżo mrożonego osocza (FFP) z dobrym re-zultatem.  W badaniach serologicznych stwierdzono obecność przeciwciał przeciwko Mycoplasma pneumo-niae w klasie IgM i włączono leczenie klarytromycyną.  Badanie składowych układu dopełniacza  (stwierdzono nieznacznie obniżone aktywności C3c i C4, bez cech wskazujących na zaburzenia w drodze alternaty-wnej) - wskazywało na aktywację drogi klasycznej. Badanie genetyczne wykazało do tej pory nie opisaną, heterozygotyczną mutację w genie CD46 (MCP) powodującą zamianę pro-liny na leucynę w kodonie 155 białka MCP (P155L). Mutacja CD46 P155L jest związana z obniżeniem ekspresji błonowego kofaktora białkowego (membrane cofactor protein – MCP, CD46)  na powierzchni granulocytów u pacjentki i członków jej rodziny. Ponadto, w tej rodzinie stwierdzono liczne dodatkowe polimorfizmy ge-netyczne  będące czynnikami ryzyka aZHM. Wnioski: Diagnostyka  aZHM jest trudna i w praktyce klinicznej opi-era się na wykluczeniu niedoboru AD-AMTS13 (czyli plamicy małopłytkowej 863Przegląd Lekarski  2016 / 73 / 11and the importance of the requirement for full differential diagnostic workup of all HUS cases is further highlighted by the current case history.zakrzepowej - TTP)  oraz typowego ZHM (STEC-ZHM). W opisanym przypadku - u dziecka miał miejsce jeden epizod ZHM w przebiegu zakażenia górnych dróg oddechowych, a szczegółowa diagnostyka genetyczna doprowadziła do identyfikacji mutacji w białku CD46.  Złożoność  etiologii aZHM oraz kliniczna waga problemu – wymagają rozważenia przeprowadzenia pełnej diagnostyki we wszystkich przy-padkach aZHM.                     BackgroundThrombotic microangiopathies (TMAs) are rare, life-threatening diseases with various etiologies. Clinical features that define HUS include the presence of acute kidney injury, microangiopathic hemolytic anemia and acute thrombocytopenia. Most cases of pediatric HUS (90% of cases) are associated with diarrhea and bacterial infections secreting Shiga-like toxins that provoke endothelial damage. Less frequent-ly various other etiologies may lead to the development of HUS, including invasive Streptococcus pneumoniae infection and Influenza A infection. According to the recently published consensus [1] the term aHUS should be reserved for patients with HUS without coexisting disease. However, the diagnosis of aHUS can be especially challenging, if aHUS triggering factors, such as infections, are also present. In the majority of aHUS patients dysreg-ulation of complement alternative pathway is based on mutations, risk variations or copy number variations in the genes encoding membrane cofactor protein (CD46), factor I (CFI), factor B (CFB), C3 (C3), factor H (CFH), factor H related proteins 1-5 (CFHR1-5) or thrombomodulin (THBD); or is based on the presence of autoantibodies against factor H (DEAP-HUS) [2].  The di-agnosis of aHUS is challenging and based on the exclusion of various clinical states. Here we present the differential diagnostic workup of a patient who presented with a single episode of HUS precipitated by an upper-airway infection caused by Mycoplas-ma pneumoniae.  Case presentation A 5 - year old, previously healthy, Cauca-sian  girl presented to Department of Pediatric Nephrology with vomiting, petechiae on the lower extremities, yellowish sclera, systolic heart murmur, weakness, catarrhal infection and oliguria which had been present for 2 days. In the medical history there had been signs of upper airway infection for 3 days, but no diarrhea. On the admission day the patient developed anuria which lasted for 1 day, there was no elevated blood pressure or signs of pneumonia present, vital param-eters were stable, the general clinical state was satisfactory. Laboratory test results were suggestive for HUS: hemolytic anemia, thrombocytopenia, acute renal failure, ele-vated LDH activity, proteinuria and haema-turia were recorded (see Tab. I). Ultrasound examination revealed slightly enlarged, hyperechogenic kidneys with maintained medullary – cortical differentiation. Microbi-ological, serological and additional specific investigations excluded STEC- or Influenza A infection and ADAMTS13 deficiency (Tab. I), but verified the presence of acute infection with Mycoplasma pneumoniae (MP) by IgM positivity on the second in-hospital day. The diagnosis of atypical HUS (aHUS) was set, the mild MP infection was considered as disease triggering factor, and FFP treatment introduced with clarithromycin therapy. Start-ing on the 5th day renal- and hematological parameters along with LDH activity improved. On the follow-up visit (11 months later) the Table IDetailed laboratory features and therapeutic activity during the whole disease course of the patient.*Further complement parameters, including alternative pathway activity and concentrations of C1q, factor H, factor I, factor B, anti-factor H IgG autoantibody were in the reference range (data not shown).Wyniki badań laboratoryjnych oraz postępowanie terapeutyczne u Pacjentki w czasie trwania choroby.*Pozostałe parametry układu dopełniacza, w tym drogi alternatywnej oraz stężenia C1q, czynnika H, czynnika I, czynnika B, przeciwciał IgG przeciw czynnikowi H  - mieściły się w zakresie wartości prawidłowych (dane nie zawarte).   Parameter/Follow-up day 1 (06-12-2014) 2 3 4 5 6 7 8 9 10 11 12 17 40 335Hemoglobin [g/l] 97 91 81 79 69 110 116 116 121 124 113 127Hematocrite [%] 28.9 25.2 27 24 20.8 32.4 35.2 35.5 37.6 36.8 33.3 38.8White Blood Cells/ml 6000 8900 8800 9000 6700 8900 10800 8200 6600 6800 5100 6240Platelet number x1000/ml 15 10 27 109 162 332 422 552 549 322 296 310BUN [mmol/l] 30 22.8 17.8 9.2 6.8 5.8 5.1 4.7Serum creatinine [µmol/l] 121.6 106.8 110 99.2 57.9 66.5 57 41.7 38 38.2 34.8 32.2 38.3LDH [U/L] 7669 4403 2350 1735 1682 1266 819 655 627Urine: Protein [g/l] 12.42 1.4 0 trace 0.71 trace 0 0 0Urine: Erythocytes/hpf 80-120 3 - 10 0-6 10-20 3 - 10 0 - 3 0-3 0-3 0-4Urine: volume [ml] 0 590 750 1330 880 1200 1410 1150 1030 1080 1349 1050 870 normal normalCystatinC [mg/l] 1.76 0.75 0.64 0.72CRP [mg/l] 12.4 28 9.6 <5.0 <5.0Direct antiglobulin test negativeBlood Pressure 116/78107/68102/63111/79108/64138/85102/65139/89143/66132/94124/86131/93112/7495/5986/68FFP transfusion [ml] 240 240 240 240  RBC transfusion [ml] 220  RRT  CAPD CAPD CAPD CAPDComplement measurements*:CH50 (48-103 CH50/ml) C3 (0.9-1.8 g/l )C4 (0.15-0.55 g/l)470.870.10         561.00.13864 M. Miklaszewska et al.patient was in a good clinical condition and her laboratory results are within normal limits. Detailed investigation of the complement profile revealed slightly decreased classical pathway activity, lower C3 and C4 serum levels, whereas alternative pathway compo-nents and regulators were in reference range (Tab. I). Disorders in the complement profile were considered as infection related, but since the diagnosis of aHUS was established, detailed genetic analysis of complement components was initiated. The genetic analyses revealed a novel missense variation in the CD46 gene in heterozygous form (c.464C>T) that causes a proline to leucine amino acid change (P155L) in the MCP protein (Figure 1). Family screening revealed that the P155L mutation was inherited maternally while the healthy sister and the father did not carry the mutation. Since all of the applied in silico prediction tools (SIFT, PROVEAN, PolyPhen-2, MutationTaster) predicted this novel variation as damaging, the functional consequences of this mutation were ex-plored by fluorescence-activated cell sorter analysis (FACS). MCP expression was examined on the white blood cells of the pa-tient and on the cells of her family members proving that the P155L mutation is linked to decreased MCP expression, especially on granulocytes (Fig. 1). Furthermore, the MCPggaac aHUS risk haplotype [3] was present in the patient and in her parents (all heterozygous) as well as in the healthy sister (homozygous). In addition, the patient was found to be homozygous variant for the c.-331C>T poly-morphism and heterozygous for the CFH H3 risk haplotype [4]. DiscussionIn 10-15% of aHUS patients mutations are found in the transmembrane alternative pathway regulator MCP [2]. MCP is ex-pressed on the surface of most nucleated cells including the kidney and endothelial cells, and is protecting cells from comple-ment damage. MCP binds C3b through its extracellular domains, and acts as a cofactor of factor I in the inactivation of C3b into iC3b [5]. Mutations and polymorphisms in CD46 gene may affect the function or expression of the protein, most of them are located in the four extracellular SCR domains and result in reduced MCP surface expression [1]. In the present case the mutation affects the SCR2 domain of MCP, and seems to be functionally relevant causing decreased cell surface expression of MCP. There are a number of considerations on the patient history. First, a complement profile without characteristic disorders of al-ternative pathway dysregulation in the acute disease stage - does not exclude comple-ment mediated aHUS. Second, the complex genetic analysis of all aHUS patients and FigureMolecular genetic and surface expression analysis of membrane cofactor protein (MCP, CD46) in the patient (arrow) and in her family members. Panel A: Parts of chromatograms showing the localization of the CD46 P155L (c.464C>T) variation. Panel B: Family tree with inheritance of the CD46 P155L mutation and additional frequent aHUS risk variations. Panels C (neutrophil granulocytes) and D (lymphocytes) show histograms of MCP protein expression as determined by fluorescence activated cell sorter (FACS) analysis.Genetyczna analiza molekularna oraz ekspresja błonowego kofaktora białkowego (membrane cofactor protein – MCP, CD46) u Pacjentki (strzałka) oraz u jej członków rodziny.Panel A. Obrazy chromatografii obrazujące lokalizację mutacji CD46 P155L (c.464C>T). Panel B. Drzewo genealogiczne obrazujące dziedziczenie mutacji CD46 P155L  oraz dodatkowe, częste czynniki ryzyka aHUS.Panel C, D. Histogramy obrazujące ekspresję MCP na  granulocytach (C) oraz  limfocytach (D) oznaczone metodą FACS (fluorescence activated cell sorter  - cytometria sortowania komórek w polu elektrycznym).  also the family members is indicated to identify predisposing factors and to allow planning of management. Third, this patient had single episodic HUS that developed af-ter Mycoplasma pneumoniae infection, and had multiple genetic predisposing factors in complement regulators. Hence, we classify the disease as complement mediated aHUS, precipitated by an infection. The therapeutic response was prompt and complete in re-sponse to plasma therapy complying with previous experiences [5]. Mycoplasma pneumoniae is a common etiologic agent of community-acquired pneumonia. Some patients may experience extrapulmonary manifestations but HUS as a complication of this infection is extremely rare. It could involve various possible mech-anisms, such as direct effects of pro-inflam-matory cytokines induced by lipoproteins in the bacterial cell and vascular occlusion due to vasculitis or thrombosis. Conclusions Our case history exemplifies the ne-cessity of genetic analysis for all patients having clinically diagnosed aHUS, even if no complement dysregulation can be iden-tified. Remarkably, rare genetic alterations are not the only factors responsible for the development of aHUS, but risk variations or haplotypes are important determinants. Our case history also demonstrates the requirement for an additional environmental factor (infection in our case) to act as dis-ease precipitating trigger in the presence of genetic predisposition.References1. Loirat C, Fakhouri F, Ariceta G, Besbas N, Bitzan M. et al: An international consensus approach to the management of atypical hemolytic uremic syndrome in children. Pediatr Nephrol. 2016; 31: 15-39.2. Sanchez-Corral P, Melgosa M: Advances in under-standing the aetiology of atypical Haemolytic Uraemic Syndrome. Brit J Haematol. 2010; 150: 529-542.3. Esparza-Gordillo J, Goicoechea de Jorge E, Buil A, Carreras Berges L, López-Trascasa M. et al: Predisposition to atypical hemolytic uremic syndrome involves the concurrence of different susceptibility al-leles in the regulators of complement activation gene cluster in 1q32. Hum Mol Genet. 2005; 14: 703-712.4. Córdoba SR, Jorge EG: Translational mini-review series on complement factor H: genetics and disease associations of human complement factor H. Clin Exp Immunol. 2008; 151: 1-13.5. Fremeaux-Bacchi V, Moulton EA, Kavanagh D, Dragon-Durey MA, Blouin J. et al: Genetic and functional analyses of membrane cofactor protein (CD46) mutations in atypical hemolytic uremic syndrome. J Am Soc Nephrol. 2006; 17: 2017-2025.Acknowledgements: We thank I. Szigeti, M. Kókai, Zs Szendrei for technical assistance. This study was supported by Hungarian National Research Fund (OTKA 100687 (ZP).Financial Disclosure: The authors have no financial relationships relevant to this article to disclose.Competing Interest: The authors have no competing interest to disclose.Consent to publish: Authors confirm that they have obtained consent to publish from the participant’s parents to report indi-vidual patient data. 

Hemolytic uremic syndrome with Mycoplasma pneumoniae infection and membrane cofactor protein mutation

Abstract

Similar works

Full text

Available Versions

Repository of the Academy's Library