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Background In atypical hemolytic-uremic syndrome (aHUS), various defects of the complement system have been reported to explain pathophysiology. Therapeutic options for complement inhibition are well-recognized; however, the links between various immune-derived diseases and aHUS are unclear, and their interference with treatment efficacy during long-term complement-blocking therapy is scarcely known. Case-diagnosis/treatment We present a pediatric patient who developed aHUS with acute kidney injury in parallel with the onset of Crohn’s disease (CD), and who required long-term complement-blocking therapy with eculizumab (ECU). Unexpectedly, during the 6-year ECU treatment, an important intra-patient variation of the degree of complement inhibition was observed. In spite of continuous and stable doses of complement-blocking therapy, periods of incomplete blockade were observed in strong association with relapses of CD. When conventional and later biological therapy with adalimumab was introduced, with CD going into remission, complement blockade became complete again. Despite periodically low ECU levels and insufficient complement inhibition, no clinical or hematological signs of aHUS recurrence were detected during CD relapses. Conclusion In aHUS cases secondary to CD, close monitoring of both complement inhibition and serum ECU levels is needed as intestinal disease can interfere with complement-blocking treatment. Increased doses of ECU may be necessary to maintain therapeutic blood levels of ECU and full complement blockade, especially if the intestinal disease is not under control.

Since the establishment of highly active antiretroviral therapy, survival rates have improved among patients with human immunodeficiency virus infection giving them the possibility to become transplant candidates. Recent publications revealed that human immunodeficiency virus‐positive heart transplant recipients' survival is similar to non‐infected patients. We present the case of a 40‐year‐old human immunodeficiency virus infected patient, who was hospitalized due to severely decreased left ventricular function with a possible aetiology of acute myocarditis, that has later been confirmed by histological investigation of myocardial biopsy. Due to rapid progression to refractory cardiogenic shock, extracorporeal membrane oxygenation implantation had been initiated, which was upgraded to biventricular assist device later. On the 35th day of upgraded support, the patient underwent heart transplantation uneventfully. Our clinical experience confirms that implementation of temporary mechanical circulatory support and subsequent cardiac transplantation might be successful in human immunodeficiency virus‐positive patients even in case of new onset, irreversible acute heart failure.

Objective.Rheumatoid arthritis (RA) and ankylosing spondylitis (AS) have been associated with cardiovascular disease. The treatment of arthritis by tumor necrosis factor-α (TNF-α) inhibitors may decrease the serum concentrations of vascular biomarkers. We determined circulating levels of oxidized low-density lipoprotein (oxLDL)/β2 glycoprotein I (β2-GPI) complexes, antibodies to 60 kDa heat shock protein (anti-Hsp60), soluble urokinase plasminogen activator receptor (suPAR), and B-type natriuretic peptide (BNP) fragment in sera of RA and AS patients undergoing anti-TNF treatment.Methods.Fifty-three patients with RA/AS were treated with etanercept or certolizumab pegol for 1 year. Circulating oxLDL/β2-GPI complex (AtherOx), anti-Hsp60 IgG, and BNP8-29 fragment levels were assessed by ELISA. suPAR levels were determined by suPARnostic Quick Triage test. Flow-mediated vasodilation (FMD), carotid intima-media thickness (CIMT), and arterial pulse wave velocity (PWV) were determined by ultrasound.Results.One-year anti-TNF treatment significantly decreased oxLDL/β2-GPI levels, as well as suPAR levels in patients with critically high suPAR levels at baseline. In RA, BNP levels were higher in seropositive vs seronegative patients. Serum levels of these vascular biomarkers variably correlated with lipids, anticitrullinated protein antibodies, rheumatoid factor, and C-reactive protein. CIMT positively correlated with BNP, and PWV with suPAR and anti-Hsp60, whereas FMD inversely associated with anti-Hsp60. In repeated measures ANOVA analysis, disease activity supported the effects of anti-TNF treatment on 12-month changes in oxLDL/β2-GPI. CIMT supported the effects of therapy on changes in anti-Hsp60 and suPAR.Conclusion.These biomarkers may be involved in the pathogenesis of atherosclerosis underlying RA/AS. TNF inhibition variably affects the serum levels of oxLDL/β2-GPI, suPAR, and BNP.

Background Pregnancies in patients with complement gene variant-mediated thrombotic microangiopathy (cTMA) are challenging, and pregnancies in such patients after kidney transplantation (KTX) are even more so. Methods We identified nine pregnancies following KTX of three genetically high-risk cTMA patients enrolled in the Vienna thrombotic microangiopathy cohort. Preventive plasma therapy was used in three pregnancies, and one patient had ongoing eculizumab (ECU) therapy during two pregnancies. Results Seven out of nine pregnancies (78%) resulted in the delivery of healthy children. The other two included one early abortion at gestational Week 12 during ongoing ECU therapy and one late foetal death at gestational Week 33 + 3, most likely not related to complement dysregulation. Kidney transplant function after delivery remained stable in all but one pregnancy. In the aforementioned case, a severe cTMA flare occurred after delivery despite use of preventive plasma infusions. Kidney graft function could be rescued in this patient by ECU. As such, successful pregnancies can be accomplished in kidney transplant recipients (KTRs) with a history of cTMA. We used preemptive plasma therapy or ongoing ECU treatment in selected cases. Conclusions Thus, becoming pregnant can be encouraged in KTRs with native kidney cTMA. Extensive preconception counselling, however, is mandatory in such cases.

In patients with pregnancy-associated complement gene variant–mediated thrombotic microangiopathy (cTMA), terminal complement blockade is used for treatment of cTMA flares during pregnancy or following delivery. We report pregnancy and delivery outcomes of 2 genetically high-risk patients with cTMA, including 1 kidney transplant recipient, during ongoing eculizumab therapy. In both patients, the first manifestation of cTMA occurred independent from pregnancy. One patient has a history of 2 uneventful pregnancies with prophylactic plasma infusions, and the other has a history of early abortion during long-term eculizumab therapy following kidney transplantation. Overall, pregnancy and delivery outcomes under ongoing eculizumab therapy in our 2 patients with preserved kidney function were excellent as compared with other patients reported in the literature. Eculizumab plasma concentrations were maintained in the therapeutic range during pregnancy and were also detectable in cord blood. Results of cord blood analysis showed deficient complement activity, with low factor and regulator levels, most likely reflecting the age of the neonates and presence of eculizumab in cord blood. In conclusion, pregnancy during ongoing eculizumab treatment appeared to be safe in 2 women with a history of high-risk genetic cTMA and excellent kidney function, even following kidney transplantation. Index Words: Atypical hemolytic uremic syndrome (aHUS), complement-mediated thrombotic microangiopathy (cTMA), complement system, complement factor, complement blockade, eculizumab, pregnancy, gravidity, childbirth, gestation, kidney transplantation, chronic kidney disease, renal failure

The inadequate adherence of patients whose hyperlipidemia is treated with atorvastatin (ATR) to medical instructions presents a serious health risk. Our aim was to develop a flexible approach based on therapeutic drug monitoring (TDM), nonparametric population pharmacokinetic modeling, and Monte Carlo simulation to differentiate adherent patients from partially and nonadherent individuals in a nonrandomized, unicentric, observational study. Sixty‐five subjects were enrolled. Nonparametric, mixed‐effect population pharmacokinetic models of the sums of atorvastatin and atorvastatin lactone concentrations (ATR+ATRL) and of the concentrations of the acid and lactone forms of ATR and its 2‐ and 4‐hydroxylated pharmacologically active metabolites (ATR+MET) were elaborated by including the TDM results obtained in 128 samples collected from thirty‐nine subjects. Monte Carlo simulation was performed based on the elaborated models to establish the probabilities of attaining a specific ATR+ATRL or ATR+MET concentration in the range of 0.002–10 nmol (mg dose)−1 L−1 at 1–24 h postdose by adherent, partially adherent, and nonadherent patients. The results of the simulations were processed to allow the estimation of the adherence of further 26 subjects who were phlebotomized at sampling times of 2–20 h postdose by calculating the probabilities of attaining the ATR+ATRL and ATR+MET concentrations measured in these subjects in adherent, partially adherent, and nonadherent individuals. The best predictive values of the estimates of adherence could be obtained with sampling at early sampling times. 61.54% and 38.46% of subjects in the adherence testing set were estimated to be fully and partially adherent, respectively, while in all cases the probability of nonadherence was extremely low. The evaluation of patient adherence to ATR therapy based on pharmacokinetic modeling and Monte Carlo simulation has important advantages over the collection of trough samples and the use of therapeutic ranges., We set forth a pharmacokinetics‐based clinical laboratory procedure and decision‐making algorithm for the therapeutic monitoring of atorvastatin concentrations at any time that has passed from drug intake to sample collection.

Complement not only plays a key role in host microbial defense but also modulates the adaptive immune response through modification of T- and B-cell reactivity. Moreover, a normally functioning complement system participates in hematopoiesis, reproduction, lipid metabolism, and tissue regeneration. Because of its powerful inflammatory potential, multiple regulatory proteins are needed to prevent potential tissue damage. In clinical practice, dysregulation and overactivation of the complement system are major causes of a variety of inflammatory and autoimmune diseases ranging from nephropathies, age-related macular degeneration (AMD), and systemic lupus erythematosus (SLE) to graft rejection, sepsis, and multi-organ failure. The clinical importance is reflected by the recent development of multiple drugs targeting complement with a broad spectrum of indications. The recognition of the role of complement in diverse diseases and the advent of complement therapeutics has increased the number of laboratories and suppliers entering the field. This has highlighted the need for reliable complement testing. The relatively rapid expansion in complement testing has presented challenges for a previously niche field. This is exemplified by the issue of cross-reactivity of complement-directed antibodies and by the challenges of the poor stability of many of the complement analytes. The complex nature of complement testing and increasing clinical demand has been met in the last decade by efforts to improve the standardization among laboratories. Initiated by the IUIS/ICS Committee for the Standardization and Quality Assessment in Complement Measurements 14 rounds of external quality assessment since 2010 resulted in improvements in the consistency of testing across participating institutions, while extending the global reach of the efforts to more than 200 laboratories in 30 countries. Worldwide trends of assay availability, usage, and analytical performance are summarized based on the past years’ experiences. Progress in complement analysis has been facilitated by the quality assessment and standardization efforts that now allow complement testing to provide a comprehensive insight into deficiencies and the activation state of the system. This in turn enables clinicians to better define disease severity, evolution, and response to therapy.

Our patient was a 37-year-old woman with Fabry disease (GLA p.R112H) with a medical history of recurrent headache, nausea, vomiting, vertigo, and tobacco use (20 cigarettes/day). Fabry disease was diagnosed in 2005 when she experienced proteinuria, preeclampsia, and hypertension (201/130 mm Hg) during pregnancy (delivered 50 cm, 3.4 kg healthy boy; GLA wild type [WT]). Enzyme replacement therapy was initiated in 2009. The patient enrolled in the phase 3 ATTRACT trial (ClinicalTrials.gov; NCT01218659) and started migalastat in May 2012 while taking hormonal contraceptives. Two years after initiating migalastat, the patient had proteinuria (2166 mg/24 h) without hypertension (131/68 mm Hg), which persisted (788 mg/24 h a month later). Kidney biopsy results were consistent with existing Fabry disease. A serum pregnancy test and ultrasound confirmed pregnancy (18 weeks’ gestation). Migalastat and hormonal contraceptives were stopped; the patient continued to smoke. Fetal MRI was normal at ~29 weeks’ gestation. In October 2014, at 37+ weeks’ gestation, the patient delivered a 45-cm, 2.29-kg healthy girl (GLA WT). Excepting low birth weight, which may be related to the patient’s smoking, pregnancy outcome was normal despite exposure to migalastat for 18 weeks. Migalastat therapy during pregnancy is not advised.

Abstract: Introduction: Previous data showed bacterial infections among diabetic patients to be more serious and frequent, with higher mortality rates in comparison with non-diabetics. Recent investigations, however, are contradictory. Aim: The goal of our prospective, observational study was to compare patients hospitalized on a general medical ward due to community-acquired bacterial infections with type 2 diabetes mellitus (T2DM) to those of non-diabetics (K) by 1) infection localization, 2) spectrum of pathogens, 3) three-month mortality rates. Method: Patients were consecutively involved (T2DM: n = 205, K: n = 202). We characterized the infections, clinical parameters, mortalities of the two groups, and matched them to international data. Results: No difference regarding clinical details of the groups were found except for glycemic parameters and BMI. In the T2DM group the skin- and soft tissue- (37.1%), in the K patients respiratory infections (37.1%) were the most common, followed by urinary ones (31.2% and 31.7%, respectively). Skin- and soft tissue infection incidence among T2DM subjects were higher compared to international results (37.1% vs. 16%). Co-presence of Gram positive and Gram negative bacteria in the skin- and soft tissue infections (23/76 vs. 5/46, p = 0.0149), and polymicrobial origin in the urinary tract infections (34.0% vs. 15.1%, p = 0.0335) were found to be more frequent in T2DM than in K. No difference regarding mortality rates were detected. In T2DM the skin- and soft tissue while in the K group the respiratory infections had the most death counts. Conclusions: We found higher rates of skin- and soft tissue infections among T2DM patients hospitalized on a general medical ward compared to international data. In total we did not find difference regarding three-month mortality between the groups. Our results highlight the importance of primary prevention and shows its inadequacy concerning skin and soft tissue infections among type 2 diabetics in Hungary. Orv Hetil. 2019; 160(41): 1623–1632.

Hereditary thrombotic thrombocytopenic purpura (hTTP) is a rare thrombotic microangiopathy characterized by severe congenital ADAMTS13 deficiency and recurring acute episodes causing morbidity and premature death. Information on the annual incidence and severity of acute episodes in patients with hTTP is largely lacking. This study reports prospective data on 87 patients from the Hereditary TTP Registry (clinicaltrials.gov #NCT01257269) for survival, frequency, and severity of acute episodes from enrollment until December 2019. The 87 patients, followed up for a median of 4.2 years (range, 0.01-15 years), had a median age at overt disease onset and at clinical diagnosis of 4.6 years and 18 years (range, 0.0-70 years for both), respectively. Forty-three patients received regular plasma prophylaxis, whereas 22 did not, and treatment changed over time or was unknown in the remaining 22. Forty-three patients experienced 131 acute episodes, of which 91 (69%) occurred in patients receiving regular prophylaxis. This resulted in an annual incidence of acute episodes of 0.36 (95% confidence interval [CI], 0.29-0.44) with regular plasma treatment and of 0.41 (95% CI, 0.30-0.56) without regular plasma treatment. More than one-third of acute episodes (n = 51) were documented in children 40 years (1.18 [95% CI, 0.88-1.55] vs 0.14 [95% CI, 0.08-0.23]). The prophylactic plasma infusion regimens used were insufficient to prevent acute episodes in many patients. Such regimens are burdensome, and caregivers, patients, and their guardians are reluctant to start regular plasma infusions, from which children particularly would benefit.

Background: Type 2 diabetes mellitus (T2DM) and obesity are risk factors of COVID-19 disease course. Retrospective observational analyses reported improvement in COVID-19 severe outcomes and mortality in T2DM patients using a DPP4 inhibitor. However, little is known about the prognostic value of circulating soluble DPP4 activity in patients with SARS-CoV-2 infection. Methods: We conducted an observational, retrospective cohort study with 184 Hungarian adults. Hospitalized patients with acute COVID-19 disease (n=102) and plasma donors (n=43) recovered from prior SARS-CoV-2 infection were enrolled from April to July, 2020 at two institutions in Budapest. “Non-COVID-19 controls” (n=39) were employed with serum sDPP4 activity determined from samples taken before September, 2019. Duplicate serum sDPP4 activity determinations by microplate base enzyme kinetic assay were successful in 182 individuals from different COVID-19 disease severity groups (acute COVID-19 patients were classified into 4 and plasma donors into 2 categories based on peak severity). Findings: A significant difference in sDPP4 activity was found among study groups (p

Background Practice patterns of eculizumab use are not well described. We examined indications for, and outcomes of, eculizumab therapy in a tertiary care nephrology center. Methods We used the “Vienna TMA cohort” and the hospital pharmacy database at the Medical University of Vienna to identify patients that received eculizumab treatment between 2012 and 2019. We describe clinical characteristics, details of eculizumab use, and outcomes of patients with complement gene-variant mediated TMA (cTMA), secondary TMA (sTMA) and C3 glomerulopathy (C3G). Results As of December 2019, 23 patients received complement blockade at the Division of Nephrology and Dialysis: 15 patients were diagnosed with cTMA, 6 patients with sTMA and 2 patients with C3G. Causes of sTMA were bone marrow transplantation (n = 2), malignant hypertension, malignant tumor, systemic lupus erythematosus, antiphospholipid syndrome and lung transplantation (each n = 1). Across all indications, patients had a median age of 31 and were predominantly female (78%) and the median duration of treatment was 227 days. Hematological recovery was seen in most patients, while renal response was best in patients with cTMA. Adverse events were recorded in 26%. Conclusions In summary, eculizumab is the treatment of choice for cTMA patients that do not respond to plasma therapy. In patients with sTMA and C3G, the response rates to therapy are much lower and therefore, the decision to start therapy needs to be considered carefully. Graphic abstract

Hematopoietic stem cell transplantation (HSCT)-associated thrombotic microangiopathy (TA-TMA) is a multifactorial complication. Complement dysregulation may play an important role in the pathogenesis of TA-TMA. Our previous observations suggested that early increase of soluble C5b-9 (sC5b-9), before the development of other complications, can predict the development of later TA-TMA. The present study aims to validate our earlier findings in an independent cohort enrolling 67 pediatric patients who underwent allogeneic HSCT during the study period (October 2015–January 2019). Five different TA-TMA diagnostic criteria were applied, and all important clinical and laboratory parameters of TA-TMA activity were registered. Complement pathway activities, components and sC5b-9 levels were systematically measured before transplantation and on days 28, 56, and 100 after HSCT. A strong and remarkable association still have been found between early increase of sC5b-9 (10 of 10 patients with TA-TMA vs. 27 of 57 without TA-TMA; P = 0.002) and the development of TA-TMA during 100 days post-transplantation. An increase in sC5b-9 concentration had 100% sensitivity and 53% specificity for TA-TMA in the cohort. All TA-TMA cases have been observed during cyclosporine immunosuppression, no TA-TMA was diagnosed during tacrolimus or mycophenolat mofetil therapy. In the majority of patients TA-TMA was mild and self-limiting, without any signs of organ damage. No additional complement parameters were closely associated with the development of TA-TMA. Early raise of the sC5b-9 activation marker was predictive for later development of TA-TMA throughout the whole study period. In patients with a marked increase, early and frequent monitoring of TA-TMA activity markers should be attempted, to facilitate subsequent therapy decisions in time. However, patients with TA-TMA were only identified during or after cyclosporine immunosuppression. Further studies enrolling higher number of patients are necessary to determine the role of immunosuppression in the pathogenesis of TA-TMA.

Background and Aims Practice patterns of Eculizumab use in patients with thrombotic microangiopathies (TMA) and C3-glomerulopathy (C3G) are not well described. Method We used the “Vienna TMA cohort” and the hospital pharmacy database at the Medical University of Vienna to identify adult patients with a history of eculizumab use between 2012 and 2019. We describe clinical characteristics, details of eculizumab use, and outcomes of patients with complement gene variant mediated TMA (cTMA), secondary TMA (sTMA) and C3G. Results As of December 2019, 212 individuals were enrolled in the Vienna TMA cohort comprising 51 cTMA, 144 sTMA, and 17 TTP patients. We included also our cohort of 14 patients with C3G for this analysis. 47 patients (22 TMA and 2 C3G, 23 other indications, i.e. paroxysmal nocturnal haemoglobinuria) received at least one dose of eculizumab at the Medical University of Vienna (Figure 1). Table 1 indicates demographic and clinical details of 15 cTMA (29.4% of all cTMA), 7 sTMA (4.9% of all sTMA) and 2 C3G (14.3% of all C3G) patients treated with eculizumab. 60% of cTMA patients showed a rare complement gene variant, while sTMA was ruled out in the remaining 40%. Causes of sTMA were bone marrow transplantation (BMT) (n=2), malignant hypertension, malignoma, systemic lupus erythematodes, antiphospholipid syndrome and lung transplantation (each n=1). One sTMA patient, a BMT recipient, had a donor with a thrombomodulin gene variant. Patients with cTMA had a greater delay from first diagnosis to treatment with eculizumab than the other groups and received maintenance therapy for a longer period of time. More female patients received eculizumab as compared to male patients. Chronic kidney disease stage improved in 60% and 43% of cTMA and sTMA patients, respectively. TMA relapses did not occur during administration of eculizumab. The 2 patients with C3G didn’t respond to eculizumab in our center. Eculizumab therapy was stopped in 66% of patients with cTMA and in all patients presenting with sTMA or C3G. In general, eculizumab was well tolerated and we did not observe life threatening infections of our patients. Three adverse drug reactions included exanthema, liver injury, and hypertensive emergency. Two patients died during therapy with eculizumab (1 cTMA, 1 C3G,) and two after cessation of eculizumab therapy (1 cTMA, 1 sTMA) resulting in a mortality of 16.7%. Conclusion Improvement of CKD stage was achieved in 60% of patients with cTMA and in 43% of patients with sTMA. In our patients with C3G, eculizumab did not improve kidney function. In general, therapy with eculizumab was well tolerated.

INTRODUCTION AND OBJECTIVES Cardiac resynchronization therapy (CRT) is beneficial for selected heart failure (HF) patients, although nonresponse to therapy is still prevalent. We investigated a set of novel biomarkers associated with various pathophysiological pathways of HF. Our purpose was to assess their ability to predict clinical outcomes after CRT. METHODS We studied 136 chronic HF patients undergoing CRT. We measured the plasma levels of fractalkine, pentraxin-3, hepatocyte growth factor (HGF), carbohydrate antigen-125, and matrix metalloproteinase-9 before and 6 months after CRT. The primary endpoint of the study was 5-year all-cause mortality, and we considered the absence of 6-month reverse remodelling (defined as at least a 15% decrease in end-systolic volume) as a secondary endpoint. RESULTS Fifty-eight patients died during the 5-year follow-up period and 66 patients were categorized as nonresponders. In multivariable models, only an increased HGF was an independent predictor of both mortality (HR, 1.35; 95%CI, 1.11-1.64; P=.003; per 1 standard deviation increase) and the absence of reverse remodelling (OR, 1.83; 95%CI, 1.10-3.04; P=.01; per 1 standard deviation increase). Applying HGF to the basic multivariable model of both mortality (net reclassification improvement=0.69; 95%CI, 0.39-0.99; P

Introduction. We shall present a case of a two-month old infant who has developed a haemolytic uremic syndrome as an atypical complication of Bordetella pertussis infection. The observation that the development of haemolytic uremic syndrome is a late complication of Bordetella pertussis infection may be a clue for further studies. Case outline. A two-month-old female infant was admitted to the hospital because of fever, intensive cough, shortness of breath and poor feeding. Real-time polymerase chain reaction (PCR) for Bordetella pertussis was positive. A macrolide was introduced in therapy. On the eighth hospital day, the infant?s condition improved, she became afebrile and eupneic. On the 16th hospital day, she developed signs of progressive respiratory distress and oliguric acute kidney injury. Hemolytic uremic syndrome (HUS) was diagnosed, so the therapy with the fresh frozen plasma (FFP) transfusion, therapeutic plasma exchange and peritoneal dialysis was initiated. Levels of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) were decreased, while the levels of factor H, factor B, and factor I were normal. Despite the full supportive and targeted care, severe multiple organ failure had developed and on the 24th hospital day the infant died. Conclusion. Further studies are necessary to identify the mechanism of potential interaction between pertussis toxins, pathophysiology of the infection and the interaction of complement activation, coagulation and the regulation of these cascades.

Sex differences among patients with complement-gene-variant-mediated thrombotic microangiopathy (cTMA) are not well established. We examined demographic and clinical data from female and male patients with a history of cTMA enrolled in the Vienna thrombotic microangiopathy (TMA) cohort. Follow-up was three years after first presentation with cTMA. In this single-center study, we identified 51 patients with a first manifestation of cTMA between 1981 and 2019, 63% were female (p = 0.09). The median age at diagnosis did not differ between females and males. There was also no disparity between the sexes with regard to renal function or the need for renal replacement therapy at presentation. Furthermore, we observed similar use of plasma or eculizumab therapy and a comparable evolution of renal function of female and male patients. More females showed risk haplotypes of complement factor H (CFH) and CD46 (97% vs. 68%, p = 0.01), but there was no difference in the prevalence of rare pathogenic variants in complement-associated genes with regard to sex. In conclusion, the majority of cTMA patients enrolled in the Vienna TMA cohort were female. Clinical presentation and renal function did not differ between the sexes, but females more frequently presented with cTMA risk haplotypes.

Hematopoietic stem cell transplantation (HSCT)–associated thrombotic microangiopathy (TA-TMA) is a multifactorial complication, and its prediction is largely unresolved. Our aim was to analyze changes of complement profile after HSCT to identify potential markers of TA-TMA development. Thirty-three consecutive pediatric patients (9.6 ± 4.4 years old) who underwent allogeneic HSCT due to malignant (n = 17) or nonmalignant (n = 16) indications were included in this study. Graft-versus-host disease (GVHD) was diagnosed using Glucksberg criteria, viral reactivation was monitored, 5 different TA-TMA diagnostic criteria were applied, and all important clinical and laboratory parameters of TA-TMA activity were registered. Complement pathway activities, components and terminal pathway activation marker (sC5b-9) levels were systematically measured before transplantation and on days 28, 56, and 100 after HSCT. During the first 100 days after HSCT, 1 of 33 patients died (day 50, multiple organ failure), whereas 10 subjects met the criteria for TA-TMA, typically on day 61 (range, 16 to 98 days). TA-TMA was preceded by acute GVHD in 3 of 10 patients, by viral reactivation in 2 of 10, or by both in 4 of 10 cases. Baseline sC5b-9 levels did not differ in patients without (200 [interquartile range, 144 to 266] ng/mL), or with (208 [interquartile range, 166 to 271] ng/mL) subsequent TA-TMA; however, on day 28 significant differences were observed (201 [interquartile range, 185 to 290] ng/mL versus 411 [interquartile range, 337 to 471] ng/mL; P = .004). Importantly, all 10 patients with TMA showed increase in sC5b-9 level from baseline level to day 28, whereas in patients without TMA the same tendency was observed for only 9 of 23 patients (P = .031). No additional complement parameters were closely associated with the development of TA-TMA. Development of TA-TMA occurred in 30% of our patients, typically after GVHD and/or viral reactivation. However, early raise of sC5b-9 activation marker was predictive for later development of TA-TMA, and should therefore be considered as an alarming sign necessitating a careful monitoring of all TA-TMA activity markers. Further studies enrolling a higher number of patients are necessary to determine if terminal pathway activation is an independent predictor of TA-TMA.

Mitochondrial mortalin and cytosolic Hsp70 are essential chaperones overexpressed in cancer cells. Our goals were to reproduce our earlier findings of elevated circulating levels of mortalin and Hsp70 in colorectal cancer (CRC) patients with a larger patient cohort, to compare death risk assessment of mortalin, Hsp70, CEA and C19-9 and to assess their prognostic value in various CRC stages. Mortalin, Hsp70, CEA and CA19-9 levels were determined in sera of 235 CRC patients enrolled in the study and followed-up 5 years after surgery. Association between their concentrations and patients' survival was analyzed by Kaplan-Meier estimator and subjected to Cox Proportional hazards analysis. Serum level of mortalin was independent of that of Hsp70, CEA and CA19-9, whereas Hsp70 level weakly correlated with CEA and CA19-9 levels. Improved short-term survival was found in early or advanced disease stages associated with lower mortalin and Hsp70 levels. Cox regression analysis showed a high mortality hazard (HR=3.7, P

Recently, we showed that ADAMTS13 circulates in an open conformation during the acute phase of immune-mediated thrombotic thrombocytopenic purpura (iTTP). Although the cause of this conformational change remains elusive, ADAMTS13 is primarily closed in iTTP patients in remission with ADAMTS13 activity >50% and undetectable anti-ADAMTS13 autoantibodies, as well as after rituximab treatment, suggesting a role for anti-ADAMTS13 autoantibodies. Therefore, immunoglobulin G from 18 acute iTTP patients was purified and added to closed ADAMTS13 in healthy donor plasma. This resulted in open ADAMTS13 in 14 of 18 (78%) samples, proving that anti-ADAMTS13 autoantibodies can induce an open ADAMTS13 conformation. To further elucidate the conformation of ADAMTS13 in iTTP patients, we studied a novel iTTP patient cohort (n = 197) that also included plasma samples from iTTP patients in remission in whom ADAMTS13 activity was 50%, although free anti-ADAMTS13 autoantibodies were not always detected. Thus, open ADAMTS13 is a hallmark of acute iTTP, as well as a novel biomarker that can be used to detect subclinical iTTP in patients in remission. Finally, a long-term follow-up study in 1 iTTP patient showed that the open conformation precedes a substantial drop in ADAMTS13 activity. In conclusion, we have shown that anti-ADAMTS13 autoantibodies from iTTP patients induce an open ADAMTS13 conformation. Most importantly, an open ADAMTS13 conformation is a biomarker for subclinical iTTP and could become an important tool in TTP management. ispartof: BLOOD vol:136 issue:3 pages:353-361 ispartof: location:United States status: published

Background Left ventricular hypertrophy and diastolic dysfunction are common echocardiographic features of both aortic valve stenosis (AS) and cardiac amyloidosis (CA). These two different entities therefore may mask each other. From recent years, there is a growing body of evidence about the relatively high incidence of wild-type transthyretin (wtTTR) amyloidosis in AS, but there are scarce data on the prevalence of AS in CA, particularly in AL-type amyloidosis. The echocardiographic approach to these patients is not obvious, and not evidence based. We aimed to study the prevalence, severity, and type of AS in patients with CA and also to evaluate the potential of echocardiography in the diagnostic process. Methods Between January 2009 and January 2019, we retrospectively analyzed the clinical and echocardiographic data, and the echocardiographic work up of 55 consecutive CA patients. Results 80% of our CA patients had AL amyloidosis. We identified 5 patients (9%) with moderate to severe AS: two with moderate AS and three with low-flow, low-grade AS (LFLG AS). Further analysis of the latter three patients with dobutamine stress echocardiography revealed pseudo-severe LFLG AS in two, and true-severe AS in one patient. Conclusion The prevalence of moderate to severe AS is 9% in our population of CA patients, the majority of whom have AL amyloidosis. Dobutamine echocardiography seems to be appropriate for the further characterization of patients with LFLG AS, even with normal ejection fraction.

Atypical haemolytic uraemic syndrome is an ultra-rare, life-threatening disease. Causative variants in genes that encode complement factors can be identified in 40-70% of cases. We performed genetic analysis of 21 Czech children with atypical haemolytic uraemic syndrome. Genetic or acquired predisposition to the disease was identified in the majority of our patients: CFHR1 and CFHR3 deletions in 14/21 (67%; 13 of them were positive for anti-complement factor H antibodies), variants in complement genes or DGKE in 13/21 (62%). Multiple genetic findings were identified in eight patients (38%). The incidence of atypical haemolytic uraemic syndrome in the Czech paediatric population was estimated to be 0.092 (CI 0.053-0.131) cases per million inhabitants and 0.92 (CI 0.53-1.32) cases per 100,000 births for the entire reporting period. Ten patients were initially treated with plasma exchange and eight with eculizumab or with a combination of eculizumab and plasma exchange. At the last follow-up, 20 patients were alive and one patient had end-stage renal disease.Conclusion: The incidence of atypical haemolytic uraemic syndrome in the Czech paediatric population corresponds to the reported incidence in Europe. We detected the unusually high rate of CFHR1/CFHR3 deletions associated with anti-complement factor H antibodies in Czech paediatric patients. Treatment by eculizumab led to superior outcomes and prevention of the disease relapses compared with plasma exchange therapy. Our results may help to understand the polygenic nature of atypical haemolytic uraemic syndrome as a disease that results from a combination of various risk factors. What is Known: • Atypical haemolytic uraemic syndrome (aHUS) is considered a polygenic and multifactorial disease. Genetic predisposition to aHUS is identified in 40-70% of children. • Anti-complement factor H antibodies are usually found in 6-25% of affected children. What is New: • Potentially causative genetic or acquired factors were confirmed in the majority of patients. The prevailing finding was the unusually high rate of CFHR1/CFHR3 deletions associated with anti-complement factor H antibodies (62% of patients). • The incidence of aHUS in Czech children is 0.092 (CI 0.053-0.131) cases per million inhabitants and 0.92 (CI 0.53-1.32) cases per 100,000 births for the entire reporting period.

Background Atypical hemolytic uremic syndrome (aHUS) frequently leads to renal failure, and kidney transplantation bears a high risk of disease recurrence and graft loss. Methods Patients who received a kidney graft in our center were retrospectively identified using our Vienna Thrombotic Microangiopathy Cohort. Since 2005, the majority of aHUS patients received perioperative plasma exchange (PE) followed by plasma infusions (PI). Patients were switched to eculizumab in case of plasma intolerance or failure. Those with no preemptive therapy served as controls. We used proportional Cox regression and logistic regression to examine predictors of graft survival. Results 19 aHUS patients received 32 grafts and had a follow-up > 1 year. Eight patients received preventive plasma therapy for eight transplants and 13 patients (including 2 patients who received plasma therapy for their last transplant) had no preventive therapy for 24 grafts. The median graft survival was 2.372 days in patients, that received preemptive therapy and 411 days in patients, that did not receive preemptive treatment (hazard ratio: 0.11; p= 0.03). Four patients were switched to eculizumab because of plasma intolerance or failure. Additionally, one patient, that was not transplanted according to the above-mentioned protocol, received eculizumab for aHUS relapse. Additionally, relapse of aHUS (p = 0.01) and year of transplantation (p Conclusions This study shows that preemptive plasma therapy and eculizumab rescue in selected cases improve graft survival among kidney transplant recipients with aHUS.

In patients with typical angina pectoris, inducible myocardial ischaemia and macroscopically normal coronaries (cardiac syndrome X (CSX)), a significantly elevated plasma level of terminal complement complex (TCC), the common end product of complement activation, has been observed without accompanying activation of the classical or the alternative pathways. Therefore, our aim was to clarify the role of the ficolin-lectin pathway in CSX. Eighteen patients with CSX, 37 stable angina patients with significant coronary stenosis (CHD) and 54 healthy volunteers (HC) were enrolled. Serum levels of ficolin-2 and ficolin-3, ficolin-3/MASP-2 complex and ficolin-3-mediated TCC deposition (FCN3-TCC) were determined. Plasma level of TCC was significantly higher in the CSX than in the HC or CHD group (5.45 versus 1.30 versus 2.04 AU/ml, P

Background:Rheumatoid arthritis (RA) and ankylosing spondylitis (AS) have been associated with inflammatory atherosclerosis, increased cardiovascular (CV) morbidity and mortality. Numerous proteins may serve as biomarkers of inflammatory atherosclerosis. The treatment of arthritis by tumour necrosis factor α (TNF-α) inhibitors may decrease the serum concentrations of these biomarkers.Objectives:In this study we wished to determine circulating levels of oxidized LDL (oxLDL) - β2 glycoprotein I (β2GPI) complexes (AtherOx), anti-hsp60 antibodies, soluble urokinase plasminogen activator receptor (sUPAR) and N-terminal B-type natriuretic peptide (NT-proBNP) in sera of RA and AS patients. We also wished to assess the effects of anti-TNF treatment on these biomarkers.Methods:Altogether 53 arthritis patients including 36 RA patients treated with either etanercept (ETN) or certolizumab pegol (CZP) and 17 AS patients treated with ETN were included in a 12-month follow-up study.Circulating oxLDL/β2gpI complexes, anti-human Hsp60 immunoglobulin G (IgG) levels and BNP8-29fragment levels were assessed by ELISA. suPAR levels were assessed by suPARnostic®Quick Triage test. All laboratory assessments were performed at baseline, as well as 6 and 12 months after treatment initiation. Results were associated with DAS28, BASDAI, CRP.Results:In the mixed cohort of 53 arthritis patients, the circulating levels of oxLDL/β2gpI significantly decreased after 12 months of anti-TNF therapy (0.20±0.11 U/ml) compared to baseline (0.24±0.10 U/ml; p=0.014). There was a tendency of non-significant decrease after 6 months (0.23±0.14 U/ml) versus baseline. Anti-Hsp60 antibody levels did not change after 6 months (158.6±138.6 AU/ml) and 12 months (167.3±143.3 AU/ml) compared to baseline (170.3±140.4 AU/ml). Among the patients, 21.2% had low, 36.4% “observe”, 9.1% high and 33.3% critical suPAR levels. suPAR levels showed a tendency of non-significant decrease after 6 months (11.3±17.7 ng/ml) and 12 months (10.3±15.3 ng/ml) versus baseline (11.5±16.4 ng/ml). However, when the four serum level categories described above were considered, suPAR concentrations exerted significant decrease in RA patients with critical suPAR levels (>9ng/ml) (p=0.04). Similarly, BNP fragment levels showed only a tendency of decrease after 6 months (518.2±422.4 pmol/l) and 12 months (484.1±418.2 pmol/l) versus baseline (530.8±441.8 pmol/l). However, serum BNP levels at baseline and after 12 months were significantly increased in CCP positive compared to CCP negative RA patients (baseline: 670.6±323.0 versus 138.0±436.4 pmol/l; p=0.030 and 12 months: 652.9±283.2 versus 456.5±423.1 pmol/l; p=0.021), as well as in RF positive compared to RF negative RA patients (baseline: 680.6±381.6 versus 292.9±198.3 pmol/l; p=0.007 and 12 months: 668.9±346.5 versus 312.2±207.0 pmol/l; p=0.001).Conclusion:One-year anti-TNF therapy significantly decreased circulating oxLDL/β2gpI complex levels. This therapy also decreased suPAR levels in patients with critically high suPAR. BNP fragment levels were associated with seropositivity in RA. These vascular biomarkers may reflect the effects of TNF inhibition on endothelial activation.Acknowledgments:This study was sponsored by an investigator-initiated grant from Pfizer.Disclosure of Interests:Anita Pusztai: None declared, Attila Hamar: None declared, Ágnes Horváth: None declared, Edit Végh: None declared, Nóra Bodnár: None declared, György Kerekes: None declared, Monika Czókolyová: None declared, Szilvia Szamosi: None declared, Levente Bodoki: None declared, Katalin Hodosi: None declared, Andrea Domjan: None declared, Gábor Nagy: None declared, Ibolya Szöllösi: None declared, Luis Lopez Employee of: Retired employee of Corgenix Inc., Eiji Matsuura: None declared, Zoltán Prohászka: None declared, Sándor Szántó: None declared, Zoltán Nagy: None declared, Yehuda Shoenfeld: None declared, Zoltán Szekanecz Grant/research support from: Pfizer, UCB, Consultant of: Sanofi, MSD, Abbvie, Pfizer, Roche, Novertis, Lilly, Gedeon Richter, Amgen, Gabriella Szücs: None declared

Patient: Male, 16 Final Diagnosis: Thrombotic thrombocytopenic purpura Symptoms: Anemia Medication: — Clinical Procedure: — Specialty: Pediatrics and Neonatology Objective: Unusual clinical course Background: Thrombotic thrombocytopenic purpura (TTP) in children is a rare life-threatening syndrome, characterized by microangiopathic hemolytic anemia, thrombocytopenia with renal dysfunction, neurologic symptoms, and fever. TTP is usually caused by deficient activity of von Willebrand factor cleaving protease (ADAMTS13), due to either gene mutations or acquired via anti-ADAMTS13 autoantibodies. It can be triggered by bone marrow or solid organ transplantation, cardiothoracic-, abdominal-, and orthopedic surgeries, infections including very rarely Helicobacter pylori infection. Case Report: Here we report a case of a 16-year-old male with TTP, who presented with thrombocytopenia before an appendectomy. Seven days after surgery, our patient started to vomit, developed melena, and was admitted to our pediatric intensive care unit (PICU) with clinical presentation of shock. Gastroscopy revealed H. pylori positive hemorrhagic gastritis. The patient was treated by erythrocyte transfusions, fresh frozen plasma, human albumin, glucose-electrolyte solutions, vitamin K, platelet transfusion before implantation of central venous catheter, and antibiotics. After 36 hours, we started plasma exchange (PEX). Blood tests showed deficiency of ADAMTS13. Due to the presence of anti-ADAMTS13 autoantibodies, rituximab was administered. Due to generalized tonic-clonic seizures, he was artificially ventilated. Brain MR angiography showed small ischemic cerebro-vascular insult in the arteria cerebri media region. Despite immunosuppressive therapy and PEX, the patient did not improve completely until the H. pylori infection was eradicated. After which, he recovered completely. Conclusions: We present a rare case of TTP accompanied with appendicitis and gastritis caused by H. pylori, where TTP improvement was dependent on H. pylori infection eradication.

BACKGROUND Hsp70 plays important role in the development and progression of cancer. Previously we described the association between serum Hsp70 levels and mortality of colorectal cancer. OBJECTIVE In this new prospective study we aimed to confirm and extend our previous findings in a larger cohort of patients, based on a longer follow-up period. METHODS Two hundred and thirty-two patients diagnosed with colorectal cancer were enrolled in the study. Baseline serum Hsp70 level and classical biomarker levels were measured. Patients were treated according to stage of the tumor and follow-up lasted for a median 46.4 months. RESULTS We found that serum Hsp70 concentrations increase significantly with stage of the disease (1.79; 2.23 and 3.21 ng/ml in stage I+II, III and IV respectively, p= 0.012 and 0.002, Mann-Whitney test) and with other known biomarkers of the disease. We managed to confirm our previous findings that high baseline serum Hsp70 level (> 1.64 ng/ml) predicted poor 5-year survival (risk of death HR: 1.94 CI: 1.294-2.909; univariate; HR: 2.418 CI: 1.373-4.258; multivariate Cox regression analysis) in the whole patient population and also in subgroups of stage IV and stage III disease. The strongest association was observed in women under age of 70 (HR: 8.12, CI: 2.02-35.84; p= 0.004; multivariate Cox regression). The power of this colorectal cancer prognostic model could be amplified by combining Hsp70 levels and inflammatory markers. Patients with high Hsp70, CRP and high baseline WBC or platelet count had 5-times higher risk of death (HR: 5.07 CI: 2.74-9.39, p< 0.0001; and HR: 4.98 CI: 3.08-8.06, p< 0.0001 respectively). CONCLUSIONS These results confirm and validate our previous findings that serum Hsp70 is a useful biomarker of colorectal cancer.

Introduction Prevoius studies have shown an increased risk of cardiovascular disease in rheumatoid arthritis (RA), due to RA-associated inflammation. Different vascular biomarkers, such as anti-hsp65 antibodies, asymmetric dimethylarginine (ADMA) and B-type natriuretic peptide (BNP) have been associated with atherosclerosis and RA. Anti-hsp65 antibodies were found in RA patients and these antibodies linked also to inflammation and atherosclerosis. ADMA is an endogenous competitive inhibitor of NOS and consequential can lead to increased nitrosative and oxidative stress. ADMA has been implicated with atherosclerosis, and also with rheumatoid arthritis. BNP also associated to cardiovascular diseases. Objectives The aim of this study was to assess the effects of anti-TNF therapy on different vascular biomarkers, such as anti-hsp65, ADMA and BNP in patients with RA and their correlation with different laboratory and clinical markers. Methods Altogether 36 RA patients were recruited and treated with either etanercept (ETN) or certolizumab pegol (CZP) in this 12 months follow-up study. Assessments were performed at baseline, at month 6 and 12. Amounts of IgG antibodies reacting with recombinant M. bovis hsp65 (Lionex, Braunschweig, Germany) were measured by ELISA. ADMA was assesed by HPLC with fluorescent detection. BNP fragments were assessed by commercially avaiable ELISA kit (Biomedica, Vienna). In addition, disease activity (DAS28), age, disease duration, CRP, IgM rheumatoid factor (RF), anti-CCP (aCCP) and plasma lipid levels were also measured. Arterial flow-mediated vasodilation (FMD), carotid intima-media thickness (cIMT) and arterial pulse-wave velocity (PWV) were assessed by ultrasound. Results There were no significant changes in the levels of anti-hsp60, ADMA and BNP due to anti-TNF therapy. However, baseline level of BNP is correlated with the levels of RF (R=0.479, p=0.004) and aCCP (R=0.591, p Conclusions BNP levels were significantly higher in RF +compared to RF- patients, which imply that BNP may associate with RF positivity. Specific biomarkers, such as ADMA, anti-hsp60 and BNP may play important role cardiovascular disease in RA. Acknowledgements The work is supported by the GINOP-2.3.2-15-2016-00015/I-KOM TEAMING project. Disclosure of interest None declared

SummaryThrombotic thrombocytopenic purpura (TTP) is characterised by the deficiency of the von Willebrand factor (VWF) cleaving protease (ADAMTS-13). Although several observations indicate an important role of endothelial activation in the pathogenesis of TTP, no reliable endothelial activation markers are available in the clinical management of TTP. Our aim was to investigate the presence of endothelial activation in TTP and to determine its connections with disease activity, therapy and complement activation. We enrolled 54 patients (median age 40.5; 44 females) and 57 healthy controls (median age 34; 30 females),VWF antigen, carboxiterminal-pro-endothelin-1 (CT-proET-1), complement Factor H and complement activation products (C3bBbP and SC5b-9) were measured. In both the acute and remission phase of TTP we found increased CT-proET-1 and VWF levels, while Factor H levels decreased compared with healthy controls. In remission, however, the elevated CT-proET-1 levels showed 22 % decrease when compared with the acute phase in paired samples (p=0.0031), whereas no changes for VWF and Factor H levels were observed. We also found positive correlations between CT-proET-1 levels and alternative pathway activation markers (C3bBbP; p=0.0360; r=0.4299). The data we present here demonstrate a role of endothelium activation in patients with acute TTP. The finding that CT-proET-1 levels decreased in remission compared with the acute phase further supports endothelial involvement. In addition, we show that endothelial activation also correlated with the activation of the alternative complement pathway. The data suggest that complement and endothelium activation jointly contribute to the development of TTP episodes in patients with predisposition to TTP.Supplementary Material to this article is available online at www.thrombosis-online.com.

Aims The low lymphocyte counts and high neutrophil leucocyte fractions have been associated with poor prognosis in chronic heart failure. We hypothesized that the baseline ratio of the neutrophil leucocytes to the lymphocytes (NL ratio) would predict the outcome of chronic heart failure patients undergoing cardiac resynchronization therapy (CRT). Methods and results The qualitative blood counts and the serum levels of N-terminal of the prohormone brain natriuretic peptide (NT-proBNP) of 122 chronic heart failure patients and 122 healthy controls were analysed prospectively in this observational study. The 2-year mortality was considered as primary endpoint and the 6-month reverse remodelling (≥15% decrease in the end-systolic volume) as secondary endpoint. Multivariable regression analyses were applied and net reclassification improvement (NRI) and integrated discrimination improvement (IDI) were calculated. The NL ratio was elevated in chronic heart failure patients when compared with the healthy controls [2.93 (2.12–4.05) vs. 2.21 (1.64–2.81), P < 0.0001]. The baseline NL ratio exceeding 2.95 predicted the lack of the 6-month reverse remodelling [n = 63, odds ratio = 0.38 (0.17–0.85), P = 0.01; NRI = 0.49 (0.14–0.83), P = 0.005; IDI = 0.04 (0.00–0.07), P = 0.02] and the 2-year mortality [n = 29, hazard ratio = 2.44 (1.04–5.71), P = 0.03; NRI = 0.63 (0.24–1.01), P = 0.001; IDI = 0.04 (0.00–0.08), P = 0.02] independently of the NT-proBNP levels or other factors. Conclusion The NL ratio is elevated in chronic heart failure and predicts outcome after CRT. According to the reclassification analysis, 4% of the patients would have been better categorized in the prediction models by combining the NT-proBNP with the NL ratio. Thus, a single blood count measurement could facilitate the optimal patient selection for the CRT.

AIM The aim of this study was to assess the influence of intraoperative cytokine adsorption on the perioperative vasoplegia, inflammatory response and outcome during orthotopic heart transplantation (OHT). METHODS Eighty-four OHT patients were separated into the cytokine adsorption (CA)-treated group or controls. Vasopressor demand, inflammatory response described by procalcitonin and C-reactive protein, and postoperative outcome were assessed performing propensity score matching. RESULTS In the 16 matched pairs, the median noradrenaline requirement was significantly less in the CA-treated patients than in the controls on the first and second postoperative days (0.14 vs 0.3 μg*kg-1 *min-1 , P = .039 and 0.06 vs 0.32 μg*kg-1 *min-1 , P = .047). The inflammatory responses were similar in the two groups. There was a trend toward shorter length of mechanical ventilation and intensive care unit (ICU) stay in the CA-treated group compared to the controls. No difference in adverse events was observed between the two groups. The frequency of renal replacement therapy was less in the CA‐treated patients than in the controls. CONCLUSIONS Intraoperative CA treatment was associated with reduced vasopressor demand with a favorable tendency in length of mechanical ventilation, ICU stay and renal replacement therapy. CA treatment was not linked to higher rates of adverse events.

Distant metastasis is a major cause of colorectal cancer–related death, but the mechanism of tumour progression is not fully understood. There is growing evidence of an interaction between tumour cells and platelets which may influence tumour progression and metastasis formation. Quality and quantity of von Willebrand factor may regulate the interaction between tumour cells and platelets. Our aim was to measure the platelet count, von Willebrand factor antigen (VWF:Ag) levels and ADAMTS13 activity in a large (n = 232) cohort of colorectal cancer patients and to examine their relationships with the stage of the disease and 5-year survival without thrombotic complications using multivariable models. Significantly higher platelet counts (p = 0.005), VWF:Ag levels (p = 0.008) and decreased ADAMTS13 activity (p = 0.006) were observed in patients with metastatic disease. Results of the Kaplan–Meier analysis showed that lower platelet counts (p 1.7, p

Background. Deficient ADAMTS13 activity (TS13:act 50%. However, also iTTP patients in remission with a persistent ( Aim. Determine ADAMTS13 conformation in plasma of iTTP patients during acute TTP and remission when TS13:act is 50% and investigate if anti-ADAMTS13 autoAbs induce conformational changes in ADAMTS13. Methods. TS13:act was determined in 120 iTTP plasma samples from 4 different centers (Marseille, Milan, Budapest, Utrecht). Samples were categorized according to the presence of clinical symptoms (acute versusremission) and their TS13:act in remission (>50%, 10-50%, Results. Of the 120 iTTP plasma samples, 46 were obtained during the acute (clinical signs present) and 74 during the remission phase (clinical signs absent). Further subdividing remission samples showed that TS13:act was >50% in 41, 10-50% in 14 and 50%, confirming our previous results. Interestingly, ADAMTS13 was open in 93% and 89% of remission samples with TS13:act 10-50% and Conclusion. We show that ADAMTS13 is not only in the open conformation in iTTP patient plasma during the acute phase but also in remission when TS13:act is Disclosures Peyvandi: Octapharma US: Honoraria; Kedrion: Consultancy; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Grifols: Speakers Bureau; Sobi: Speakers Bureau; Shire: Speakers Bureau; Novo Nordisk: Speakers Bureau; Octapharma US: Honoraria; Octapharma US: Honoraria; Novo Nordisk: Speakers Bureau; Kedrion: Consultancy; Roche: Speakers Bureau; Novo Nordisk: Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Shire: Speakers Bureau; Sobi: Speakers Bureau; Roche: Speakers Bureau; Roche: Speakers Bureau; Shire: Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Octapharma US: Honoraria; Roche: Speakers Bureau; Roche: Speakers Bureau; Grifols: Speakers Bureau; Sobi: Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Sobi: Speakers Bureau; Shire: Speakers Bureau; Sobi: Speakers Bureau; Novo Nordisk: Speakers Bureau; Shire: Speakers Bureau; Kedrion: Consultancy; Novo Nordisk: Speakers Bureau; Grifols: Speakers Bureau; Grifols: Speakers Bureau; Kedrion: Consultancy; Kedrion: Consultancy; Grifols: Speakers Bureau; Octapharma US: Honoraria. Coppo:Ablynx: Consultancy. Veyradier:LFB: Other: Investigator. Vanhoorelbeke:Shire: Consultancy; Ablynx: Consultancy.

There is a difficulty in the molecular diagnosis of congenital adrenal hyperplasia (CAH) due to the c.955C>T (p.(Q319*), formerly Q318X, rs7755898) variant of the CYP21A2 gene. Therefore, a systematic assessment of the genetic and evolutionary relationships between c.955C>T, CYP21A2 haplotypes and the RCCX copy number variation (CNV) structures, which harbor CYP21A2, was performed. In total, 389 unrelated Hungarian individuals with European ancestry (164 healthy subjects, 125 patients with non-functioning adrenal incidentaloma and 100 patients with classical CAH) as well as 34 adrenocortical tumor specimens were studied using a set of experimental and bioinformatic methods. A unique, moderately frequent (2%) haplotypic RCCX CNV structure with three repeated segments, abbreviated to LBSASB, harboring a CYP21A2 with a c.955C>T variant in the 3'-segment, and a second CYP21A2 with a specific c.*12C>T (rs150697472) variant in the middle segment occurred in all c.955C>T carriers with normal steroid levels. The second CYP21A2 was free of CAH-causing mutations and produced mRNA in the adrenal gland, confirming its functionality and ability to rescue the carriers from CAH. Neither LBSASB nor c.*12C>T occurred in classical CAH patients. However, CAH-causing CYP21A2 haplotypes with c.955C>T could be derived from the 3'-segment of LBSASB after the loss of functional CYP21A2 from the middle segment. The c.*12C>T indicated a functional CYP21A2 and could distinguish between non-pathogenic and pathogenic genomic contexts of the c.955C>T variant in the studied European population. Therefore, c.*12C>T may be suitable as a marker to avoid this genetic confound and improve the diagnosis of CAH.

Background Hereditary angioedema (HAE) caused by C1-inhibitor (C1-INH) deficiency (HAE–C1-INH) is a potentially life-threatening rare disease caused by the decreased activity of C1-INH. Lack of C1-INH leads to overproduction of bradykinin, a potent vasoactive peptide. Although angioedema is induced by bradykinin, the function and activation of endothelial cells (ECs), the targets of bradykinin, have not yet been studied during HAE attacks. Objective We studied whether EC function is altered during HAE attacks in comparison with attack-free intervals. Methods Forty-six consecutive samples obtained during attacks from 18 patients with HAE–C1-INH were compared with inter-attack samples of the same patients. The patients' sera were tested for von Willebrand factor (VWF) antigen, VWF collagen-binding activity, soluble E-selectin, and endothelin-1 levels by using ELISA and BRAHMS Kryptor technologies. Results Levels of all 4 EC markers (VWF antigen, VWF collagen-binding activity, soluble E-selectin, and endothelin-1) were significantly increased during HAE attacks. Their increases were even more obvious in the subgroup of patients without any pre-existing risk factors for endothelial dysfunction. Conclusion In this study we demonstrated that ECs are activated during HAE attacks. Our results might suggest the need for revising the knowledge on the pathogenesis of HAE–C1-INH and for reconsidering the role of ECs as a possible novel therapeutic target in patients with this disease.

Cardiac arrest causes generalized ischemia/hypoxia, and subsequent resuscitation inflicts reperfusion injury, the pathology of which is not fully understood. Moreover, predicting the prognosis of comatose, post-cardiac-arrest patients is a complex clinical challenge. We hypothesized that the extent of complement activation might be a reliable predictor of mortality in this population. Forty-six comatose cardiac arrest patients were enrolled into our prospective cohort study, conducted in a tertiary care university clinic. All subjects were cooled to 32-34 degrees C body temperature for 24 hours and then, allowed to rewarm to normothermia. All patients underwent diagnostic coronary angiography. On admission, at 6 and at 24 hrs, blood samples were taken from the arterial catheter. In these, complement products (C3a, C3, C4d, C4, SC5b9, and Bb) were measured by ELISA in blood samples. Patients were followed-up for 30 days; 22 patients (47.8%) died by the end of this period. We observed that complement activation (determined as the C3a to C3 ratio) was higher in non-survivors than in survivors at each time point. In the multivariate Cox regression analysis, the C3a/C3 ratio determined 24 hours after the initiation of therapeutic hypothermia predicted 30-day mortality regardless of age, sex, and the APACHE II score. Complement activation occurs in post-cardiac arrest patients, and its extent correlates with 30-day survival. The C3a/C3 ratio might prove useful for estimating the prognosis of comatose post-cardiac-arrest patients. This article is protected by copyright. All rights reserved.

Red blood cell distribution width (RDW) is a biomarker quantifying the variability of red blood cell size in peripheral blood. Elevated RDW has been found to be an independent prognostic factor for cardiovascular events. SSc is characterized by generalized micro- and macroangiopathy. Our aim was to investigate RDW as a potential biomarker for the assessment of the severity of vascular involvement.One hundred and sixty-eight consecutive SSc patients--62 with dcSSc and 106 with lcSSc--were investigated at baseline and after 1-year of follow-up. Measurements in 93 patients with primary RP and 40 healthy subjects served as controls.The median RDW value of patients with SSc was higher [14.2% (25th-75th percentiles 13.5-14.8%) compared with the group of primary RP patients [13.9% (13.4-14.4%); P0.05) and healthy volunteers [13.6% (13.2-13.8%; P0.01]. dcSSc and anti-topoisomerase antibody-positive cases showed elevated RDW values compared with lcSSc and anti-topoisomerase antibody-negative cases (P0.05). RDW showed a positive correlation with inflammatory markers, including ESR (P0.05) and CRP (P0.05), and a negative correlation with forced vital capacity (P0.05) and diffusing capacity of the lung for carbon monoxide (DLCO) (P0.05) during the follow-up. An increase in RDW of5% during follow-up was associated with an average 8.9% decrease in left ventricular ejection fraction (LVEF) and 7% in DLCO and these associations were independent of each other.RDW in SSc may represent an integrative measure of multiple pathological processes including extensive vasculopathy, fibrosis or ongoing inflammation. An increase in RDW may indicate an impairment of cardiorespiratory function.

Mortalin/GRP75 is a ubiquitous mitochondrial chaperone related to the cytosolic heat shock protein 70 (HSP70). It protects cells from senescence and apoptosis and is overexpressed in cancer cells. Cell resistance to complement-dependent cytotoxicity depends on mortalin and during complement attack mortalin is released from cells. Our goal was to determine whether cancer patients have circulating mortalin in blood. The significance of mortalin in blood to survival prospects of colorectal cancer patients was evaluated. Occurrence of extracellular soluble HSP70 (sHSP70) is documented. We developed a sensitive ELISA for mortalin. The association between mortalin level and survival was subjected to the Cox proportional hazards analysis (univariate and multivariate analyses). Mortalin concentration in serum of colorectal cancer patients was 10-214 ng/ml. Survival data of the patients were known from an earlier study of sHSP70 in these samples. Cox regression analysis indicated that high mortalin (>60 ng/ml) is a risk factor for shorter survival. Serum levels of sHSP70 and mortalin in patients were independent variables. Concurrence of high sHSP70 and mortalin was associated with rapid disease progression (HR = 4, 2.04-8.45, p < 0.001). Addition of high sHSP70 and mortalin to a baseline model of age, sex and TNM stage, significantly (p < 0.001) enhanced the risk score to 8 (3.26-20.46). This is the first demonstration of circulating mortalin in cancer patients. Analysis of mortalin in blood, and even more so of mortalin and sHSP70, adds a high prognostic value to the TNM stage and will identify colorectal cancer patients at high risk of poor survival.

The recent discovery of mutations in the gene encoding diacylglycerol kinase epsilon (DGKE) identified a novel pathophysiologic mechanism leading to HUS and/or MPGN. We report ten new patients from eight unrelated kindreds with DGKE nephropathy. We combined these cases with all previously published cases to characterize the phenotypic spectrum and outcomes of this new disease entity. Most patients presented with HUS accompanied by proteinuria, whereas a subset of patients exhibited clinical and histologic patterns of MPGN without TMA. We also report the first two patients with clinical and histologic HUS/MPGN overlap. DGKE-HUS typically manifested in the first year of life but was not exclusively limited to infancy, and viral triggers frequently preceded HUS episodes. We observed signs of complement activation in some patients with DGKE-HUS, but the role of complement activation remains unclear. Most patients developed a slowly progressive proteinuric nephropathy: 80% of patients did not have ESRD within 10 years of diagnosis. Many patients experienced HUS remission without specific treatment, and a few patients experienced HUS recurrence despite complete suppression of the complement pathway. Five patients received renal allografts, with no post-transplant recurrence reported. In conclusion, we did not observe a clear genotype-phenotype correlation in patients with DGKE nephropathy, suggesting additional factors mediating phenotypic heterogeneity. Furthermore, the benefits of anti-complement therapy are questionable but renal transplant may be a feasible option in the treatment of patients with this condition.

Atypical HUS (aHUS) is a disorder most commonly caused by inherited defects of the alternative pathway of complement, or the proteins that regulate this pathway, and life-threatening episodes of aHUS can be provoked by pregnancy. We retrospectively and prospectively investigated 27 maternal and fetal pregnancy outcomes in 14 women with aHUS from the Vienna Thrombotic Microangiopathy Cohort. Seven pregnancies (26%) were complicated by pregnancy-associated aHUS (p-aHUS), of which three appeared to be provoked by infection, bleeding, and curettage, and three individuals were considered to have preeclampsia/HELLP syndrome before the definitive diagnosis of p-aHUS was made. Mutations in genes that encode the complement alternative pathway proteins or the molecules that regulate this pathway were detected in 71% of the women, with no relationship to pregnancy outcome. Twenty-one pregnancies (78%) resulted in a live birth, two preterm infants were stillborn, and four pregnancies resulted in early spontaneous abortions. Although short-term renal outcome was good in most women, long-term renal outcome was poor; among the 14 women, four had CKD stage 1-4, five had received a renal allograft, and three were dialysis-dependent at study end. We prospectively followed nine pregnancies of four women and treated six of these pregnancies with prophylactic plasma infusions (one pregnancy resulted in p-aHUS, one intrauterine fetal death occurred, and seven pregancies were uneventful). Our study emphasizes the frequency of successful pregnancies in women with aHUS. Close monitoring of such pregnancies for episodes of thrombotic microangiopathy is essential but, the best strategy to prevent these episodes remains unclear.

BACKGROUND Human fetuin A (AHSG) has been associated with the development of obesity, insulin resistance, type 2 diabetes mellitus, and atherosclerosis. Observations on the role of AHSG rs4918 single-nucleotide polymorphism are contradictory. We investigated the association between variants of rs4918 and parameters of obesity, lipid status, tumor necrosis factor-α (TNFα), adipokines (adiponectin, resistin, leptin), and insulin resistance in healthy persons and in patients with previous myocardial infarction. MATERIAL AND METHODS This was a cross-sectional study comprising cohort 1 (81 healthy individuals) and cohort 2 (157 patients with previous myocardial infarction). We used the allele-specific KASP genotyping assay to detect rs4918 polymorphism. RESULTS In cohort 1, G-nucleotide carriers had significantly lower serum TNFα, adiponectin, and higher leptin concentrations than in non-G carriers. These differences, however, were not observed in cohort 2. In cohort 2, G-carriers had lower BMI and waist circumferences than in non-G carriers. The G allele was more frequent among lean than obese patients (RR=1.067, 95%CI=1.053-2.651, p=0.015). An association between BMI and rs4918 polymorphism was observed among patients without diabetes (CC/CG/GG genotypes: p=0.003, G vs. non-G allele: p=0.008) but not in diabetics. In addition, a strong linearity between BMI and the CC/CG/GG genotypes (association value: 4.416, p=0.036) and the frequency of the G allele (7.420, p=0.006) could be identified. In cohort 2, non-obese, non-diabetic G-carriers still had lower BMI and waist circumferences than in non-G carriers. CONCLUSIONS The rs4918 minor variant is associated with lower TNFα and adiponectin, higher leptin levels in healthy persons, and more favorable anthropomorphic parameters of obesity in cohort 2.

• ANP level is lower in C1-INH HAE patients than in control subjects, and it remains unchanged during edematous attacks.

The chronic inflammation plays an important role in heart failure and complement components might be useful markers of the prognosis. We set out to evaluate their predictive value in the clinical outcomes of patients with cardiac resynchronization therapy (CRT).We determined the complement levels C3, C3a, sC5b-9 and also the N-terminus of the prohormone brain natriuretic peptide (NT-proBNP) of 126 heart failure patients in a prospective, single-center observational study before and 6 months after CRT implantation.CRT reduced the C3a [212.5 (148.2-283.6) vs. 153 (119.8-218.3) ng/mL, p 0.0001] and the sC5b-9 levels [296.9 (234.2-358.8) vs. 255.1 (210.1-319.0) ng/mL, p = 0.0006], but not the total C3 levels [1.43 (1.26-1.61) vs. 1.38 (1.23-1.57) g/L, p = 0.57]. C3a predicted the 5-year mortality of the patients [C3a 165 ng/mL hazard ratio = 4.21 (1.65-10.72), p = 0.003] independent of the NT-proBNP and other factors. After reclassification, we observed a significant net reclassification improvement [NRI = 0.71 (0.43-0.98), p 0.0001] and integrated discrimination improvement [IDI = 0.08 (0.03-0.12), p = 0.0002].In patients with CRT, elevated C3a levels increase the risk of mortality independent of the NT-proBNP levels or other factors. CRT exerts anti-inflammatory effect by reducing the complement activation.

Studies investigating serum ghrelin level in atherosclerosis yielded contradictory results. Interaction of ghrelin with adipocytokines is obscure in cardiovascular diseases. We undertook this study to determine which molecules influence ghrelin level and to see whether post-myocardial infarction (MI) patients have decreased ghrelin levels.In this cross-sectional study, acyl-ghrelin concentration was determined by radioimmunoassay in sera of 171 patients (ages 62 ± 6 years, mean ± SD) with previous MI and 81 age-matched referent subjects. We evaluated the associations of ghrelin with insulin, adiponectin, leptin, resistin, fetuin-A and tumor necrosis factor-alpha (TNF-α).Patients had lower ghrelin levels compared to referent subjects (240.55 ± 59.33 vs. 337.96 ± 30.75 pg/mL, p0.001) even after excluding diabetic and obese patients (240.63 ± 54.08 vs. 337.96 ± 30.75, p0.001). In multivariate analysis, insulin (β = -0.327, p0.001) and adiponectin (β = 0.301, p0.001) determined ghrelin level (R(2) = 0.199, p0.001). There was no association between ghrelin and TNF-α levels. In discriminant analysis using ghrelin, adiponectin, leptin, fetuin-A, resistin and TNF-α, the structure matrix revealed ghrelin and TNF-α as strongest predictors for belonging to the patient group (0.760 and -0.569, respectively). Using these two parameters, 89.7% of cases were correctly classified. Subjects with high TNF-α/ghrelin ratio had 11.25 times higher chance for belonging to the patient group (95% CI 5.80-21.80; χ(2) (1) = 215.6, p0.001)Acylated ghrelin levels are decreased in patients with coronary atherosclerosis, independently of body weight and the presence of type 2 diabetes mellitus. Ghrelin level is determined by elevated insulin and decreased adiponectin levels. Ghrelin alone or in combination with TNF-α may prove to be a novel indicator of coronary atherosclerosis.

Rationale: This article describes a child with a life-threatening multiorgan failure with disseminated intravascular coagulation (DIC) and massive complement consumption. To our knowledge this therapeutic approach was for the first time effectively applied in a pediatric patient. Patient concerns: A 14-month-old boy was presented with a severe, rapidly progressing, life-threatening disease because of sudden onset of fever, hemathemesis, hematuria, and bloody diarrhoea alongside fast spreading hematomas and general corporeal edema. Diagnosis: The most plausible diagnosis in our patient is Clostridium difficile sepsis-induced thrombotic microangiopathy alongside with DIC and consumption coagulopathy. The diagnosis was confirmed by positive C difficile bacteria strain in coproculture, clinical, and laboratory tests affirming DIC and global complement activation and consumption. Interventions: The patient was treated with antibiotics (Metronidazole, Vancomycin), plasmapheresis, dialysis, methylprednisolone, mycophenolate mofetil, and Eculizumab. Outcomes: The child is in fair overall condition in a 2 year follow-up with no complications save chronic renal failure. Lessons: In rare cases of sepsis with massive complement consumption, a case-sensitive Eculizumab therapy may be at least considered after the resolution of life-threatening multiorgan failure. The application of this drug can be performed only after sepsis induced disease is put under control. A fast withdrawal of Eculizumab after control of massive complement consumption is recommended to prevent triggering of second sepsis reactivation.