MAP-2:CD55 chimeric construct effectively modulates complement activation.

The complement system is a complex, tightly regulated protein cascade involved in pathogen defense and the pathogenesis of several diseases. Thus, the development of complement modulators has risen as a potential treatment for complement-driven inflammatory pathologies. The enzymatically inactive MAP-2 has been reported to inhibit the lectin pathway by competing with its homologous serine protease MASP-2. The membrane-bound complement inhibitor CD55 acts on the C3/C5 convertase level. Here, we fused MAP-2 to the four N-terminal domains of CD55 generating a targeted chimeric inhibitor to modulate complement activation at two different levels of the complement cascade. Its biological properties were compared in vitro with the parent molecules. While MAP-2 and CD55 alone showed a minor inhibition of the three complement pathways when co-incubated with serum (IC50MAP-2+CD55 1-4 = 60.98, 36.10, and 97.01 nM on the classical, lectin, and alternative pathways, respectively), MAP-2:CD551-4 demonstrated a potent inhibitory activity (IC50MAP-2:CD55 1-4 = 2.94, 1.76, and 12.86 nM, respectively). This inhibitory activity was substantially enhanced when pre-complexes were formed with the lectin pathway recognition molecule mannose-binding lectin (IC50MAP-2:CD55 1-4 = 0.14 nM). MAP-2:CD551-4 was also effective at protecting sensitized sheep erythrocytes in a classical hemolytic assay (CH50 = 13.35 nM). Finally, the chimeric inhibitor reduced neutrophil activation in full blood after stimulation with Aspergillus fumigatus conidia, as well as phagocytosis of conidia by isolated activated neutrophils. Our results demonstrate that MAP-2:CD551-4 is a potent complement inhibitor reinforcing the idea that engineered fusion proteins are a promising design strategy for identifying and developing drug candidates to treat complement-mediated diseases.

Storage of Transfusion Platelet Concentrates Is Associated with Complement Activation and Reduced Ability of Platelets to Respond to Protease-Activated Receptor-1 and Thromboxane A2 Receptor.

Platelet activation and the complement system are mutually dependent. Here, we investigated the effects of storage time on complement activation and platelet function in routinely produced platelet concentrates. The platelet concentrates (n = 10) were stored at 22 °C for seven days and assessed daily for complement and platelet activation markers. Additionally, platelet function was analyzed in terms of their responsiveness to protease-activated receptor-1 (PAR-1) and thromboxane A2 receptor (TXA2R) activation and their capacity to adhere to collagen. Complement activation increased over the storage period for all analyzed markers, including the C1rs/C1-INH complex (fold change (FC) = 1.9; p < 0.001), MASP-1/C1-INH complex (FC = 2.0; p < 0.001), C4c (FC = 1.8, p < 0.001), C3bc (FC = 4.0; p < 0.01), and soluble C5b-9 (FC = 1.7, p < 0.001). Furthermore, the levels of soluble platelet activation markers increased in the concentrates over the seven-day period, including neutrophil-activating peptide-2 (FC = 2.5; p < 0.0001), transforming growth factor beta 1 (FC = 1.9; p < 0.001) and platelet factor 4 (FC = 2.1; p < 0.0001). The ability of platelets to respond to activation, as measured by surface expression of CD62P and CD63, decreased by 19% and 24% (p < 0.05) for PAR-1 and 69-72% (p < 0.05) for TXA2R activation, respectively, on Day 7 compared to Day 1. The extent of platelet binding to collagen was not significantly impaired during storage. In conclusion, we demonstrated that complement activation increased during the storage of platelets, and this correlated with increased platelet activation and a reduced ability of the platelets to respond to, primarily, TXA2R activation.

Natural Autoantibodies in Biologic-Treated Rheumatoid Arthritis and Ankylosing Spondylitis Patients: Associations with Vascular Pathophysiology.

Cardiovascular (CV) morbidity and mortality have been associated with rheumatoid arthritis (RA) and ankylosing spondylitis (AS). Natural autoantibodies (nAAb) are involved in innate immunity, as well as autoimmunity, inflammation, and atherosclerosis. There have not been any studies assessing the effects of biologics on nAAbs in RA and AS, also in relation to vascular pathophysiology. Fifty-three anti-TNF-treated RA and AS patients were included in a 12-month follow-up study. Anti-citrate synthase (CS) and anti-topoisomerase I fragment 4 (TOPO-F4) IgM and IgG levels were determined by ELISA. Ultrasonography was performed to assess brachial artery flow-mediated vasodilation (FMD), common carotid intima-media thickness (ccIMT), and arterial pulse-wave velocity (PWV). Other variables were also evaluated at baseline and 6 and 12 months after treatment initiation. Anti-TNF therapy improved FMD in RA and PWV in AS and stabilized ccIMT. TNF inhibition increased anti-CS IgM and IgG, and possibly also anti-TOPO-F4 IgG levels. Various correlation analyses revealed that nAAbs might be independently involved in autoimmunity as well as changes in inflammation and vascular pathology over time in biologic-treated patients (p < 0.05). We also found associations between anti-TOPO-F4 IgG and anti-Hsp60 IgG (p < 0.05). Baseline nAAb levels or nAAb level changes might determine changes in CRP, disease activity, FMD, PWV, and ccIMT over time (p < 0.05). The interplay between arthritis and inflammatory atherosclerosis, as well as the effects of anti-TNF biologics on these pathologies, might independently involve nAAbs.

Differentiating between activation via the lectin or the classical complement pathway in patients with systemic lupus erythematosus.

Complement activation is a hallmark of systemic lupus erythematosus (SLE) and can proceed through the classical (CP), lectin (LP), or alternative pathway (AP). When managing SLE patients, pathway-specific complement activation is rarely monitored as clinical assays are unavailable. In this study, we aim to differentiate between CP- or LP-mediated complement activation in SLE patients by quantifying pathway-specific protein complexes, namely C1s/C1-inhibitor (C1-INH) (CP-specific activation) and MASP-1/C1-INH (LP-specific activation). Levels for both complexes were assessed in 156 SLE patients and 50 controls using two newly developed ELISAs. We investigated whether pathway-specific complement activation was associated with disease activity and lupus nephritis (LN). Disease activity stratification was performed using SLEDAI scores assessed at inclusion. C1s/C1-INH concentrations were significantly increased in active SLE patients (SLEDAI ≥6) when compared with SLE patients with low disease activity (SLEDAI <6, P < 0.01) and correlated with SLEDAI score (r = .29, P < 0.01). In active LN, MASP-1/C1-INH plasma concentrations were significantly increased compared with nonactive LN (P = 0.02). No differences in MASP-1/C1-INH plasma concentrations were observed between active SLE patients and patients with low disease activity (P = 0.11) nor did we observe a significant correlation with disease activity (r = 0.12, P = 0.15). Our data suggest that the CP and the LP are activated in SLE. The CP is activated in active SLE disease, whereas activation of the LP might be more specific to disease manifestations like LN. Our results warrant further research into specific complement pathway activation in SLE patients to potentially improve specific-targeted and tailored-treatment approaches.

Annual incidence and severity of acute episodes in hereditary thrombotic thrombocytopenic purpura.

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 <10 years of age at enrollment and were often triggered by infections. Their annual incidence of acute episodes was significantly higher than in patients aged >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.

The Rationale of Complement Blockade of the MCPggaac Haplotype following Atypical Hemolytic Uremic Syndrome of Three Southeastern European Countries with a Literature Review.

We present eight cases of the homozygous MCPggaac haplotype, which is considered to increase the likelihood and severity of atypical hemolytic uremic syndrome (aHUS), especially in combination with additional risk aHUS mutations. Complement blockade (CBT) was applied at a median age of 92 months (IQR 36-252 months). The median number of relapses before CBT initiation (Eculizumab) was two. Relapses occurred within an average of 22.16 months (median 17.5, minimum 8 months, and maximum 48 months) from the first subsequent onset of the disease (6/8 patients). All cases were treated with PI/PEX, and rarely with renal replacement therapy (RRT). When complement blockade was applied, children had no further disease relapses. Children with MCPggaac haplotype with/without additional gene mutations can achieve remission through renal replacement therapy without an immediate need for complement blockade. If relapse of aHUS occurs soon after disease onset or relapses are repeated frequently, a permanent complement blockade is required. However, the duration of such a blockade remains uncertain. If complement inhibition is not applied within 4-5 relapses, proteinuria and chronic renal failure will eventually occur.

Immunosuppressive Therapy of Antibody-Mediated aHUS and TTP.

The recent classification of pediatric thrombotic microangiopathies (TMA) takes into consideration mechanisms of disease for guidance to targeted therapies. We present our experience with seven patients with antibody mediated atypical hemolytic uremic syndrome (aHUS) and thrombotic thrombocytopenic purpura (TTP). Five children had aHUS with antibodies against complement factor H (CFH-ab) and two with TTP with antibodies against metalloproteinase ADAMTS13. In the aHUS cases diagnosed and treated before the eculizumab era, CFH-ab was detected using the ELISA assay. Mutational analysis of selected complement genes was performed. TTP was diagnosed if, in addition to microangiopathic hemolytic anemia and thrombocytopenia, ischemic organ involvement and severe deficiency in ADAMTS13 activity were present. Treatment protocol consisted of plasma exchanges (PE) and steroid pulses, followed by the combination of cyclophosphamide and rituximab to achieve long-term immunosuppression. Four patients with CFH-ab and the TTP patients with ADAMTS13 antibodies came into sustained remission. After a median follow-up of 11.7 (range 7.7-12.9) years without maintenance therapy, no disease recurrence was observed; nevertheless, six patients, two had hypertension and two had proteinuria as a late consequence. One patient, with late diagnosis of CFH-ab and additional genetic risk factors who was treated only with PE and plasma substitution, reached end-stage renal disease and was later successfully transplanted using eculizumab prophylaxis. In the cases of antibody-mediated TMAs, PE and early immunosuppressive treatment may result in sustained remission with preserved kidney function. Further data are needed to establish optimal treatment of anti-FH antibody-associated HUS.

Open ADAMTS13, induced by antibodies, is a biomarker for subclinical immune-mediated thrombotic thrombocytopenic purpura.

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%. The open ADAMTS13 conformation was found during acute iTTP, as well as in patients in remission with ADAMTS13 activity <50% and in half of the patients with ADAMTS13 activity >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.

Exploring red cell distribution width as a biomarker for treatment efficacy in home mechanical ventilation.

BACKGROUND: With the growing practice of home mechanical ventilation, there is a need to identify biological markers for adequate follow-up. Red cell distribution width (RDW) is a promising candidate because it is convenient, objective and may reflect treatment effect over a long period of time. The aim of this study was to explore the possible role of RDW as a marker for home mechanical ventilation in real-life, unselected chronic respiratory patient populations. METHODS: First, we identified characteristic RDW values for mixed case, unselected chronic respiratory failure and home mechanical ventilated patients through retrospective review within our institutional database. Next, we conducted a prospective observational study to identify RDW changes during the first six months of optimized home mechanical ventilation treatment. Adult patients starting home mechanical ventilation were included. Factors affecting RDW change during the first 6 months of treatment were analysed. RESULTS: RDW was elevated in both chronic respiratory failure and home mechanical ventilation patients compared to healthy individuals in the retrospective review. In the prospective study of 70 patients, we found that 55.4% of patients starting home mechanical ventilation have abnormal RDW values which are reduced from 14.7 (IQR = 13.2-16.2)% to 13.5 (IQR = 13.1-14.6)% during the first 6 months of HMV treatment (p < 0.001). RDW improvement correlates with improvement in self-reported health-related quality of life and sleepiness scale scores, as well as physical functional status during the same time frame. RDW proved to be a comparable marker to other parameters traditionally used to evaluate treatment efficacy. CONCLUSIONS: RDW is elevated in chronic respiratory failure patients and is significantly reduced in the first six months of optimized home mechanical ventilation. Although further research is needed to verify if RDW change reflects outcome and how comorbidities influence RDW values, our results suggest that RDW is a promising marker of home mechanical ventilation efficacy. Trial registration This study was approved by and registered at the ethics committee of Semmelweis University (TUKEB 250/2017 and TUKEB 250-1/2017, 20th of December 2017 and 1st of October 2019).

Serum fetuin-A level is independent of Helicobacter pylori postinfection status in systemic lupus erythematosus.

Helicobacter pylori is a common pathogen causing gastric inflammation and malignancy. Fetuin-A is a multifunctional protein that is involved in the regulation of calcification, insulin resistance and inflammation. Reports on serum levels of fetuin-A in acute H. pylori infection are contradictory. We intended to see whether H. pylori post-infection status has a long-term effect on serum fetuin-A levels in a well-characterized series of systemic lupus erythematosus cases. In this cross-sectional study 117 patients with systemic lupus erythematosus were enrolled. Helicobacter infection status and serum fetuin-A concentration were determined by ELISA and radial immunodiffusion, respectively. H. pylori positive patients had higher serum fetuin-A concentration than negative ones: 517 (456-603) vs. 476 (408-544) mg L-1, median (25-75% percentiles), P = 0.020. No other parameters differed between these groups. During univariate regression analysis fetuin-A levels were associated with Erythrocyte sedimentation rate (ESR), White blood cell count (WBC), C-reactive protein (CRP), serum total protein, albumin, and the SLEDAI index at the time of diagnosis but only serum albumin remained a significant determinant in multivariate regression study.

Atypical HUS and Crohn's disease-interference of intestinal disease activity with complement-blocking treatment.

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.

Molecular basis and outcomes of atypical haemolytic uraemic syndrome in Czech children.

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.

["Risk factors of childhood depression" research grant - past, present, future].

The authors summarize the last 10 years of an ongoing collaborative study between the Universities of Szeged and Pittsburgh on early onset major depression. First, the "Risk factors of childhood depression" grant is presented briefly as an initial research study in which the subjects of the current studies were recruited. This is a prominently large clinical sample in the field of child psychiatry even on an international level. In addition to the follow-up of the prognosis of the disorder, recent studies continue to explore the early onset depression in two directions. On the one hand, two studies investigate the role of biobehavioral inflexibility markers in the development of major depression ("Biobehavioral inflexibility and risk for juvenile-onset depression" and "Biobehavioral inflexibility and risk for juvenile-onset depression - renewal grant"). On the other hand, the authors would like to have a better understanding of the possible relationship between the major depression and cardiovascular diseases ("Pediatric depression and subsequent cardiac risk factors: a longitudinal study"). The most significant aims of the three studies will be demonstrated, as well as how the studies were prepared and organized along with the already existing experience concerning research management and involvement of new collaborating partners and experts.

FHR-1 Binds to C-Reactive Protein and Enhances Rather than Inhibits Complement Activation.

Factor H-related protein (FHR) 1 is one of the five human FHRs that share sequence and structural homology with the alternative pathway complement inhibitor FH. Genetic studies on disease associations and functional analyses indicate that FHR-1 enhances complement activation by competitive inhibition of FH binding to some surfaces and immune proteins. We have recently shown that FHR-1 binds to pentraxin 3. In this study, our aim was to investigate whether FHR-1 binds to another pentraxin, C-reactive protein (CRP), analyze the functional relevance of this interaction, and study the role of FHR-1 in complement activation and regulation. FHR-1 did not bind to native, pentameric CRP, but it bound strongly to monomeric CRP via its C-terminal domains. FHR-1 at high concentration competed with FH for CRP binding, indicating possible complement deregulation also on this ligand. FHR-1 did not inhibit regulation of solid-phase C3 convertase by FH and did not inhibit terminal complement complex formation induced by zymosan. On the contrary, by binding C3b, FHR-1 allowed C3 convertase formation and thereby enhanced complement activation. FHR-1/CRP interactions increased complement activation via the classical and alternative pathways on surfaces such as the extracellular matrix and necrotic cells. Altogether, these results identify CRP as a ligand for FHR-1 and suggest that FHR-1 enhances, rather than inhibits, complement activation, which may explain the protective effect of FHR-1 deficiency in age-related macular degeneration.

Coexistence of aortic valve stenosis and cardiac amyloidosis: echocardiographic and clinical significance.

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.

Maternal and Fetal Outcomes of Pregnancies in Women with Atypical Hemolytic Uremic Syndrome.

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.

Early Increase in Complement Terminal Pathway Activation Marker sC5b-9 Is Predictive for the Development of Thrombotic Microangiopathy after Stem Cell Transplantation.

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.

Impact of intraoperative cytokine adsorption on outcome of patients undergoing orthotopic heart transplantation-an observational study.

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 CA­treated 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.

The Phenotypic Spectrum of Nephropathies Associated with Mutations in Diacylglycerol Kinase ε.

The recent discovery of mutations in the gene encoding diacylglycerol kinase ε (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.

Circulating mitochondrial stress 70 protein/mortalin and cytosolic Hsp70 in blood: Risk indicators in colorectal cancer.

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 < 0.001) in patients with both high mortalin and Hsp70 circulating levels. Multivariate analysis showed that high mortalin and Hsp70 significantly enhances risk score over a baseline model of age, number of affected lymph nodes, CEA, CA19-9, disease stage and perioperative therapy. Analysis of mortalin and Hsp70 in CRC patients' sera adds a high prognostic value to TNM stage and to CEA and CA19-9 and identifies patients with lower or higher survival probability in all CRC stages. Determination of mortalin and Hsp70 in blood could be a useful additive prognostic tool in guiding clinical management of patients.