Bioactive adrenomedullin (bio-ADM) is associated with endothelial dysfunction in infants and children with complex congenital heart disease undergoing open-heart surgery on cardiopulmonary bypass.

OBJECTIVES: Children with congenital heart disease (CHD) undergoing cardiac surgery on cardiopulmonary bypass (CPB) are at risk for systemic inflammation leading to endothelial dysfunction associated with increased morbidity. Bioactive adrenomedullin (bio-ADM) is a peptide regulating vascular tone and endothelial permeability. The aim of this study was to evaluate the dynamics of plasma bio-ADM in this patient cohort and its role in capillary leak. METHODS: Plasma samples from 73 pediatric CHD patients were collected for bio-ADM measurement at five different timepoints (TP) in the pre-, intra-, and post-operative period. The primary endpoint was a net increase in bio-ADM levels after surgery on CPB. Secondary endpoints included association of bio-ADM levels with clinical signs for endothelial dysfunction. RESULTS: Bio-ADM levels increased after surgery on CPB from pre-operative median of 12 pg/mL (IQR [interquartile range] 12.0-14.8 pg/mL) to a maximum post-operative median of 48.8 pg/mL (IQR 34.5-69.6 pg/mL, p<0.001). Bio-ADM concentrations correlated positively with post-operative volume balance, (r=0.341; p=0.005), increased demand for vasoactive medication (duration: r=0.415; p<0.001; quantity: TP3: r=0.415, p<0.001; TP4: r=0.414, p<0.001), and hydrocortisone treatment for vasoplegia (bio-ADM median [IQR]:129.1 [55.4-139.2] pg/mL vs. 37.9 [25.2-64.6] pg/mL; p=0.034). Patients who required pleural effusion drainage revealed higher bio-ADM levels compared to those who did not (median [IQR]: 66.4 [55.4-90.9] pg/mL vs. 40.2 [28.2-57.0] pg/mL; p<0.001). CONCLUSIONS: Bio-ADM is elevated in children after cardiac surgery and higher levels correlate with clinical signs of capillary leakage. The peptide should be considered as biomarker for endothelial dysfunction and as potential therapeutic target in this indication.

Population-based screening in children for early diagnosis and treatment of familial hypercholesterolemia: design of the VRONI study.

BACKGROUND: Heterozygous familial hypercholesterolemia (FH) represents the most frequent monogenic disorder with an estimated prevalence of 1:250 in the general population. Diagnosis during childhood enables early initiation of preventive measures, reducing the risk of severe consecutive atherosclerotic manifestations. Nevertheless, population-based screening programs for FH are scarce. METHODS: In the VRONI study, children aged 5-14 years in Bavaria are invited to participate in an FH screening program during regular pediatric visits. The screening is based on low-density lipoprotein cholesterol measurements from capillary blood. If exceeding 130 mg/dl (3.34 mmol/l), i.e. the expected 95th percentile in this age group, subsequent molecular genetic analysis for FH is performed. Children with FH pathogenic variants enter a registry and are treated by specialized pediatricians. Furthermore, qualified training centers offer FH-focused training courses to affected families. For first-degree relatives, reverse cascade screening is recommended to identify and treat affected family members. RESULTS: Implementation of VRONI required intensive prearrangements for addressing ethical, educational, data safety, legal and organizational aspects, which will be outlined in this article. Recruitment started in early 2021, within the first months, more than 380 pediatricians screened over 5200 children. Approximately 50 000 children are expected to be enrolled in the VRONI study until 2024. CONCLUSIONS: VRONI aims to test the feasibility of a population-based screening for FH in children in Bavaria, intending to set the stage for a nationwide FH screening infrastructure. Furthermore, we aim to validate genetic variants of unclear significance, detect novel causative mutations and contribute to polygenic risk indices (DRKS00022140; August 2020).

Systematic screening for novel, serologically reactive Hepatitis E Virus epitopes.

BACKGROUND: The National Institutes of Health classified Hepatitis E as an emerging disease since Hepatitis E Virus (HEV) is the major cause of acute hepatitis in developing countries. Interestingly, an increasing number of sporadic cases of HEV infections are described in industrialized countries as zoonosis from domestic livestock. Despite the increasing relevance of this pathogen in clinical virology, commercial antibody assays are mainly based on fragments of HEV open reading frame (ORF) 2 and ORF3. The largest ORF1 (poly-)protein, however, is not part of current testing formats. METHODS: From a synthesized full length HEV genotype 1 cDNA-bank we constructed a complete HEV gene library consisting of 15 respective HEV ORF domains. After bacterial expression and purification of nine recombinant HEV proteins under denaturating conditions serum profiling experiments using 55 sera from patients with known infection status were performed in microarray format. SPSS software assessed the antigenic potential of these nine ORF domains in comparison to seven commercial HEV antigens (genotype 1 and 3) by performing receiver operator characteristics, logistic regression and correlation analysis. RESULTS: HEV antigens produced with our method for serum profiling experiments exhibit the same quality and characteristics as commercial antigens. Serum profiling experiments detected Y, V and X domains as ORF1-antigens with potentially comparable diagnostic significance as the well established epitopes of ORF2 and ORF3. However no obvious additional increase in sensitivity or specificity was achieved in diagnostic testing as revealed by bioinformatic analysis. Additionally we found that the C-terminal domain of the potential transmembrane protein ORF3 is responsible for IgG and IgM seroreactivity. Data suggest that there might be a genotype specific seroreactivity of homologous ORF2-antigens. CONCLUSIONS: The diagnostic value of identified ORF1 epitopes might not necessarily improve sensitivity and specificity, but broaden the overall quality of existing test systems. ORF2 and ORF3-antigens are still commonly used in diagnostic assays and possibly hold the potential to serologically differentiate between genotype 1 and 3 infections. Our systematic approach is a suitable method to investigate HEV domains for their serologic antigenicity. Epitope screening of native viral domains could be a preferable tool in developing new serologic test components.

Population-based screening in children for early diagnosis and treatment of familial hypercholesterolemia: design of the VRONI study.

Familial hypercholesterolemia (FH) is the most frequent monogenic disorder (prevalence 1:250) in the general population. Early diagnosis during childhood enables pre-emptive treatment, thus reducing the risk of severe atherosclerotic manifestations later in life. Nonetheless, FH screening programs are scarce. VRONI offers all children aged 5-14 years in Bavaria a FH screening in the context of regular pediatric visits. LDL-cholesterol (LDL-C) is measured centrally, followed by genetic analysis for FH if exceeding the age-specific 95th percentile (130 mg/dl, 3.34 mmol/l). Children with FH pathogenic variants are treated by specialized pediatricians and offered a FH-focused training course by a qualified training center. Reverse cascade screening is recommended for all first-degree relatives. VRONI aims to prove the feasibility of a population-based FH screening in children and to lay the foundation for a nationwide screening program.

A systematic approach for the identification of novel, serologically reactive recombinant Varicella-Zoster Virus (VZV) antigens.

BACKGROUND: Varicella-Zoster virus causes chickenpox upon primary infection and shingles after reactivation. Currently available serological tests to detect VZV-specific antibodies are exclusively based on antigens derived from VZV-infected cells. RESULTS: We present a systematic approach for the identification of novel, serologically reactive VZV antigens. Therefore, all VZV open reading frames were cloned into a bacterial expression vector and checked for small scale recombinant protein expression. Serum profiling experiments using purified VZV proteins and clinically defined sera in a microarray revealed 5 putative antigens (ORFs 1, 4, 14, 49, and 68). These were rearranged in line format and validated with pre-characterized sera. CONCLUSIONS: The line assay confirmed the seroreactivity of the identified antigens and revealed its suitability for VZV serodiagnostics comparable to commercially available VZV-ELISA. Recombinant ORF68 (gE) proved to be an antigen for high-confidence determination of VZV serostatus. Furthermore, our data suggest that a serological differentiation between chickenpox and herpes zoster may be possible by analysis of the IgM-portfolio against individual viral antigens.

Serologic analysis of the IgG antibody response in children with varicella zoster virus wild-type infection and vaccination.

INTRODUCTION: In contrast to varicella zoster virus (VZV) primary infection, VZV vaccination does not seem to provide lifelong immunity against varicella. Because more people get vaccinated every year, the development of sensitive serological test systems for the detection of protective anti-VZV IgG will become important in the future. METHODS: We have previously developed a novel VZV line assay based on 5 different recombinant VZV antigens. In this study, we compared this novel assay with a commercially available glycoprotein enzyme immunoassay (RIDASCREEN VZV IgG) in detecting anti-VZV IgG of children with previous varicella infection and VZV vaccination. RESULTS: One hundred twenty-five children were included in this study, 72 with a history of varicella infection and 53 with VZV vaccination. Both assays detected anti-VZV IgG antibodies in both study groups with similar sensitivities. The VZV line assay revealed striking differences in the anti-VZV IgG composition against the VZV open reading frames, 4, 14 and 49, between both study groups, indicating that wild-type varicella infection causes a more diverse immune response against VZV than does vaccination. The exploitation of these results enabled the discrimination of both study groups with a sensitivity of 0.93 and a specificity of 0.83, indicating that the serologic differentiation of children with previous varicella infection and VZV vaccination might be possible. CONCLUSION: The VZV line assay enables the detection of anti-VZV IgG with sensitivities comparable to glycoprotein enzyme immunoassays and might be suitable for the serologic discrimination between children with a history of varicella infection and VZV vaccination.