Abstract DNA damage is a constant event in every cell caused by exogenous factors such as ultraviolet and ionizing radiation (UVR/IR) and intercalating drugs, or endogenous metabolic and replicative stress. Proteins of the DNA damage response (DDR) network sense DNA lesions and induce cell cycle arrest, DNA repair, and apoptosis. Genetic defects of DDR or DNA repair proteins can be associated with immunodeficiency, bone marrow failure syndromes, and cancer susceptibility. Although various diagnostic tools are available to evaluate DNA damage, their quality to identify DNA repair deficiencies differs enormously and depends on affected pathways. In this study, we investigated the DDR biomarkers γH2AX (Ser139), p-ATM (Ser1981), and p-CHK2 (Thr68) using flow cytometry on peripheral blood cells obtained from patients with combined immunodeficiencies due to non-homologous end-joining (NHEJ) defects and ataxia telangiectasia (AT) in response to low-dose IR. Significantly reduced induction of all three markers was observed in AT patients compared to controls. However, delayed downregulation of γH2AX was found in patients with NHEJ defects. In contrast to previous reports of DDR in cellular models, these biomarkers were not sensitive enough to identify ARTEMIS deficiency with sufficient reliability. In summary, DDR biomarkers are suitable for diagnosing NHEJ defects and AT, which can be useful in neonates with abnormal TREC levels (T cell receptor excision circles) identified by newborn screening. We conclude that DDR biomarkers have benefits and some limitations depending on the underlying DNA repair deficiency.
Hintergrund Eine geringe Diffusionsbarriere ist essentiell für die Funktion des pulmonalen Gasaustausches. Sowohl bei der alveolären kapillären Dysplasie (ACD) als auch bei der nicht-spezifischen interstitiellen Pneumonie (NSIP) ist diese Funktion kompromittiert durch einen Gewebsumbau an der Schnittstelle von Alveole und Kapillare. Eine direkte molekulare Gegenüberstellung dieser beiden Erkrankungen ist bislang nicht erfolgt.
Background: Primary ciliary dyskinesia (PCD) represents a group of rare hereditary disorders characterized by deficient ciliary airway clearance that can be associated with laterality defects. We aimed to describe the underlying gene defects, geographical differences in genotypes and their relationship to diagnostic findings and clinical phenotypes. Methods: Genetic variants and clinical findings (age, sex, body mass index, laterality defects, FEV1) were collected from 19 countries using the ERN LUNG International PCD Registry. Genetic data were evaluated according to ACMG guidelines. We assessed regional distribution of implicated genes and genetic variants as well as genotype correlations with laterality defects and FEV1. Findings: 1,236 individuals carried 910 distinct pathogenic DNA-variants in 46 PCD genes. We found considerable variation in the distribution of PCD genotypes across countries due to the presence of distinct founder variants. The prevalence of PCD genotypes associated with pathognomonic ultrastructural defects (mean 72%; 47-100%) and laterality defects (mean 42%; 28-69%) varied widely among the countries. The prevalence of laterality defects was significantly lower in PCD individuals without pathognomonic ciliary ultrastructure defects (18%). The PCD cohort had a reduced median FEV1 z-score (-1·66). In the group of individuals with CCNO (-3·26), CCDC39 (-2·49), and CCDC40 (-2·96) variants, FEV1 z-scores were significantly lower, while the group of DNAH11 (-0·83) and ODAD1 (-0·85) variant individuals had significantly milder FEV1 z-score reductions compared to the whole PCD cohort. Interpretation: This unprecedented multinational dataset of DNA variants and information on their distribution across countries facilitates interpretation of genetic epidemiology of PCD and provides prediction of diagnostic and phenotypic features such as the course of lung function. Funding: Deutsche Forschungsgemeinschaft and Registry Warehouse (Germany).Declaration of Interest: I have no conflict of interest.
Die Präeklampsie stellt einen unabhängigen Risikofaktor für zukünftige Herz-Kreislauf-Erkrankungen der Mutter dar und geht ebenfalls mit kardiovaskulären Veränderungen der Nachkommen einher. Wir konnten zeigen, dass bereits zum Zeitpunkt der Geburt funktionelle Einschränkungen fetaler Endothelzellen aus präeklamptischen Schwangerschaften nachweisbar sind. Wir untersuchten nun, ob es außerdem zur Beeinträchtigung von Zell-Zell-Interaktionen zwischen Trophoblasten und Endothelzellen kommt und diese durch den Einsatz von 1,25 (OH)2 Vitamin D3 aufhebbar sind.
The pregnancy complication preeclampsia represents an independent risk factor for cardiovascular disease. Our previous research shows a diminished function of fetal endothelial colony-forming cells (ECFC), a proliferative subgroup of endothelial progenitor cells (EPC) in preeclampsia. The aim of this study was to further investigate whether DNA methylation of fetal EPC is affected in preeclampsia.The genomic methylation pattern of fetal ECFC from uncomplicated and preeclamptic pregnancies was compared for 865918 CpG sites, and genes were classified into gene networks. Low and advanced cell culture passages were compared to explore whether expansion of fetal ECFC in cell culture leads to changes in global methylation status and if methylation characteristics in preeclampsia are maintained with increasing passage.A differential methylation pattern of fetal ECFC from preeclampsia compared to uncomplicated pregnancy was detected for a total of 1266 CpG sites in passage 3, and for 2362 sites in passage 5. Key features of primary networks implicated by methylation differences included cell metabolism, cell cycle and transcription and, more specifically, genes involved in cell-cell interaction and Wnt signaling. We identified an overlap between differentially regulated pathways in preeclampsia and cardiovascular system development and function. Cell culture passages 3 and 5 showed similar gene network profiles, and 1260 out of 1266 preeclampsia-associated methylation changes detected in passage 3 were confirmed in passage 5.Methylation modification caused by preeclampsia is stable and detectable even in higher cell culture passages. An epigenetically modified endothelial precursor may influence both normal morphogenesis and postnatal vascular repair capacity. Further studies on epigenetic modifications in complicated pregnancies are needed to facilitate development of EPC based therapies for cardiovascular alterations.
Abstract Rare genetic diseases are a major cause of severe illnesses and deaths in new-borns and infants. Disease manifestation in critically ill children may be atypical or incomplete, making a monogenetic disease difficult to diagnose clinically. Rapid exome or genome (“genomic”) sequencing in critically ill children demonstrated profound diagnostic and clinical value, and there is growing evidence that the faster a molecular diagnosis is established in such children, the more likely clinical management is influenced positively. An early molecular diagnosis enables treatment of critically ill children with precision medicine, has the potential to improve patient outcome and leads to healthcare cost savings. In this review, we outline the status quo of rapid genomic sequencing and possible future implications.
BackgroundHuman tapasin deficiency is reported to cause an autosomal-recessive inborn error of immunity characterized by substantially reduced cell surface expression of major histocompatibility complex class I (MHC-I).ObjectiveWe evaluated the immunologic and clinical consequences of tapasin deficiency.MethodsA novel homozygous variant in TAPBP was identified by means of whole genome sequencing. The expression of tapasin and both subunits of the transporter associated with antigen presentation (TAP) were evaluated by Western blot analysis. Cell surface and intracellular expression of MHC-I were evaluated by flow cytometry. Small interfering RNAs were used for silencing TAPBP expression in HEK293T cells.ResultsWe identified a deletion in TAPBP (c.312del, p.(K104Nfs∗6)) causing tapasin deficiency in a patient with bronchiectasis and recurrent respiratory tract infections as well as herpes zoster. Besides substantial reduction in TAP1 and TAP2 expression, peripheral blood mononuclear cells from this patient and TAPBP-knockdown HEK293T cells, displayed reduced cell surface expression of MHC-I, while reduction in intracellular expression of MHC-I was less prominent, suggesting a defect in MHC-I trafficking to the plasma membrane. IFN-α improved cell surface expression of MHC-I in tapasin deficient lymphocytes and TAPBP-knockdown HEK293T cells, representing a possible therapeutic approach for tapasin deficiency.ConclusionTapasin deficiency is a very rare inborn error of immunity, the pathomechanism and clinical spectrum of which overlaps with TAP deficiencies.
