Auxilin is a novel susceptibility gene for congenital heart block which directly impacts fetal heart function

Objective: Neonatal lupus erythematosus (NLE) may develop after transplacental transfer of maternal autoantibodies with cardiac manifestations (congenital heart block, CHB) including atrioventricular block, atrial and ventricular arrhythmias, and cardiomyopathies. The association with anti-Ro/SSA antibodies is well established, but a recurrence rate of only 12%–16% despite persisting maternal autoantibodies suggests that additional factors are required for CHB development. Here, we identify fetal genetic variants conferring risk of CHB and elucidate their effects on cardiac function. Methods: A genome-wide association study was performed in families with at least one case of CHB. Gene expression was analysed by microarrays, RNA sequencing and PCR and protein expression by western blot, immunohistochemistry, immunofluorescence and flow cytometry. Calcium regulation and connectivity were analysed in primary cardiomyocytes and cells induced from pleuripotent stem cells. Fetal heart performance was analysed by Doppler/echocardiography. Results: We identified DNAJC6 as a novel fetal susceptibility gene, with decreased cardiac expression of DNAJC6 associated with the disease risk genotype. We further demonstrate that fetal cardiomyocytes deficient in auxilin, the protein encoded by DNAJC6, have abnormal connectivity and Ca2+ homoeostasis in culture, as well as decreased cell surface expression of the Cav1.3 calcium channel. Doppler echocardiography of auxilin-deficient fetal mice revealed cardiac NLE abnormalities in utero, including abnormal heart rhythm with atrial and ventricular ectopias, as well as a prolonged atrioventricular time intervals. Conclusions: Our study identifies auxilin as the first genetic susceptibility factor in NLE modulating cardiac function, opening new avenues for the development of screening and therapeutic strategies in CHB.publishedVersio

A heteroglycan from the cyanobacterium Nostoc commune modulates LPS-induced inflammatory cytokine secretion by THP-1 monocytes through phosphorylation of ERK1/2 and Akt.

To access publisher's full text version of this article click on the hyperlink at the bottom of the pageCyanobacteria (blue-green algae) have been consumed as food and used in folk medicine since ancient times to alleviate a variety of diseases. Cyanobacteria of the genus Nostoc have been shown to produce complex exopolysaccharides with antioxidant and antiviral activity. Furthermore, Nostoc sp. are common in cyanolichen symbiosis and lichen polysaccharides are known to have immunomodulating effects. Nc-5-s is a heteroglycan isolated from free-living colonies of Nostoc commune and its structure has been characterized in detail. The aim of this study was to determine the effects of Nc-5-s on the inflammatory response of lipopolysaccharide (LPS)-stimulated human THP-1 monocytes and how the effects are mediated. THP-1 monocytes primed with interferon-γ and stimulated with LPS in the presence of Nc-5-s secreted less of the pro-inflammatory cytokine interleukin (IL)-6 and more of the anti-inflammatory cytokine IL-10 than THP-1 monocytes stimulated without Nc-5-s. In contrast, Nc-5-s increased LPS-induced secretion of the pro-inflammatory cytokines tumor necrosis factor (TNF)-α and IL-8. Nc-5-s decreased LPS-induced phosphorylation of the extracellular regulated kinase (ERK)1/2 and Akt kinase, but did not affect phosphorylation of the p38 kinase, activation of the nuclear factor kappa B pathway, nor DNA binding of c-fos. These results show that Nc-5-s has anti-inflammatory effects on IL-6 and IL-10 secretion by THP-1 monocytes, but its effects are pro-inflammatory when it comes to TNF-α and IL-8. Furthermore, they show that the effects of Nc-5-s may be mediated through the ERK1/2 pathway and/or the Akt/phosphoinositide 3-kinase pathway and their downstream effectors. The ability of Nc-5-s to decrease IL-6 secretion, increase IL-10 secretion and moderate ERK1/2 activation indicates a potential for its development as an anti-inflammatory agent.Icelandic Research Fund (110418023), the Icelandic Research Fund for Graduate Students (R10-0008), the University of Iceland Research Fund, the Landspitali University Hospital Research Fund and the Bergthora Magnusdóttir and Jakob Bjarnason Memorial Fun

Protein and DNA methylation-based scores as surrogate markers for interferon system activation in patients with primary Sjögren's syndrome

Objective: Standard assessment of interferon (IFN) system activity in systemic rheumatic diseases depends on the availability of RNA samples. In this study, we describe and evaluate alternative methods using plasma, serum and DNA samples, exemplified in the IFN-driven disease primary Sjogren's syndrome (pSS). Methods: Patients with pSS seropositive or negative for anti-SSA/SSB and controls were included. Protein-based IFN (pIFN) scores were calculated from levels of PD-1, CXCL9 and CXCL10. DNA methylation-based (DNAm) IFN scores were calculated from DNAm levels at RSAD2, IFIT1 and IFI44L. Scores were compared with mRNA-based IFN scores measured by quantitative PCR (qPCR), Nanostring or RNA sequencing (RNAseq). Results: mRNA-based IFN scores displayed strong correlations between B cells and monocytes (r=0.93 and 0.95, p<0.0001) and between qPCR and Nanostring measurements (r=0.92 and 0.92, p<0.0001). The pIFN score in plasma and serum was higher in patients compared with controls (p<0.0001) and correlated well with mRNA-based IFN scores (r=0.62-0.79, p<0.0001), as well as with each other (r=0.94, p<0.0001). Concordance of classification as 'high' or 'low' IFN signature between the pIFN score and mRNA-based IFN scores ranged from 79.5% to 88.6%, and the pIFN score was effective at classifying patients and controls (area under the curve, AUC=0.89-0.93, p<0.0001). The DNAm IFN score showed strong correlation to the RNAseq IFN score (r=0.84, p<0.0001) and performed well in classifying patients and controls (AUC=0.96, p<0.0001). Conclusions: We describe novel methods of assessing IFN system activity in plasma, serum or DNA samples, which may prove particularly valuable in studies where RNA samples are not available

Type i IFN system activation in newborns exposed to Ro/SSA and La/SSB autoantibodies in utero

Objective In utero exposure of the fetus to Ro/La autoantibodies may lead to congenital heart block (CHB). In the mother, these autoantibodies are associated with activation of the type I interferon (IFN)-system. As maternal autoantibodies are transferred to the fetus during pregnancy, we investigated whether the type I IFN-system is activated also in newborns of anti-Ro/La positive mothers, and whether fetal IFN activation is affected by maternal immunomodulatory treatment. Methods Blood drawn at birth from anti-Ro/La positive mothers, their newborns and healthy control pairs was separated into plasma and peripheral blood mononuclear cells (PBMC). PBMC were analysed directly or cultured. mRNA expression was analysed by microarrays, cell surface markers by flow cytometry, and IFNα levels by immunoassays. Results We observed increased expression of IFN-regulated genes and elevated plasma IFNα levels not only in anti-Ro/La positive women, but also in their newborns. CD14 + monocytes of both anti-Ro/La positive mothers and their neonates showed increased expression of Sialic acid-binding Ig-like lectin-1, indicating cellular activation. Notably, the IFN score of neonates born to mothers receiving immunomodulatory treatment was similar to that of controls, despite persistent IFN activation in the mothers. In both maternal and neonatal PBMC, IFNα production was induced when cells were cultured with anti-Ro/La positive plasma. Conclusions Ro/La autoantibody-exposed neonates at risk of CHB have signs of an activated immune system with an IFN signature. This study further demonstrates that neonatal cells can produce IFNα when exposed to autoantibody-containing plasma, and that maternal immunomodulatory treatment may diminish the expression of IFN-regulated genes in the fetus