Vijole Dzikaite

Development of heart block in children of SSA/SSB-autoantibody-positive women is associated with maternal age and displays a season-of-birth pattern

Objective Congenital heart block may develop in the fetuses of Ro/SSA-positive and La/SSB-positive mothers. Recurrence rates of only 10–20% despite persisting maternal antibodies indicate that additional factors are critical for the establishment of heart block. The authors investigated the influence of other maternal and fetal factors on heart block development in a Swedish population-based cohort. Methods The influence of fetal gender, maternal age, parity and time of birth on heart block development was analysed in 145 families, including Ro/La-positive (n=190) and Ro/La-negative (n=165) pregnancies. Results There was a recurrence rate of 12.1% in Ro/La-positive women, and no recurrence in Ro/La-negative women. Fetal gender and parity did not influence the development of heart block in either group. Maternal age in Ro/La-positive pregnancies with a child affected by heart block was, however, significantly higher than in pregnancies resulting in babies without heart block (p<0.05).Seasonal timing of pregnancy influenced the outcome. Gestational susceptibility weeks 18–24 occurring during January–March correlated with a higher proportion of children with heart block and lower vitamin D levels during the same period in a representative sample of Swedish women and a corresponding higher proportion of children with heart block born in the summer (p<0.02). Maternal age or seasonal timing of pregnancy did not affect the outcome in Ro/La-negative pregnancies. Conclusion This study identifies maternal age and seasonal timing of pregnancy as novel risk factors for heart block development in children of Ro/La-positive women. These observations may be useful for counselling when pregnancy is considered.

Structure and liver cell expression pattern of the HFE gene in the rat

BACKGROUND/AIMS Very little is known about the HFE gene in the rat. The aim of the present study was to determine: (1) the structure of the rat HFE gene; and (2) the tissue expression of the HFE mRNA in the rat, with special emphasis on the liver. METHODS Cloning of the rat HFE gene was performed using library screening and PCR. Exon-intron borders were assigned by DNA sequencing. Parenchymal and non-parenchymal liver cells were isolated by fractionation of normal rat liver. HFE mRNA levels were determined by Northern blot (tissues) and real-time PCR (isolated liver cells). RESULTS The rat HFE gene contained six exons and five introns. The HFE gene is expressed in multiple tissues in the rat, including bone marrow, with the highest expression in the liver. We observed HFE transcripts in several categories of isolated rat liver cells. Unexpectedly, expression also occurred in rat hepatocytes. CONCLUSIONS The exon-intron pattern of the HFE gene is strongly conserved between rat and mouse. The pattern of tissue expression of the HFE gene is rather similar in humans and rodents. The finding of HFE gene expression in rat hepatocytes raises interesting questions regarding its role in the hepatocyte iron metabolism.

Expression of iron regulatory genes in a rat model of hepatocellular carcinoma.

Abstract: Background/Aims: The altered iron metabolism in hepatocellular carcinomas (HCCs), characterized by the iron‐deficient phenotype, is suggested to be of importance for tumour growth. However, the underlying molecular mechanisms remain poorly understood. We asked whether these iron perturbations would involve altered expression of genes controlling iron homeostasis.

Anti-Ro52 monoclonal antibodies specific for amino acid 200–239, but not other Ro52 epitopes, induce congenital heart block in a rat model

Background Congenital heart block (CHB) may develop in fetuses of women with anti-Ro/La autoantibodies following placental transfer of maternal autoantibodies and disruption of the fetal atrioventricular (AV) conduction system. Animal models of CHB currently rely on immunisation or transfer of anti-Ro/La antibodies purified from mothers of children with CHB, which does not allow precise identification of the disease-inducing antibody specificity. Objective To determine the ability of different anti-Ro52 monoclonal antibodies to induce cardiac electrophysiological abnormalities in vivo and affect the calcium homoeostasis of cardiomyocytes in vitro. Methods Monoclonal antibodies recognising different domains of Ro52 were generated and injected into pregnant rats, and ECG was recorded on newborn pups. Cultures of rat neonatal cardiomyocytes were established to assess the effect of the different anti-Ro52 monoclonal antibodies on calcium homoeostasis. Results First-degree AV block and bradycardia developed after maternal transfer of antibodies specific for amino acids 200–239 of Ro52 (p200), while pups exposed to antibodies targeting N- or C-terminal epitopes of Ro52 did not show any electrocardiogram abnormalities. Addition of an anti-p200 antibody to cultured cardiomyocytes induced calcium dyshomoeostasis in a time- and dose-dependent manner, while addition of other Ro52 antibodies had no effect. Conclusion These data for the first time show unambiguously that antibodies specific for amino acids 200–239 of Ro52 can induce cardiac conduction defects in the absence of other autoantibodies, and may therefore be the main initiators of cardiac pathology in the pool of anti-Ro52 antibodies in mothers of children with CHB.

A population-based investigation of the autoantibody profile in mothers of children with atrioventricular block.

The objective of the study was to investigate the antigen specificity and occurrence of individual autoantibodies in mothers of children diagnosed with atrioventricular (AV) block in a nation‐wide setting. Patients with AV block detected before 15 years of age were identified using national quality registries as well as a network of pediatric and adult cardiologists and rheumatologists at the six university hospitals in Sweden. Patients with gross heart malformations, surgically or infectiously induced blocks were excluded. Blood samples were obtained from the mothers and maternal autoantibody profile, including the occurrence of antibodies against Ro52, Ro60, La, SmB, SmD, RNP‐70k, RNP‐A, RNP‐C, CENP‐C, Scl‐70, Jo‐1, ribosomal RNP and histones was investigated in 193 mothers of children with AV block by immunoblotting and ELISA. Autoantibody reactivity was detected in 48% (93/193) of the mothers of children with AV block. In autoantibody‐positive mothers, the vast majority, 95% (88/93), had antibodies against Ro52, while 63% (59/93) had autoantibodies to Ro60 and 58% (54/93) had autoantibodies to La. In addition, 13% (12/93) of the autoantibody‐positive mothers had antibodies to other investigated antigens besides Ro52, Ro60 and La, and of these anti‐histone antibodies were most commonly represented, detected in 8% (7/93) of the mothers. In conclusion, this Swedish population‐based study confirms that maternal autoantibodies may associate with heart block in the child. Further, our data demonstrate a dominant role of Ro52 antibodies in association with AV block.

Maternal MHC Regulates Generation of Pathogenic Antibodies and Fetal MHC-Encoded Genes Determine Susceptibility in Congenital Heart Block

Congenital heart block develops in fetuses of anti-Ro52 Ab-positive women. A recurrence rate of 20%, despite the persistence of maternal autoantibodies, indicates that there are additional, yet unidentified, factors critical for development of congenital heart block. In this study, we demonstrate that besides the maternal MHC controlling Ab specificity, fetal MHC-encoded genes influence fetal susceptibility to congenital heart block. Using MHC congenic rat strains, we show that heart block develops in rat pups of three strains carrying MHC haplotype RT1av1 (DA, PVG.AV1, and LEW.AV1) after maternal Ro52 immunization, but not in LEW rats (RT1l). Different anti-Ro52 Ab fine specificities were generated in RT1av1 versus RT1l animals. Maternal and fetal influence was determined in an F2 cross between LEW.AV1 and LEW strains, which revealed higher susceptibility in RT1l than RT1av1 pups once pathogenic Ro52 Abs were present. This was further confirmed in that RT1l pups more frequently developed heart block than RT1av1 pups after passive transfer of RT1av1 anti-Ro52 sera. Our findings show that generation of pathogenic Ro52 Abs is restricted by maternal MHC, whereas the fetal MHC locus regulates susceptibility and determines the fetal disease outcome in anti-Ro52–positive pregnancies.

Development of heart block in SSA/SSB autoantibody-positive pregnancies is associated with maternal age and display a season-of-birth pattern

Background and objectives Congenital heart block (CHB) may develop in the fetus of anti-Ro/SSA and anti-La/SSB positive mothers. Reported recurrence rates of only 10–20% despite persisting maternal antibodies indicate that additional factors are critical for establishment of the heart block. The authors therefore investigated the influence of other maternal and fetal factors on heart block development in a Swedish population-based cohort. Material and methods The influence of fetal gender, maternal age, parity and time of birth on heart block development was analysed in 145 families including Ro/La-positive (n= 190) and Ro/La-negative (n=165) pregnancies. Results The authors observed a recurrence rate for heart block of 12.1% in Ro/La-positive women, and no recurrence in Ro/La-negative women. Fetal gender and parity did not influence the development of heart block in either group. Maternal age in Ro/La-positive pregnancies with a child affected by heart block was however significantly higher than in pregnancies resulting in babies without heart block (p<0.01). Seasonal timing of pregnancy appeared to be an important factor in pregnancy outcome in Ro/La-positive women as the ratio of affected to healthy births in the summer (June–August) was significantly different from the rest of the year, corresponding to a higher proportion of children with heart block born in the summer compared to the rest of the year. This in turn corresponds to a higher proportion of affected pregnancies for which the susceptible gestational weeks 18–24 occur during the late winter (January–March). Maternal age or seasonal timing of pregnancy did not affect the outcome in Ro/La-negative pregnancies. Conclusions This study identifies maternal age and seasonal timing of pregnancy as novel risk factors for CHB development in Ro/La positive pregnancy. These observations will be important for counselling when a pregnancy is considered.

MHC genes determine fetal susceptibility in a rat model of congenital heart block

Objective Congenital heart block (CHB) develops in fetuses of anti-Ro52 antibody positive women. A recurrence rate of 12–25%, despite the persistance of maternal autoantibodies indicates that there are additional, yet unidentified, factors critical for development of CHB. The authors hypothesised that fetal susceptibility to the maternal autoantibodies could be determined by fetal genes. Methods A passive antibody transfer model was used in several different rat strains and MHC congenics (DA.AV1; Lew.1AV1; Lew.L; Lew.N). Two-four mg of a Ro52 monoclonal antibody (7.8C7) was injected intraperitonially before day 7 of gestation. On the day of delivery, three-lead ECG were recorded from conscious pups. Results After maternal passive antibody transfer, significant PR prolongation was induced in the DA.AV1 pups by Ro52-p200 specific antibodies at the dose of 4 mg. In Lew.1AV1 rats, PR prolongation was induced in the pups already at 2 mg injections, indicating that fetal non-MHC genes can increase susceptability to CHB. In Lew.L rats, PR intervals were significantly more prolonged after 2 mg injections, indicating that fetal L MHC genes conferred to highest susceptibility to CHB. In Lewis.N rats however, no difference in the PR intervals were observed between the PBS-injected control group and groups injected with 2 or 4 mg of Ro52 monoclonal antibody, suggesting that specific fetal MHC genes can confer the resistance and play a protective role in the development of the CHB. Conclusions Our findings show that both fetal MHC and non-MHC genes regulate susceptibility to CHB and determine the fetal disease outcome in anti-Ro52 positive pregnancies. Different fetal MHC alleles can either render the fetus susceptible or play a protective role in the disease development.