Comprehensive molecular screening of the FBN1 gene favors locus homogeneity of classical Marfan syndrome.

In order to estimate the contribution of mutations at the fibrillin-1 locus (FBN1) to classical Marfan syndrome (MFS) and to study possible phenotypic differences between patients with an FBN1 mutation vs. without, a comprehensive molecular study of the FBN1 gene in a cohort of 93 MFS patients fulfilling the clinical diagnosis of MFS according to the Ghent nosology was performed. The initial mutation screening by CSGE/SSCP allowed identification of an FBN1-mutation in 73 patients. Next, sequencing of all FBN1-exons was performed in 11 mutation-negative patients, while in nine others, DHPLC was used. This allowed identification of seven and five additional mutations, respectively. Southern blot analysis revealed an abnormal hybridization pattern in one more patient. A total of 23 out of the 85 mutations identified here are reported for the first time. Phenotypic comparison of MFS patients with cysteine-involving mutations vs. premature termination mutations revealed significant differences in ocular and skeletal involvement. The phenotype of the eight patients without proven FBN1 mutation did not differ from the others with respect to the presence of major cardiac, ocular, and skeletal manifestations or positive familial history. Most likely, a portion of FBN1-mutations remains undetected because of technical limitations. In conclusion, the involvement of the FBN1-gene could be demonstrated in at least 91% of all MFS patients (85/93), which strongly suggests that this gene is the predominant, if not the sole, locus for MFS.

Homozygosity mapping of a gene for arterial tortuosity syndrome to chromosome 20q13.

BACKGROUND: Arterial tortuosity syndrome (ATS) is an uncommon connective tissue disorder of unknown aetiology. The most prominent feature is tortuosity of the large arteries, but lengthening, stenosis, and aneurysm formation are also frequent. METHODS: We performed a genomewide screen by homozygosity mapping of three consanguineous multiplex families, two from Morocco, and one from Italy, which included 11 ATS patients. The two families from Morocco may possibly have a common ancestor. RESULTS: We mapped the ATS gene to chromosome 20q13. Recombinations within an extended haplotype of 11 microsatellite markers localised the ATS gene between markers D20S836 and D20S109, an interval of 9.5 cM. CONCLUSIONS: Cloning and completing functional and structural analysis of the ATS gene may provide new insights into the molecular mechanisms of elastogenesis.

Strategies for prenatal and preimplantation genetic diagnosis in Marfan syndrome (MFS).

Marfan syndrome (MFS) is an autosomal dominant disorder with a prevalence of 2-3 per 10 000 individuals. Symptoms range from skeletal overgrowth, cutaneous striae to ectopia lentis and aortic dilatation leading to dissection. Prenatal diagnosis was until recently mainly performed in familial cases by linkage analysis. However, mutation detection has become available with thorough screening methods. The phenotypic variability observed in MFS makes reproductive options difficult, as molecular diagnosis cannot predict clinical severity of the disease. Data are presented on 15 prenatal and/or preimplantation genetic diagnoses (PGD) in nine families, originating from Belgium, the Netherlands, Spain and France. In four families data from linkage analysis were used, whereas in five other families the causative FBN1 mutation was characterised. Four PGD cycles in two couples led to one ongoing pregnancy. In addition, two amniocenteses and nine chorionic villus (CV) samplings were performed. In five pregnancies an affected fetus was diagnosed. In one of them, the couple chose to continue the pregnancy and an affected child was born, whereas the other four couples decided to terminate the pregnancy. It is expected that the greater availability of mutation testing of the FBN1 gene will increase requests for prenatal diagnosis. PGD appears to be an acceptable alternative for couples facing ethical reproductive dilemmas.

Characterization of mutations leading to recessive dystrophic epidermolysis bullosa and Marfan syndrome in a single patient.

Dystrophic epidermolysis bullosa (DEB) is a rare genetic skin disorder. In this report we have investigated an Italian child affected with recessive DEB (RDEB) and demonstrated that he was homozygous for the mutation R226X in the type VII collagen gene (COL7A1), leading to absence of type VII collagen at the dermal-epidermal junction. There was no family history of inherited skin blistering but the child's father was affected by Marfan syndrome, an autosomal dominant connective tissue disorder that results from mutations in the fibrillin-1 gene (FBN1). Analysis of this gene showed that the RDEB patient and his father were both heterozygous for a novel FBN1 mutation, C1971Y. This mutation affects one of the six obligate cysteine residues within one of the calcium-binding epidermal growth factor-like regions of the protein. At the age of 2-years the RDEB patient showed signs of early aortic dilatation, suggesting that he is likely to develop a Marfan syndrome phenotype in the future. This is a unique case of these two coexisting inherited disorders.

Low-density lipoprotein receptor gene mutations in a Southeast Asian population with familial hypercholesterolemia.

The aim of this study was to detect mutations in the genes coding for the low-density lipoprotein receptor and apolipoprotein B in patients of Southeast Asian origin with clinically diagnosed familial hypercholesterolemia (FH) and to relate these findings with the observed lower incidence of coronary heart disease in this part of the world. A total of 86 unrelated patients with FH were selected on clinical grounds, and complete DNA analysis of the low-density lipoprotein (LDL)-receptor and apolipoprotein B (apoB) genes by DGGE and DNA-sequencing was performed. In the majority (73%) of the cohort studied, no mutations could be detected, even after extensive analysis of the LDL-receptor and apoB genes. However, the 22 patients with a mutation had significantly more xanthomas and a higher incidence of coronary heart disease and levels of low-density lipoproteins were also significantly different. There was no correlation between the type of the mutation and lipoprotein levels or clinical signs of atherosclerosis. The fact that the majority of the FH patients studied had no detectable mutation and that this group had a significant milder phenotype, suggests the presence of a third gene in the Southeast Asian population, predominantly leading to a disorder resembling a milder form of FH. A similar, but less frequent, trait has recently been described in a number of European families.

Genetic causes of familial hypercholesterolaemia in a Malaysian population.

A total of 86 unrelated Malaysian patients with familial hypercholesterolaemia (FH) were studied for mutations in their low-density lipoprotein receptor (LDL-R) gene. Amongst them, 23 had a LDL-R gene mutation, while none having an Apolipoprotein B-3500 (Apo B-3500) mutation. Patients with the LDL-R gene defect appeared to have a higher level of low-density lipoprotein cholesterol (LDL-C), an increased incidence of xanthomas and coronary heart disease (CHD), but no relationships were found between the type of LDL-R gene mutations and their lipid levels or clinical signs of CHD. In contrast to Western data, our findings seemed to indicate a predominance of mutations in the ligand binding domain and an absence of Apo B-3500 gene mutation. The latter finding may offer a genetic basis as to why Asian patients with familial hypercholesterolaemia have lower LDL-C levels and less premature CHD than their Western counterparts.

High prevalence of a novel mutation in the exon 4 of the low-density lipoprotein receptor gene causing familial hypercholesterolemia in Belgium.

In a cohort of 70 unrelated patients living in Southern Belgium with autosomal dominantly inherited hypercholesterolemia, 11 had a hitherto undescribed mutation in exon 4. It consisted in a C-->A mutation at nucleotide 366, resulting in a stop codon at residue Cys122. This C122X mutation is expected to cause a class I receptor defect. The biochemical and clinical data collected from the patients carrying the mutation were consistent with a severe form of familial hypercholesterolemia (FH). Some differences between generations were noted. Amongst the C122X carriers, those born after 1926 had cardiovascular complications earlier than those born before 1926. This raises the possibility that changes in environmental factors during the course of the century have had an unfavorable impact on the prognosis of the disease. The mutation was found in 16% of the suspected FH patients and less frequently (less than 3% of suspected FH) in Northern Belgium. The haplotype of the chromosomes carrying the mutation was the same in all C122X families, but extensive genealogical studies failed to reveal a common ancestor. We conclude that C122X is an old and common cause of FH in Belgium. Screening for this mutation may be useful in the diagnosis of FH in Belgium.

The effect of semi-automatic external defibrillation by emergency medical technicians on survival after out-of-hospital cardiac arrest: an observational study in urban and rural areas in Belgium.

The introduction of semi-automatic external defibrillators (SAEDs) allowed emergency medical technicians (EMTs) to deliver electroshocks in cases of out-of-hospital ventricular fibrillation (VF) or ventricular tachycardia (VT), often many minutes before the arrival of the mobile intensive care unit (MICU) team. In this observational study we report on the results obtained by the EMTs from the fire departments of Gent, Aalter and Brugge. In Gent, an SAED project started in May 1991. By December 1995, the SAED's electrodes had been attached in 367 cardiac arrest patients. The first rhythm detected by the device was asystole or electromechanical dissociation (EMD) in 241 patients (66%): only 5 of these patients survived to hospital discharge (2%). In the remaining 126 VF/VT cases (34%) the survival rate was 21% (26/126). In 14 of these 26 patients the shock(s) delivered by the EMTs restored spontaneous circulation before the arrival of the MICU team, with only venous cannulation and/or intubation being performed by the MICU team. In 4 other VF patients, the shock(s) delivered by EMTs converted the VF, with the MICU team successfully taking care of VF/VT relapses or postcountershock EMD. In the remaining 8 VF/VT cases, only the MICU attempts could resuscitate the patient. The SAED project in Aalter was set up in April 1993. By December 1995, care was taken for only 21 patients. None of the 4 VF/VT patients and the 17 asystole/EMD patients survived. In Brugge, there were 240 cardiac arrest cases treated with SAED between January 1991 and December 1995. Among the 89 VF/VT cases, there were 20 survivors (22%): 8 cases survived thanks to SAED shock(s) delivered by EMTs, in 3 cases survival was due to the combination of SAED shock(s) by EMTs and extensive ALS treatment by the MICU team, and in 9 cases restoration of spontaneous circulation was only obtained after application of ALS techniques by the MICU team. This observational study seems to show a beneficial effect of the introduction of SAED in Gent and Brugge. In Aalter the number of treated cases is tool low to draw conclusions. Anyhow, the global survival rate in the three areas remains low. Therefore, more efforts are needed to strengthen the other links of the chain of survival (early access to the emergency medical services-system, early basic cardiopulmonary resuscitation and early advanced life support.

Retinoic acid induction of human tissue-type plasminogen activator gene expression via a direct repeat element (DR5) located at -7 kilobases.

All-trans-retinoic acid (RA) and retinoids induce synthesis of tissue-type plasminogen activator (t-PA) in endothelial and neuroblastoma cells in vitro and in rats in vivo. In HT1080 fibrosarcoma cells, induction of t-PA-related antigen secretion and t-PA mRNA steady state levels by RA were found to depend on de novo protein and mRNA synthesis. Fragments derived from the 5'-flanking region of the t-PA gene (+197 to -9578 base pairs (bp)) were linked to the chloramphenicol acetyltransferase gene. Transfection studies demonstrated that the region spanning bp -7145 to -9578 mediated induction by RA. A functional retinoic acid response element (RARE), consisting of a direct repeat of the GGGTCA motif spaced by 5 nucleotides (t-PA/DR5), was localized at -7.3 kilobases. The t-PA/DR5 element interacted with the heterodimer composed of retinoic acid receptor alpha and retinoid X receptor alpha in vitro, whereas its mutation abolished induction by RA in transient expression. In human EA.hy926 hybrid endothelial and in SK-N-SH neuroblastoma cells, the activity of t-PA/DR5 was found to be independent of the intervening sequence (-632 to -7144 bp) and of its distance from the transcription initiation site. Staurosporine, an inhibitor of protein kinase activity, inhibited induction by RA, suggesting that it required protein phosphorylation.

Determination of ascorbic acid and isoascorbic acid by capillary zone electrophoresis: application to fruit juices and to a pharmaceutical formulation.

Capillary zone electrophoresis was applied to the determination of ascorbic and isoascorbic acid, analysing the various parameters of influence such as the separation voltage, the buffer pH and concentration, the type of separation capillary or the loading conditions. Both analytes could be adequately determined within 5 min. The proposed method uses a 20 cm x 25 microns i.d. coated column, 0.1 M phosphate buffer pH 5.0, 8 kV separation voltage and light absorption detection at 265 nm. Linear calibration curves were obtained in the 0-1 mg mi-1 range, with detection limits of 0.5 micrograms ml-1. This method proved to be very rapid, simple and practical for the qualitative and quantitative determination of ascorbic acid in lemon and orange juices, as well as in a commercially available pharmaceutical formulation.

Should semi-automatic defibrillators be used by emergency medical technicians in Belgium? The Belgian Cerebral Resuscitation Study Group.

Early external defibrillation is the single most effective intervention in patients with out-of-hospital cardiac arrest. Literature data indicate that instructing emergency medical technicians (EMTs) to use defibrillators is beneficial, provided the local emergency medical system is well organized. We tried to estimate the potential benefit of early defibrillation in some centres in Belgium by retrospectively analyzing the data from the Belgian Cardio-Pulmonary-Cerebral Resuscitation Registry collected between 1983 and 1987 in Belgian centres with a Mobile Intensive Care Unit (MICU). The data show that 2310 out of 3371 patients (69%) were first attended by the EMTs; on subsequent arrival of the MICU-teams, 584 of these 2310 patients i.e. 17% of the whole study population, presented with ventricular fibrillation. Analysis of estimated time factors in these 2310 patients revealed that the median time interval between collapse and start of resuscitation by EMTs was 8 min; the median time interval between collapse and start of MICU-resuscitation attempts was 16 min. The duration of EMT-resuscitation before MICU-arrival was probably more than 5 min and 10 min in 58% and 23% of the cases respectively. It is concluded that EMTs can be expected to reach a substantial number of ventricular fibrillation victims within a few minutes after the collapse and many minutes before arrival of the MICU. Therefore, training of EMTs in the use of semi-automatic defibrillators seems worthwhile in MICU-served regions in Belgium.

Bristow-Latarjet repair for recurrent anterior shoulder instability; an eight-year study.

A follow-up study was made of 44 patients who underwent 47 should operations as described by Bristow and Latarjet. The group consisted of 32 recurrent shoulder dislocations and 15 so-called spontaneous shoulder instabilities. The average follow-up was 3.7 years. No significant complications occurred either per- or postoperatively and relapse of luxation was not seen. Only one patient had objectively confirmed shoulder instability after the operation. The average limitation of external rotation at 90 degrees abduction was 12 degrees.

[Haemagnioma of the liver in newborn infant (author's transl)]. / Haemagnioom van de lever bij een prematuur

A case of a giant cavernous and capillary haemangioma of the liver in newborn infant occurred. Resection of the tumor by right hepatic lobectomy was carried out without morbidity or permanent derangement of liver function. There was regeneration of the liver.