X-linked congenital ptosis and associated intellectual disability, short stature, microcephaly, cleft palate, digital and genital abnormalities define novel Xq25q26 duplication syndrome.

Submicroscopic duplications along the long arm of the X-chromosome with known phenotypic consequences are relatively rare events. The clinical features resulting from such duplications are various, though they often include intellectual disability, microcephaly, short stature, hypotonia, hypogonadism and feeding difficulties. Female carriers are often phenotypically normal or show a similar but milder phenotype, as in most cases the X-chromosome harbouring the duplication is subject to inactivation. Xq28, which includes MECP2 is the major locus for submicroscopic X-chromosome duplications, whereas duplications in Xq25 and Xq26 have been reported in only a few cases. Using genome-wide array platforms we identified overlapping interstitial Xq25q26 duplications ranging from 0.2 to 4.76 Mb in eight unrelated families with in total five affected males and seven affected females. All affected males shared a common phenotype with intrauterine- and postnatal growth retardation and feeding difficulties in childhood. Three had microcephaly and two out of five suffered from epilepsy. In addition, three males had a distinct facial appearance with congenital bilateral ptosis and large protruding ears and two of them showed a cleft palate. The affected females had various clinical symptoms similar to that of the males with congenital bilateral ptosis in three families as most remarkable feature. Comparison of the gene content of the individual duplications with the respective phenotypes suggested three critical regions with candidate genes (AIFM1, RAB33A, GPC3 and IGSF1) for the common phenotypes, including candidate loci for congenital bilateral ptosis, small head circumference, short stature, genital and digital defects.

Corpus callosum abnormalities, intellectual disability, speech impairment, and autism in patients with haploinsufficiency of ARID1B.

Corpus callosum abnormalities, intellectual disability, speech impairment, and autism in patients with haploinsufficiency of ARID1B. Corpus callosum abnormalities are common brain malformations with a wide clinical spectrum ranging from severe intellectual disability to normal cognitive function. The etiology is expected to be genetic in as much as 30-50% of the cases, but the underlying genetic cause remains unknown in the majority of cases. By next-generation mate-pair sequencing we mapped the chromosomal breakpoints of a patient with a de novo balanced translocation, t(1;6)(p31;q25), agenesis of corpus callosum (CC), intellectual disability, severe speech impairment, and autism. The chromosome 6 breakpoint truncated ARID1B which was also truncated in a recently published translocation patient with a similar phenotype. Quantitative polymerase chain reaction (Q-PCR) data showed that a primer set proximal to the translocation showed increased expression of ARID1B, whereas primer sets spanning or distal to the translocation showed decreased expression in the patient relative to a non-related control set. Phenotype-genotype comparison of the translocation patient to seven unpublished patients with various sized deletions encompassing ARID1B confirms that haploinsufficiency of ARID1B is associated with CC abnormalities, intellectual disability, severe speech impairment, and autism. Our findings emphasize that ARID1B is important in human brain development and function in general, and in the development of CC and in speech development in particular.

Complement factor H deficiency and endocapillary glomerulonephritis due to paternal isodisomy and a novel factor H mutation.

Complement factor H (CFH) is a regulator of the alternative complement activation pathway. Mutations in the CFH gene are associated with atypical hemolytic uremic syndrome, membranoproliferative glomerulonephritis type II and C3 glomerulonephritis. Here, we report a 6-month-old CFH-deficient child presenting with endocapillary glomerulonephritis rather than membranoproliferative glomerulonephritis (MPGN) or C3 glomerulonephritis. Sequence analyses showed homozygosity for a novel CFH missense mutation (Pro139Ser) associated with severely decreased CFH plasma concentration (<6%) but normal mRNA splicing and expression. The father was heterozygous carrier of the mutation, but the mother was a non-carrier. Thus, a large deletion in the maternal CFH locus or uniparental isodisomy was suspected. Polymorphic markers across chromosome 1 showed homozygosity for the paternal allele in all markers and a lack of the maternal allele in six informative markers. This combined with a comparative genomic hybridization assay demonstrated paternal isodisomy. Uniparental isodisomy increases the risk of homozygous variations in other genes on the affected chromosome. Therefore, we analyzed other susceptibility genes on chromosome 1 and found no sequence variation in membrane cofactor protein, but homozygosity for the common deletion of CFH-related proteins 1 and 3, which may contribute to the early onset of disease.

Diagnostic yield by supplementing prenatal metaphase karyotyping with MLPA for microdeletion syndromes and subtelomere imbalances.

OBJECTIVE: The aim of the study was to retrospectively assess the relevance of using multiplex ligation-dependent probe amplification (MLPA) for detection of selected microdeletion syndromes (22q11, Prader-Willi/Angelman, Miller-Dieker, Smith-Magenis, 1p-, Williams), the reciprocal microduplication syndromes and imbalance at the subtelomere regions of chromosomes in a routine prenatal setting. METHOD: A total of 530 prenatal samples were analysed by commercial MLPA kits (SALSA P064, P036 and P069) in addition to rapid aneuploidy testing and G-band karyotyping. RESULTS: Among the prenatal samples with a normal metaphase karyotype, nine submicroscopic imbalances were detected: seven 22q11 deletions (Velocardiofacial/DiGeorge syndrome), one 15q11 deletion (Prader-Willi syndrome) and one terminal deletion of the short arm of chromosome 4 (Wolf-Hirschhorn syndrome). All imbalances were found in amniocentesis (AC) taken due to fetal structural malformation and/or other ultrasound scan (US) detected abnormality. The diagnostic yield was 4.1% in the subgroup with structural malformation and 1.6% in the subgroup with other US abnormality. CONCLUSION: The data set substantiates that additional MLPA analyses for selected microdeletions and subtelomere imbalances are valuable in routine prenatal diagnostics, when a malformation(s) and/or other abnormalities are detected by US. In contrast, the additional MLPA analyses gave no diagnostic yield in case of increased nuchal translucency (NT).

A new microduplication syndrome encompassing the region of the Miller-Dieker (17p13 deletion) syndrome.

BACKGROUND: The use of array comparative genome hybridisation (CGH) analyses for investigation of children with mental retardation has led to the identification of a growing number of new microdeletion and microduplication syndromes, some of which have become clinically well characterised and some that await further delineation. This report describes three children with de novo 17p13.1 duplications encompassing the PAFAH1B1 gene, who had similar phenotypic features, including mild to moderate developmental delay, hypotonia and facial dysmorphism, and compares them to the few previously reported cases with this duplication. METHODS: Multiplex ligation-dependent probe amplification (MLPA) or array-CGH was used to diagnose three developmentally delayed children with duplications of 17p13. The duplications were characterised further using Agilent array technology, revealing duplication sizes from 1.8 to 4.0 Mb, with a region of overlap corresponding to 1.8 Mb. Detailed clinical information was obtained from patient files and personal examinations. RESULTS: The developmental delay and similar clinical features in the three patients were most likely due to a common microduplication of 17p13. CONCLUSIONS: In contrast to patients with deletion of the region (Miller-Dieker syndrome) the patients reported here had mild to moderate retardation and displayed no lissencephaly or gross brain malformations. Further cases with similar duplications are expected to be diagnosed, and will contribute to the delineation of a potential new microduplication syndrome of 17p13.

Chromosomal deletion unmasking a recessive disease: 22q13 deletion syndrome and metachromatic leukodystrophy.

A deletion on one chromosome and a mutant allele on the other may cause an autosomal recessive disease. We report on two patients with mental retardation, dysmorphic features and low catalytic activity of arylsulfatase A. One patient had a pathogenic mutation in the arylsulfatase A gene (ARSA) and succumbed to metachromatic leukodystrophy (MLD). The other patient had a pseudoallele, which does not lead to MLD. The presenting clinical features and low arylsulfatase A activity were explained, in each patients, by a deletion of 22q13 and, thereby, of one allele of ARSA.

Increased nuchal translucency with normal karyotype: a follow-up study of 100 cases supplemented with CGH and MLPA analyses.

OBJECTIVE: To evaluate whether high-resolution comparative genomic hybridization (HR-CGH) and subtelomeric and syndrome-specific multiplex ligation-dependent probe amplification (MLPA) would detect minor chromosomal aberrations in fetuses with increased nuchal translucency thickness (NT) and normal karyotype on conventional karyotyping. METHODS: Chorionic villus samples from 100 fetuses with NT > or = 99(th) percentile and normal G-banding analysis and MLPA for detection of aneuploidies for chromosomes 13, 18, 21, X and Y were included. Examinations were supplemented by HR-CGH and MLPA for syndromes and subtelomeric regions. Pregnancy outcome was followed up. RESULTS: Among 80 liveborn children who were followed up, three (4%) had syndromes involving mental retardation, including a case of Sotos syndrome caused by a de novo mutation. 15% of fetuses were lost during pregnancy due to abnormalities and termination. The rate of adverse outcome overall was 18%. HR-CGH and MLPA did not detect any chromosomal aberrations associated with the syndromes. CONCLUSION: The rate of adverse outcome was similar to levels recorded in the literature. Using CGH and MLPA did not increase the detection rate of genetic disease, which supports the current approach of repeated ultrasound examinations in these high-risk pregnancies.

Impact of low copy repeats on the generation of balanced and unbalanced chromosomal aberrations in mental retardation.

Low copy repeats (LCRs) are stretches of duplicated DNA that are more than 1 kb in size and share a sequence similarity that exceeds 90%. Non-allelic homologous recombination (NAHR) between highly similar LCRs has been implicated in numerous genomic disorders. This study aimed at defining the impact of LCRs on the generation of balanced and unbalanced chromosomal rearrangements in mentally retarded patients. A cohort of 22 patients, preselected for the presence of submicroscopic imbalances, was analysed using submegabase resolution tiling path array CGH and the results were compared with a set of 41 patients with balanced translocations and breakpoints that were mapped to the BAC level by FISH. Our data indicate an accumulation of LCRs at breakpoints of both balanced and unbalanced rearrangements. LCRs with high sequence similarity in both breakpoint regions, suggesting NAHR as the most likely cause of rearrangement, were observed in 6/22 patients with chromosomal imbalances, but not in any of the balanced translocation cases studied. In case of chromosomal imbalances, the likelihood of NAHR seems to be inversely related to the size of the aberration. Our data also suggest the presence of additional mechanisms coinciding with or dependent on the presence of LCRs that may induce an increased instability at these chromosomal sites.

High resolution comparative genomic hybridisation in clinical cytogenetics.

High resolution comparative genomic hybridisation (HR-CGH) is a diagnostic tool in our clinical cytogenetics laboratory. The present survey reports the results of 253 clinical cases in which 47 abnormalities were detected. Among 144 dysmorphic and mentally retarded subjects with a normal conventional karyotype, 15 (10%) had small deletions or duplications, of which 11 were interstitial. In addition, a case of mosaic trisomy 9 was detected. Among 25 dysmorphic and mentally retarded subjects carrying apparently balanced de novo translocations, four had deletions at translocation breakpoints and two had deletions elsewhere in the genome. Seventeen of 19 complex rearrangements were clarified by HR-CGH. A small supernumerary marker chromosome occurring with low frequency and the breakpoint of a mosaic r(18) case could not be clarified. Three of 19 other abnormalities could not be confirmed by HR-CGH. One was a Williams syndrome deletion and two were DiGeorge syndrome deletions, which were apparently below the resolution of HR-CGH. However, we were able to confirm Angelman and Prader-Willi syndrome deletions, which are about 3-5 Mb. We conclude that HR-CGH should be used for the evaluation of (1) dysmorphic and mentally retarded subjects where normal karyotyping has failed to show abnormalities, (2) dysmorphic and mentally retarded subjects carrying apparently balanced de novo translocations, (3) apparently balanced de novo translocations detected prenatally, and (4) for clarification of complex structural rearrangements.

Further delineation of the 22q13 deletion syndrome.

A chromosomal deletion syndrome associated with a 22q13 microdeletion has previously been reported in approximately 75 children. We report six cases from Denmark with a deletion of 22q13. One was cytogenetically visible by conventional karyotyping, one was diagnosed by high resolution karyotyping after the demonstration of low arylsulfatase A activity. Two were diagnosed by high resolution CGH analysis, one was diagnosed by multisubtelomeric FISH analysis and one was diagnosed serendipitously as lack of the control signal in a FISH analysis for 22q11 deletion. One of the cases was a mosaic with 16% of cells showing two signals. The phenotype of the children included: generalized developmental delay, compromised language development, hypotonia, normal or accelerated growth and minor facial dysmorphism. Other features were partial agenesis of the corpus callosum, bilateral ureteropelvic stricture, gastroesophageal reflux and hearing loss. One case had a different phenotype, and showed a deletion as well as a duplication. The extent of the deletion was studied by quantitative PCR analysis of a number of DNA markers in the 22q13 region. The deletions varied in size, extending from 4.0 to 9.0 Mb. The clinical phenotype seemed rather similar although some specific features might be attributable to differences in deletions.

High resolution comparative genomic hybridisation analysis reveals imbalances in dyschromosomal patients with normal or apparently balanced conventional karyotypes.

A sensitive technique is needed for screening whole genome imbalances in dyschromosomal patients when G-banding shows normal karyotypes or apparently balanced translocations. In this study we performed highly sensitive comparative genomic hybridisation analysis on a number of such cases and revealed chromosomal imbalances in all.

Activation of Raf-1 is defective in annexin 6 overexpressing Chinese hamster ovary cells.

Annexin 6 is a Ca2+-dependent phospholipid-binding protein involved in membrane trafficking. In this study we demonstrate the association of Raf-1 with recombinant rat annexin 6. Raf-annexin 6 interaction was shown to be independent of cell activation by epidermal growth factor (EGF) or phorbol esters (12-O-tetradecanoyl-phorbol-13-acetate (TPA)). A stable Chinese hamster ovary (CHO)-anx6 cell line overexpressing annexin 6 was established to examine the function of annexin 6. In these cells, no increase of Ras-GTP levels, induced by EGF or TPA, was detected. In addition, the activity of Raf was completely inhibited, whereas the mitogen-activated protein kinase-P was unaffected.

The most common chromosome aberration detected by high-resolution comparative genomic hybridization in vulvar intraepithelial neoplasia is not seen in vulvar squamous cell carcinoma.

We analyzed genetic changes in condylomas (four cases), vulvar intraepithelial neoplasia I-III (VIN I-III, eleven cases), and primary vulvar squamous cell carcinomas (VSCC, ten cases) by high-resolution comparative genomic hybridization (HR-CGH) and flowcytometry. All samples were also human papilloma virus (HPV)-genotyped. Gain of chromosome 1, the aberration most often seen in VIN III (67%), was not seen in HPV-positive or -negative VSCCs (0%). Both VIN III and VSCC frequently showed gain of 3q (56 and 70%, respectively). The VIN III samples often demonstrated gain of 20q (56%) and 20p (44%), and the VSCC samples gain of 8q (60%), loss of 3p (50%), and 8p (40%). None of the four most frequent changes in the VSCC samples occurred exclusively in the HPV-positive or -negative samples. As expected, we did not find any cytogenetic changes in condylomas and nearly any changes in VIN I-II.

Casual blood pressure in a general Danish population. Relation to age, sex, weight, height, diabetes, serum lipids and consumption of coffee, tobacco and alcohol.

A population survey was conducted on 3608 randomly selected Danes aged 30, 40, 50 and 60 years respectively. Of these, 3400 were not in medical treatment for arterial hypertension. The following parameters were investigated: sex, age, serum lipid levels (total cholesterol, HDL cholesterol, triglycerides), presence of diabetes mellitus, height, body mass index (BMI), and average daily consumption of coffee, tobacco and alcohol. Analysis with multiple linear regression showed that all variables with the exception of triglycerides, HDL cholesterol and height were significantly associated with systolic blood pressure. Likewise all factors except diabetes, triglycerides and height were significantly associated with diastolic blood pressure. Further analysis in which the effect of each parameter was corrected for by the effects of the remaining variables, demonstrated that apart from age and sex only BMI and high alcohol consumption were positively associated with differences in blood pressure greater than a few mmHg. However, the variation in both systolic and diastolic blood pressures was only partly accounted for by the parameters studied--in the covariates analysis R2 for systolic blood pressure was 0.28 and R2 for diastolic blood pressure was 0.30. In conclusion, this investigation demonstrated that blood pressure is relatively independent of other factors important in the development of cardiovascular disease. Of the above-mentioned factors with some influence on blood pressure only age, BMI and high alcohol consumption have potential clinical importance.

Incidence of myocardial infarction in the Danish MONICA population 1982-1991.

BACKGROUND: Cardiovascular mortality has been declining in Denmark over the past 20 years. Trends in incidence of myocardial infarction (MI) over the period 1982-1991 are described within the framework of the World Health Organization MONICA Project. METHODS: The DAN-MONICA heart register included all cases of MI in 25-74-year-old men and women living in 11 municipalities around Glostrup County Hospital evolving over a period of 10 years. They were identified retrospectively based mainly on relevant ICD diagnoses in death certificates and hospital discharge reports. Cases meeting WHO-MONICA criteria for definite or possible MI, recurrent as well as first-ever MI, were registered. Subsequent tracing of cases through national registers on deaths and hospitalizations by means of the patient's civil registration number ensured the completeness of the registration. RESULTS: A total of 6025 cases of MI occurred in the period, 4532 among men and 1493 among women. A total of 2923 men and 1047 women had a first-ever MI in the period. The age-standardized rates show a definite decline over the registration period for men and a less distinct decline for women. CONCLUSIONS: The DAN-MONICA heart register meets the requirements for completeness and uniformity throughout the registration period. Causes and magnitude of bias are well described. Even when possible sources of bias are taken into account, the incidence of MI decreased significantly over the 10-year-period 1982-1991 by an average of 5.0% per year for men and 3.5% per year for women.

Comparative genomic hybridization reveals non-random chromosomal aberrations in early preinvasive cervical lesions.

We performed CGH analysis on 34 cervical lesions, which included 8 cases of koilocytosis, 6 mild dysplasias and 20 moderate dysplasias. Chromosome aberrations were detected in 11 cases of which 9 were moderate dysplasias. A total of 55 chromosome arms were involved. The most frequent aberrations were losses of 5p and Xq, each of which was present in 5/34 cases. Gain of 3q was detected in two moderate dysplasias. This aberration is the most frequent copy number change in advanced-stage cervical carcinoma. A considerable number of the aberrations found in the preinvasive cases of this study are frequently present in invasive cervical tumors. The presence of apparently non-random chromosome aberrations in early preinvasive cervical lesions has not previously been described.

Postphlebitic syndrome and general surgery: an epidemiologic investigation.

Of a random sample comprising 4581 subjects from The Copenhagen County, 3608 (79%) attended an interview and a general health examination. The subjects were defined as suffering from subjective postphlebitic syndrome if they claimed of lower extremity pain or cramps at rest and from objective postphlebitic syndrome if varicose veins, edema, lower extremity ulcers, or skin changes were present. By means of logistic regression analysis, subjective postphlebitic syndrome was found independently associated with previous thromboembolism, obesity, increasing age, female sex, hormonal therapy, varicose veins, and previous major abdominal surgery. Objective postphlebitic syndrome was associated with previous thromboembolism, obesity, former birthgiving, and high social status. The findings support the view that subclinical deep venous thrombosis in connection with previous surgery may give rise to symptoms in the lower extremities.

[Changes in smoking habits in a Danish population from 1983 to 1988]. / Aendringer i rygevaner i en dansk befolkning fra 1983 til 1988.

Changes in smoking habits are described for 2,986 danes aged 30, 40, 50 and 60 examined in 1983 as part of the MONICA survey and reexamined five years later. In this period the prevalence declined by 8.1% for men and 5.9% for women. Smoking was related to social class, with an increasing proportion of smokers with decreasing social class. In 1988 the social differences had increased for females. The prevalence of cessation was 17.6% for males and 17.2% for females. The highest proportion of cessation was found among 60 year-old males. The average consumption of tobacco in the group of smokers increased over the period, the increment was higher the younger the person. The prevalence of heavy smokers (20 grams or more daily) increased especially in younger persons. The proportion of young heavy smokers was highest in the lowest social class. By logistic regression a low tobacco consumption was the most important factor associated with cessation. In addition, high social class was associated with cessation for women.

[The correlation between serum ferritin, alcohol consumption and social status in a population of 2235 Danes]. / Relationen mellem serumferritin, alkoholforbrug og social status i en population af 2.235 danskere.

The objective was to examine the relationships between serum ferritin, alcohol consumption and sociodemographic factors (school education, occupational education, occupation, income, marital status, cohabituary status, housing, social class) in a survey performed in Copenhagen County during 1982-1984. The participants, selected at random, comprised 2235 Danish Caucasian individuals, all non-blood-donors (1044 men, 1191 non-pregnant women) in cohorts 30, 40, 50 and 60 years old. In all age groups, men had significantly higher serum ferritin and alcohol consumption than women. In men, there was no relationship between serum ferritin and social class. Significant relationships were observed between ferritin and occupation (unemployed and self-employed men had the highest ferritin and ferritin and income (ferritin increased with income). None of the social variables were associated with the prevalence of small or absent iron stores (serum ferritin < or = 30 micrograms/l) or abundant iron stores (serum ferritin > 300 micrograms/l). Alcohol consumption displayed relationships to occupation and become, but not to social class. In women, none of the social variables showed any relationship to ferritin levels or iron overload. The prevalence of small iron stores was lower and the make of alcohol was higher in women from high social classes. In both men and women, serum ferritin displayed significant positive correlations with alcohol consumption. Likewise, the prevalence of iron overload was closely correlated to alcohol consumption. In conclusion, sociodemographic factors per se had a minor influence on serum ferritin levels and iron status in Danes. The distinct association between alcohol consumption and serum ferritin should be considered in future epidemiological iron status surveys.

[Iron levels in 1359 Danish women in relation to menstruation, use of oral contraceptives and parity]. / Jerndepoterne hos 1.359 danske kvinder set i relation til menstruationsforhold, p-pilleforbrug og fødsler.

Iron status was assessed by measuring serum ferritin and haemoglobin in a population survey in Copenhagen County, comprising 1359 nonpregnant women in age cohorts of 30, 40, 50, and 60 years; 809 were premenopausal and 550 postmenopausal. Premenopausal women had lower serum ferritin (median 37 micrograms/l) than postmenopausal women (median 71 micrograms/l), p < 0.0001. Of premenopausal women, 17.7% had ferritin < 15 micrograms/l (depleted iron stores), and 23.1% had ferritin levels of 15-30 micrograms/l (small iron stores). Corresponding figures in postmenopausal women were 3.3% and 10.3%. Iron deficiency anaemia (ferritin < 15 micrograms/l and haemoglobin < 121 g/l) was observed in 2.6% of pre- and 0.36% of postmenopausal women. After menstruations had ceased, there was a steep rise in ferritin levels, being most pronounced during the first 7 postmenopausal years. Pre- and postmenopausal multipara had lower ferritin than nulli- and unipara (p < 0.04), indicating that pregnancy and childbirth had a long lasting reducing influence on iron stores. The use of oral hormonal contraceptives had a marked increasing effect on iron stores, being correlated to the number of years the women had been taking the pill.