A new approach to germinal mutation surveillance: pair-wise evaluation of component elements in unidentified multiple congenital abnormalities

In the Hungarian population-based surveillance of germinal mutations, 3 indicator conditions of offspring are being followed, namely 15 sentinel anomalies, Down syndrome and unidentified multiple congenital abnormality. The latter is discussed here as a possible indicator of germinal dominant gene and chromosomal mutations. The component congential abnormalities of unidentified multiple congenital abnormalities are classified into 45 groups. The component congenital abnormalities were reduced to pairs. A pair is a set of 2 independent component congenital abnormalities in index patients with 2 or more congenital abnormalities. Baseline figures of all component congenital abnormality pairs in 3722 unidentified multiple congenital abnormalities were determined in the study period 1973–1982. The observed data for 1983 were compared with expected occurrences based on baseline figures. This pair-wise evaluation of component elements within unidentified multiple congenital abnormalities seems to be an adequate surveillance method to detect any time cluster of congenital abnormality pairs due to environmental factors including germinal mutagens.     

Hungarian surveillance of germinal mutations

Summary The Hungarian surveillance of germinal mutations is based on three indicator conditions seen in offspring, i.e., 15 sentinel anomalies, Down syndrome and component anomaly pairs of unidentified multiple congenital anomalies. It is an opportunistic program, because the necessary data are available from the Hungarian Congenital Malformation Registry. This system is described and the criteria of a good registry are summarized. The analysis of indicator conditions caused by germinal mutations did not reveal any measurable mutagenic effects in Hungary following the accident at the Chernobyl nuclear power plant. The pros and cons of germinal mutation surveillance are discussed.     

Congenital abnormalities and indicators of germinal mutations in the vicinity of an acrylonitrile producing factory.

The results of an environmental mutation and teratologic epidemiological study are presented which was performed in inhabitants living in the surrounding region of an acrylonitrile factory in Nyergesújfalu. The endpoint of the study was congenital abnormalities in 46,326 infants born to mothers living in the 30 settlements of the study region within a 25 km radius of the acrylonitrile factory between 1980 and 1996. The ascertainment of cases with congenital abnormalities was based on the dataset of the Hungarian Congenital Abnormality Registry complemented with the review of pediatric, pathology and cytogenetic records. A particular attention was paid to the indicators of germinal mutations as sentinel anomalies, Down syndrome and unidentified multiple congenital abnormalities and the indicators of teratogens as the specific pattern of multiple congenital abnormalities. Three congenital abnormalities: pectus excavatum in Tata, 1990-1992 (OR with 95%CI: 78.5, 8.4-729.6), undescended testis in Nyergesújfalu between 1980 and 1983 (8.6, 1.4-54.3) and in Esztergom, 1981-1982 (4. 2, 1.3-13.5) and clubfoot in Tata, 1980-1981 (5.5, 1.5-20.3) showed significant time-space clusters in the study region. There was a decrease in risk of undescended testis with increasing distance from the acrylonitrile factory. An unusual increase was found in the combination of oral cleft and cardiac septal defects in multimalformed babies in Tatabánya, 1990. The detailed analysis of congenital abnormalities in all settlements of a given territory may help to detect clusters of congenital abnormalities and their possible relation to the environmental hazards.     

Incidence of chromosomal rearrangements in couples with reproductive loss

Summary We report on 50 couples with reproductive loss who did not have any detectable chromosome abnormality. A history of a previous child with multiple congenital abnormalities may be significant in identifying couples with a structural rearrangement. Only by studying more families can this hypothesis be tested. Studies of abortus tissue reveal a high percentage of chromosome abnormalities but a very low incidence of unbalanced translocations. Cytogenetic studies are indicated in a couple which has a past history of spontaneous abortions and a previous child with multiple congenital anomalies.     

What proportion of congenital abnormalities can be prevented?

OBJECTIVE--To estimate the proportion of preventable congenital abnormalities in Hungary. DESIGN--Analysis of available Hungarian data-bases and of the effectiveness of primary, secondary, and tertiary preventive methods. SETTING--Databases of ad hoc epidemiological studies and of the Hungarian congenital abnormality registry. MAIN OUTCOME MEASURES--Prevalence at birth and prevalence after prevention in 73 congenital abnormality types or groups. RESULTS--Preventive methods are available for 51 (70%) of the 73 congenital abnormality types or groups evaluated. The birth prevalence of all congenital abnormalities could be reduced from 65 to 26 per 1000; thus 39 per 1000 (60%) are preventable. Without congenital dislocation of the hip, which is unusually common in Hungary, the preventable proportion of congenital abnormalities is 52%. CONCLUSION--Many congenital abnormalities can be prevented, but as they do not represent a single pathological category there is no single strategy for their prevention.     

Congenital hemihypertrophy. Tendency to association with other abnormalities and/or tumors

Nine cases of congenital hemihypertrophy of an entire half of the body ("true" hemihypertrophy) were investigated in relation to literature data. The left side was more often affected (7:2) and the abnormality was more frequent in females (5:4). Associated abnormalities were present in all cases: a single abnormality in 3 cases (mental retardation, iris heterochromia and, respectively, left foot gigantism) and 3 to 13 abnormalities in the other 6 cases. Prominent hemiface, asymmetric macroglossia on the affected side, unequally developed breasts, unilateral gigantism of the foot, iris heterochromia, duplication of the renal pelvis, were considered important signs which easily draw attention. Out of nine cases investigated, three presented associated tumors: two tumors, one cerebral (malignant ependymoma) and the other hepatic (cavernous hemangioma) in one case, hepatoblastoma in another case, and multiple skin angiomas in the affected side of the third one. The case presenting two tumors had also the greatest number (twelve) of associated abnormalities.     

PATTERNS OF MITOSIS AND DIFFERENTIATION IN CELLS DERIVED FROM PEA ROOT PROTOPLASTS

Mitotic figures of diploid, tetraploid, octaploid and 16-ploid nuclei were observed in cultures of pea root protoplasts whose initial DNA content was apparently 2C and 4C. The distribution of these mitotic figures in the different ploidy levels paralleled the distribution of mitotic figures in the culture of intact root explants and may be related to the hormonal stimulation of mitoses in these cultures. The patterns of the time course of both DNA synthesis and cell division in the protoplast cultures were similar to such patterns observed in the culture of intact root explants, although longer lag periods were observed in the protoplast cultures. Mitotic abnormalities including both chromosome breakage and spindle disfunction were observed in protoplast cultures. A large portion of the cell pairs derived from mitoses (27 % in one experiment) contained Feulgen-positive micronuclei. An accumulation of an as yet unidentified differentiation product termed dense cytoplasmic protoplast derivative was observed. Some of the conditions influencing the development of these derivatives are reported.     

Identification of somatic chromosomal abnormalities in hypothalamic hamartoma tissue at the GLI3 locus

Hypothalamic hamartomas (HH) are rare, benign congenital tumors associated with intractable epilepsy. Most cases are sporadic and nonsyndromic. Approximately 5% of HH cases are associated with Pallister-Hall syndrome (PHS), which is caused by haploinsufficiency of GLI3. We have investigated the possibility that HH pathogenesis in sporadic cases is due to a somatic (tumor-only) mutation in GLI3. We isolated genomic DNA from peripheral blood and surgically resected HH tissue in 55 patients with sporadic HH and intractable epilepsy. A genome-wide screen for loss of heterozygosity (LOH) and chromosomal abnormalities was performed with parallel analysis of blood and HH tissue with Affymetrix 10K SNP microarrays. Additionally, resequencing and fine mapping with SNP genotyping were completed for the GLI3 gene with comparisons between peripheral blood and HH tissue pairs. By analyzing chromosomal copy-number data for paired samples on the Affymetrix 10K array, we identified a somatic chromosomal abnormality on chromosome 7p in one HH tissue sample. Resequencing of GLI3 did not identify causative germline mutations but did identify LOH within the GLI3 gene in the HH tissue samples of three patients. Further genotyping of 28 SNPs within and surrounding GLI3 identified five additional patients exhibiting LOH. Together, these data provide evidence that the development of chromosomal abnormalities within GLI3 is associated with the pathogenesis of HH lesions in sporadic, nonsyndromic patients with HH and intractable epilepsy. Chromosomal abnormalities including the GLI3 locus were seen in 8 of 55 (15%) of the resected HH tissue samples. These somatic mutations appear to be highly variable.     

Description and mission evaluation of the Hungarian case-control surveillance of congenital abnormalities, 1980-1996

BACKGROUND: The Hungarian Case-Control Surveillance of Congenital Abnormalities was established in 1980. This article describes how the Hungarian Case-Control Surveillance of Congenital Abnormalities was first organized and is currently maintained. The baseline statistics are provided and potential venues of postmarketing surveillance of drug teratogenicity and other public health tasks and research are proposed. METHODS: Cases with congenital abnormalities and patient controls with Down syndrome were selected from the Hungarian Congenital Abnormality Registry. Population controls without congenital abnormalities were selected from the National Birth Registry on the basis of three matching criteria: sex, week of birth, and district of parent's residence. Three sources of information concerning drug exposures, maternal disorders, and pregnancy complications, among others, were used: (1) prospective and medically recorded data from antenatal care logbooks and discharge summaries; (2) retrospective maternal self-reported data obtained with a structured questionnaire in all the three study groups; and (3) data collected by regional nurse in house visits to nonrespondent cases and patient controls. Twenty-five congenital abnormality groups were evaluated. During the 17-year period of data collection, 22,843 cases, 38,151 population controls, and 834 patient controls were incorporated into the data set, constituting the largest population-based case-control data set of congenital abnormalities to date. RESULTS: Demographic features of pregnant women and informative offspring are presented along with the distribution of 25 main groups of congenital abnormalities. CONCLUSIONS: This system is appropriate for postmarketing the surveillance of drug teratogenicity, for the improvement of congenital abnormality diagnosis, to get informed consent, to have a communication with parents and to provide material for research.     

根据在癌突变谱中的共突变基因对识别癌基因

癌是一种涉及多基因变异的遗传异质性疾病,涉及多种生物学功能通路中不同基因的遗传变异。因此,识别癌基因是一项富有挑战性的工作。提出通过寻找在癌样本中突变显著共发生的基因筛选候选癌基因的方法。应用该方法,通过分析蛋白激酶基因在癌组织中的突变谱数据,发现了167个显著共发生突变的基因对,包含85个基因。分析这167个基因对发现:(1)发生共突变的基因富集已知的癌基因;(2)共突变基因对倾向于共扰动与癌症相关的通路对。以上结果提示,在癌样本中显著共发生突变的基因倾向于候选癌基因;在癌发生过程中起重要作用的基因倾向于协同扰动不同的癌相关细胞生物学过程。     

Congenitally caused fused labia in the common marmoset (Callithrix jacchus)

In this paper, the occurrence of an external genital abnormality in female marmoset monkeys (fused labia) is discussed. This malformation was detected, for the first time, in a group of animals at the German Primate Center (GPC), Goettingen. The malformed vulva was completely sealed except for an opening of 1.5-2.5 mm around the urethra sufficient for urination. Because of this defect the animals were not able to copulate. As a consequence, the affected females were functionally infertile although they had a normal genital tract and a regular cycle. This vulvar abnormality was found in 12 females, offspring of 10 pairs in which either one or both came to the German Primate Center from two genetically related colonies in Munich, Germany, and one colony in Basel, Switzerland. The abnormality appeared to be recessive and inheritable from either parent. In pairs in which both animals were from one of the mentioned colonies, 45% of the female offspring were affected. In pairs where only one partner came from these colonies, 26% of female offspring had the malformation. These results indicate that avoidance of inbreeding, which is frequently performed in primate colonies, may reduce, but not eliminate the expression of abnormalities of genetic origin. Therefore selective breeding is required, and, in colonies where these recessive mutations are widespread, the development of genetic screening tests would be advantageous.     

Whole genome sequencing of pairwise human subjects reveals DNA mutations specific to developmental dysplasia of the hip

Developmental dysplasia of the hip (DDH) is a common congenital malformation characterized by mismatch in shape between the femoral head and acetabulum, and leads to hip dysplasia. To date, the pathogenesis of DDH is poorly understood and may involve multiple factors, including genetic predisposition. However, comprehensive genetic analysis has not been applied to investigate a genetic component of DDH. In the present study, 10 pairs of healthy fathers and DDH daughters were enrolled to identify genetic hallmarks of DDH using high throughput whole genome sequencing. The DDH-specific DNA mutations were found in each patient. Overall 1344 genes contained DDH-specific mutations. Functional enrichment analysis showed that these genes played important roles in the cytoskeleton, microtubule cytoskeleton, sarcoplasm and microtubule associated complex. These functions affected osteoblast and osteoclast development. Therefore, we proposed that the DDH-specific mutations might affect bone development, and caused DDH. Our pairwise high throughput sequencing results comprehensively delineated genetic hallmarks of DDH. Further research into the biological impact of these mutations may inform the development of DDH diagnostic tools and allow neonatal gene screening.     

Impaired affinity maturation in Cr2-/- mice is rescued by adjuvants without improvement in germinal center development

Cr2-/- mice have an impairment in humoral immunity, as shown by the decrease in the Ab titers against T cell-dependent Ags and abnormalities in germinal center formation. Germinal centers are present, but they are decreased in size and number, indicating problems in their development. In this study, we investigated whether this abnormality in germinal center development is associated with problems in the establishment of optimal affinity maturation and the generation of memory B cells, processes closely related to the germinal center reaction. We immunized the Cr2-/- animals with different Ags with or without adjuvants. We showed that, when immunized without adjuvants, complement receptors are absolutely required for optimal affinity maturation. Although limited affinity maturation is elicited in the Cr2-/- Ab response, it is decreased as compared with normal animals. Memory B cell generation is also impaired. In the presence of adjuvants, germinal center development in the Cr2-/- mice is still abnormal, as demonstrated by their decreased size and number. Surprisingly, adjuvants establish optimal affinity maturation and partially restore the amount of Ab produced during the primary response and memory B cell generation. However, adjuvants cannot improve the ability of follicular dendritic cells to retain Ags in the form of immune complexes. These observations indicate that immunization with inflammatory Ags offset some of the immunological abnormalities found in the Cr2-/- mice and show that optimal affinity maturation in the Cr2-/- mice can be achieved in the absence of normal germinal centers.     

Prevention of congenital abnormalities by periconceptional multivitamin supplementation.

OBJECTIVE--To study the effect of periconceptional multivitamin supplementation on neural tube defects and other congenital abnormality entities. DESIGN--Randomised controlled trial of supplementation with multivitamins and trace elements. SETTING--Hungarian family planning programme. SUBJECTS--4156 pregnancies with known outcome and 3713 infants evaluated in the eighth month of life. INTERVENTIONS--A single tablet of a multivitamin including 0.8 mg of folic acid or trace elements supplement daily for at least one month before conception and at least two months after conception. MAIN OUTCOME MEASURES--Number of major and mild congenital abnormalities. RESULTS--The rate of all major congenital abnormalities was significantly lower in the group given vitamins than in the group given trace elements and this difference cannot be explained totally by the significant reduction of neural tube defects. The rate of major congenital abnormalities other than neural tube defects and genetic syndromes was 9.0/1000 in pregnancies with known outcome in the vitamin group and 16.6/1000 in the trace element group; relative risk 1.85 (95% confidence interval 1.02 to 3.38); difference, 7.6/1000. The rate of all major congenital abnormalities other than neural tube defects and genetic syndromes diagnosed up to the eighth month of life was 14.7/1000 informative pregnancies in the vitamin group and 28.3/1000 in the trace element group; relative risk 1.95 (1.23 to 3.09); difference, 13.6/1000. The rate of some congenital abnormalities was lower in the vitamin group than in the trace element group but the differences for each group of abnormalities were not significant. CONCLUSIONS--Periconceptional multivitamin supplementation can reduce not only the rate of neural tube defects but also the rate of other major non-genetic syndromatic congenital abnormalities. Further studies are needed to differentiate the chance effect and vitamin dependent effect.     

International Commission for Protection Against Environmental Mutagens and Carcinogens. ICPEMC working paper no. 4. Mutation surveillance of sentinel anomalies in Hungary, 1980-1984

Sentinel phenotypes are indicators of germinal dominant gene mutations. 23 sentinel abnormalities and 2 sentinel childhood tumours (bilateral retinoblastoma and Wilms' tumour), i.e., 25 sentinel anomalies of autosomal dominant origin were selected from the material of the Hungarian Congenital Malformation Registry, 1980-1984. Furthermore, cases of sentinel childhood tumours from the Hungarian Childhood Tumour Registry and some extra notifications of sentinel abnormalities were also included. Experts examined index patients and their parents in order to confirm or exclude nosological diagnosis, to separate sporadic and familial cases, to obtain environmental history and to give genetic counselling. The revised total observed prevalence of 25 sentinel anomalies was 3.80 per 10,000 livebirths. Only 12% of cases examined were familial. According to the statistical power calculation, 47,500 livebirths are needed to detect a doubling of mutation rate with probabilities of type I and II errors of 0.05 level. In Hungary the average number of yearly births was 135,548 in the study period.     

Rates of mutant structural chromosome rearrangements in human fetuses: data from prenatal cytogenetic studies and associations with maternal age and parental mutagen exposure.

In 27,225 prenatal cytogenetic studies of amniotic fluid reported to the New York State Chromosome Registry and the United States Interregional Chromosome Register System, there were 61 cases with a structural chromosomal abnormality not known inherited, a rate per 1,000 of 2.24. Of these 33, 1.21 per 1,000 were known de novo and nonmosaic; consequently, the rate of events resulting from germinal mutation is highly likely to be between these two limits. The rates per 1,000 of unbalanced abnormalities were 0.59-1.29; of balanced abnormalities, 0.62-0.96; of balanced Robertsonian translocations, 0.22-0.29; and of unbalanced Robertsonian translocations, 0.07-0.11. The rates of fetuses with supernumerary markers and fragments were unexpectedly high: 0.26-0.70 per 1,000. These abnormalities were associated with increased maternal age (38.0 +/- 5.4 to 38.4 +/- 3.6 compared to 35.6 +/- 4.3 in controls), but even after adjustment for the bias to preferential study of older women, the observed rates of these supernumerary abnormalities were greater than would be expected from live-birth studies or rates estimated in all recognized conceptuses. There were trends to elevated maternal age for the group of all balanced rearrangements, and to diminished maternal age for the nonsupernumerary, non-Robertsonian unbalanced rearrangements. In 136 women studied primarily because of exposure to a putative mutagen, a de novo deletion and an inversion not known inherited were detected. The rate of abnormality in these 136, 1.47%, was significantly greater than the rate of abnormality in the remainder: 0.14%-0.22%.     

An infantile case of Zellweger syndrome presented with Kabuki-like phenotype

Zellweger syndrome is a peroxisomal disorder resulting from the mutations in PEX genes generally presenting in the neonatal period with profound hypotonia seizures, inability to feed, liver cysts with hepatic dysfunction, chondrodysplasia punctata. Kabuki make-up syndrome is a multiple congenital anomalies and mental retardation syndrome with characteristic facial appearance, skeletal abnormalities, dermatoglyphic abnormalities, mental retardation and short stature. Abnormal liver functions and some atypical findings were also reported in some patients with Kabuki syndrome. In this report a case with late onset Zellweger syndrome who had some phenotypical findings which are also seen in Kabuki Syndrome will be presented. The inclusion of Zellweger syndrome into the differential diagnosis of the patients with Kabuki-like phenotype in addition to abnormal liver functions is emphasized.     

A novel mutation in the putative DNA helicase XH2 is responsible for male-to-female sex reversal associated with an atypical form of the ATR-X syndrome.

We describe a pedigree presenting X-linked severe mental retardation associated with multiple congenital abnormalities and 46,XY gonadal dysgenesis, leading in one family member to female gender assignment. Female carriers are unaffected. The dysmorphic features are similar to those described in the alpha-thalassemia and mental retardation (ATR-X) syndrome, although there is no clinical evidence of alpha-thalassemia in this family. In addition, the family had other clinical features not previously observed in the ATR-X syndrome, including partial optic-nerve atrophy and partial ocular albinism. Mutations in a putative DNA helicase, termed XH2, have been reported to give rise to the ATR-X syndrome. We screened the XH2 gene for mutations in affected members of the family and identified a 4-bp deletion at an intron/exon boundary that removes an invariant 3' splice-acceptor site. The mutation cosegregates with the syndrome. The genomic deletion causes missplicing of the pre-mRNA, which results in the loss of 8 bp of coding sequence, thereby generating a frameshift and a downstream premature stop codon. Our finding increases the range of clinical features associated with mutations in the XH2 gene.