Novel mutation of the initiation codon of PAX9 causes oligodontia.

Tooth development is under strict genetic control. Oligodontia is defined as the congenital absence of 6 or more permanent teeth, excluding the third molar. The occurrence of non-syndromic oligodontia is poorly understood, but in recent years several cases have been described where a single gene mutation is associated with oligodontia. Several studies have shown that MSX1 and PAX9 play a role in early tooth development. We screened one family with non-syndromic oligodontia for mutations in MSX1 and PAX9. The pedigree showed an autosomal-dominant pattern of inheritance. Direct sequencing and restriction enzyme analysis revealed a novel heterozygous A to G transition mutation in the AUG initiation codon of PAX9 in exon 1 in the affected members of the family. This is the first mutation found in the initiation codon of PAX9, and we suggest that it causes haploinsufficiency.

Disease associated balanced chromosome rearrangements: a resource for large scale genotype-phenotype delineation in man.

Disease associated balanced chromosomal rearrangements (DBCRs), which truncate, delete, or otherwise inactivate specific genes, have been instrumental for positional cloning of many disease genes. A network of cytogenetic laboratories, Mendelian Cytogenetics Network (MCN), has been established to facilitate the identification and mapping of DBCRs. To get an estimate of the potential of this approach, we surveyed all cytogenetic archives in Denmark and southern Sweden, with a population of approximately 6.6 million. The nine laboratories have performed 71 739 postnatal cytogenetic tests. Excluding Robertsonian translocations and chromosome 9 inversions, we identified 216 DBCRs ( approximately 0.3%), including a minimum estimate of 114 de novo reciprocal translocations (0.16%) and eight de novo inversions (0.01%). Altogether, this is six times more frequent than in the general population, suggesting a causal relationship with the traits involved in most of these cases. Of the identified cases, only 25 (12%) have been published, including 12 cases with known syndromes and 13 cases with unspecified mental retardation/congenital malformations. The remaining DBCRs were associated with a plethora of traits including mental retardation, dysmorphic features, major congenital malformations, autism, and male and female infertility. Several of the unpublished DBCRs defined candidate breakpoints for nail-patella, Prader-Willi, and Schmidt syndromes, ataxia, and ulna aplasia. The implication of the survey is apparent when compared with MCN; altogether, the 292 participating laboratories have performed >2.5 million postnatal analyses, with an estimated approximately 7500 DBCRs stored in their archives, of which more than half might be causative mutations. In addition, an estimated 450-500 novel cases should be detected each year. Our data illustrate that DBCRs and MCN are resources for large scale establishment of phenotype-genotype relationships in man.

BRCA1 and BRCA2 mutation status and cancer family history of Danish women affected with multifocal or bilateral breast cancer at a young age.

INTRODUCTION: A small fraction of breast cancer is the result of germline mutations in the BRCA1 and BRCA2 cancer susceptibility genes. Mutation carriers frequently have a positive family history of breast and ovarian cancer, are often diagnosed at a young age, and may have a higher incidence of double or multiple primary breast tumours than breast cancer patients in general. OBJECTIVES: To estimate the prevalence and spectrum of BRCA1 and BRCA2 mutations in young Danish patients affected with bilateral or multifocal breast cancer and to determine the relationship of mutation status to family history of cancer. SUBJECTS: From the files of the Danish Breast Cancer Cooperative Group (DBCG), we selected 119 breast cancer patients diagnosed before the age of 46 years with either bilateral (n=59) or multifocal (n=61) disease. METHODS: DNA from the subjects was screened for BRCA1 and BRCA2 mutations using single strand conformation analysis (SSCA) and the protein truncation test (PTT). Observed and expected cancer incidence in first degree relatives of the patients was estimated using data from the Danish Cancer Registry. RESULTS: Twenty four mutation carriers were identified (20%), of whom 13 had a BRCA1 mutation and 11 carried a BRCA2 mutation. Two mutations in BRCA1 were found repeatedly in the material and accounted for seven of the 24 (29%) mutation carriers. The mutation frequency was about equal in patients with bilateral (22%) and multifocal breast cancer (18%). The incidence of breast and ovarian cancer was greatly increased in first degree relatives of BRCA1 and BRCA2 mutation carriers, but to a much lesser degree in relatives of non-carriers. An increased risk of cancer was also noted in brothers of non-carriers. CONCLUSIONS: A relatively broad spectrum of germline mutations was observed in BRCA1 and BRCA2 and most of the mutations are present in other populations. Our results indicate that a diagnosis of bilateral and multifocal breast cancer is predictive of BRCA1 and BRCA2 mutation status, particularly when combined with information on the patients' age at diagnosis and family history of breast/ovarian cancer.

Sister chromatid exchange in patients treated with methotrexate for psoriasis.

Sister chromatid exchange (SCE), which is considered a sensitive method in evaluation of mutagenicity and may be an indicator of carcinogenicity, was determined in lymphocytes from 8 patients with severe psoriasis who underwent treatment with oral methotrexate (MTX). During the treatment period (median 118 days, range 50-298 days) a significant increase in SCE compared to the pretreatment value was found in 1 patient only. It is concluded that no long-term damage as measured by SCE occurs in lymphocytes from psoriasis patients treated with MTX.

Lymphocyte function and chromosome aberrations in patients with early mycosis fungoides and parapsoriasis en plaques.

Thirteen patients with stage I or II mycosis fungoides (MF) and 10 patients with large-plaque parapsoriasis en plaques (PEP) were examined for immunologic and cytogenetic disturbances. Total lymphocyte counts and immunoglobulin concentrations in the blood were normal. In vitro lymphocyte responses to polyclonal activators and various antigens in standard concentrations were normal. However, titration of phytohemagglutinin and concanavalin A (ConA) disclosed significantly lowered responses to suboptimal concentrations in the patient group, most pronounced in patients with MF II. ConA-induced leukocyte migration inhibitory factor (LIF) production, tested in an indirect leukocyte migration inhibitory assay, was low in the patient group. Furthermore spontaneous LIF production in vitro and small amounts of serum LIF were demonstrated in a few patients. The chromosomal banding pattern, sister chromatid exchange, and break frequency were within normal limits except for 3 translocations in the MF group. It is concluded that even in early-stage MF a pathologic function of blood lymphocytes can be demonstrated, when sensitive methods are applied. The findings might be important for monitoring disease activity and effect of treatment.

Studies of the cranial base in 23 patients with cri-du-chat syndrome suggest a cranial developmental field involved in the condition.

The purpose of the present study was to investigate the cranial base on profile radiographs of patients with cri-du-chat syndrome and to relate the findings to current knowledge of brain malformation in an attempt to localize the developmental field affected in cri-du-chat syndrome. The material of profile radiographs of 23 patients was collected in Denmark in the 1970s. Twenty-two patients had terminal deletions of chromosome 5 (5p13.3, 5p14.1, 5p14.2, and 5p14.3), and one patient had an interstitial deletion. The cranial base angle (n-s-ba) was in most cases reduced and in no cases increased compared to age-related standards for normal individuals. Malformations in the bony contours of the sella turcica and the clivus occurred in cri-du-chat patients with terminal deletions. This specific cranial base region develops around the notochord at the location from where the rhombencephalic-derived brainstem, pons, and cerebellum have developed dorsally, and from where the neurons to the larynx have migrated ventrally. As the cranial base, the cerebellum and the larynx are involved in cri-du-chat syndrome, and attention is drawn to a new developmental field which comprises the dorsum sellae, clivus, cerebellum, and larynx. This field seemingly originates from the same notochordal location. The study has demonstrated a cranial base malformation in cri-du-chat patients, which ought to be elucidated in future research and combined with neurological and chromosomal investigations.

Parental origin of chromosome 5 deletions in the cri-du-chat syndrome.

The parental origin of de novo deletions leading to the cri-du-chat syndrome has been investigated. Since the cri-du-chat syndrome is correlated with deletions involving the short arm of chromosome 5 (5p), DNA fragments known to detect restriction fragment length polymorphisms (RFLPs) along 5p were used to establish whether the paternal or the maternal chromosome had suffered the deletion. In cases where only one parent was available, somatic cell hybrids were used in conjunction with RFLP analysis to determine the origin of the deleted chromosome. The deleted chromosome 5 was of paternal origin in 20/25 cases.

Normal sister chromatid exchange in lymphocytes from patients with multiple epidermal cancer?

Exposure to toxins in the environment and due to personal habits, e.g., tobacco smoking, may increase the rate of spontaneous sister chromatid exchange (SCE). The SCE in lymphocytes from a group of 31 patients with multiple epidermal cancer, who in the past had been exposed to various skin carcinogens, as a whole exceeded that of a control group--matched by sex, age, and smoking habits--but the difference was not statistically significant (p = 0.08). The individual SCE in these patients was also statistically independent of the nature of the carcinogenic exposure. We were unable to detect correlations between the SCE and UVC-radiation induced DNA synthesis, UVC-radiation tolerance, or rate of X-ray damage repair. This suggests that the molecular mechanisms involved in SCE induction and in repair of radiation damage are basically independent.

Sister-chromatid exchange in childhood in relation to age and sex.

Small children have been found to have a lower SCE/cell than adults and in recent reports females have had higher SCEs/cell than males. We here describe the relationship between SCE/cell and age and sex in 46 girls and 39 boys with an age range of 1.4-19.2 years and 2.6-18.7 years, respectively. For the calculation a transformation y = (sum SCE)1/2 + (sum SCE + 1)1/2 was used. The best fit to our material was represented by the equation y = b0 + b1 X log age. A common slope (b1) could be used for the boys and girls. This slope was significantly different from zero (P less than 0.0005). The levels of the regression lines for the two sexes were different (P = 0.0006). The girls had a 0.55-0.7 higher SCE/cell than the boys, depending on age. The following equations were found: Girls: y = 22.49 + 6.53 X log age. Boys: y = 21.11 + 6.53 X log age. By this model 43% of the variation in y could be explained. As a consequence of the result it is absolutely essential, when planning studies of children, to use age-matched groups to decrease the variability of the test system.

Sister-chromatid exchanges in lymphocytes in women with cancer of the breast.

Examination of sister-chromatid exchanges (SCE) in lymphocytes may be useful for the evaluation of exposure to mutagens/carcinogens. Information of a possible association between SCE and cancer is scarce. We therefore examined SCE in peripheral lymphocytes in 131 women, aged 17-90 years (median 51.8 years), coming to operation because of a tumor of the breast. Venous blood samples were cultivated during PHA stimulation in the presence of BrdU. After treatment with colcemid (R), fixation, treatment with bisbenzimide and staining with Giemsa, 30 metaphases were scored in each specimen. 52 patients with peroperatively demonstrated carcinoma of the breast had 9.39 +/- 0.17 SCE/cell and the remaining 79 women with non-malignant fibroadenomatosis had 9.88 +/- 0.18 SCE/cell. By multiple regression analysis it appeared that the character of the tumor, the patient's age, hormone treatment and preoperative examination by mammography all were without significant influence on the SCE rate. A statistically significant correlation was found between SCE and cigarette smoking. THe 45 cigarette-smoking patients had 10.49 +/- 0.23 SCE/cell compared with 9.26 +/- 0.13 SCE/cell in the 86 non-smokers. It was concluded that spontaneous SCE in lymphocytes is not an indicator of carcinoma of the breast.

Normal sister-chromatid exchanges in oral contraceptive users.

Sister-chromatid exchanges in peripheral lymphocytes were examined in 115 healthy women, aged 15-43 years. 63 women were on no medication at all, and 52 women used oral contraceptives. 30 metaphases from each culture were scored for SCE. In accord with previous studies, cigarette smoking increased SCE. The use of oral contraceptives had no influence on SCE on non-smoking and in cigarette-smoking women. Thus the results did not indicate any potential mutagenic effect of oral contraceptives.

Sister-chromatid exchanges in cannabis smokers.

The genotoxicity of cannabis smoking was evaluated by means of the sister-chromatid exchange (SCE) test. The SCE test is considered to be a sensitive tool for the discovery of genotoxic agents in the environment. Twenty-two tobacco smokers and 22 persons smoking both tobacco and cannabis were compared. Our findings showed that smoking in itself enhanced the SCE level significantly (18.5%) compared to a group of non-smokers, but adding smoking of cannabis to tobacco smoking did not affect the SCE level further. Based on our observations cannabis smoking could not be considered genotoxic.

SCE in lymphocytes of patients treated with single, large doses of diazepam.

When using the SCE test for evaluation of exposure in vivo to potential mutagens/carcinogens, it is necessary to consider possible confounding factors. In studies of possible mutagenic effects of pharmacological treatment the effect of concomitant administration of other agents such as sedatives may have to be considered. In order to assess whether diazepam per se influences SCE we have examined SCE in peripheral lymphocytes in 34 persons before and after oral administration of a single large dose of diazepam. 18 men and 16 women undergoing minor surgery of the hand received diazepam 0.2 mg kg-1 body weight orally on the day of operation, and venous blood samples were drawn on the day before the operation and again 2-5 h after the administration of diazepam. Both within cigarette smokers and non-smokers there was no statistically significant change of SCE following diazepam. It was concluded that there was no indication, from the SCE test, of an immediate mutagenic effect of a single large dose of diazepam and that such medication is not a confounding factor in studies by the SCE test.

Very low sister-chromatid exchange rate in Seventh-Day Adventists.

42 Seventh-Day Adventists (SDAs) and 42 controls matched for sex, age and occupation had their sister-chromatid exchange (SCE) examined in peripheral blood lymphocytes. This was done to examine if the SCE frequency was lower in this group of people, who are known to have a decreased cancer risk compared to the general population. The average SCE/cell in 30 cells from each person was 5.54 +/- 0.07 (mean +/- standard error of the mean) for the SDAs and 8.00 +/- 0.15 for the controls, the difference being statistically significant (p less than 0.00001). No difference in SCE frequency was found between SDAs eating only an ovo-lacto-vegetarian diet and those eating some fish or meat. The mitotic index (MI) was significantly higher and the replication index (RI) was significantly lower in SDAs than in controls. No correlation was found between gamma (a statistical transformation of SCEs/cell) and MI or RI within the groups of SDAs or controls. In the pooled data there was a negative correlation of gamma and MI and a positive correlation of gamma and RI. Of the interpersonal variation in gamma 8% and 14% could be explained by MI and RI. The finding of a lower SCE frequency in a group of SDAs who have a low risk of cancer might indirectly indicate a relation between SCE and cancer and encourages further studies of SCE and diet.

Distribution of SCEs in lymphocytes in persons with normal, slightly increased, and heavily increased SCEs.

The distribution of SCEs in lymphocytes was examined for 165 healthy persons (58 non-smokers and 107 smokers with cigarette consumption ranging from 1 to greater than 20 per day), and for 1 patient treated with melphalan, a cytostatic drug. The data from the healthy persons did not follow a Poisson distribution. A mixed Poisson that allowed different lambda values for the 30 cells scored from each person and postulated a gamma distribution for the lambda s within the 30 cells fitted all the data examined including those from the melphalan-treated patient. In the latter case the 7 samples taken at various times after the treatment could all be represented satisfactorily with a common parameter, c, in the gamma distribution for the lambda s, even though the mean SCEs/cell varied from 9.8 to 36.8. Because the c parameter determines the spread of lambda values within the 30 cells, this suggested that the effect of the cytostatic drug was to increase all the lambda s by a constant amount. The sum of the SCEs taken over all 30 cells in a sample is a convenient summary statistic, and the transformation y = square root s + square root s + 1 behaves as a normal variate with a constant variance within a group.

Sister chromatid exchange (SCE) in Greenlandic Eskimos. Dose-response relationship between SCE and seal diet, smoking, and blood cadmium and mercury concentrations.

The mutagenicity of the chromosomes of the peripheral lymphocytes of 147 Greenlandic Eskimos living in the district of Angmagssalik, Greenland, and in Denmark, was evaluated by means of the sister chromatid exchange (SCE) test. Thirty cells from each person were examined. The purpose of the investigation was to determine if there was any relationship between mutagenic activity and diet, and hence the elements selenium, cadmium, mercury and lead. The probands were divided into three groups according to their intake of seal meat or industrially prepared food: group 1, those eating seal at least six times per week; group 2, two to five times per week; and group 3 once each week or not at all. The statistical analysis was performed by means of multiple linear regression analyses, with diet, living district, sex, age, tobacco smoking, and blood lead and mercury concentrations as variables. Forty-eight percent of the variation in SCE could be explained by differences in diet, living district, age, and tobacco consumption. Groups 1 and 2 had a 1.7 and 0.65 times higher SCE/cell, respectively, than group 3. For every additional 10 years of age of the probands, the SCE/cell increased by 0.4, and for every 10 g of tobacco smoked per day the SCE/cell was 0.7 higher compared to non-smokers. When priority was given to blood Hg concentration in the calculation, 16.3% of the total variation in SCE/cell could be explained. An increase in the blood Hg concentration of 10 micrograms l-1 corresponded to an increase of 0.3 SCE/cell. In 92 individuals blood Se and Cd concentrations were also analysed. The variables, tobacco smoking, diet, living district and Cd explained 53% of the total variation in SCE. Giving priority to the blood Hg and Cd concentrations, explained 21.4% of the total variation in SCE/cell. An increase of 10 micrograms l-1 in blood Cd and Hg corresponded to an increase in SCE/cell of 0.7 and 0.2, respectively. No influence on the SCE/cell could be attributed to the blood Pb and Se concentrations. Evaluated by the SCE test, seal diet, smoking, living district and blood Hg and Cd concentrations all contribute to mutagenicity in Greenlandic Eskimos, with seal diet as the most important of the factors examined.

Sister chromatid exchanges in lymphocytes of dentists and chairside assistants: no indication of a mutagenic effect of exposure to waste nitrous oxide.

Cytogenetic methods are used increasingly for monitoring occupational exposure to potential mutagenic agents. By one such method, the sister chromatid exchange (SCE) test, previous studies of hospital operating room personnel did not indicate any mutagenic effect from exposure to waste anesthetic gases. Dentists and their assistants are exposed to much higher levels of nitrous oxide than hospital personnel, and in studies of dentists it is possible to eliminate bias from concomitant exposure to halogenated anesthetics by including unexposed persons engaged in the same professional activities as the exposed persons. SCE was examined in lymphocytes of venous blood from 38 female dentists and 74 chairside assistants, all working in the Child Dental Service, and 30 male dentists, of whom 23 worked in private practice. From studies of the model y = bo + b1 X age + b2 X cig./day + b3 X hours of exposure to nitrous oxide/week, by multiple linear regression, there was no indication of any influence of exposure to nitrous oxide on SCE. It was concluded that, from the SCE test, there is no indication of a mutagenic effect from exposure to waste nitrous oxide, and that possible health hazards from such exposure would be induced by other effects of nitrous oxide.

[Preclinical and prenatal diagnosis of familial adenomatous polyposis]. / Praeklinisk og praenatal diagnostik af familiaer adenomatøs polypose.

In order to investigate the possibility of preclinical and prenatal genetic diagnosis of familial adenomatous polyposis (FAP) by means of DNA-systems and other markers, blood samples were collected from 246 persons in 29 families, including 90 with the clinical diagnosis FAP and 73 clinically unaffected first degree relatives (persons at risk). The material was studied with up to 4 DNA-marker systems located in the region around the disease gene. Among the first degree relatives eight (11%) had probably inherited the disease gene, while 31 persons (42%) in this risk group had probably not inherited the gene. It was not possible to evaluate the risk in the remaining 34 persons (47%). In 45 (85%) out of 53 persons under 40 years the DNA-systems were informative, so that it would be possible to offer the option of prenatal diagnosis. It is concluded that preclinical and possibly prenatal genetic diagnosis may be offered; but the current practice of prophylactic proctosigmoidoscopic surveillance should be maintained.