Prenatal exposure to cigarette smoke or alcohol and cerebellum volume in attention-deficit/hyperactivity disorder and typical development.

Prenatal exposure to teratogenic substances, such as nicotine or alcohol, increases the risk of developing attention-deficit/hyperactivity disorder (ADHD). To date, studies examining this relationship have used symptom scales as outcome measures to assess the effect of prenatal exposure, and have not investigated the neurobiological pathways involved. This study explores the effect of prenatal exposure to cigarettes or alcohol on brain volume in children with ADHD and typically developing controls. Children with ADHD who had been exposed prenatally to either substance were individually matched to children with and without ADHD who had not been. Controls who had been exposed prenatally were also individually matched to controls who had not been. For prenatal exposure to both smoking and alcohol, we found a pattern where subjects with ADHD who had been exposed had the smallest brain volumes and unexposed controls had the largest, with intermediate volumes for unexposed subjects with ADHD. This effect was most pronounced for cerebellum. A similar reduction fell short of significance for controls who had been exposed to cigarettes, but not alcohol. Our results are consistent with an additive effect of prenatal exposure and ADHD on brain volume, with the effects most pronounced for cerebellum.

Morbidity and quality of life after continent ileostomy in the Netherlands.

BACKGROUND: Continent ileostomy (CI) after proctocolectomy is an alternative technique compared to an ileal pouch-anal anastomosis (IPAA). The question arises as to whether this technique is valuable. The aim of this study was to evaluate the role of the continent ileostomy, by patient follow-up satisfaction and quality of life assessment. METHOD: Twenty-eight patients with a continent ileostomy operated between 1996 and 2007 were compared with patients who received an IPAA or a conventional ileostomy. SF-36 and EORTC QLC-CR38 questionnaires and a specific continent ileostomy questionnaire were used to assess differences and patient satisfaction. RESULTS: The quality of life in patients with a CI is not significant better or worse than patients with either a conventional ileostomy or an IPAA. On three scales (sexual enjoyment, gastro-intestinal tract symptoms and male sexual problems) statistically significant differences were reported. Overall, nearly all patients are very satisfied with the CI. All patients would make the same decision again and would recommend this procedure to other patients. CONCLUSION: The continent ileostomy remains to be a suitable alternative for the preservation of continence after a proctocolectomy, especially when an ileal pouch-anal anastomosis is not an option. If a choice has to be made between a CI and conventional ileostomy good preoperative counselling is necessary to make a well founded decision. To minimize complications, these procedures have to be performed in centres with specific expertise. Therefore, knowledge about the CI should be preserved for the future.

Chemotherapy-induced chromosomal damage in peripheral blood lymphocytes of cancer patients supplemented with antioxidants or placebo.

A total of 27 patients with various types of cancer were treated with cisplatin-based combination chemotherapy. Out of these, 13 patients were randomized to receive supplementation treatment with a beverage containing the antioxidants vitamins C and E, plus selenium, during chemotherapy. The antioxidant mixture was administered to investigate whether it could reduce the potential genotoxic and nephrotoxic effect of the applied chemotherapy. A placebo group of 14 cancer patients received a beverage without selenium or antioxidants. Micronuclei (MN) in cytochalasin B-blocked binucleate (BN) peripheral blood lymphocytes (PBLs) and hypoxanthine phosphoribosyl transferase (HPRT) mutants in PBLs were studied before, during and after chemotherapy as a measure for chemotherapy-induced genotoxic effects. Before chemotherapy, patients mean frequencies of MN and HPRT mutants did not differ from those in a group of 10 healthy subjects. The mean frequency of MN in patients increased significantly after one cycle of chemotherapy (P=0.002). This frequency was still elevated at 2 months after the completion of chemotherapy (not significantly). There was no significant difference in micronuclei frequency (MNF) between the antioxidant and placebo group of patients. Chemotherapy-induced frequencies of MN after three cycles of chemotherapy correlated significantly with the cumulative dose of cisplatin (r=0.58, P=0.012) and the cisplatin-mediated loss of renal function (r=0.53, P=0.03). No consistent change in HPRT mutant frequency following chemotherapy was observed in the placebo and antioxidant group of patients. In conclusion, cisplatin-combination chemotherapy resulted in a cisplatin dose-related increase of the frequency of chromosomal damage. Supplementation with antioxidants did not prevent or reduce this effect.

Association between genetic polymorphisms and biomarkers in styrene-exposed workers.

A comprehensive approach to evaluate genotoxic effects induced by styrene exposure was employed in 44 hand-lamination workers in comparison with 18 unexposed controls. The acquired data on single-strand breaks in DNA (SSBs), frequency of chromosomal aberrations and HPRT mutant frequency in peripheral blood lymphocytes were compared to the results on genotyping of some of the xenobiotic-metabolising enzymes (CYP1A1, CYP2E1, epoxide hydrolase and GSTM1, GSTP1 and GSTT1). Multifactorial regression analysis indicated that SSB in DNA were significantly associated with styrene exposure and with heterozygosity in CYP2E1 (5'-flanking region and intron 6; r(2)=0.614). The frequency of chromosomal aberrations (CA), as analysed by linear multiple regression analysis, significantly correlated with years of employment (P=0.004) and with combinations of epoxide hydrolase (EPHX) genotypes (exon 3, Tyr/His and exon 4, His/Arg), where individuals with low and medium activity EPHX genotypes exhibited higher frequencies of CA than those with high activity genotypes (P=0.044, r(2)=0.563). Moderately higher HPRT mutant frequencies were detected in styrene-exposed individuals (20.2 +/- 25.8 x 10(-6)) as compared to controls (13.3 +/- 6.3 x 10(-6)), but this difference was not significant. ANOVA (in the whole set of data) revealed that mutant frequencies at the HPRT gene were significantly associated with years of employment (F=6.9, P=0.0001), styrene in blood (F=10.1, P=0.0001), and heterozygosity in CYP2E1 (intron 6; F=13.5, P=0.0008) and GSTP1 (exon 5; F=3.6, P=0.038). In conclusion, our present data suggest that analysed biomarkers of DNA damage may be modulated by polymorphic CYP2E1, EPHX and GSTP1. In our study, styrene-specific DNA and haemoglobin adducts are under investigation. Completing these data with the results of genotyping of metabolising enzymes may provide a useful tool for individual genotoxic risk assessment.

Radiation effects in lymphocytes of children living in a Chernobyl contaminated region of Belarus.

PURPOSE: To investigate cytogenetic and mutational effects in lymphocytes from individuals chronically exposed to radiation from the Chernobyl catastrophe. MATERIALS AND METHODS: Nine years after the Chernobyl accident (1986), peripheral blood lymphocytes from 20 Kalinkovichi children (age 10-15) and 10 Minsk children (age 10-17) were analysed for genetic damage by several assays. Radiation damage in exposed children was investigated in descendants of progenitor cells that were irradiated during a short period immediately after the accident. In the time-span between the accident and blood sampling the cells were also irradiated chronically by internal radiation originating from ingested radionuclides and, to a smaller extent, by external radiation from radionuclides. The parameters measured in whole blood smears were the frequency of micronucleated mononucleated lymphocytes and binucleated lymphocytes with nucleoplasmic bridges and associated micronuclei. Cultures of cytokinesis-blocked lymphocytes were used to analyse mononuclear and binuclear cells for the presence of micronuclei, also cell killing effects. A colony assay was used to study induction of recessive mutations in the HPRT gene. RESULTS: The analysis of whole-blood smears indicated a doubling of the frequency of micronuclei per 100 mononuclear lymphocytes in exposed children compared with unirradiated children. Small numbers of binucleated lymphocytes with nucleoplasmic bridges and associated micronuclei were found in blood smears from exposed children. Analysis of cytokinesis-blocked cultures indicated in mononuclear cells of exposed children a statistically significant increase in the frequency of micronuclei. When the same parameters were studied in binucleated cells there was no difference between exposed and unexposed children. Results of the dye-exclusion assay showed a four-fold increase in the percentage of dead cells between exposed and unexposed children. There was no evidence for induction of HPRT mutations in exposed children. CONCLUSION: These results indicate that the frequently advocated procedure of simply analysing micronuclei in cytokinesis-blocked binucleated lymphocytes can result in an underestimate of genetic damage induced by radiation accidents. Biodosimetric studies should therefore employ a battery of assays for the detection of several types of genetic damage in different generations of lymphocytes.

Validation of the human T-lymphocyte cloning assay--ring test report from the EU concerted action on HPRT mutation (EUCAHM).

The T-cell cloning assay, which enables the enumeration and molecular analysis of 6-thioguanine resistant (HPRT-negative) mutant T-cells, has been extensively used for studying human somatic gene mutation in vivo. However, large inter-laboratory variations in the HPRT mutant frequency (MF) call for further investigation of inter-laboratory differences in the experimental methodology, and development of an optimal but easy uniform cloning protocol. As part of the EU Concerted Action on HPRT Mutation (EUCAHM), we have carried out two Ring tests for the T-cell cloning assay. For each test, duplicate and coded samples from three buffy coats were distributed to five laboratories for determination of MF using six different protocols. The results indicated a good agreement between split samples within each laboratory. However, both the cloning efficiencies (CEs) and MFs measured for the same blood donors showed substantial inter-laboratory variations. Also, different medium compositions used in one and the same laboratory resulted in a remarkable difference in the level of MF. A uniform operating protocol (UOP) was proposed and compared with the traditional protocols in the second Ring test. The UOP (preincubation) increased the CE in laboratories traditionally using preincubation, but decreased the CE in laboratories traditionally using priming. Adjusted for donor, use of different protocols contributed significantly to the overall variation in lnCE (P = 0.0004) and lnMF (P = 0.03), but there was no significant laboratory effect on the lnCE (P = 0.38) or lnMF (P = 0.14) produced by the UOP alone. Finally, a simplified version of the UOP using the serum-free medium X-Vivo 10 and PMA was tested in one laboratory, and found to produce a considerable increase in CE. This modified UOP needs to be further evaluated in order to be used for future databases on HPRT MFs in various populations.

Measurement of HPRT mutations in splenic lymphocytes and haemoglobin adducts in erythrocytes of Lewis rats exposed to ethylene oxide.

Young adult male Lewis rats were exposed to ethylene oxide (EO) via single intraperitoneal (i.p.) injections (10-80 mg kg-1) or drinking water (4 weeks at concentrations of 2, 5, and 10 mM) or inhalation (50, 100 or 200 ppm for 4 weeks, 5 days week-1, 6 h day-1) to measure induction of HPRT mutations in lymphocytes from spleen by means of a cloning assay. N-ethyl-N-nitrosourea (ENU) and N-(2-hydroxyethyl)-N-nitrosourea (HOENU) were used as positive controls. Levels of N-(2-hydroxyethyl)valine (HOEtVal) adducts in haemoglobin (expressed in nmol g-1 globin) were measured to determine blood doses of EO (mmol kg-1 h, mM h). Blood doses were used as a common denominator for comparison of mutagenic effects of EO administered via the three routes. The mean HPRT mutant frequency (MF) of the historical control was 4.3 x 10(-6). Maximal mean MFs for ENU (100 mg kg-1) and HOENU (75 mg kg-1) were 243 x 10(-6) and 93 x 10(-6), respectively. In two independent experiments, EO injections led to a statistically significant dose-dependent induction of mutations, with a maximal increase in MF by 2.3-fold over the background. Administration of EO via drinking water gave statistically significant increases of MFs in two independent experiments. Effects were, at most, 2.5-fold above the concurrent control. Finally, inhalation exposure also caused a statistically significant maximal increase in MF by 1.4-fold over the background. Plotting of mutagenicity data (i.e., selected data pertaining to expression times where maximal mutagenic effects were found) for the three exposure routes against blood dose as common denominator indicated that, at equal blood doses, acute i.p. exposure led to higher observed MFs than drinking water treatment, which was more mutagenic than exposure via inhalation. In the injection experiments, there was evidence for a saturation of detoxification processes at the highest doses. This was not seen after subchronic administration of EO. The resulting HPRT mutagenicity data suggest that EO is a relatively weak mutagen in T-lymphocytes of rats following exposure(s) by i.p. injection, in drinking water or by inhalation.

Comparison of induction of hprt mutations by 1,3-butadiene and/or its metabolites 1,2-epoxybutene and 1,2,3,4-diepoxybutane in lymphocytes from spleen of adult male mice and rats in vivo.

Induction of hprt mutations by 1,3-butadiene (BD) and its metabolites 1,2-epoxybutene (EB) and 1,2,3,4-diepoxybutane (DEB) was studied in lymphocytes from spleens of 6- to 14-week-old mice and 10- to 11-week-old rats. For unknown reasons, results from experiments with mice that received inhalation exposure to BD were quite variable. In the first experiment, mice were exposed for 5 days to 200, 500 or 1300 ppm and this resulted in a statistically significant, dose-dependent, induction of mutations. When the experiment was repeated and an extra expression time for mutations was included, it was not possible to detect induction of mutations. In a third experiment, a 6-day exposure to 500 ppm was mutagenic when mice with zero mutants were not excluded from the statistical analysis of the data. The monofunctional metabolite EB appeared to be mutagenic in mice (3 x 33 and 3 x 100 mg/kg), but not in rats (3 x 33 and 100 mg/kg or 30 days drinking water with 0.1, 0.3, or 1.0 mM EB). Contrary to expectations, there was no induction of mutations in mice and rats exposed to the bifunctional metabolite DEB (mice, 3 x 7, 21, 3 x 14, or 42 mg/kg; rats, 20 or 40 mg/kg or 30 days drinking water with 0.3 or 1 mM DEB), although in our earlier studies with mice and rats, DEB treatment significantly enhanced frequencies of micronuclei in splenocytes and in early spermatids of mice and rats. Some of these results differ from findings reported by other investigators. It is now becoming evident that these differences are, to a large extent, due to differences in age of the animals at the time of treatment. For example, the mutagenic potency of BD, EB and DEB was stronger in preweanling mice or 4-week-old mice than in 8- to 12-week-old adult mice.

Biological effect monitoring in industrial workers from the Czech Republic exposed to low levels of butadiene.

Blood samples were collected twice (in 1993 and 1994) from 19 workers exposed to 1,3-butadiene and 19 matched controls. Three exposed and three control subjects were the same in 1993 and 1994. Personal passive dosimetry was performed in 1993 and twice in 1994 on the day preceding blood sampling. Mean exposure level in 1994 was 1.76 +/- 4.20 ppm (S.D.) and individual exposure levels ranged between 0.012 ppm (detection limit) and 19.77 ppm. Using the clonal assay, geometric mean of hprt mutant frequencies adjusted for cloning efficiency, age and smoking were, respectively, 7.85 (+/- 7.09) x 10(-6) and 10.14 (+/- 9.16) x 10(-6) in pooled (1993 plus 1994) exposed and control subjects. The difference was not statistically significant indicating that 1,3-butadiene did not induce a detectable increase in mutations at the hprt locus. A similar result was obtained for the 1994 subjects alone. There was no difference between adjusted geometric mean mutant frequencies of exposed and unexposed non-smokers or between exposed and unexposed smokers. Analysis of chromosomal aberrations in lymphocytes from 1994 subjects indicated that the percentage of aberrant cells was significantly enhanced in exposed subjects. In 1993 (data not shown), it was impossible to demonstrate a significant increase of aberrant cells in subjects exposed to 1,3-butadiene. Frequencies of micronuclei in cytochalasin-B blocked binucleate lymphocytes in exposed and unexposed 1994 subjects were not significantly different. This was also the case for earlier samples analyzed in the same plant. Using the comet assay for 1994 subjects, no statistically significant difference was found between the whole group of exposed and unexposed subjects. This was true for both the comet tail length and the percentage of DNA in the tail. In exposed smokers, however, the comet tail length was significantly longer than in unexposed smokers. Unexpectedly, in unexposed smokers the tail length was significantly shorter than in unexposed non-smokers. It was also unexpected that the percentage of DNA in the comet tail was significantly lower in exposed non-smokers than in unexposed non-smokers.

In vivo sensitivity of Plasmodium falciparum to chloroquine in rural areas of Côte d'Ivoire.

In vivo testing of Plasmodium falciparum sensitivity to chloroquine was carried out in four rural sites of differing socio-geographical environment in Côte d'Ivoire. Of a total of 1282 patients of all ages with fever or previous history of fever, 649 were slide positive, with 435 patients with a pure P. falciparum infection; 191 fulfilled all the criteria for inclusion in this study, and 113 completed it. Treatment failure rates ranged from 9.7% (Djébonoua) to 38.1% (Tiéviéssou), and were most often associated with higher degrees of resistance (RII = 54.2%; RIII = 37.5%). Blood chloroquine levels measured by ELISA test suggest that many people take chloroquine routinely; furthermore 37.5% of resistance cases occurred in subjects who had high blood chloroquine concentrations on day 0. Twenty-three out of 24 cases of resistance were found in children under 7 years of age. Nearly all children with persisting parasitaemia were afebrile on day 7, even those (7/8) with RIII resistance. Children aged < 7 years represent the the best sentinel group for monitoring P. falciparum sensitivity to chloroquine in Côte d'Ivoire.

Measurement of frequencies of HPRT mutants, chromosomal aberrations, micronuclei, sister-chromatid exchanges and cells with high frequencies of SCEs in styrene/dichloromethane-exposed workers.

Frequencies of HPRT mutants (MFs), chromosomal aberrations with or without gaps (CA+; CA-), aberrant cells (AC), micronuclei (MN), sister-chromatid exchanges (SCEs) and cells with high frequencies of SCEs (HFCs) were measured in lymphocytes collected from 46 workers occupationally exposed to styrene and dichloromethane (DCM = methylene chloride). These parameters were also determined in 23 controls. Time-weighted average (TWA) values for styrene and DCM exposure during an 8-h working day were respectively 70 mg/m3 (range: 0-598) and 108 mg/m3 (range: 0-742). These values correspond to TWA values of 17 ppm styrene and 31 ppm DCM. In exposed workers, all cytogenetic parameters were significantly enhanced (P < 0.0001; one-sided), but, due to the lack of appropriate control data, no definite conclusions could be drawn concerning the mutagenicity of styrene/DCM exposure. Duration of exposure was not correlated with genetic effects analyzed. The TWA value for styrene was not correlated with the extent of genetic damage detected, but the TWA value for DCM was positively correlated with the frequencies of chromosome aberrations (with gaps) and aberrant cells. These observations make it difficult to decide whether styrene or DCM, or both chemicals, induced the cytogenetic effects observed in exposed workers. Using the present styrene/DCM data, earlier ethylene oxide data and unpublished epichlorohydrin data, the relative sensitivity of the genetic endpoints to detect genotoxic exposure was: HFC > CA- > CA+ > SCE > MN > HPRT.

Development of a cloning assay with high cloning efficiency to detect induction of 6-thioguanine-resistant lymphocytes in spleen of adult mice following in vivo inhalation exposure to 1,3-butadiene.

A cloning assay with high cloning efficiency has been developed to detect spontaneous and induced 6-thioguanine-resistant T-lymphocytes (HPRT mutants) from the spleen of adult mice. The mean cloning efficiency in untreated male mice of 20-22 weeks old was 34.5 +/- 11.2% (SD) and the corresponding mutant frequency 0.7 +/- 0.8 (SD) x 10(-6). The cloning efficiencies obtained in this study are substantially higher than those reported previously by other investigators. Using this assay, it could be demonstrated that inhalation exposure of mice to 200, 500 or 1300 ppm of 1,3-butadiene for 6 h/day on 5 consecutive days caused a statistically significant induction of 6-thioguanine-resistant mutations in T-lymphocytes from spleens of adult mice exposed to 1300 ppm. The exposure to 1300 ppm resulted in a three-fold increase of the spontaneous mutant frequency. The mutant frequency after exposure to 500 ppm was higher than the control but the increase was not significant.

Psychoeducation of partners of bipolar-manic patients.

The partners of 14 bipolar-manic patients attending psychoeducation sessions were compared with 12 controls. After the sessions and 6 months later they showed more knowledge of the disease, medication and social strategies. Patient compliance did not change over the next year. The risk of this procedure is the initially increased level of anxiety of the patients.

To what extent does age influence the reporting of life events?

A study of life events reporting was carried out among 64 depressed and 74 non-depressed patients. The results show that age as well as depression can affect the number and type of reported life events. The younger age group (less than 45 years) reported more events than the older group, and the depressed patients reported more events than their non-depressed counterparts. The younger group also listed more desirable events than their elders. There was no evidence of an interaction between diagnosis and age.

Extraction with methyl tert-butyl ether overcomes erratic elution patterns of 6-keto-prostaglandin F1 alpha on high pressure liquid chromatography.

Solvent extraction of 6-keto-PGF1 alpha from aqueous solutions with ethyl acetate was found to result in variable and irreproducible elution patterns, when the extracts were subjected to high pressure liquid chromatography. These problems could not be resolved satisfactorily by using ethyl acetate from different suppliers, nor by changing acids or pH for acidification. After a number of unsuccessful attempts to resolve this problem, we found that variable and irreproducible elution patterns could be avoided by using methyl t-butyl ether as extraction solvent.

Anhedonia, suicide ideation and dexamethasone nonsuppression in depressed patients.

In the search for a valid analysis of a number of operationalised symptoms common to depressive behaviour, a study was performed comprising 46 patients showing depressive symptoms, according to operationalised criteria and as part of which all agreed to undergo the following tests: (a) psychiatric: Present State Examination; (b) psychological: Hamilton Rating Scale, Montgomery-Asberg Rating Scale, State-Trait Anxiety Inventory, Beck Suicide Ideation Scale, Chapman Anhedonia Scale, Mood Scale, Sleep Quality Scale, Activities Scale, Social Support Scale, Questionnaire on Recently Experienced Events and the Paykel Life Events Interview; and (c) biochemical: Dexamethasone Suppression (DEX) Test. After gathering different depressive subgroups, based on operationalised symptoms, a dichotomy was made in the distributions of the (an)hedonia, suicide ideation and DEX-(non) suppression scores. This study may indicate that anhedonia, suicide ideation and DEX-nonsuppression are the opening to the identification of a subgroup of depressed patients. This symptom complex could not definitely be identified on the basis of existing DSM-III diagnostic entities, because of the known fact that this method of classification is not appropriate for our purposes in revealing pathophysiological processes. It is suggested, therefore, that these symptoms might prove to be the anchor-point from which to reach a better insight into the aetiology and pathogenesis (i.e. the final common pathway) of depression.

Changes in plasma levels of PGF2 alpha and PGI2 metabolites at and after delivery at term.

Plasma levels of 6-keto-PGF1 alpha and 13,14-dihydro-15-keto-PGF2 alpha (PGFM) were measured by high pressure liquid chromatography and radioimmunoassay during and up to 48 hours after term labor. PGFM levels increased during labor to reach values which at full dilatation, at delivery of the fetal head and at placental separation were each time higher than levels obtained earlier. In all women (n = 10) PGFM levels reached their maximum and started to decline within 10 min. after placental separation. Levels decreased to prelabor values within 2 to 3 hours after delivery and no temporary increases were observed within the first 2 days. Levels of 6-keto-PGF1 alpha on the other hand, showed no consistent trends throughout labor and the early puerperium. The observed changes are believed to be of relevance for ensuring adequate hemostasis after birth.

Changes in urinary 6-keto-prostaglandin F1 alpha excretion during pregnancy and labor.

Urinary excretion of 6-keto-PGF1 alpha was measured by high pressure liquid chromatography and radioimmunoassay at various stages of pregnancy and labor. In the first trimester of pregnancy, urinary 6-keto-PGF1 alpha concentrations were not different from those measured before pregnancy, but they showed a significant increase in the second trimester of pregnancy (p less than 0.001). The levels rose further in the third trimester, although this increase was not statistically significant when compared to levels obtained in the second trimester. There was no evidence for a significant change in 6-keto-PGF1 alpha excretion with the onset of labor. During well-established, progressive labor mean values of 6-keto-PGF1 alpha excretion were about twice as high as before the onset of labor, but the range of values during labor was so wide that there was no statistical difference with values obtained in the second half of pregnancy. It is concluded that the increase in the urinary excretion of 6-keto-PGF1 alpha occurs later in pregnancy than the increase in TXB2 excretion and that labor at term is not associated with marked changes in 6-keto-PGF1 alpha excretion.