Carcinogen-specific mutation pattern in the p53 tumour suppressor gene in UV radiation-induced basal cell carcinoma.

OBJECTIVES: Basal cell carcinoma (BCC) is mainly caused by high and long-term UV radiation. UV radiation causes DNA damage in various genes. Mutations of the p53 tumour suppressor gene have been identified in a wide variety of human cancers. The aim of the study was to analyse specific p53 mutations in BCCs in workers exposed to high and long-term UV radiation. METHODS: The mutation pattern of the p53 tumour suppressor gene was analysed in tissue from 12 patients with UV-related BCC. All patients had a suspected occupational disease notified within the period 1995-1999. As a control, 20 BCC skin samples removed from areas definitively unexposed to sunlight were analysed. The specific mutations were determined by direct sequencing of codon 4 to 9 of the p53 gene in carcinomatous and adjacent non-neoplastic tissue after microdissection. Immunohistochemical analysis was performed to detect p53 protein. RESULTS: p53 mutations were detected in 7/12 cases (58%). Point mutations were found in six cases (50%). In one case a deletion of 24 base pairs was observed. The most frequent mutations we found were CC-->TT base-pair changes in four and C-->T mutations in two cases. Within the control group specific p53 mutations were found in 11 cases (55%) without any C-->T predominance. No case showed CC-->TT mutations. CONCLUSIONS: Mutations of the p53 tumour suppressor gene in UV-associated BCC are frequent events. A predominance of C-->T mutations and tandem CC-->TT base-pair changes were observed in the sunlight-exposed cases only supporting the idea of site-directed mutagenesis by UV radiation in human BCC.

p16INK4A-alterations in primary angiosarcoma of the liver.

BACKGROUND/AIMS: Alterations in the p16 (CDKN2/MTS-1/INK4A) gene have been implicated in the tumorigenesis of different human cancers. Recent evidence shows that transcriptional silencing as a consequence of hypermethylation of CpG islands is the predominant mechanism of p16INK4a gene inactivation in malignant epithelial tumors. This study was performed to determine whether alterations of p16 are involved in the development of angiosarcoma of the liver. METHODS: The status of p16 was evaluated in 17 angiosarcomas of the liver by methylation-specific PCR (MSP), microsatellite analysis, DNA sequencing and immunohistochemical staining. The results obtained were correlated with histopathological variables and with patient survival. RESULTS: Hypermethylation of the 5' CpG island of the p16 gene was found in 12 out of 17 (71%) angiosarcomas examined. Homozygous deletion at the p16 region was present in one case (6%), and loss of heterozygosity was present in two cases (12%). We failed to detect p16 gene missense mutations. The status of p16 correlated with neither histopathological factors nor with the prognosis of the patients with angiosarcomas. CONCLUSIONS: These data suggest that inactivation of the p16 gene is a frequent event in angiosarcomas of the liver. The most common somatic alteration is promotor methylation of the p16 gene. We failed to establish p16 as independent prognostic factors in these tumors.

Frequent k- ras -2 mutations and p16(INK4A)methylation in hepatocellular carcinomas in workers exposed to vinyl chloride.

Vinyl chloride (VC) is a know animal and human carcinogen associated with liver angiosarcomas (LAS) and hepatocellular carcinomas (HCC). In VC-associated LAS mutations of the K- ras -2 gene have been reported; however, no data about the prevalence of such mutations in VC associated HCCs are available. Recent data indicate K- ras -2 mutations induce P16 methylation accompanied by inactivation of the p16 gene. The presence of K- ras -2 mutations was analysed in tissue from 18 patients with VC associated HCCs. As a control group, 20 patients with hepatocellular carcinoma due to hepatitis B (n = 7), hepatitis C (n = 5) and alcoholic liver cirrhosis (n = 8) was used. The specific mutations were determined by direct sequencing of codon 12 and 13 of the K- ras -2 gene in carcinomatous and adjacent non-neoplastic liver tissue after microdissection. The status of p16 was evaluated by methylation-specific PCR (MSP), microsatellite analysis, DNA sequencing and immunohistochemical staining. All patients had a documented chronic quantitated exposure to VC (average 8883 ppmy, average duration: 245 months). K- ras -2 mutations were found in 6 of 18 (33%) examined VC-associated HCCs and in 3 cases of adjacent non-neoplastic liver tissue. There were 3 G --> A point mutations in the tumour tissue. All 3 mutations found in non-neoplastic liver from VC-exposed patients were also G --> A point mutations (codon 12- and codon 13-aspartate mutations). Hypermethylation of the 5' CpG island of the p16 gene was found in 13 of 18 examined carcinomas (72%). Of 6 cancers with K- ras -2 mutations, 5 specimens also showed methylated p16. Within the control group, K- ras -2 mutation were found in 3 of 20 (15%) examined HCC. p16 methylation occurred in 11 out of 20 (55%) patients. K- ras -2 mutations and p16 methylation are frequent events in VC associated HCCs. We observed a K- ras -2 mutation pattern characteristic of chloroethylene oxide, a carcinogenic metabolite of VC. Our results strongly suggest that K- ras -2 mutations play an important role in the pathogenesis of VC-associated HCC.

Internal exposure to polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDDs/PCDFs) of Bavarian chimney sweeps.

This study was carried out to evaluate the internal exposure to polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDDs/PCDFs) of chimney sweeps in Bavaria compared to a control group without occupational exposure. The PCDD/PCDF concentrations in the blood fat of 227 chimney sweeps were compared with the concentrations in samples from 60 controls. Using an internal standard containing 17 (13)C(12)-labeled PCDD/F congeners, the samples were cleaned up after fat elution using standard methods. The statistical analysis was adjusted to account for demographic differences, dietary habits, smoking status, and both occupational and nonoccupational contact with chlorinated hydrocarbons. Detailed information on the type of heating in the households swept, the length of time the chimney sweeps had carried out the profession (min 34, med 195, max 466 months) and the protective measures employed, were used to examine the influence of the working conditions specific to chimney sweeps on the internal PCDD/PCDF exposure. The correlation between blood-fat PCB concentrations as well as urinary chlorophenol concentrations and the exposure to PCDDs/PCDFs was evaluated. The sum of PCDD/PCDF components in chimney sweeps, expressed by International Toxic Equivalents (I-TEQ), was significantly increased compared to the control group (median: 26.36 versus 20.75 pg I-TEQ/g blood fat). For 37 chimney sweeps (16.3%) the sum of PCDDs/PCDFs exceeded the 95th percentile of the control group, i.e., 38.23 pg I-TEQ/g blood fat. Multiple regression analysis revealed that in addition to occupation, the variables age, district, and proximity to a waste incineration plant seem to have an effect on the internal PCDD/PCDF exposure. An additional influence on the internal exposure could not be determined for any of the special aspects of the work. We identified no high correlations between the concentrations of PCBs and chlorophenols and PCDDs/PCDFs. This study revealed significantly higher internal exposure to PCDDs/PCDFs in chimney sweeps than in the control group. The differences are small and within the range of the internal exposure to PCDDs/PCDFs in blood found in the general population in Germany since 1989. Further investigations in to PCDD/PCDF-related diseases in these study groups were not carried out.

Biological monitoring of workers exposed to N,N-dimethylformamide in the synthetic fibre industry.

OBJECTIVES: Monitoring of workplace air and biological monitoring of 23 workers exposed to N,N-dimethylformamide (DMF) in the polyacrylic fibre industry was carried out on 4 consecutive days. The main focus of the investigation was to study the relationship between external and internal exposure, the suitability of the metabolites of DMF for biological monitoring and their toxicokinetic behaviour in humans. METHODS: Air samples were collected using personal air samplers. The limit of detection (LOD) for DMF using an analytical method recommended by the Deutsche Forschungsgemeinschaft (DFG) was 0.1 ppm. The urinary metabolites, N-hydroxymethyl-N-methylformamide (HMMF), N-methylformamide (NMF), and N-acetyl-S-(N-methylcarbamoyl)-cysteine (AMCC), were determined in one analytical run by gas chromatography with thermionic sensitive detection (GC/TSD). The total sum of HMMF and NMF was determined in the form of NMF. The LOD was 1.0 mg/l for NMF and 0.5 mg/l for AMCC. RESULTS AND CONCLUSIONS: The external exposure to DMF vapour varied greatly depending on the workplace (median 1.74 ppm, range < 0.1-159.77 ppm). Urinary NMF concentrations were highest in post-shift samples. They also covered a wide range (< 1.0-108.7 mg/l). This variation was probably the result of different concentrations of DMF in the air at different workplaces, dermal absorption and differences in the protective measures implemented by each individual (gloves, gas masks etc.). The urinary NMF concentrations had decreased almost to zero by the beginning of the next shift. The median half-time for NMF was determined to be 5.1 h. The concentrations of AMCC in urine were determined to be in the range from < 0.5 to 204.9 mg/l. Unlike the concentrations of NMF, the AMCC concentrations did not decrease during the intervals between the shifts. For the exposure situation investigated in our study, a steady state was found between the external exposure to DMF and the levels of AMCC excreted in urine about 2 days after the beginning of exposure. AMCC is therefore excreted more slowly than NMF. The half-time for AMCC is more than 16 h. Linear regression analysis for external exposure and urinary excretion of metabolites was carried out for a sub-group of 12 workers. External exposure to 10 ppm DMF in air (the current German MAK value) corresponds to an average NMF concentration of about 27.9 mg/l in post-shift urine from the same day and an average AMCC concentration of 69.2 mg/l in pre-shift urine from the following day. NMF in urine samples therefore represents an index of daily exposure to DMF, while AMCC represents an index of the average exposure over the preceding working days. AMCC is considered to be better suited for biomonitoring purposes because (1) it has a longer half-time than NMF and (2) its formation in humans is more closely related to DMF toxicity.

Liver function in workers exposed to N,N-dimethylformamide during the production of synthetic textiles.

OBJECTIVE: In a factory producing synthetic fibers the hepatotoxic effects of the solvent N,N-dimethylformamide (DMF) were investigated in 126 male employees, especially with regard to the combination effects of DMF exposure and ethyl alcohol consumption. A collective of similar structure from the same factory served as a control collective. METHODS: Reference is made to the results of air measurements and biological monitoring presented in a previous publication. The DMF concentrations in the air ranged from < 0.1 (detection limit) to 37.9 ppm (median 1.2 ppm). Concentrations of the DMF metabolite N-methylformamide (NMF) in urine were 0.05-22.0 mg/l (preshift) and 0.9-100.0 mg/l (postshift), corresponding to 0.02-44.6 mg/g creatinine (preshift) and 0.4-62.3 mg/g creatinine (postshift). A standardized anamnesis was drawn up for relevant previous illnesses and other factors influencing liver function. The laboratory tests included parameters especially relevant to the liver (e.g., AST, ALT, gamma-GT, hepatitis B and C antibodies, and carbohydrate-deficient transferrin). RESULTS: The results indicate a statistically significant toxic influence of DMF on liver function. Alcohol has a synergistic effect. The effects of DMF and those of alcohol are dose-dependent. Under the existing workplace conditions the hepatotoxic effects of alcohol are more severe than those of DMF. In the exposed group there was a statistically significantly greater number of persons who stated that they had drunk less since the beginning of exposure (13% versus 0). This corresponded with the data on symptoms occurring after alcohol consumption (71% versus 4%). In the work areas with lower-level exposure to DMF there was greater alcohol consumption. It corresponded to that of the control collective not exposed to DMF. CONCLUSION: In this study we tried to differentiate and quantify the interaction between DMF exposure and alcohol consumption and the influence of both substances on liver function. The experience gained from former occupational health surveillance in DMF-exposed persons and from the present study show that there are individual differences in tolerance of interactions between DMF and ethyl alcohol. Further studies are necessary for the evaluation of these individual degrees of susceptibilitiy.

Occupational chronic exposure to organic solvents XVII. Ambient and biological monitoring of workers exposed to xylenes.

OBJECTIVE: Ambient-air and biological monitoring of occupational xylene exposure were carried out on 2 groups of workers (13 and 10 men, respectively) exposed to a mixture of xylenes during the production of paints or during spraying. METHODS: Personal ambient-air monitoring was performed for one complete work shift. Blood and urine samples were collected directly at the end of the shift. Biological monitoring was based on the determination of the concentration of xylenes in blood and on the quantification of the sum of the three methylhippuric acids in urine. RESULTS: Average xylene ambient-air concentrations were 29 ppm (production) and 8 ppm (spraying), ranging from 5 to 58 ppm and from 3 to 21 ppm, respectively. The concentrations of xylenes in blood ranged from 63 to 715 microg/l and from 49 to 308 microg/l, with average values being 380 and 130 microg/l, respectively. Accordingly, the workers engaged in paint production also excreted more methylhippuric acids in their urine (average 1221 mg/l, range 194 2333 mg/l) than did the sprayers (average 485 mg/l, range 65-1633 mg/l). DISCUSSION: Our results as well as a literature review indicate that occupational xylene exposure on average barely exceeds the threshold limit value of 100 ppm as proposed by both American and German institutions. Biological monitoring based on the determination of xylenes in blood and of methylhippuric acids in urine provides sufficient sensitivity and specificity for occupational health surveillance. The results also confirm the current limit values (BAT values) proposed by the Deutsche Forschungsgemeinschaft for xylenes in blood (1500 microg/l) and methylhippuric acids in urine (2000 mg/l).

N,N-dimethylformamide--influence of working conditions and skin penetration on the internal exposure of workers in synthetic textile production.

OBJECTIVES: This study examined the external and internal exposure to the solvent N,N-dimethylformamide (DMF) of 126 workers from a factory producing synthetic fibers. METHODS: Air measurements were carried out using personal air samplers with diffusion tubes (Drager, ORSA 5). For the purpose of biological monitoring the levels of N-methylformamide (NMF) in urine were measured in preshift and postshift samples. Determinations were carried out using gas chromatography. Anamnestic data were collected with standardized questionnaires, including personal data, working history and current working conditions, and former and current illness with regard to the effects of DMF. Skin diseases were documented by a dermatologist. RESULTS: DMF concentrations measured in the air ranged between <0.1 and 37.9 ppm (median 1.2 ppm). Concentrations of NMF varied from 0.05 to 22.0 mg/l (preshift values) and from 0.9 to 100.0 mg/l (postshift values). The creatinine-related values (0.02-44.6 mg/g preshift; 0.4-62.3 postshift) were subject to less variation and therefore represented the level of exposure better than the values related to volume. Additional investigation of a subcollective (n=31) over a period of 4 days showed that NMF did not accumulate in the organism. The positive but relatively weak association observed between the DMF concentrations measured in the workplace air and the values recorded for internal exposure in this study can be explained by influencing factors such as dermal absorption or protective clothing. Interindividual differences in internal exposure were found for the specific work areas. The German BAT value (15 mg NMF/l urine) was exceeded in 36 persons (29%) despite the use of breathing protection and protective gloves, without increased values being measured in the air. Increased absorption without higher-level exposure could particularly also be observed in employees with eczema. CONCLUSIONS: From the point of view of the prevention of disease, biological monitoring is the best instrument for exposure assessment of workers exposed to DMF.

Occupational health in Germany and other countries of the European Union.

Occupational medicine is affected to a much greater extent by national legal and social conditions than by clinical issues. The different preconditions specific to each country serve to restrict the scientific dialogue on issues of occupational medicine. Therefore, in this paper are described the organisation and the under- and postgraduate education of occupational medicine in Germany and other European countries (Austria, Belgium, Denmark, Italy, United Kingdom). In summary one can state that in many member states of the EU there is a lack of undergraduate training in occupational medicine for students and there are distinct systems for the postgraduate training and assessment of occupational physicians. The practice of occupational medicine in the EU countries probably has many similarities. The responsibilities of occupational physicians are rather comparable, though in some countries the approaches of occupational medical prevention and the interest in evaluating the structure, process and outcome of many of the activities of occupational medicine seem to recede something into the background.

Internal exposure of waste incineration workers to organic and inorganic substances.

One hundred and twenty-two persons employed in an industrial waste incineration plant were examined with respect to organic and inorganic substances which may be produced during the combustion of different waste. The employees were divided into three groups: persons with contact with the incinerator (WI workers, n = 45), periphery workers (n = 54) and management (n = 23). For the evaluation of internal exposure, the levels of lead, cadmium, mercury, benzene, toluene, ethylbenzene and m-xylene in blood, chromium in the erythrocytes, polychlorinated biphenyls, hexachlorobenzene and pentachlorophenol in plasma, and arsenic, chromium, nickel, vanadium, chlorophenols and hydroxypyrene in urine were determined. The internal exposures of the three groups were tested against each other and were compared with the reference values of the general population. Differences between the groups investigated were tested using the U test according to Wilcoxon, Mann and Whitney (P < 0.05). The biological exposure limits valid in Germany (BAT values) were not exceeded in any cases. Compared with the background levels of the German population, certain parameters were exceeded in several employees. Significantly higher levels of the WI workers in comparison to both periphery workers and management were found for toluene in blood (median: 1.1 vs 0.9 vs 0.6 microgram/l). For the lead and cadmium levels in blood and for the urinary excretion of arsenic, 2,4-dichlorophenol and tetrachlorophenols, statistical differences were found only between WI workers and one of the other groups. However, in all cases the elevations were very small and of interest more from the environmental than from the occupational point of view. It must be stressed that this waste incineration plant is very modern in terms of worker health and safety. At older plants without corresponding health and safety measures, higher internal exposure of the employees to hazardous substances may exist.

Early reaction type allergies and diseases of the respiratory passages in employees from persulphate production.

The prevalence of positive skin-prick test reactions ammonium persulphate and potassium persulphate (1% and 5% solutions) was tested in a cross-sectional study on 52 employees of a company producing persulphates after a case of "persulphate asthma" was observed. A random test of 13 persons without occupational exposure to persulphates served as controls; among them all the skin-prick test reactions were negative. Eight company employees showed a positive skin-prick test reaction to at least one of the persulphate solutions tested. Employees showed lower lung function results with a positive prick test reaction than did employees with a negative result. The positive skin-prick reactions correspond well to the anamnestic data and indicate a possible relationship to obstructive ventilation disorders. The results therefore suggest an IgE-induced, allergic pathomechanism of "persulphate asthma" triggered by persulphates.

[Chlorophenols in urine as an environmental medicine monitoring parameter]. / Chlorphenole im Harn als umweltmedizinische Untersuchungsparameter.

Chlorophenols occur ubiquitously in the environment. They are taken up as such in man or are formed in intermediary metabolism e.g. from chlorobenzenes. In particular pentachlorophenol (PCP) is one of those chemical substances used up to the early 70's as a component of wood preservatives also indoors; for many years it has been at the centre of discussion about the environment. Mono, di, tri and tetrachlorophenols as well as pentachlorophenol occur in the urine in the general population often in surprisingly high concentrations. An increased chlorophenol excretion under certain circumstances also indicates an increased dioxin exposure (pre-dioxins). Possible sources of emission and routes of absorption for corresponding organochlorine compounds can be found in industry, agriculture and also private households. Environmental analysis in the air, in earth or dust do not allow any evaluation of health risks. Only biological monitoring with qualitative and quantitative determination of the actual concentration of the substance taken up by the organism allows a reliable estimation of the individual health risk. The background exposure of the general population not occupationally exposed to organochlorine compounds can be used for the determination of so-called norm values. For the determination of the chlorophenol spectrum 50 ml urine are necessary. The following values can be given as reference values for the most important chlorophenols: 4-monochlorophenol: 7.5 micrograms/l, 2.4-dichlorophenol and 2.5-dichlorophenol: 33.6 micrograms/l, 2.4.6-trichlorophenol: 4.7 micrograms/l, 2.4.5-trichlorophenol: 4.5 micrograms/l, 2.3.4.6-tetrachlorophenol and 2.3.5.6-tetrachlorophenol: 22.0 micrograms/l, pentachlorophenol 9.0 micrograms/l urine.