5-Aminolevulinate dehydratase porphyria: Update on hepatic 5-aminolevulinic acid synthase induction and long-term response to hemin.

BACKGROUND: 5-Aminolevulinic acid dehydratase (ALAD) porphyria (ADP) is an ultrarare autosomal recessive disease, with only eight documented cases, all of whom were males. Although classified as an acute hepatic porphyria (AHP), induction of the rate limiting hepatic enzyme 5-aminolevulinic acid synthase-1 (ALAS1) has not been demonstrated, and the marrow may also contribute excess 5-aminolevulinic acid (ALA). Two patients have died and reported follow up for the others is limited, so the natural history of this disease is poorly understood and treatment experience limited. METHODS: We report new molecular findings and update the clinical course and treatment of the sixth reported ADP patient, now 31 years old and the only known case in the Americas, and review published data regarding genotype-phenotype correlation and treatment. RESULTS: Circulating hepatic 5-aminolevulinic acid synthase-1 (ALAS1) mRNA was elevated in this case, as in other AHPs. Gain of function mutation of erythroid specific ALAS2 - an X-linked modifying gene in some other porphyrias - was not found. Seven reported ADP cases had compound heterozygous ALAD mutations resulting in very low residual ALAD activity and symptoms early in life or adolescence. One adult with a germline ALAD mutant allele developed ADP in association with a clonal myeloproliferative disorder, polycythemia vera. CONCLUSIONS: Elevation in circulating hepatic ALAS1 and response to treatment with hemin indicate that the liver is an important source of excess ALA in ADP, although the marrow may also contribute. Intravenous hemin was effective in most reported cases for treatment and prevention of acute attacks of neurological symptoms.

Riboflavin as an independent and accurate biomarker for adherence in a randomized double-blind and placebo-controlled clinical trial.

BACKGROUND: Medication adherence is critical for success of clinical trials. OBJECTIVE: To assess oral riboflavin is an adherence marker. METHODS: Riboflavin was incorporated into active treatment and placebo pills for a clinical trial lasting for 2 years. RESULTS: The accuracy (area under the receiver operating curve) of urinary riboflavin was 0.91 as a binary classifier of adherence, and was similar or better than for two active study ingredients daidzein (0.92) and genistein (0.87) (all p < 0.0001). Decreased adherence over time was similar in the two study groups. CONCLUSION: Riboflavin is an accurate and useful biomarker for study pill ingestion.

Porphyrin Accumulation and Biliary Lithiasis Causing Diffusely Black Livers in Broiler Chickens.

Two 7-wk-old broiler chickens presented with uniformly black livers upon postslaughter examination, while all other organs as well as their carcasses were grossly normal. No clinical signs were reported by the field veterinarian prior to slaughter. Other broiler chickens within the same flock were unaffected. Microscopically, the liver exhibited variably sized, globoid concrements that were dark brown to green-brown and birefringent under polarized light. Ultrastructurally, concrements consisted of radially arranged electron-dense crystal spicules. Concrements were located in hepatocytes, within ecstatic bile canaliculi, or surrounded by small clusters of macrophages. Liquid chromatography assay determined the presence of protoporphyrin IX in the affected liver.Two 7-wk-old broiler chickens presented with uniformly black livers upon postslaughter examination, while all other organs as well as their carcasses were grossly normal. No clinical signs were reported by the field veterinarian prior to slaughter. Other broiler chickens within the same flock were unaffected. Microscopically, the liver exhibited variably sized, globoid concrements that were dark brown to green-brown and birefringent under polarized light. Ultrastructurally, concrements consisted of radially arranged electron-dense crystal spicules. Concrements were located in hepatocytes, within ecstatic bile canaliculi, or surrounded by small clusters of macrophages. Liquid chromatography assay determined the presence of protoporphyrin IX in the affected liver.

Vivid biofluorescence discovered in the nocturnal Springhare (Pedetidae).

Biofluorescence has been detected in several nocturnal-crepuscular organisms from invertebrates to birds and mammals. Biofluorescence in mammals has been detected across the phylogeny, including the monotreme duck-billed platypus (Ornithorhyncus anatinus), marsupial opossums (Didelphidae), and New World placental flying squirrels (Gluacomys spp.). Here, we document vivid biofluorescence of springhare (Pedetidae) in both museum specimens and captive individuals-the first documented biofluorescence of an Old World placental mammal. We explore the variation in biofluorescence across our sample and characterize its physical and chemical properties. The striking visual patterning and intensity of color shift was unique relative to biofluorescence found in other mammals. We establish that biofluorescence in springhare likely originates within the cuticle of the hair fiber and emanates, at least partially, from several fluorescent porphyrins and potentially one unassigned molecule absent from our standard porphyrin mixture. This discovery further supports the hypothesis that biofluorescence may be ecologically important for nocturnal-crepuscular mammals and suggests that it may be more broadly distributed throughout Mammalia than previously thought.

Bioaccumulation and toxicity of gold nanoparticles after repeated administration in mice.

Gold nanoparticles (GNPs) offer a great promise in biomedicine. Currently, there is no data available regarding the accumulation of nanoparticles in vivo after repeated administration. The purpose of the present study was to evaluate the bioaccumulation and toxic effects of different doses (40, 200, and 400 microg/kg/day) of 12.5 nm GNPs upon intraperitoneal administration in mice every day for 8 days. The gold levels in blood did not increase with the dose administered, whereas in all the organs examined there was a proportional increase on gold, indicating efficient tissue uptake. Although brain was the organ containing the lowest quantity of injected GNPs, our data suggest that GNPs are able to cross the blood-brain barrier and accumulate in the neural tissue. Importantly, no evidence of toxicity was observed in any of the diverse studies performed, including survival, behavior, animal weight, organ morphology, blood biochemistry and tissue histology. The results indicate that tissue accumulation pattern of GNPs depend on the doses administered and the accumulation of the particles does not produce sub-acute physiological damage.

Human retinal cadmium accumulation as a factor in the etiology of age-related macular degeneration.

Cadmium is a naturally occurring, highly toxic, metallic element. It pollutes the environment as a result of industrial activity and accumulates in human tissues with a long biological half-life. Cadmium content has been demonstrated to increase in human retinal tissues as a function of age and tobacco smokers have approximately twice as much cadmium in retinal tissues than non-smokers. Smoking is also a key environmental risk factor for the retinal disease age-related macular degeneration (AMD). Recent studies have shown that urinary cadmium levels (a measure of Cd body burden) are higher in smokers who have AMD. We now report the Cd measurements in human retinal tissues from eyes afflicted with AMD compared to non-diseased eyes (controls) from age-matched donors. Human donor eyes frozen under argon gas were assessed for AMD severity using color stereoscopic fundus photographs and the Minnesota Grading System. Cadmium, zinc and, copper levels were measured in retinal tissues (neural retina, retinal pigment epithelium and choroid) using inductively coupled plasma mass spectrometry and graphite furnace spectrophotometry and values were normalized to tissue protein levels. Higher Cd levels were found in the neural retina and RPE for eyes afflicted with AMD compared to controls in males, differences were not statistically significant in females. The results indicate that higher retinal cadmium burdens are associated with the presence of AMD at least in males and suggest possible gender differences in the metabolism of metals in the human retina.

Harderoporphyria: Case of lifelong photosensitivity associated with compound heterozygous coproporphyrinogen oxidase (CPOX) mutations.

A 78-year-old man with a history of neonatal anemia and jaundice and life-long photosensitivity was found to have harderoporphyria, as evidenced by increased porphyrins in urine, plasma, erythrocytes and feces including large amounts of harderoporphyrin in feces and erythrocytes. Two previously undescribed coproporphyrinogen oxidase (CPOX) mutations were identified, including a deletion of four amino acids in a region of the enzyme mutated in 7 of the 8 previously reported cases. This case increases the molecular heterogeneity of this rare porphyria, and illustrates that it should be considered as a cause of chronic photosensitivity and porphyrin elevation at any age.

Cystathionine-γ-lyase (CSE) deficiency increases erythropoiesis and promotes mitochondrial electron transport via the upregulation of coproporphyrinogen III oxidase and consequent stimulation of heme biosynthesis.

BACKGROUND: Hydrogen sulfide (H2S) is an endogenous gasotransmitter produced by mammalian cells. The current study investigated the potential role of H2S in the regulation of heme biosynthesis using mice deficient in cystathionine gamma-lyase (CSE), one of the three major mammalian H2S-producing enzymes. METHODS: Wild-type and global CSE-/- mice, as well as mitochondria prepared from their liver were used. In vivo, arterial and venous blood gases were measured, and survival of the mice to severe global hypoxia was monitored. Ex vivo, expression of various heme biosynthetic enzymes including coproporphyrinogen oxidase (CPOX) was measured, and mitochondrial function was evaluated using Extracellular Flux Analysis. Urine samples were collected to measure the oxidized porphyrinogen intermediates. The in vivo/ex vivo studies were complemented with mitochondrial bioenergetic studies in hepatocytes in vitro. Moreover, the potential effect of H2S on the CPOX promoter was studied in cells expressing a CPOX promoter construct system. RESULTS: The main findings are as follows: (1) CSE-/- mice exhibit elevated red blood cell counts and red blood cell mean corpuscular volumes compared to wild-type mice; (2) these changes are associated with elevated plasma and liver heme levels and (3) these alterations are likely due to an induction of CPOX (the sixth enzyme involved in heme biosynthesis) in CSE-/- mice. (4) Based on in vitro promoter data the promoter activation of CPOX is directly influenced by H2S, the product of CSE. With respect to the potential functional relevance of these findings, (5) the increased circulating red blood cell numbers do not correspond to any detectable alterations in blood gas parameters under resting conditions, (6) nor do they affect the hypoxia tolerance of the animals in an acute severe hypoxia model. However, there may be a functional interaction between the CSE system and the CPOX system in terms of mitochondrial bioenergetics: (7) CSE-/- hepatocytes and mitochondria isolated from them exhibit increased oxidative phosphorylation parameters, and (8) this increase is partially blunted after CPOX silencing. Although heme is essential for the biosynthesis of mitochondrial electron chain complexes, and CPOX is required for heme biosynthesis, (9) the observed functional mitochondrial alterations are not associated with detectable changes in mitochondrial electron transport chain protein expression. CONCLUSIONS: The CSE system regulates the expression of CPOX and consequent heme synthesis. These effects in turn, do not influence global oxygen transport parameters, but may regulate mitochondrial electron transport.

Copper and zinc distribution in the human retina: relationship to cadmium accumulation, age, and gender.

The essential metals copper and zinc play vital roles in retinal cell survival and are crucial for the normal functioning of antioxidant enzymes. Retinal zinc deficiencies and decreased cellular antioxidative capacity have been linked to human retinal diseases including age-related macular degeneration (AMD). We recently reported that cadmium (a toxic metal with no known physiological function that interferes with copper and zinc metabolism) accumulates in human retinal tissues during aging. Moreover, cadmium content was higher in specific retinal tissues of aged women compared to men. Since cadmium, zinc and copper bind to similar proteins, we hypothesized that Cu and Zn content of human retinal tissues change as functions of cadmium accumulation during aging. Thus, we assessed the distribution of zinc and copper in the neural retina, retinal pigment epithelium (RPE) and choroid (Bruch's membrane-choroid; BMC) in male and female donors aged 1.5-87 years. Two independent methods, graphite furnace atomic absorption spectrometry and inductively-coupled plasma mass spectrometry, were used to measure Cd, Zn, and Cu in retinal tissues in human eyes from donors aged 1.5 to 87 years and the resulting values were normalized to protein concentration. Zn levels were approximately 5 times higher than Cu levels in the same tissues. The relative tissue distributions of these metals were: BMC>RPE>neural retina (Zn) and BMC>RPE=neural retina (Cu). In the choroid, mean Cu and Zn levels were higher in aged donors (>or=55 years old) than young donors (<55 years) and levels of these metals were strongly correlated with each other (r=0.90). In the neural retina, Cu and Zn both significantly decreased as a function of age. Several sex-related differences were found in the RPE. Specifically, copper levels were significantly higher in males than in females. In addition, both Zn and Cu levels in males were positively correlated with cadmium content, whereas this association did not occur in females. The results are consistent with co-regulation of zinc and copper stores in retinal tissues and suggest that the balance of these metals is associated with cadmium accumulation and gender. Thus, the roles of cadmium and gender differences in retinal metal balance warrant further investigation as factors in age-related retinal disease.

Novel effects of phytoestrogenic soy isoflavones on serum calcium and chloride in premenopausal women: A 2-year double-blind, randomized, placebo-controlled study.

BACKGROUND: Soy phytoestrogens are potential alternatives to postmenopausal hormone replacement therapy (HRT). Adverse effects of HRT such as myocardial infarction, stroke, and pulmonary embolism are mediated by calcium-induced signaling. OBJECTIVE: To determine whether soy isoflavones affect serum calcium in healthy female subjects. DESIGN: In a double-blind trial, 197 premenopausal women were randomly assigned to either isoflavone (N = 99) or placebo pills (N = 98) 5 days per week for up to 2 years, plus prenatal vitamins. Isoflavone pills contained 60 mg genistein, 60 mg daidzein and 16.6 mg glycitein (expressed as aglycone equivalents). All pills contained 15 mg riboflavin as an adherence marker. Blood chemistries and urinary daidzein, genistein and riboflavin were measured multiple times during the luteal phase before and during treatment. RESULTS: Analysis of the adherent population (N = 83 per group), revealed significantly strong associations between urinary levels of isoflavones and serum concentrations of calcium (regression coefficients 0.082 for daidzein and 0.229 for genistein, all P < 0.01) and chloride (regression coefficient, -1.537 for genistein, P < 0.0001), mediated in part by albumin. The effects amounted to mean changes of +0.24 mg/dL for calcium and -1.45 mEq/L for chloride, with each visit for subjects excreting the most vs. the least amounts of isoflavones. These associations were not evident in the intention-to-treat analysis (N = 197) that did not assess expected variations in isoflavone levels within and between subjects from metabolism and adherence. CONCLUSIONS: These novel and strong effects of soy isoflavones on calcium homeostasis have important implications for long term effects of these natural substances on cardiovascular diseases.

Mercury, lead, and zinc in baby teeth of children with autism versus controls.

This study determined the level of mercury, lead, and zinc in baby teeth of children with autism spectrum disorder (n = 15, age 6.1 +/- 2.2 yr) and typically developing children (n = 11, age = 7 +/- 1.7 yr). Children with autism had significantly (2.1-fold) higher levels of mercury but similar levels of lead and similar levels of zinc. Children with autism also had significantly higher usage of oral antibiotics during their first 12 mo of life, and possibly higher usage of oral antibiotics during their first 36 mo of life. Baby teeth are a good measure of cumulative exposure to toxic metals during fetal development and early infancy, so this study suggests that children with autism had a higher body burden of mercury during fetal/infant development. Antibiotic use is known to almost completely inhibit excretion of mercury in rats due to alteration of gut flora. Thus, higher use of oral antibiotics in the children with autism may have reduced their ability to excrete mercury, and hence may partially explain the higher level in baby teeth. Higher usage of oral antibiotics in infancy may also partially explain the high incidence of chronic gastrointestinal problems in individuals with autism.

A 27-amino-acid synthetic peptide corresponding to the NH2-terminal zinc-binding domain of endostatin is responsible for its antitumor activity.

The first recombinant endostatin that elicited strong antitumor activity was expressed in Escherichia coli and administered as a suspension. Under these conditions, the protein retained its full antiangiogenic activity. Lack of requirement for a folded structure prompted us to investigate antitumor properties of synthetic peptides corresponding to different regions of endostatin. Here, we show that the entire antitumor, antimigration, and antipermeability activities of endostatin are mimicked by a 27-amino-acid peptide corresponding to the NH2-terminal domain of endostatin. This peptide contains three histidines that are responsible for zinc binding. Mutations of the zinc-binding histidines abolished its antitumor and antimigration activities, but not antipermeability properties.

Iron release and oxidative DNA damage in splenic toxicity of aniline.

Mechanisms by which aniline produces selective toxicity to the spleen are not well understood. Previously, studies showed that aniline exposure induces lipid peroxidation and protein oxidation in the spleen. The present study was aimed to determine the release of free iron and oxidative DNA damage in the spleen following aniline exposure. To achieve this, male SD rats were orally administered 1 mmol/kg/d aniline for 7 d, while controls received the vehicle only. Total splenic iron content showed a significant increase of 200% in the aniline-treated rats, whereas free iron (low-molecular-weight chelatable iron) showed a marked increase of 375% in comparison to controls. Oxidative DNA damage, measured in terms of 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels, showed a remarkable increase of 83% in the aniline-treated rats. These results suggest an association between release of free iron and oxidative DNA damage, which could lead to mutagenic and/or carcinogenic responses in the spleen.

A simplified method to quantify isoflavones in commercial soybean diets and human urine after legume consumption.

Reliable and economical quantification of micronutrients in diets and human is a critical component of successful epidemiological studies to establish relationships between dietary constituents and chronic disease. Legumes are one of the major dietary components consumed by populations worldwide. Consumption of legumes is thought to play a major role in lowering breast and prostate cancer risk. In this study, a simplified method that uses solid-phase extraction and gas chromatography was developed to measure isoflavones at levels down to 10 micrograms/5ml. With the use of this method, 12.5 g miso (a soybean paste), 12 ounces Isomil, and 12 ounces soymilk had daidzin/daidzein levels of 2, 5, and 12.4 mg, respectively, and genistin/genistein levels of 3, 6.5, and 13.7 mg, respectively. In these products, most of the isoflavones were present as glucosides. With the same method, urinary levels of isoflavones in six 15-17-year-old subjects were determined after soymilk ingestion. Each subject was placed on unrestricted nonsoya diets, and three 12-ounce portions of soymilk were given at 12-h intervals. Males excreted 15.02 +/- 2.74 (SD) mg of daidzein glucuronides/sulfates [mean recovery, 40.4 +/- 7.4% (SD)] by 24 h after the third soymilk ingestion, whereas females excreted 25.56 +/- 5.10 mg (68.7 +/- 13.7%) of daidzein conjugates, which was more than males (P = 0.02). Males and females excreted 7.73 +/- 1.95 mg and 9.11 +/- 0.84 mg of genistein glucuronides/sulfates (20% recovery of genistin intake), respectively, in the urine. Most of the isoflavones were excreted within 24 h after ingestion.(ABSTRACT TRUNCATED AT 250 WORDS)

Genetic effects of benzene and radiation in ICR and X/Gf mice.

X/Gf mice (a tumor-resistant strain) were compared with ICR mice (moderately tumor-sensitive) for their sensitivity to chromosomal damage caused by benzene, cyclophosphamide (CP), benzo(a)pyrene (BP) and radiation. There was no difference between strains in the level of micronucleus formation caused by BP, CP or radiation. Although X/Gf mice metabolized somewhat less of the dose of benzene per weight than ICR mice, and had somewhat higher levels of genetic damage, it is not known whether X/Gf mice would be measurably more resistant to benzene carcinogenicity. Short-term genotoxicity tests are used as indicators of initiation, therefore, equal sensitivity to a set of standard clastogens suggests that tumor resistance in X/Gf mice is a function of later stages of carcinogenesis.

Persistence of micronuclei in peripheral blood normochromatic erythrocytes of subchronically benzene-treated male mice.

The kinetics of micronucleus (MN) induction and decline in blood normochromatic erythrocytes (NCE) of mice subchronically exposed to benzene was investigated during and after exposure. Swiss (ICR) male mice (10/group) were given 0.0, 36.6, 73.2, and 146.4 mg/kg body weight benzene by gavage daily for 14 days, except for days 5 and 10. The frequency of MN increased significantly (P less than .001) during benzene treatment as a function of both concentration and time. Eleven days after exposure the levels of MN were higher than those observed at the end of exposure. After an initial rapid decline in the frequency of MN from 11 to 18 days postexposure, the decline became linear with time through 60 days postexposure. Using linear regression analysis, the MN level in each treatment group was predicted to reach control levels by approximately 85 days post-treatment. Dose-dependent suppression and recovery of erythropoiesis, estimated by polychromatic erythrocyte frequency, were observed in the 1st and 2nd weeks of exposure, respectively. Red blood cell (RBC) production was markedly increased in the first 3 weeks after benzene treatment. At later times the rate of production of the RBC returned to normal and may account for the linear decline observed in MN frequency. This research indicates that the frequency of MN is dose and duration dependent, while the decline in MN frequency after the end of benzene exposure can be related to changes in the kinetics of erythropoiesis.

Effect of acute acrylonitrile exposure on metrazol induced seizures in the rat.

The effects of acute exposure to acrylonitrile (ACN), 10, 20, or 40 mg/kg by gavage, on the ability of metrazol (MTZ) to induce seizures was studied in adult, male Sprague-Dawley rats. The frequency of seizure occurrence and the frequency of a lethal seizure was greater when the high ACN dosage was given in combination with metrazol. This dosage of ACN was not lethal when given alone. Examination of brain tissue in these animals revealed no difference in cyanide levels when MTZ was combined with ACN. However, brain cytochrome c was significantly lower in animals given ACN+MTZ and brain cholinesterase was significantly higher. These results suggest that the enhanced lethality occurring in animals exposed to the combination of ACN+MTZ is not due to cyanide, a metabolic product of ACN, but rather to a potentiation of other effects of ACN perhaps involving cholinergic neurotransmission.

Induction of chromosome aberrations in lymphocytes of mice after subchronic exposure to benzene.

The induction of chromosome aberrations in lymphocytes of mice after subchronic exposure to benzene was investigated. 4 groups of 5 Swiss (ICR) male mice were given orally a solution of benzene every day for 14 days except days 5 and 10. The daily doses were 0, 36.6, 73.2 and 146.4 mg/kg. Mice were sacrificed on day 15, lymphocytes were obtained by perfusion of the spleen and the cells were cultured in RPMI 1640 medium. After 48 h of culture, cells were harvested for cytogenetic analysis. A significant dose-dependent increase in the frequency of cells with chromatid aberrations were found (p less than 0.001). A significant increase in polyploid cells were also observed (p less than or equal to 0.05). This study represents the first report on the induction of chromosome aberrations and polyploid cells in lymphocytes of mice after subchronic exposure to benzene. Such dual activity of benzene suggests that benzene may be responsible for more human health problems than currently estimated.

The influence of simple aromatics on benzene clastogenicity.

The micronucleus test was performed in male ICR Swiss mice following modification of benzene metabolism by co-administration of aniline, pyridine or naphthalene, or by prior injection of alpha-naphthoflavone. HPLC profiles of urinary metabolites were compared to the effects of these compounds on clastogenicity. Pyridine inhibited both benzene clastogenicity and its metabolism. Aniline and naphthalene increased the clastogenicity and slightly altered the metabolism of benzene. alpha-Naphthoflavone inhibited benzene clastogenicity and metabolism only at high doses. Since 3-methylcholanthrene and phenobarbital both increase the metabolism of benzene but only 3-methylcholanthrene increases benzene clastogenicity, specific P450 isozymes may be responsible for different biological effects of benzene, and alterations in these effects might be caused by a shift from one isozyme to another.

Population monitoring: experience with residents exposed to uranium mining/milling waste.

More emphasis should be placed upon using biomarkers to address potential health risk among populations exposed to high concentrations of environmental toxicants. Among these studies, those which integrate exposure measurements with analyses of validated biomarkers may provide more reliable information for risk assessment and disease prevention. We have used a multidisciplinary approach to elucidate potential health hazards in a population living around uranium mining/milling facilities. The study included 24 target and 24 control residents who were matched for age and gender and selected based on time of residence in the study areas and proximity to mining/milling sites. Environmental samples were analyzed for uranium-238 (238U) concentrations and lead isotope ratios using inductively coupled plasma-mass spectrometry (ICP-MS) procedures, and blood samples were collected for cytogenetic analysis. We found that the 238U concentrations in soil samples were significantly higher than those in the control areas. In addition, the concentrations in the surface soil were significantly higher than in the subsurface soil (p<0.05) from target areas indicating environmental contamination by the mining/milling activities. Lead isotope data from soil samples taken near a railroad transfer location was significantly different from those of other sites, indicating contamination by non-native ore transported from sources outside of the region to local milling facilities for processing. Therefore, local residents have been exposed to low levels of radioactive contamination from the mining/milling activities on a daily basis for many years. From our cytogenetic analysis, the target population had more chromosome aberrations than the controls, although the differences were not significant (p<0.05). However, using our challenge assay, cells from the target population had a significantly abnormal DNA repair response, compared to cells from the same control population. In conclusion, the observed environmental contamination by uranium is consistent with the observed genotoxic effects in the target residents. Therefore, the residents have increased health risk and some of the health problems will most likely be related to exposure to the radioactive contaminants. Since the chromosome aberration frequency revealed increased, but not significant differences between the exposed and the control populations, we conclude that the health risk among the exposed residents is similar to those among nuclear workers.