Relevance of the deletion polymorphisms of the glutathione S-transferases GSTT1 and GSTM1 in pharmacology and toxicology

Although cytosolic glutathione S-transferase (GST) enzymes occupy a key position in biological detoxification processes, two of the most relevant human isoenzymes, GSTT1-1 and GSTM1-1, are genetically deleted (non-functional alleles GSTT1*0 and GSTM1*0) in a high percentage of the human population, with major ethnic differences. The structures of the GSTT and GSTM gene areas explain the underlying genetic processes. GSTT1-1 is highly conserved during evolution and plays a major role in phase-II biotransformation of a number of drugs and industrial chemicals, e.g. cytostatic drugs, hydrocarbons and halogenated hydrocarbons. GSTM1-1 is particularly relevant in the deactivation of carcinogenic intermediates of polycyclic aromatic hydrocarbons. Several lines of evidence suggest that hGSTT1-1 and/or hGSTM1-1 play a role in the deactivation of reactive oxygen species that are likely to be involved in cellular processes of inflammation, ageing and degenerative diseases. There is cumulating evidence that combinations of the GSTM1*0 state with other genetic traits affecting the metabolism of carcinogens (CYP1A1, GSTP1) may predispose the aero-digestive tract and lung, especially in smokers, to a higher risk of cancer. The GSTM1*0 status appears also associated with a modest increase in the risk of bladder cancer, consistent with a GSTM1 interaction with carcinogenic tobacco smoke constituents. Both human GST deletions, although largely counterbalanced by overlapping substrate affinities within the GST superfamily, have consequences when the organism comes into contact with distinct man-made chemicals. This appears relevant in industrial toxicology and in drug metabolism.

Occupational and non-occupational risk factors in bladder cancer patients in an industrialized area located in former East-Germany [Berufliche und außerberufliche Risikofaktoren für das Harnblasenkarzinom in einem ehemaligen Industriegebiet der neuen Bundesländer]

Purpose: Several occupational carcinogens are metabolized by polymorphic enzymes. The distribution of the polymorphic enzymes N-acetyltransferase 2 (NAT2; substrates: aromatic amines), glutathione S-transferase M1 (GSTM1; substrates: e.g., reactive metabolites of polycyclic aromatic hydrocarbons), and glutathione S-transferase T1 (GSTT1; substrates: small molecules with 1-2 carbon atoms) were investigated. Material and Methods: At the urological department in Lutherstadt Wittenberg, 136 patients with a histologically proven transitional cell cancer of the urinary bladder were investigated for all occupations performed for more than 6 months. Several occupational and non-occupational risk factors were asked. The genotypes of NAT2, GSTM1, and GSTT1 were determined from leucocyte DNA by PCR. Results: Compared to the general population in Middle Europe, the percentage of GSTT1 negative persons (22.1 %) was ordinary; the percentage of slow acetylators (59.6%) was in the upper normal range, while the percentage of GSTM1 negative persons (58.8%) was elevated in the entire group. Shifts in the distribution of the genotypes were observed in subgroups who had been exposed to asbestos (6/6 GSTM1 negative, 5/6 slow acetylators), rubber manufacturing (8/10 GSTM1 negative), and chlorinated solvents (9/15 GSTM1 negative). Conclusions: The overrepresentation of GSTM1 negative bladder cancer patients also in this industrialized area and more pronounced in several occupationally exposed subgroups points to an impact of the GSTM1 negative genotype in bladder carcinogenesis. [Article in German]

Influence of polymorphisms of the human glutathione transferases and cytochrome P450 2E1 enzyme on the metabolism and toxicity of ethylene oxide and acrylonitrile

A cohort of 59 persons with industrial handling of low levels of acrylonitrile is being studied as part of a medical surveillance programme. Previously, an extended haemoglobin adduct monitoring (N-(cyanoethyl)valine and N-(hydroxyethyl)-valine) was performed regarding the glutathione transferases hGSTM1 and hGSTT1 polymorphisms but no influence of hGSTM1 or hGSTT1 polymorphisms on specific adduct levels was found. A compilation of case reports of human accidental poisonings had pointed to significant individual differences in human acrylonitrile metabolism and toxicity. Therefore, a re-evaluation of the industrial cohort included known polymorphisms of the glutathione transferases hGSTM3 and hGSTP1 as well as of the cytochrome P450 CYP2E1. A detailed statistical analysis revealed that exposed carriers of the allelic variants of hGSTP1, hGSTP1*B/hGSTP1*C, characterized by a single nucleotide polymorphism at nucleotide 313 which results in a change from Ile to Val at codon 104, had higher levels of the acrylonitrile-specific haemoglobin adduct N-(cyanoethyl)valine compared to the carriers of the codon 113 alleles hGSTP1*A and hGSTP1*D. The single nucleotide polymorphism at codon 113 of hGSTP1 (hGSTP1*A/hGSTP1*B versus hGSTP1*C/hGSTP1*D) did not show an effect, and also no influence was seen on specific haemoglobin adduct levels of the polymorphisms of hGSTM3 or CYP2E1. The data, therefore, point to a possible influence of a human enzyme polymorphism of the GSTP1 gene at codon 104 on the detoxication of acrylonitrile which calls for experimental toxicological investigation. The study also confirmed the impact of GSTT1 polymorphism on background N-(hydroxyethyl)-valine adduct levels in haemoglobin which are caused by endogenous ethylene oxide.

Markers of genetic susceptibility in human environmental hygiene and toxicology : the role of selected CYP, NAT and GST genes

Inherited genetic traits co-determine the susceptibility of an individual to a toxic chemical. Special emphasis has been put on individual responses to environmental and industrial carcinogens, but other chronic diseases are of increasing interest. Polymorphisms of relevant xenobiotic metabolising enzymes may be used as toxicological susceptibility markers. A growing number of genes encoding enzymes involved in biotransformation of toxicants and in cellular defence against toxicant-induced damage to the cells has been identified and cloned, leading to increased knowledge of allelic variants of genes and genetic defects that may result in a differential susceptibility toward environmental toxicants. "Low penetrating" polymorphisms in metabolism genes tend to be much more common in the population than allelic variants of "high penetrating" cancer genes, and are therefore of considerable importance from a public health point of view. Positive associations between cancer and CYP1A1 alleles, in particular the *2C I462V allele, were found for tissues following the aerodigestive tract. Again, in most cases, the effect of the variant CYP1A1 allele becomes apparent or clearer in connection with the GSTM1 null allele. The CYP1B1 codon 432 polymorphism (CYP1B1*3) has been identified as a susceptibility factor in smoking-related head-and-neck squameous cell cancer. The impact of this polymorphic variant of CYP1B1 on cancer risk was also reflected by an association with the frequency of somatic mutations of the p53 gene. Combined genotype analysis of CYP1B1 and the glutathione transferases GSTM1 or GSTT1 has also pointed to interactive effects. Of particular interest for the industrial and environmental field is the isozyme CYP2E1. Several genotypes of this isozyme have been characterised which seem to be associated with different levels of expression of enzyme activity. The acetylator status for NAT2 can be determined by genotyping or by phenotyping. In the pathogenesis of human bladder cancer due to occupational exposure to "classical" aromatic amines (benzidine, 4-aminodiphenyl, 1-naphthylamine) acetylation by NAT2 is regarded as a detoxication step. Interestingly, the underlying European findings of a higher susceptibility of slow acetylators towards aromatic amines are in contrast to findings in Chinese workers occupationally exposed to aromatic amines which points to different mechanisms of susceptibility between European and Chinese populations. Regarding human bladder cancer, the hypothesis has been put forward that genetic polymorphism of GSTM1 might be linked with the occurrence of this tumour type. This supports the hypothesis that exposure to PAH might causally be involved in urothelial cancers. The human polymorphic GST catalysing conjugation of halomethanes, dihalomethanes, ethylene oxide and a number of other industrial compounds could be characterised as a class theta enzyme (GSTT1) by means of molecular biology. "Conjugator" and "non-conjugator" phenotypes are coincident with the presence and absence of the GSTT1 gene. There are wide variations in the frequencies of GSTT1 deletion (GSTT1 *0/0) among different ethnicities. Human phenotyping is facilitated by the GST activity towards methyl bromide or ethylene oxide in erythrocytes which is representative of the metabolic GSTT1 competence of the entire organism. Inter-individual variations in xenobiotic metabolism capacities may be due to polymorphisms of the genes coding for the enzymes themselves or of the genes coding for the receptors or transcription factors which regulate the expression of the enzymes. Also, polymorphisms in several regions of genes may cause altered ligand affinity, transactivation activity or expression levels of the receptor subsequently influencing the expression of the downstream target genes. Studies of individual susceptibility to toxicants and gene-environment interaction are now emerging as an important component of molecular epidemiology.

Haemoglobin adducts of acrylonitrile and ethylene oxide in acrylonitrile workers, dependent on polymorphisms of the glutathione transferases GSTT1 and GSTM1

Fifty-nine persons with industrial handling of low levels of acrylonitrile (AN) were studied. As part of a medical surveillance programme an extended haemoglobin adduct monitoring [N-(cyanoethyl)valine, CEV; N- (methyl)valine, MV; N-(hydroxyethyl)valine, HEV] was performed. Moreover, the genetic states of the polymorphic glutathione transferases GSTM1 and GSTT1 were assayed by polymerase chain reaction (PCR). Repetitive analyses of CEV and MV in subsequent years resulted in comparable values (means, 59.8 and 70.3 μg CEV/1 blood; 6.7 and 6.7 μg MV/1 blood). Hence, the industrial AN exposures were well below current official standards. Monitoring the haemoglobin adduct CEV appears as a suitable means of biomonitoring and medical surveillance under such exposure conditions. There was also no apparent correlation between the CEV and HEV or CEV and MV adduct levels. The MV and HEV values observed represented background levels, which apparently are not related to any occupational chemical exposure. There was no consistent effect of the genetic GSTM1 or GSTT1 state on CEV adduct levels induced by acrylonitrile exposure. Therefore, neither GSTM1 nor GSTT1 appears as a major AN metabolizing isoenzyme in humans. The low and physiological background levels of MV were also not influenced by the genetic GSTM1 state, but the MV adduct levels tended to be higher in GSTT1- individuals compared to GSTT1 + persons. With respect to the background levels of HEV adducts observed, there was no major influence of the GSTM1 state, but GST- individuals displayed adduct levels that were about 1/3 higher than those of GSTT1+ individuals. The coincidence with known differences in rates of background sister chromatid exchange between GSTT1- and GSTT1 + persons suggests that the lower ethylene oxide (EO) detoxification rate in GSTT1- persons, indicated by elevated blood protein hydroxyethyl adduct levels, leads to an increased genotoxic effect of the physiological EO background.

Differential substrate behaviours of ethylene oxide and propylene oxide towards human glutathione transferase theta hGSTT1-1

The transformation of ethylene oxide (EO), propylene oxide (PO) and 1- butylene oxide (1-BuO) by human glutathione transferase theta (hGSTT1-1) was studied comparatively using 'conjugator' (GSTT1 + individuals) erythrocyte lysates. The relative sequence of velocity of enzymic transformation was PO > EO >> 1-BuO. The faster transformation of PO compared to EO was corroborated in studies with human and rat GSTT1-1 (hGSTT1-1 and rGSTT1-1, respectively) expressed by Salmonella typhimurium TA1535. This sequence of reactivities of homologous epoxides towards GSTT1-1 contrasts to the sequence observed in homologous alkyl halides (methyl bromide, MBr; ethyl bromide, EtBr; n-propyl bromide, PrBr) where the relative sequence MeBr >> EtBr > PrBr is observed. The higher reactivity towards GSTT1-1 of propylene oxide compared to ethylene oxide is consistent with a higher chemical reactivity. This is corroborated by experimental data of acid-catalysed hydrolysis of a number of aliphatic epoxides, including ethylene oxide and propylene oxide and consistent with semi-empirical molecular orbital modelings.

Effects of HNO3 molecular doping in graphene/Si Schottky barrier solar cells

Schottky barrier solar cells based on graphene/n-silicon heterojunction have been fabricated and characterized and the effect of graphene molecular doping by HNO3 on the solar cells performances have been analyzed. Different doping conditions and thermal annealing processes have been tested to asses and optimize the stability of the devices. The PCE of the cells increases after the treatment by HNO3 and reaches 5% in devices treated at 200 °C immediately before the exposition to the oxidant. Up to now our devices retain about 80% of efficiency over a period of two weeks, which represents a good stability result for similar devices.

Keeping our nursing and midwifery workforce : factors that support non-practising clinicians to return to practice

BACKGROUND: Within Australia and internationally (Health Workforce Australia, 2012) an increasing and on-going nursing workforce shortage is documented. Recent international estimates indicate that there will be ongoing and significant gaps in the supply of a nursing workforce; the United Kingdom is predicted to have a reduction of 12.12% nurses over the coming eight years if a current 'steady state' is maintained (Buchan and Seacombe, 2011); Canada is predicted to have a shortage of 60,000 nurses by 2022 (Tomblin et al., 2012) with Australia's anticipated nursing shortage reported as over 90,000 by the year 2025 (Health Workforce Australia, 2012). Queensland Health in response to their tracked emerging nursing and midwifery workforce shortages developed a nursing and midwifery refresher programme to return registered staff back to the workforce. A study was undertaken between 2008 and 2010 to provide an understanding of how non-practising nurses and midwives maybe supported back into the workforce. METHODS: Programme applicants (444) were invited to respond to an on-line survey designed to understand what aspects of the programme supported their learning and ability to return to the workforce. This number represents those who applied but not all completed or commenced the programme. Descriptive statistics (Polit and Beck, 2008) were used to collate quantifiable survey responses and free text and unsolicited responses were themed. RESULTS: The survey received a 35.5% response rate (n=158) with a return of 20% of unsolicited comments in the form of e-mail responses which were included in the themed results. Key themes supporting participants' learning and ability to return to the workforce were: Respondents were 94% female and 6% male, with 37.7% >51 years of age. Child rearing was the foremost reason for female staff relinquishing workforce roles (36.6%). The primary reason for returning to the workforce was maintenance of registration (40.5%). Both theory and clinical placement components were seen by participants as contributing to their confidence to return to the health workforce. CONCLUSION: The Queensland Nursing and Midwifery Refresher Programs provided a structured programme for registered, non-practising nurses and midwives to return to the Queensland Health workforce. Responses indicated that clinical supervision and contract learning should be central to a return to workforce induction programme for registered but non-practising nurses and midwives. The majority of nurses and midwives returning to the workforce were approaching retirement age in 10-15 years.