The Effect of Microstructural Changes on Mechanical and Electrochemical Corrosion Properties of Duplex Stainless Steel Aged for Short Periods.

This work reports the effects of microstructural changes due to the secondary phases, in particular sigma (σ), on the mechanical properties and electrochemical behavior of thermally aged duplex stainless steel (DSS). Structural, morphological, mechanical, and electrochemical characterizations were performed. Sigma phase content increased with increasing aging treatment time. It had a net-like shape, as observed by electron backscatter diffractometry (EBSD). Its presence directly damaged mechanical properties. The corrosion assessment included electrochemical impedance spectroscopy (EIS) in 1 M NaCl solution at temperatures of 25, 40, and 65 °C. EIS results demonstrate that an increase in the σ phase content decreased the corrosion resistance (21.1-0.8, 3.5-0.3, and 3.1-0.2 kΩ cm2 at 25, 40, and 60 °C, respectively).

A genome-wide association study identifies risk loci for childhood acute lymphoblastic leukemia at 10q26.13 and 12q23.1.

Genome-wide association studies (GWASs) have shown that common genetic variation contributes to the heritable risk of childhood acute lymphoblastic leukemia (ALL). To identify new susceptibility loci for the largest subtype of ALL, B-cell precursor ALL (BCP-ALL), we conducted a meta-analysis of two GWASs with imputation using 1000 Genomes and UK10K Project data as reference (totaling 1658 cases and 7224 controls). After genotyping an additional 2525 cases and 3575 controls, we identify new susceptibility loci for BCP-ALL mapping to 10q26.13 (rs35837782, LHPP, P=1.38 × 10-11) and 12q23.1 (rs4762284, ELK3, P=8.41 × 10-9). We also provide confirmatory evidence for the existence of independent risk loci at 9p21.3, but show that the association marked by rs77728904 can be accounted for by linkage disequilibrium with the rare high-impact CDKN2A p.Ala148Thr variant rs3731249. Our data provide further insights into genetic susceptibility to ALL and its biology.

c-MYC is a radiosensitive locus in human breast cells.

Ionising radiation is a potent human carcinogen. Epidemiological studies have shown that adolescent and young women are at increased risk of developing breast cancer following exposure to ionising radiation compared with older women, and that risk is dose-dependent. Although it is well understood which individuals are at risk of radiation-induced breast carcinogenesis, the molecular genetic mechanisms that underlie cell transformation are less clear. To identify genetic alterations potentially responsible for driving radiogenic breast transformation, we exposed the human breast epithelial cell line MCF-10A to fractionated doses of X-rays and examined the copy number and cytogenetic alterations. We identified numerous alterations of c-MYC that included high-level focal amplification associated with increased protein expression. c-MYC amplification was also observed in primary human mammary epithelial cells following exposure to radiation. We also demonstrate that the frequency and magnitude of c-MYC amplification and c-MYC protein expression is significantly higher in breast cancer with antecedent radiation exposure compared with breast cancer without a radiation aetiology. Our data also demonstrate extensive intratumor heterogeneity with respect to c-MYC copy number in radiogenic breast cancer, suggesting continuous evolution at this locus during disease development and progression. Taken together, these data identify c-MYC as a radiosensitive locus, implicating this oncogenic transcription factor in the aetiology of radiogenic breast cancer.

Common genetic variation contributes significantly to the risk of childhood B-cell precursor acute lymphoblastic leukemia.

Recent genome-wide association studies (GWAS) have provided the first unambiguous evidence that common genetic variation influences the risk of childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL), identifying risk single-nucleotide polymorphisms (SNPs) localizing to 7p12.2, 9p21.3, 10q21.2 and 14q11.2. The testing of SNPs individually for an association in GWA studies necessitates the imposition of a very stringent P-value to address the issue of multiple testing. While this reduces false positives, real associations may be missed and therefore any estimate of the total heritability will be negatively biased. Using GWAS data on 823 BCP-ALL cases by considering all typed SNPs simultaneously, we have calculated that 24% of the total variation in BCP-ALL risk is accounted for common genetic variation (95% confidence interval 6-42%). Our findings provide support for a polygenic basis for susceptibility to BCP-ALL and have wider implications for future searches for novel disease-causing risk variants.

Polymorphic MLH1 and risk of cancer after methylating chemotherapy for Hodgkin lymphoma.

BACKGROUND AND OBJECTIVE: Methylating agents are effective chemotherapy agents for Hodgkin lymphoma, but are associated with the development of second primary cancers. Cytotoxicity of methylating agents is mediated primarily by the DNA mismatch repair (MMR) system. Loss of MLH1, a major component of DNA MMR, results in tolerance to the cytotoxic effects of methylating agents and persistence of mutagenised cells at high risk of malignant transformation. We hypothesised that a common substitution in the basal promoter of MLH1 (position -93, rs1800734) modifies the risk of cancer after methylating chemotherapy. METHODS: 133 patients who developed cancer following chemotherapy and/or radiotherapy (n = 133), 420 patients diagnosed with de novo myeloid leukaemia, 242 patients diagnosed with primary Hodgkin lymphoma, and 1177 healthy controls were genotyped for the MLH1 -93 polymorphism by allelic discrimination polymerase chain reaction (PCR) and restriction fragment length polymorphism assay. Odds ratios and 95% confidence intervals for cancer risk by MLH1 -93 polymorphism status, and stratified by previous exposure to methylating chemotherapy, were calculated using unconditional logistic regression. RESULTS: Carrier frequency of the MLH1 -93 variant was higher in patients who developed therapy related acute myeloid leukaemia (t-AML) (75.0%, n = 12) or breast cancer (53.3%. n = 15) after methylating chemotherapy for Hodgkin lymphoma compared to patients without previous methylating exposure (t-AML, 30.4%, n = 69; breast cancer patients, 27.2%, n = 22). The MLH1 -93 variant allele was also over-represented in t-AML cases when compared to de novo AML cases (36.9%, n = 420) and healthy controls (36.3%, n = 952), and was associated with a significantly increased risk of developing t-AML (odds ratio 5.31, 95% confidence interval 1.40 to 20.15), but only in patients previously treated with a methylating agent. CONCLUSIONS: These data support the hypothesis that the common polymorphism at position -93 in the core promoter of MLH1 defines a risk allele for the development of cancer after methylating chemotherapy for Hodgkin lymphoma. However, replication of this finding in larger studies is suggested.

Deregulation of homologous recombination DNA repair in alkylating agent-treated stem cell clones: a possible role in the aetiology of chemotherapy-induced leukaemia.

Chemotherapeutic regimes involving alkylating agents, such as methylators and crosslinking nitrogen mustards, represent a major risk factor for acute myeloid leukaemia. A high frequency of microsatellite instability and evidence of MSH2 loss in alkylating chemotherapy-related acute myeloid leukaemia (t-AML) suggests that DNA mismatch repair (MMR) dysfunction may be an initiating event in disease evolution. Subsequent accumulation of secondary genetic changes as a result of DNA MMR loss may ultimately lead to the gross chromosomal abnormalities seen in t-AML. Homologous recombination repair (HRR) maintains chromosomal stability by the repair of DNA double-strand breaks, and is therefore a possible target for deregulation in MMR dysfunctional t-AML. In order to test this hypothesis Msh2- proficient and -deficient murine embryonic stem (ES) cells were used to examine the effects of MMR status and methylating agent treatment on cellular expression of DNA double-strand break repair genes. HRR gene expression was significantly deregulated in Msh2 null ES cell clones compared to wild-type clones. Furthermore, some Msh2 null clones expressed high levels of Rad51 specifically, a critical component of HRR. Such Rad51 superexpressing clones were also observed when expression was determined in monocytic myeloid cells differentiated from ES cells. A deregulated HRR phenotype could be partially recapitulated in MMR-competent wild-type cells by treatment with the methylating agent, N-methyl-N-nitrosourea. Furthermore, treatment with melphalan, a leukaemogenic DNA crosslinking chemotherapy nitrogen mustard predicted to elicit HRR, selected against cells with deregulated HRR. These data suggest a t-AML mechanism whereby DNA MMR loss promotes the emergence of HRR gene superexpressing clones, with concomitant chromosomal instability. However, melphalan selection against clones with deregulated HRR suggests that persistence and expansion of unstable clones may require additional genetic alterations that promote cell survival.

Polymorphism in glutathione S-transferase P1 is associated with susceptibility to chemotherapy-induced leukemia.

Glutathione S-transferases (GSTs) detoxify potentially mutagenic and toxic DNA-reactive electrophiles, including metabolites of several chemotherapeutic agents, some of which are suspected human carcinogens. Functional polymorphisms exist in at least three genes that encode GSTs, including GSTM1, GSTT1, and GSTP1. We hypothesize, therefore, that polymorphisms in genes that encode GSTs alter susceptibility to chemotherapy-induced carcinogenesis, specifically to therapy-related acute myeloid leukemia (t-AML), a devastating complication of long-term cancer survival. Elucidation of genetic determinants may help to identify individuals at increased risk of developing t-AML. To this end, we have examined 89 cases of t-AML, 420 cases of de novo AML, and 1,022 controls for polymorphisms in GSTM1, GSTT1, and GSTP1. Gene deletion of GSTM1 or GSTT1 was not specifically associated with susceptibility to t-AML. Individuals with at least one GSTP1 codon 105 Val allele were significantly over-represented in t-AML cases compared with de novo AML cases [odds ratio (OR), 1.81; 95% confidence interval (CI), 1.11-2.94]. Moreover, relative to de novo AML, the GSTP1 codon 105 Val allele occurred more often among t-AML patients with prior exposure to chemotherapy (OR, 2.66; 95% CI, 1.39-5.09), particularly among those with prior exposure to known GSTP1 substrates (OR, 4.34; 95% CI, 1.43-13.20), and not among those t-AML patients with prior exposure to radiotherapy alone (OR,1.01; 95% CI, 0.50-2.07). These data suggest that inheritance of at least one Val allele at GSTP1 codon 105 confers a significantly increased risk of developing t-AML after cytotoxic chemotherapy, but not after radiotherapy.

Genetic alterations in bronchial mucosa and plasma DNA from individuals at high risk of lung cancer.

Evidence suggests that the majority of lung cancer patients have tumour-derived genetic alterations in circulating plasma DNA, and that this may be developed as a diagnostic tool. To this end, we have studied 60 individuals attending bronchoscopy clinic, with symptoms suspicious of lung cancer, for genetic alterations in bronchial mucosa biopsy (n = 47) and plasma (n = 40) DNA. Thirteen of 47 individuals from whom biopsies were taken displayed allelic loss of heterozygosity (LOH) in biopsy DNA for at least 1 of 4 markers. All 13 of these individuals had neoplastic tumour cells in their biopsies and were subsequently diagnosed with cancer. Thirteen of 40 individuals from whom plasma was taken displayed a plasma DNA LOH, and 12 of these 13 individuals were subsequently diagnosed with cancer. LOH in plasma was generally representative of LOH in the corresponding biopsy. In terms of sensitivity, using just 4 markers, biopsy LOH and plasma LOH were found in 13 of 44 (30%) and 12 of 29 (41%), respectively, of those patients subsequently diagnosed with cancer. Two patients were positive for LOH in plasma samples that pre-dated a diagnosis of cancer by several months. These data suggest that assay of genetic alterations in circulating plasma DNA may be developed as a useful addition to conventional techniques for the diagnosis of lung cancer.

Bone formation into surface phosphonylated polymeric implants.

Through the use of two animal models, the present study demonstrates the ability of phosphonylated surfaces to bind bone. In one model, surface-treated polypropylene (PP) and polyethylene (PE) were implanted in the medial cortex of the goat tibia. In the second model, surface-treated poly(ether-ether ketone) (PEEK) and carbon fiber-reinforced PEEK (CFR-PEEK) were implanted through both cortices of the goat mandible. Selected rods of all material types were microtextured using crystallization induced microphase separation, a method for the formation of continuous, open-cell microporous surfaces in thermoplastic polymers. Microtextured and smooth rods were phosphonylated, and calcium was subsequently introduced to the phosphonylated surface by incubating the samples in a saturated solution of calcium oxide. For all substrate materials tested, phosphonylation and calcium posttreatment resulted in an increased propensity for bone binding and apposition, as measured by push out test. Microtextured PP, PE, and CFR-PEEK surfaces that were further phosphonylated and calcium treated resulted in test samples with an increased interfacial strength.

The effect of hOGG1 and glutathione peroxidase I genotypes and 3p chromosomal loss on 8-hydroxydeoxyguanosine levels in lung cancer.

Polymorphic genes for the peroxide scavenger glutathione peroxidase I (GPX1) and 8-hydroxydeoxyguanosine (8-OHdG) DNA glycosylase/apurinic (AP) lyase (hOGG1) map to loci on chromosome 3p which are subject to frequent loss of heterozygosity (LOH) in lung tumours. Levels of the pro-mutagenic, oxidative DNA lesion 8-OHdG, were measured in 37 paired normal and tumorous lung specimens using HPLC with electrochemical detection. Lung tumours were also analysed for 3p LOH by fluorescent PCR with Genescan analysis. No significant difference was observed between 8-OHdG levels in tumour [7.7 +/- 6.7 (mean +/- SE) 8-OHdG/10(6) 2'-deoxyguanosine (dG)] and normal (8.1 +/- 8.8 8-OHdG/10(6) dG) lung tissue. Adduct levels in normal lung tissue DNA were not associated with constitutive hOGG1 genotype although there was a trend towards lower 8-OHdG levels in individuals possessing the ALA6 GPX1 polymorphism. Lung tumours exhibiting 3p LOH (40%) contained higher levels of 8-OHdG adducts (10.9 +/- 2.6 8-OHdG/10(6) dG) (P = 0.05) and lower GPX1 enzyme activity [45.5 nmol glutathione (GSH)/min/mg] (P = 0.09) when compared with tumours without LOH at these sites (5.55 +/- 0.87 8-OHdG/10(6) dG and 63.6 nmol GSH/min/mg, respectively). In conclusion, tumours with 3p LOH at loci associated with hOGG1 and GPX1 appear to have compromised oxidative defence mechanisms as measured by reduced GPX1 enzyme activity and elevated 8-OHdG levels and this may affect the prognosis of lung cancer patients.

3-methyladenine DNA glycosylases: structure, function, and biological importance.

The genome continuously suffers damage due to its reactivity with chemical and physical agents. Finding such damage in genomes (that can be several million to several billion nucleotide base pairs in size) is a seemingly daunting task. 3-Methyladenine DNA glycosylases can initiate the base excision repair (BER) of an extraordinarily wide range of substrate bases. The advantage of such broad substrate recognition is that these enzymes provide resistance to a wide variety of DNA damaging agents; however, under certain circumstances, the eclectic nature of these enzymes can confer some biological disadvantages. Solving the X-ray crystal structures of two 3-methyladenine DNA glycosylases, and creating cells and animals altered for this activity, contributes to our understanding of their enzyme mechanism and how such enzymes influence the biological response of organisms to several different types of DNA damage.

DNA repair methyltransferase (Mgmt) knockout mice are sensitive to the lethal effects of chemotherapeutic alkylating agents.

We have generated mice deficient in O6-methylguanine DNA methyltransferase activity encoded by the murine Mgmt gene using homologous recombination to delete the region encoding the Mgmt active site cysteine. Tissues from Mgmt null mice displayed very low O6-methylguanine DNA methyltransferase activity, suggesting that Mgmt constitutes the major, if not the only, O6-methylguanine DNA methyltransferase. Primary mouse embryo fibroblasts and bone marrow cells from Mgmt -/- mice were significantly more sensitive to the toxic effects of the chemotherapeutic alkylating agents 1,3-bis(2-chloroethyl)-1-nitrosourea, streptozotocin and temozolomide than those from Mgmt wild-type mice. As expected, Mgmt-deficient fibroblasts and bone marrow cells were not sensitive to UV light or to the crosslinking agent mitomycin C. In addition, the 50% lethal doses for Mgmt -/- mice were 2- to 10-fold lower than those for Mgmt +/+ mice for 1,3-bis(2chloroethyl)-1-nitrosourea, N-methyl-N-nitrosourea and streptozotocin; similar 50% lethal doses were observed for mitomycin C. Necropsies of both wild-type and Mgmt -/mice following drug treatment revealed histological evidence of significant ablation of hematopoietic tissues, but such ablation occurred at much lower doses for the Mgmt -/- mice. These results demonstrate the critical importance of O6-methylguanine DNA methyltransferase in protecting cells and animals against the toxic effects of alkylating agents used for cancer chemotherapy.

Mammalian 3-methyladenine DNA glycosylase protects against the toxicity and clastogenicity of certain chemotherapeutic DNA cross-linking agents.

DNA repair status is recognized as an important determinant of the clinical efficacy of cancer chemotherapy. To assess the role that a mammalian DNA glycosylase plays in modulating the toxicity and clastogenicity of the chemotherapeutic DNA cross-linking alkylating agents, we compared the sensitivity of wild-type murine cells to that of isogenic cells bearing homozygous null mutations in the 3-methyladenine DNA glycosylase gene (Aag). We show that Aag protects against the toxic and clastogenic effects of 1,3-bis(2-chloroethyl)-1-nitrosourea and mitomycin C (MMC), as measured by cell killing, sister chromatid exchange, and chromosome aberrations. This protection is accompanied by suppression of apoptosis and a slightly reduced p53 response. Our results identify 3-methyladenine DNA glycosylase-initiated base excision repair as a potentially important determinant of the clinical efficacy and, possibly, the carcinogenicity of these widely used chemotherapeutic agents. However, Aag does not contribute significantly to protection against the toxic and clastogenic effects of several chemotherapeutic nitrogen mustards (namely, mechlorethamine, melphalan, and chlorambucil), at least in the mouse embryonic stem cells used here. We also compare the Aag null phenotype with the Fanconi anemia phenotype, a human disorder characterized by cellular hypersensitivity to DNA cross-linking agents, including MMC. Although Aag null cells are sensitive to MMC-induced growth delay and cell cycle arrest, their sensitivity is modest compared to that of Fanconi anemia cells.

A chemical and genetic approach together define the biological consequences of 3-methyladenine lesions in the mammalian genome.

DNA-damaging agents produce a plethora of cellular responses that include p53 induction, cell cycle arrest, and apoptosis. It is generally assumed that it is the DNA damage produced by these agents that triggers such responses, but there is limited direct evidence to support this assumption. Here, we used DNA alkylation repair proficient and deficient isogenic mouse cell lines to demonstrate that the signal to trigger p53 induction, cell cycle arrest, and apoptosis in response to alkylating agents does emanate from DNA damage. Moreover, we established that 3-methyladenine, a relatively minor DNA lesion produced by most methylating agents (which form mainly 7-methylguanine), can specifically induce sister chromatid exchange, chromatid and chromosome gaps and breaks, S phase arrest, the accumulation of p53, and apoptosis. This study was made possible by the generation of 3-methyladenine DNA glycosylase null mutant cells by targeted homologous recombination and by the chemical synthesis of a methylating agent that almost exclusively produces 3-methyladenine DNA lesions. The combined use of these two experimental tools has defined the biological consequences of 3-methyladenine, a DNA lesion produced by endogenous cellular metabolites, environmental carcinogens, and chemotherapeutic alkylating agents.

Infusing a teacher preparation program in learning disabilities with assistive technology.

A recent trend in the fields of special education, rehabilitation, and technology is the development and implementation of assistive technology (AT) devices and services to assist individuals in compensating for disabilities and/or utilizing functional capabilities to meet environmental demands. AT devices and services have major implications for individuals with learning disabilities (LD) regarding life span issues, environmental and curricular accessibility, and compensatory strategies. Faculty members in higher education who are responsible for designing teacher preparation programs in LD must explore ways to structure curricula, methodologies, and practica to better prepare teachers to work with students who use AT devices to compensate for their specific learning disabilities. The purpose of this article is to describe curriculum design steps and barriers to and solutions for infusing LD teacher preparation programs with assistive technology.