Polymorphic low penetrance genes and breast cancer : The role of genes involved in metabolism of xenobiotics, estrogens and reactive oxygen species

Various endogenous and exogenous factors have been reported to increase the risk of breast cancer. Many of those are related to prolonged lifetime exposure to estrogens. Furthermore, a positive family history of breast cancer and certain benign breast diseases are known to increase the risk of breast cancer. The role of lifestyle factors, such as use of alcohol and smoking has been an area of intensive study. Alcohol has been found to increase the risk of breast cancer, whereas the role of smoking has remained obscure. A multitude of enzymes are involved in the metabolism of estrogens and xenobiotics including the carcinogens found in tobacco smoke. Many of the metabolic enzymes exhibit genetic polymorphisms that can lead to inter-individual differences in their abilities to modify hazardous substrates. Therefore, in presence of a given chemical exposure, one subgroup of women may be more susceptible to breast carcinogenesis, since they carry unfavourable forms of the polymorphic genes involved in the metabolism of the chemical. In this work, polymorphic genes encoding for cytochrome P450 (CYP) 1A1 and 1B1, N-acetyl transferase 2 (NAT2), sulfotransferase 1A1 (SULT1A1), manganese superoxide dismutase (MnSOD) and vitamin D receptor (VDR) were investigated in relation to breast cancer susceptibility in a Finnish population. CYP1A1, CYP1B1 and SULT1A1 are involved in the metabolism of both estrogens and xenobiotics, whereas NAT2 is involved only in the latter. MnSOD is an antioxidant enzyme protecting cells from oxidative damage. VDR, in turn, mediates the effects of the active form of vitamin D (1,25(OH)2D3, calcitriol) on maintenance of calcium homeostasis and it has anti-proliferative effects in many cancer cells. A 1.3-fold (95% CIs 1.01-1.73) increased risk of breast cancer was seen among women who carried the NAT2 slow acetylator genotype and a 1.5-fold (95% CI 1.1-2.0) risk was found in women with a MnSOD variant A allele containing genotypes compared to women with the NAT2 rapid acetylator genotype or to those with the MnSOD VV genotype, respectively. Instead, women with the VDR a allele containing genotypes were found to be at a decreased risk for breast cancer (OR 0.73; 95% CI 0.54-0.98) compared to women with the AA genotype. No significant overall associations were found between SULT1A1 or CYP genotypes and breast cancer risk, whereas a combination of the CYP1B1 432Val allele containing genotypes with the NAT2 slow acetylator genotypes posed a 1.5-fold (95% CI 1.03-2.24) increased risk. Moreover, NAT2 slow acetylator genotype was found to be confined to women with an advanced stage of breast cancer (stages III and IV). Further evidence for the association of xenobiotic metabolising genes with breast cancer risk was found when active smoking was taken into account. Women who smoked less than 10 cigarettes/day and carried at least one CYP1B1 432Val variant allele, were at 3.1-fold (95% CI 1.32-7.12) risk of breast cancer compared to women who smoked the same amount but did not carry the variant allele. Furthermore, the risk was significantly increased with increasing number of the CYP1B1 432Val alleles (p for trend 0.005). In addition, women who smoked less than 5 pack-years and carried the NAT2 slow acetylator genotype were at a 2.6-fold (95% CI 1.01-6.48) increased risk of breast cancer compared to women who smoked the same amount but carried the NAT2 rapid acetylator genotype. Furthermore, the combination of the CYP1B1 432Val allele and the NAT2 slow acetylator genotype increased the risk of breast cancer by 2.5-fold (95% CI 1.11-5.45) among ever smokers. Instead, the MnSOD A allele was found to be a risk factor among postmenopausal long-term smokers (>15 years of smoking) (OR 5.1; 95% CI 1.4-18.4) or among postmenopausal women who had smoked more than 10 cigarettes/day (OR 5.5; 95% CI 1.3-23.4) compared to women who had similar smoking habits but carried the MnSOD V/V genotype. Similarly, within subgroups of postmenopausal women who were using oral contraceptives, hormone replacement therapy or alcohol, women carrying the MnSOD A allele genotypes seemed to be at increased risk of breast cancer compared to women with the MnSOD V/V genotype. A positive family history of breast cancer and high parity were shown to be inversely associated with breast cancer risk among women carrying the VDR ApaI a allele or among premenopausal women carrying the SULT1A1*2 allele, respectively.

Rad51-Rad52 Mediated Maintenance of Centromeric Chromatin in Candida albicans

Specification of the centromere location in most eukaryotes is not solely dependent on the DNA sequence. However, the non-genetic determinants of centromere identity are not clearly defined. While multiple mechanisms, individually or in concert, may specify centromeres epigenetically, most studies in this area are focused on a universal factor, a centromere-specific histone H3 variant CENP-A, often considered as the epigenetic determinant of centromere identity. In spite of variable timing of its loading at centromeres across species, a replication coupled early S phase deposition of CENP-A is found in most yeast centromeres. Centromeres are the earliest replicating chromosomal regions in a pathogenic budding yeast Candida albicans. Using a 2-dimensional agarose gel electrophoresis assay, we identify replication origins (ORI7-LI and ORI7-RI) proximal to an early replicating centromere (CEN7) in C. albicans. We show that the replication forks stall at CEN7 in a kinetochore dependent manner and fork stalling is reduced in the absence of the homologous recombination (HR) proteins Rad51 and Rad52. Deletion of ORI7-RI causes a significant reduction in the stalled fork signal and an increased loss rate of the altered chromosome 7. The HR proteins, Rad51 and Rad52, have been shown to play a role in fork restart. Confocal microscopy shows declustered kinetochores in rad51 and rad52 mutants, which are evidence of kinetochore disintegrity. CENP-A(CaCse4) levels at centromeres, as determined by chromatin immunoprecipitation (ChIP) experiments, are reduced in absence of Rad51/Rad52 resulting in disruption of the kinetochore structure. Moreover, western blot analysis reveals that delocalized CENP-A molecules in HR mutants degrade in a similar fashion as in other kinetochore mutants described before. Finally, co-immunoprecipitation assays indicate that Rad51 and Rad52 physically interact with CENP-A(CaCse4) in vivo. Thus, the HR proteins Rad51 and Rad52 epigenetically maintain centromere functioning by regulating CENP-A(CaCse4) levels at the programmed stall sites of early replicating centromeres.

Epithelial atrophy in oral submucous fibrosis is mediated by copper (II) and arecoline of areca nut

Exposure of oral cavity to areca nut is associated with several pathological conditions including oral submucous fibrosis (OSF). Histopathologically OSF is characterized by epithelial atrophy, chronic inflammation, juxtaepithelial hyalinization, leading to fibrosis of submucosal tissue and affects 0.5% of the population in the Indian subcontinent. As the molecular mechanisms leading to atrophied epithelium and fibrosis are poorly understood, we studied areca nut actions on human keratinocyte and gingival fibroblast cells. Areca nut water extract (ANW) was cytotoxic to epithelial cells and had a pro-proliferative effect on fibroblasts. This opposite effect of ANW on epithelial and fibroblast cells was intriguing but reflects the OSF histopathology such as epithelial atrophy and proliferation of fibroblasts. We demonstrate that the pro-proliferative effects of ANW on fibroblasts are dependent on insulin-like growth factor signalling while the cytotoxic effects on keratinocytes are dependent on the generation of reactive oxygen species. Treatment of keratinocytes with arecoline which is a component of ANW along with copper resulted in enhanced cytotoxicity which becomes comparable to IC50 of ANW. Furthermore, studies using cyclic voltammetry, mass spectrometry and plasmid cleavage assay suggested that the presence of arecoline increases oxidation reduction potential of copper leading to enhanced cleavage of DNA which could generate an apoptotic response. Terminal deoxynucleotidyl transferase dUTP Nick End Labeling assay and Ki-67 index of OSF tissue sections suggested epithelial apoptosis, which could be responsible for the atrophy of OSF epithelium.

A new genus and species of arboreal toad with phytotelmonous larvae, from the Andaman Islands, India (Lissamphibia, Anura, Bufonidae)

A new bufonid amphibian, belonging to a new monotypic genus, is described from the Andaman Islands, in the Bay of Bengal, Republic of India, based on unique external morphological and skeletal characters which are compared with those of known Oriental and other relevant bufonid genera. Blythophryne gen. n. is distinguished from other bufonid genera by its small adult size (mean SVL 24.02 mm), the presence of six presacral vertebrae, an absence of coccygeal expansions, presence of an elongated pair of parotoid glands, expanded discs at digit tips and phytotelmonous tadpoles that lack oral denticles. The taxonomic and phylogenetic position of the new taxon (that we named as Blythophryne beryet gen. et sp. n.) was ascertained by comparing its 12S and 16S partial genes with those of Oriental and other relevant bufonid lineages. Resulting molecular phylogeny supports the erection of a novel monotypic genus for this lineage from the Andaman Islands of India.

Caracterización de aislamientos atípicos de Candida dubliniensis

241 p. : il., gráf

Clinical factors associated with a Candida albicans Germ Tube Antibody positive test in Intensive Care Unit patients

Background: Poor outcomes of invasive candidiasis (IC) are associated with the difficulty in establishing the microbiological diagnosis at an early stage. New scores and laboratory tests have been developed in order to make an early therapeutic intervention in an attempt to reduce the high mortality associated with invasive fungal infections. Candida albicans IFA IgG has been recently commercialized for germ tube antibody detection (CAGTA). This test provides a rapid and simple diagnosis of IC (84.4% sensitivity and 94.7% specificity). The aim of this study is to identify the patients who could be benefited by the use of CAGTA test in critical care setting. Methods: A prospective, cohort, observational multicentre study was carried out in six medical/surgical Intensive care units (ICU) of tertiary-care Spanish hospitals. Candida albicans Germ Tube Antibody test was performed twice a week if predetermined risk factors were present, and serologically demonstrated candidiasis was considered if the testing serum dilution was >= 1: 160 in at least one sample and no other microbiological evidence of invasive candidiasis was found. Results: Fifty-three critically ill non-neutropenic patients (37.7% post surgery) were included. Twenty-two patients (41.5%) had CAGTA-positive results, none of them with positive blood culture for Candida. Neither corrected colonization index nor antifungal treatment had influence on CAGTA results. This finding could corroborate that the CAGTA may be an important biomarker to distinguish between colonization and infection in these patients. The presence of acute renal failure at the beginning of the study was more frequent in CAGTA-negative patients. Previous surgery was statistically more frequent in CAGTA-positive patients. Conclusions: This study identified previous surgery as the principal clinical factor associated with CAGTA-positive results and emphasises the utility of this promising technique, which was not influenced by high Candida colonization or antifungal treatment. Our results suggest that detection of CAGTA may be important for the diagnosis of invasive candidiasis in surgical patients admitted in ICU.

A remarkable new species of Psednos (Teleostei: Liparidae) from the western North Atlantic Ocean

Psednos rossi new species (Teleostei: Liparidae) is described from two specimens collected in the North Atlantic Ocean off Cape Hatteras, North Carolina, at depths of 500–674 m. Psednos rossi belongs to the P. christinae group, which includes six other species and is characterized by 46–47 vertebrae and the absence of a coronal pore. Psednos rossi differs from those six species by having characters unique within the genus: straight spine, body not humpbacked at the occiput, and a very large mouth with a vertical oral cleft. Other distinguishing characters include a notched pectoral fin with 15–16 rays, eye 17–19% SL, and color in life orange-rose. With P. rossi, the genus Psednos as currently known includes 26 described and five undescribed species of small meso- or bathypelagic liparids from the Atlantic, Pacific, and Indian Oceans.

Lymph Node Colonization Dynamics after Oral Salmonella Typhimurium Infection in Mice

An understanding of how pathogens colonize their hosts is crucial for the rational design of vaccines or therapy. While the molecular factors facilitating the invasion and systemic infection by pathogens are a central focus of research in microbiology, the population biological aspects of colonization are still poorly understood. Here, we investigated the early colonization dynamics of Salmonella enterica subspecies 1 serovar Typhimurium (S. Tm) in the streptomycin mouse model for diarrhea. We focused on the first step on the way to systemic infection - the colonization of the cecal lymph node (cLN) from the gut - and studied roles of inflammation, dendritic cells and innate immune effectors in the colonization process. To this end, we inoculated mice with mixtures of seven wild type isogenic tagged strains (WITS) of S. Tm. The experimental data were analyzed with a newly developed mathematical model describing the stochastic immigration, replication and clearance of bacteria in the cLN. We estimated that in the beginning of infection only 300 bacterial cells arrive in the cLN per day. We further found that inflammation decreases the net replication rate in the cLN by 23%. In ccr7-/- mice, in which dendritic cell movement is impaired, the bacterial migration rate was reduced 10-fold. In contrast, cybb-/- mice that cannot generate toxic reactive oxygen species displayed a 4-fold higher migration rate from gut to cLN than wild type mice. Thus, combining infections with mixed inocula of barcoded strains and mathematical analysis represents a powerful method for disentangling immigration into the cLN from replication in this compartment. The estimated parameters provide an important baseline to assess and predict the efficacy of interventions. © 2013 Kaiser et al.

Cytocidal amino acid starvation of Saccharomyces cerevisiae and Candida albicans acetolactate synthase (ilv2{Delta}) mutants is influenced by the carbon source and rapamycin.

The isoleucine and valine biosynthetic enzyme acetolactate synthase (Ilv2p) is an attractive antifungal drug target, since the isoleucine and valine biosynthetic pathway is not present in mammals, Saccharomyces cerevisiae ilv2Delta mutants do not survive in vivo, Cryptococcus neoformans ilv2 mutants are avirulent, and both S. cerevisiae and Cr. neoformans ilv2 mutants die upon isoleucine and valine starvation. To further explore the potential of Ilv2p as an antifungal drug target, we disrupted Candida albicans ILV2, and demonstrated that Ca. albicans ilv2Delta mutants were significantly attenuated in virulence, and were also profoundly starvation-cidal, with a greater than 100-fold reduction in viability after only 4 h of isoleucine and valine starvation. As fungicidal starvation would be advantageous for drug design, we explored the basis of the starvation-cidal phenotype in both S. cerevisiae and Ca. albicans ilv2Delta mutants. Since the mutation of ILV1, required for the first step of isoleucine biosynthesis, did not suppress the ilv2Delta starvation-cidal defects in either species, the cidal phenotype was not due to alpha-ketobutyrate accumulation. We found that starvation for isoleucine alone was more deleterious in Ca. albicans than in S. cerevisiae, and starvation for valine was more deleterious than for isoleucine in both species. Interestingly, while the target of rapamycin (TOR) pathway inhibitor rapamycin further reduced S. cerevisiae ilv2Delta starvation viability, it increased Ca. albicans ilv1Delta and ilv2Delta viability. Furthermore, the recovery from starvation was dependent on the carbon source present during recovery for S. cerevisiae ilv2Delta mutants, reminiscent of isoleucine and valine starvation inducing a viable but non-culturable-like state in this species, while Ca. albicans ilv1Delta and ilv2 Delta viability was influenced by the carbon source present during starvation, supporting a role for glucose wasting in the Ca. albicans cidal phenotype.

PKC signaling regulates drug resistance of the fungal pathogen Candida albicans via circuitry comprised of Mkc1, calcineurin, and Hsp90.

Fungal pathogens exploit diverse mechanisms to survive exposure to antifungal drugs. This poses concern given the limited number of clinically useful antifungals and the growing population of immunocompromised individuals vulnerable to life-threatening fungal infection. To identify molecules that abrogate resistance to the most widely deployed class of antifungals, the azoles, we conducted a screen of 1,280 pharmacologically active compounds. Three out of seven hits that abolished azole resistance of a resistant mutant of the model yeast Saccharomyces cerevisiae and a clinical isolate of the leading human fungal pathogen Candida albicans were inhibitors of protein kinase C (PKC), which regulates cell wall integrity during growth, morphogenesis, and response to cell wall stress. Pharmacological or genetic impairment of Pkc1 conferred hypersensitivity to multiple drugs that target synthesis of the key cell membrane sterol ergosterol, including azoles, allylamines, and morpholines. Pkc1 enabled survival of cell membrane stress at least in part via the mitogen activated protein kinase (MAPK) cascade in both species, though through distinct downstream effectors. Strikingly, inhibition of Pkc1 phenocopied inhibition of the molecular chaperone Hsp90 or its client protein calcineurin. PKC signaling was required for calcineurin activation in response to drug exposure in S. cerevisiae. In contrast, Pkc1 and calcineurin independently regulate drug resistance via a common target in C. albicans. We identified an additional level of regulatory control in the C. albicans circuitry linking PKC signaling, Hsp90, and calcineurin as genetic reduction of Hsp90 led to depletion of the terminal MAPK, Mkc1. Deletion of C. albicans PKC1 rendered fungistatic ergosterol biosynthesis inhibitors fungicidal and attenuated virulence in a murine model of systemic candidiasis. This work establishes a new role for PKC signaling in drug resistance, novel circuitry through which Hsp90 regulates drug resistance, and that targeting stress response signaling provides a promising strategy for treating life-threatening fungal infections.

Use of 16S ribosomal RNA gene analyses to characterize the bacterial signature associated with poor oral health in West Virginia.

BACKGROUND: West Virginia has the worst oral health in the United States, but the reasons for this are unclear. This pilot study explored the etiology of this disparity using culture-independent analyses to identify bacterial species associated with oral disease. METHODS: Bacteria in subgingival plaque samples from twelve participants in two independent West Virginia dental-related studies were characterized using 16S rRNA gene sequencing and Human Oral Microbe Identification Microarray (HOMIM) analysis. Unifrac analysis was used to characterize phylogenetic differences between bacterial communities obtained from plaque of participants with low or high oral disease, which was further evaluated using clustering and Principal Coordinate Analysis. RESULTS: Statistically different bacterial signatures (P<0.001) were identified in subgingival plaque of individuals with low or high oral disease in West Virginia based on 16S rRNA gene sequencing. Low disease contained a high frequency of Veillonella and Streptococcus, with a moderate number of Capnocytophaga. High disease exhibited substantially increased bacterial diversity and included a large proportion of Clostridiales cluster bacteria (Selenomonas, Eubacterium, Dialister). Phylogenetic trees constructed using 16S rRNA gene sequencing revealed that Clostridiales were repeated colonizers in plaque associated with high oral disease, providing evidence that the oral environment is somehow influencing the bacterial signature linked to disease. CONCLUSIONS: Culture-independent analyses identified an atypical bacterial signature associated with high oral disease in West Virginians and provided evidence that the oral environment influenced this signature. Both findings provide insight into the etiology of the oral disparity in West Virginia.

New genus and two new species of the family Ethmolaimidae (Nematoda: Chromadorida), found in two different cold-seep environments

This study describes a new genus Dystomanema gen. nov. with two new species, D. cadizensis sp. nov. and D. brandtae sp. nov. within the family Ethmolaimidae, subfamily Neotonchinae, based on specimens from two low-activity cold-seep environments at distant geographical locations. The new genus was first identified in samples from the Darwin mud volcano (1100 m depth) in the Gulf of Cadiz and later on also found in samples from a low-activity seep in the Larsen B embayment (820m depth) off the eastern Antarctic Peninsula. Until now, the family Ethmolaimidae contained nine genera: Ethmolaimus and Paraethmolaimus in the subfamily Ethmolaiminae, and Comesa, Filitonchoides, Filitonchus, Gomphionchus, Gomphionema, Nannolaimus, and Neothonchus in the subfamily Neotonchinae. The most important family characteristics are: an annulated cuticle bearing transverse rows of dots, cephalic sensilla arrangement of 6+6+4, a spiral amphid, an oesophagus with muscular posterior bulb, paired gonads and males with cup-shaped precloacal supplements. The new genus resembles Comesa and Neotonchus, but is typified by a ventrally displaced oral opening with three very small teeth that are easily overlooked. D. cadizensis gen. nov. sp. nov. is characterized by the 1401-2123 mu m long body; cuticle transversally striated with fine punctation; head conical; low lips; amphid spiralled 3 turns, oral opening ventrally displaced, male with outstretched testes; spicules of equal size; gubernaculum plate-like and ten to twelve conspicuous cup-shaped precloacal supplements with external longitudinal articulated flange. D. brandtae gen. nov. sp. nov. can be distinguished by the 2438-3280 mu m long body; cuticle transversally striated with fine punctuation; head conical; low lips; amphid spiraled 3+ turns; oral opening ventrally displaced; male with anterior testes outstretched and posterior one smaller and reflexed; spicules of equal size; gubernaculum plate-like and twenty conspicuous cup-shaped precloacal supplements with external longitudinal articulated flange. Notes on the ecology and habitat of the new genus are provided in light of its discovery in cold-seep environments.

Conditioning film and environmental effects on the adherence of Candida spp. to silicone and poly(vinylchloride) biomaterials.

The reported incidence of colonization of oropharyngeal medical devices with Candida spp. has increased in recent years, although few studies that have systematically examined the adherence of yeast cells to such biomaterials, the primary step in the process of colonization. This study, therefore, examined the effects of oropharyngeal atmospheric conditions (5% v/v carbon dioxide) and the presence of a salivary conditioning film on both the surface properties and adherence of Candida albicans, Candida krusei and Candida tropicalis to PVC and silicone. Furthermore, the effects of the salivary conditioning film on the surface properties of these biomaterials are reported. Growth of the three Candida spp. in an atmosphere containing 5% v/v CO2 significantly increased their cell surface hydrophobicity and reduced the zeta potential of C. albicans and C. krusei yet increased the zeta potential of C. tropicalis (p < 0.05). Furthermore, growth in 5% v/v CO2 decreased the adherence of C. tropicalis and C. albicans to both PVC and silicone, however, increased adherence of C. krusei (p < 0.05). Pre-treatment of the microorganisms with pooled human saliva significantly decreased their cell surface hydrophobicity and increased their adherence to either biomaterial in comparison to yeast cells that had been pre-treated with PBS (p < 0.05). Saliva treatment of the microorganisms had no consistent effect on microbial zeta potential. Interestingly, adherence of the three, saliva-treated Candida spp. to saliva-treated silicone and PVC was significantly lower than whenever the microorganisms and biomaterials had been treated with PBS (p < 0.05). Treatment of silicone and PVC with saliva significantly altered the surface properties, notably reducing both the advancing and receding contact angles and, additionally, the microrugosity. These effects may contribute to the decreased adherence of saliva-treated microorganisms to these biomaterials. In conclusion, this study has demonstrated the effects of physiological conditions within the oral cavity on the adherence of selected Candida spp. to biomaterials employed as oropharyngeal medical devices. In particular, this study has ominously shown that these materials act as substrates for yeast colonization, highlighting the need for advancements in biomaterial design. Furthermore, it is important that physiological conditions should be employed whenever biocompatibility of oropharyngeal biomaterials is under investigation. © 2001 Kluwer Academic Publishers.

'What say you to [this] book? Is it yours?'::Oral and Collaborative Narrative Trajectories in the Mediated Writings of Anna Trapnel

This article argues that, when a printed page is initially orally generated and then transcribed, either at the time or on a subsequent occasion by a listener or an interlocutor, there are important critical implications for the “I” of the account. It takes as a case study Anna Trapnel's first published works. Appearing within a few weeks of each other in 1654, The Cry of a Stone and Strange and Wonderful News are both mediated texts, large parts of which depend on the agency of a relater. The article begins by examining the textual traces of the relater, arguing for the centrality of his role and other agencies in the shaping of the works which bear Trapnel's name. Situating itself in relation to a current orientation in feminist autobiographical theory that places emphasis on the external requirement to narrate one's life, rather than on the spontaneous production of autobiography by an inner self, the article emphasizes notions of coaxing, witnessing and intersubjectivity to point up an appreciation of women's life writing as a species of cultural production in which various historical actors—male and female—participate. This dialogic process, which persists into the afterlife of transcription, owes part of its genesis to the political vagaries of 1654 and precipitates two contrasting—but equally “authentic”—versions of Trapnel's life and self. Mapping this movement, discussion concentrates on the ways in which a critical confrontation with women's oral narrative is as much an activity of disentangling as it is of reconstructing, an activity which is revealing of the extent to which a spectrum of social and cultural networks participates in and facilitates the female writing act.