Cr Isotopes and the Engineered Attenuation of Cr(VI)-Rich Runoff.

The leaching of lateritic soils can result in drainage waters with high concentrations of Cr(VI). Such Cr(VI)-rich waters have developed in streams that drain lateritic soils in Central Sulawesi Island, Indonesia. Chromium in this lateritic drainage system is removed by reduction of Cr(VI) to Cr(III) through two faucets delivering an FeSO4 solution to the drainage waters. Cr stable isotope compositions from both water and sediment samples along the drainage path were used to evaluate the efficacy of this remediation strategy. Overall, dissolved [Cr(VI)] decreased moving downstream, but there was an increase in [Cr(VI)] after the first faucet that was effectively removed at the second faucet. This intermittent increase in [Cr(VI)] was the likely result of oxidative remobilization of sediment Cr(III) through reaction with Mn oxides. Cr isotope distributions reflect near quantitative reduction associated with the FeSO4 faucets but also reveal that Cr isotope fractionation is imparted due to Cr redox cycling, downstream. During this redox cycling, fractionation appeared to accompany oxidation, with the product Cr(VI) becoming enriched in 53Cr relative to the reactant Cr(III) with an apparent fractionation factor of 0.7 ± 0.3‰. This study suggests that while FeSO4 effectively removes Cr(VI) from the drainage, the presence of Mn oxides can confound attenuation and improvements to Cr(VI) remediation should consider means of preventing the back reaction of Cr(III) with Mn oxides.

Effect of community population size on breast cancer screening, stage distribution, treatment use and outcomes.

OBJECTIVE: Residents of rural communities have decreased access to cancer screening and treatments compared to urban residents, though use of resources and patient outcomes have not been assessed with a comprehensive population-based analysis. The objectives of this study were to investigate whether breast cancer screening and treatments were utilized less frequently in rural BC and whether this translated into differences in outcomes. METHODS: All patients diagnosed with breast cancer in British Columbia (BC) during 2002 were identified from the Cancer Registry and linked to the Screening Mammography database. Patient demographics, pathology, stage, treatments, mammography use and death data were abstracted. Patients were categorized as residing in large, small and rural local health authorities (LHAs) using Canadian census information. Use of resources and outcomes were compared across these LHA size categories. We hypothesized that mastectomy rates (instead of breast-conserving surgery) would be higher in rural areas, since breast conservation is standardly accompanied by adjuvant radiotherapy, which has limited availability in rural BC. In contrast we hypothesized that cancer screening and systemic therapy use would be similar, as they are more widely dispersed across BC. Exploratory analyses were performed to assess whether disparities in screening and treatment utilization translated into differences in survival. RESULTS: 2,869 breast cancer patients were included in our study. Patients from rural communities presented with more advanced disease (p=0.01). On multivariable analysis, patients from rural, compared to urban, LHAs were less likely to be screening mammography attendees (OR=0.62; p<0.001). Women from rural communities were less likely to undergo breast-conserving surgery (multivariable OR=0.47; p<0.001). There was no significant difference in use of chemotherapy (p=0.54) or hormonal therapy (p=0.36). The 5-year breast cancer-specific survival for large, small and rural LHAs was 90%, 88% and 86%, respectively (p=0.08), while overall survival was 84%, 81% and 77%, respectively (p=0.01). On multivariable analysis with 7.4 years of median follow-up, neither breast cancer-specific survival (HR=1.16; 0.76-1.76; p=0.49) nor overall survival (HR=1.25; 0.92-1.70; p=0.16) was significantly worse for patients from rural compared to large LHAs. CONCLUSION: There was a significant difference in screening mammography use, stage distribution and loco-regional treatments use by population size of LHA. After controlling for differences in patient and tumour factors by LHA, survival was not significantly different.

Predictors of Treatment Response to Multidisciplinary Care for Persistent Symptoms after Pediatric Concussion.

PURPOSE: To assess which pediatric patients experiencing persistent post-concussive symptoms (PCS) benefit most from multidisciplinary treatment including specialists in Neurology, Neuropsychology, Physical Therapy, and Athletic Training, and to explore the effectiveness of this approach. METHODS: A retrospective chart review of 56 adolescents 10-20 years old (M = 15.0 ± 2.1) receiving multidisciplinary care for PCS (>30 days) was conducted. RESULTS: Systolic blood pressure and Body Mass Index predicted time to concussion resolution (p < .05), such that higher values were associated with slower resolution. PCS scores significantly decreased between participants' initial and final clinic visits, p < .01, and among the 25 participants for whom pre-intervention PCS scores were available, symptom severity scores significantly declined following multidisciplinary intervention compared to pre-referral values (p < .01). CONCLUSIONS: Exploratory analyses reveal that multidisciplinary treatment is a promising approach for reducing symptoms among adolescents with PCS, and that those with greater levels of physical fitness may benefit most.

Chromatin-level regulation of biosynthetic gene clusters.

Loss-of-function Aspergillus nidulans CclA, a Bre2 ortholog involved in histone H3 lysine 4 methylation, activated the expression of cryptic secondary metabolite clusters in A. nidulans. One new cluster generated monodictyphenone, emodin and emodin derivatives, whereas a second encoded two anti-osteoporosis polyketides, F9775A and F9775B. Modification of the chromatin landscape in fungal secondary metabolite clusters allows for a simple technological means to express silent fungal secondary metabolite gene clusters.

Do anthracyclines still have a role in adjuvant chemotherapy of breast cancer?

Anthracycline-based regimens became the standard of care for early breast cancer patients based on the survival advantage they provide over nonanthracycline-containing regimens. The addition of taxanes, and subsequently trastuzumab in HER2-overexpressing patients, to anthracyclines further improved their efficacy in several studies involving high-risk early breast cancer patients. Concern over toxicity initially surfaced after anthracyclines were reported to carry an increased risk of cardiotoxicity and secondary leukemia. Trastuzumab has since been shown to compound the risk of cardiotoxicity in patients who have received an anthracycline. This has led to the development of regimens featuring a taxane without an anthracycline; these protocols vary in design and have different toxicity and efficacy profiles. Ongoing investigations are centered on the optimization of nonanthracycline regimens, prospective exploration of molecular markers to identify populations of patients who will derive maximal benefit from anthracycline-based chemotherapy, and the identification of less cardiotoxic formulations of existing anthracycline agents. Perhaps most importantly, a rapidly growing understanding of the biological heterogeneity of breast cancer is likely to lead to an individualized standard of care guided by particular patient and tumor characteristics.

Characterization of the Aspergillus nidulans monodictyphenone gene cluster.

Deletion of cclA, a component of the COMPASS complex of Aspergillus nidulans, results in the production of monodictyphenone and emodin derivatives. Through a set of targeted deletions in a cclA deletion strain, we have identified the genes required for monodictyphenone and emodin analog biosynthesis. Identification of an intermediate, endocrocin, from an mdpHDelta strain suggests that mdpH might encode a decarboxylase. Furthermore, by replacing the promoter of mdpA (a putative aflJ homolog) and mdpE (a putative aflR homolog) with the inducible alcA promoter, we have confirmed that MdpA functions as a coactivator. We propose a biosynthetic pathway for monodictyphenone and emodin derivatives based on bioinformatic analysis and characterization of biosynthetic intermediates.

Evaluating cimetidine for GFR estimation in liver transplant recipients.

Background. Serum creatinine (Scr)-based equations lack accuracy in predicting glomerular filtration rate (GFR) in patients with liver disease. Cimetidine has been shown to improve the performance of Scr-based GFR formulae. Methods. We evaluated the use of cimetidine on the performance of GFR-estimating equations in 39 liver transplant recipients. The patients received oral cimetidine (800 mg tid) during a 24-h urine collection. The next day, the patients underwent radionucleotide GFR (rGFR) determination and Scr was measured for creatinine clearance (CrCl) and GFR estimation using the Cockcroft-Gault, Nankivell and modified diet in renal disease (MDRD) equations. Data were analysed using the Pearson correlation statistic and Bland-Altman plots. Results. The mean rGFR was 65 +/- 26.4 mL/min. The use of cimetidine increased the bias between rGFR and the Nankivell and MDRD equations. The combined root mean square error for the CrCl, Cockcroft-Gault, Nankivell and MDRD equations without cimetidine were 20.2, 15.6, 17.0 and 15.5 and cimetidine-aided were 28.2, 23.2, 23.7 and 24.3, respectively. Conclusions. All the tested equations without using cimetidine predicted GFR with modest accuracy. The addition of cimetidine decreased the precision and increased the bias of all the GFR-estimating equations. In the absence of accurate GFR-estimating equations, rGFR should be used to monitor kidney function in liver transplant recipients.

A gene cluster containing two fungal polyketide synthases encodes the biosynthetic pathway for a polyketide, asperfuranone, in Aspergillus nidulans.

The genome sequencing of Aspergillus species including A. nidulans reveals that the products of many of the secondary metabolism pathways in these fungi have not been elucidated. Our examination of the 27 polyketide synthases (PKS) in A. nidulans revealed that one highly reduced PKS (HR-PKS, AN1034.3) and one nonreduced PKS (NR-PKS, AN1036.3) are located next to each other in the genome. Since no known A. nidulans secondary metabolites could be produced by two PKS enzymes, we hypothesized that this cryptic gene cluster produces an unknown natural product. Indeed after numerous attempts we found that the products from this cluster could not be detected under normal laboratory culture conditions in wild type strains. Closer examination of the gene cluster revealed a gene with high homology to a citrinin biosynthesis transcriptional activator (CtnR, 32% identity/47% similarity), a fungal transcription activator located next to the two PKSs. We replaced the promoter of the transcription activator with the inducible alcA promoter, which enabled the production of a novel polyketide that we have named asperfuranone. A series of gene deletions has allowed us to confirm that the two PKSs together with five additional genes comprise the asperfuranone biosynthetic pathway and leads us to propose a biosynthetic pathway for asperfuranone. Our results confirm and substantiate the potential to discover novel compounds even from a well-studied fungus by using a genomic mining approach.

Molecular genetic mining of the Aspergillus secondary metabolome: discovery of the emericellamide biosynthetic pathway.

The recently sequenced genomes of several Aspergillus species have revealed that these organisms have the potential to produce a surprisingly large range of natural products, many of which are currently unknown. We have found that A. nidulans produces emericellamide A, an antibiotic compound of mixed origins with polyketide and amino acid building blocks. Additionally, we describe the discovery of four previously unidentified, related compounds that we designate emericellamide C-F. Using recently developed gene targeting techniques, we have identified the genes involved in emericellamide biosynthesis. The emericellamide gene cluster contains one polyketide synthase and one nonribosomal peptide synthetase. From the sequences of the genes, we are able to deduce a biosynthetic pathway for the emericellamides. The identification of this biosynthetic pathway opens the door to engineering novel analogs of this structurally complex metabolite.

Identification and characterization of the asperthecin gene cluster of Aspergillus nidulans.

The sequencing of Aspergillus genomes has revealed that the products of a large number of secondary metabolism pathways have not yet been identified. This is probably because many secondary metabolite gene clusters are not expressed under normal laboratory culture conditions. It is, therefore, important to discover conditions or regulatory factors that can induce the expression of these genes. We report that the deletion of sumO, the gene that encodes the small ubiquitin-like protein SUMO in A. nidulans, caused a dramatic increase in the production of the secondary metabolite asperthecin and a decrease in the synthesis of austinol/dehydroaustinol and sterigmatocystin. The overproduction of asperthecin in the sumO deletion mutant has allowed us, through a series of targeted deletions, to identify the genes required for asperthecin synthesis. The asperthecin biosynthesis genes are clustered and include genes encoding an iterative type I polyketide synthase, a hydrolase, and a monooxygenase. The identification of these genes allows us to propose a biosynthetic pathway for asperthecin.

Isolation, culture, and characterization of canine Sertoli cells.

Primary Sertoli cell cultures have been established from several animals including the sheep and rhesus monkey; however, not for the domestic dog, Canis familiaris. Sertoli cells are the only readily accessible cell type in the body which expresses all six type IV collagens. These collagens play key roles in tissue structure, basement membrane formation, and filtration. The study of these genes is necessary to determine their exact roles and regulation in the aforementioned functions and to investigate diseases associated with mutations in these genes. For such studies, a cell culture system is a requisite tool. Therefore, Sertoli cells were targeted, and a culture was established from cells isolated from canine testes. Cultures maintained consistent morphology and steady growth for up to seven passages. Cultured cells were identified as Sertoli cells through positive Western blot results for SOX9 and Clusterin B proteins and transcript sequence verification of SOX9 as well as the presence of type IV collagen transcripts. Primary cultures of canine Sertoli cells will provide a useful tool for study of the function and regulation of collagen genes and will permit new research pertaining to canine health while also serving as a model for the study of human diseases.

Genetic cause of autosomal recessive hereditary nephropathy in the English Cocker Spaniel.

BACKGROUND: Autosomal recessive hereditary nephropathy (ARHN) in the English Cocker Spaniel is caused by a type IV collagen defect, but the underlying mutation is unknown. ANIMALS: One hundred thirty-four English Cocker Spaniels (12 with ARHN, 8 obligate carriers, and 114 others), 3 mixed breed dogs with X-linked hereditary nephropathy (XLHN), and 7 other dogs without hereditary nephropathy were included. METHODS: Diagnosis of ARHN was based on transmission electron microscopy and immunostaining of kidney. Quantitative real time reverse transcriptase polymerase chain reaction (qRT-PCR) was used to compare COL4A3, COL4A4, and COL4A5 mRNA concentrations in the renal cortex from ARHN-affected English Cocker Spaniels, XLHN-affected dogs, and dogs without hereditary nephropathy. The entire coding region of COL4A4 was sequenced in 2 ARHN-affected dogs, 2 obligate carriers, 2 English Cocker Spaniels of unknown status, and 2 healthy mixed breed dogs. The exon containing the mutation was sequenced for all 134 English Cocker Spaniels. RESULTS: Quantitative real time RT-PCR implicated COL4A4 as the gene harboring the mutation, and sequencing identified a single nucleotide substitution at base 115 as the cause of ARHN in English Cocker Spaniels. This mutation, which causes a premature stop codon in exon 3 of COL4A4, was segregated with clinical status in all affected dogs and obligate carriers. The mutation also was identified in 39 of 114 other English Cocker Spaniels with previously unknown status. CONCLUSIONS AND CLINICAL IMPORTANCE: The cause of this disease has been identified, and use of a test for the mutation will permit eradication of ARHN in the English Cocker Spaniel.

Facilitating practice learning through a clinical academy.

This article reports on a project that identified a range of solutions to the challenges of facilitating induction and mandatory training within an acute general hospital. This involved developing a 'blended' approach to delivering induction and mandatory training/education.

Stage, treatment and outcomes for patients with breast cancer in British Columbia in 2002: a population-based cohort study.

BACKGROUND: There are very few long-term Canadian data on breast cancer outcomes by stage. We described the stage, treatment and outcomes of breast cancer at a population level for patients in British Columbia. METHODS: This population-based cohort study included almost all patients with incident breast cancer registered in 2002 (about 97.6% registry case completeness). For these patients, information on stage, primary local surgery, radiotherapy, chemotherapy, hormone therapy and survival outcome (based on registry date and cause-of-death data) were available. We calculated Kaplan-Meier curves for breast cancer-specific survival and overall survival by stage and analyzed prognostic and treatment factors with a multivariable Cox model. RESULTS: The 2927 incident cases of breast cancer identified in 2002 had the following distribution by stage: stage 0 (in situ), 424 (14%); stage I, 1118 (38%); stage II, 938 (32%); stage III, 233 (8%); stage IV, 123 (4%); unknown, 91 (3%). The distribution of patients' ages was < 40 years, 127 (4%); 40-49, 538 (18%); 50-59, 719 (25%); 60-69, 660 (23%); 70-79, 583 (20%); ≥ 80, 300 (10%). Within the first year after diagnosis, radiotherapy was provided to 1649 patients (56%), chemotherapy to 928 (32%) and hormone therapy to 1664 (57%). Ten-year breast cancer-specific survival rates by stage were > 99% for stage 0, 95% for stage I, 81% for stage II, 55% for stage III and 4% for stage IV. Ten-year overall survival rates were 89% for stage 0, 81% for stage I, 68% for stage II, 43% for stage III and 2% for stage IV. INTERPRETATION: This analysis provides a Canadian benchmark for treatment rates and 10-year outcomes by stage for all incident cases of breast cancer in a single province. Outcomes in British Columbia compared well with published rates for the United States and Europe.

Molecular genetic analysis of the orsellinic acid/F9775 gene cluster of Aspergillus nidulans.

F-9775A and F-9775B are cathepsin K inhibitors that arise from a chromatin remodelling deletant strain of Aspergillus nidulans. A polyketide synthase gene has been determined to be responsible for their formation and for the simpler, archetypical polyketide orsellinic acid. We have discovered simple culture conditions that result in the production of the three compounds, and this facilitates analysis of the genes responsible for their synthesis. We have now analysed the F9775/orsellinic acid gene cluster using a set of targeted deletions. We find that the polyketide synthase alone is required for orsellinic acid biosynthesis and only two additional genes in the cluster are required for F9775 A and B synthesis. Our deletions also yielded the bioactive metabolites gerfelin and diorcinol.