Peri-operative tobacco cessation interventions: a systematic review and meta-analysis.

Tobacco smoking is associated with a substantially increased risk of postoperative complications. The peri-operative period offers a unique opportunity to support patients to stop tobacco smoking, avoid complications and improve long-term health. This systematic review provides an up-to-date summary of the evidence for tobacco cessation interventions in surgical patients. We conducted a systematic search of randomised controlled trials of tobacco cessation interventions in the peri-operative period. Quantitative synthesis of the abstinence outcomes data was by random-effects meta-analysis. The primary outcome of the meta-analysis was abstinence at the time of surgery, and the secondary outcome was abstinence at 12 months. Thirty-eight studies are included in the review (7310 randomised participants) and 26 studies are included in the meta-analysis (5969 randomised participants). Studies were pooled for subgroup analysis in two ways: by the timing of intervention delivery within the peri-operative period and by the intensity of the intervention protocol. We judged the quality of evidence as moderate, reflecting the degree of heterogeneity and the high risk of bias. Overall, peri-operative tobacco cessation interventions increased successful abstinence both at the time of surgery, risk ratio (95%CI) 1.48 (1.20-1.83), number needed to treat 7; and 12 months after surgery, risk ratio (95%CI) 1.62 (1.29-2.03), number needed to treat 9. More work is needed to inform the design and optimal delivery of interventions that are acceptable to patients and that can be incorporated into contemporary elective and urgent surgical pathways. Future trials should use standardised outcome measures.

Scoping review of mode of anaesthesia in emergency surgery.

BACKGROUND: Emergency surgery encompasses more than 50 per cent of the surgical workload; however, research efforts are disproportionally low. The mode of anaesthesia used during emergency surgery may affect outcomes, but the extent of research and the impact of the different modes of anaesthesia used are unclear. METHODS: MEDLINE and Embase were searched using scoping review methodology with a rapid systematic search strategy, identifying any study comparing locoregional (local, nerve block, subarachnoid, epidural) anaesthesia with general anaesthesia. All studies describing outcomes of emergency surgery with differing modes of anaesthesia were identified. Excluded were: studies published before 2003, studies enrolling patients aged less than 18 years and studies using sedation only. RESULTS: Forty-two studies were identified, describing 11 surgical procedures. Most publications were retrospective cohort studies (32). A very broad range of clinical and patient-reported outcomes were described, with wide variation in the outcomes reported in different studies. CONCLUSION: Reporting of mode of anaesthesia is inconsistent across different procedures and is often absent. There is a need for directed research efforts to improve the reporting standards of anaesthesia interventions, to understand the role of different modes of anaesthesia in specific emergency surgical procedures, and to standardize outcome reporting using core outcome sets.

ANTECEDENTES: La cirugía de urgencias constituye > 50% de la carga de trabajo quirúrgico, aunque los esfuerzos realizados en investigación en este ámbito son desproporcionadamente bajos. La modalidad de anestesia utilizada durante la cirugía de urgencias puede afectar a los resultados, sin embargo, la investigación realizada y el impacto de los diferentes tipos de anestesia utilizados no están claros. MÉTODOS: Se realizaron búsquedas en Medline y Embase utilizando una metodología enfocada a la recuperación de revisiones, con una estrategia de búsqueda sistemática rápida, identificando cualquier estudio que comparara la anestesia locorregional (local, bloqueo nervioso, subaracnoidea, epidural) con la anestesia general. Se identificaron todos los estudios que describían los resultados de la cirugía de urgencias con diferentes tipos de anestesia. Se excluyeron los estudios publicados antes del 2003, los estudios que reclutaron pacientes < 18 años y los estudios que solo usaron sedación. RESULTADOS: Se identificaron 42 estudios que describían 11 procedimientos quirúrgicos. La mayoría de publicaciones fueron estudios de cohortes retrospectivos (n = 32). Se describió una gama muy amplia de resultados clínicos y resultados aportados por los pacientes, con una amplia variación en los resultados de los diferentes estudios. CONCLUSIÓN: Los resultados publicados respecto a la modalidad de anestesia empleada en diferentes procedimientos quirúrgicos son inconsistentes, a menudo esta información está ausente y no se pueden establecer conclusiones sobre el impacto del tipo de anestesia en los resultados. Es necesario realizar esfuerzos dirigidos a la investigación para mejorar la notificación de los estándares de los procedimientos de anestesia, comprender el papel de los diferentes tipos de anestesia en los procedimientos quirúrgicos específicos de urgencias, y estandarizar la presentación de los resultados obtenidos utilizando un conjunto de datos principales.

Validation and development of MTH1 inhibitors for treatment of cancer.

BACKGROUND: Previously, we showed cancer cells rely on the MTH1 protein to prevent incorporation of otherwise deadly oxidised nucleotides into DNA and we developed MTH1 inhibitors which selectively kill cancer cells. Recently, several new and potent inhibitors of MTH1 were demonstrated to be non-toxic to cancer cells, challenging the utility of MTH1 inhibition as a target for cancer treatment. MATERIAL AND METHODS: Human cancer cell lines were exposed in vitro to MTH1 inhibitors or depleted of MTH1 by siRNA or shRNA. 8-oxodG was measured by immunostaining and modified comet assay. Thermal Proteome profiling, proteomics, cellular thermal shift assays, kinase and CEREP panel were used for target engagement, mode of action and selectivity investigations of MTH1 inhibitors. Effect of MTH1 inhibition on tumour growth was explored in BRAF V600E-mutated malignant melanoma patient derived xenograft and human colon cancer SW480 and HCT116 xenograft models. RESULTS: Here, we demonstrate that recently described MTH1 inhibitors, which fail to kill cancer cells, also fail to introduce the toxic oxidized nucleotides into DNA. We also describe a new MTH1 inhibitor TH1579, (Karonudib), an analogue of TH588, which is a potent, selective MTH1 inhibitor with good oral availability and demonstrates excellent pharmacokinetic and anti-cancer properties in vivo. CONCLUSION: We demonstrate that in order to kill cancer cells MTH1 inhibitors must also introduce oxidized nucleotides into DNA. Furthermore, we describe TH1579 as a best-in-class MTH1 inhibitor, which we expect to be useful in order to further validate the MTH1 inhibitor concept.

Improving molecular detection of fungal DNA in formalin-fixed paraffin-embedded tissues: comparison of five tissue DNA extraction methods using panfungal PCR.

DNA extraction from formalin-fixed paraffin-embedded (FFPE) tissues is difficult and requires special protocols in order to extract small amounts of DNA suitable for amplification. Most described methods report an amplification success rate between 60 and 80%; therefore, there is a need to improve molecular detection and identification of fungi in FFPE tissue. Eighty-one archived FFPE tissues with a positive Gomori methenamine silver (GMS) stain were evaluated using five different commercial DNA extraction kits with some modifications. Three different panfungal PCR assays were used to detect fungal DNA, and two housekeeping genes were used to assess the presence of amplifiable DNA and to detect PCR inhibitors. The sensitivities of the five extraction protocols were compared, and the quality of DNA detection (calculated for each kit as the number of housekeeping gene PCR-positive samples divided by the total number of samples) was 60 to 91% among the five protocols. The efficiencies of the three different panfungals used (calculated as the number of panfungal-PCR-positive samples divided by the number of housekeeping gene PCR-positive samples) were 58 to 93%. The panfungal PCR using internal transcribed spacer 3 (ITS3) and ITS4 primers yielded a product in most FFPE tissues. Two of the five DNA extraction kits (from TaKaRa and Qiagen) showed similar and promising results. However, one method (TaKaRa) could extract fungal DNA from 69 of the 74 FFPE tissues from which a housekeeping gene could be amplified and was also cost-effective, with a nonlaborious protocol. Factors such as sensitivity, cost, and labor will help guide the selection of the most appropriate method for the needs of each laboratory.

MIPS: a database for genomes and protein sequences.

The Munich Information Center for Protein Sequences (MIPS-GSF, Neuherberg, Germany) continues to provide genome-related information in a systematic way. MIPS supports both national and European sequencing and functional analysis projects, develops and maintains automatically generated and manually annotated genome-specific databases, develops systematic classification schemes for the functional annotation of protein sequences, and provides tools for the comprehensive analysis of protein sequences. This report updates the information on the yeast genome (CYGD), the Neurospora crassa genome (MNCDB), the databases for the comprehensive set of genomes (PEDANT genomes), the database of annotated human EST clusters (HIB), the database of complete cDNAs from the DHGP (German Human Genome Project), as well as the project specific databases for the GABI (Genome Analysis in Plants) and HNB (Helmholtz-Netzwerk Bioinformatik) networks. The Arabidospsis thaliana database (MATDB), the database of mitochondrial proteins (MITOP) and our contribution to the PIR International Protein Sequence Database have been described elsewhere [Schoof et al. (2002) Nucleic Acids Res., 30, 91-93; Scharfe et al. (2000) Nucleic Acids Res., 28, 155-158; Barker et al. (2001) Nucleic Acids Res., 29, 29-32]. All databases described, the protein analysis tools provided and the detailed descriptions of our projects can be accessed through the MIPS World Wide Web server (http://mips.gsf.de).

hMYH and hMTH1 cooperate for survival in mismatch repair defective T-cell acute lymphoblastic leukemia.

hMTH1 is an 8-oxodGTPase that prevents mis-incorporation of free oxidized nucleotides into genomic DNA. Base excision and mismatch repair pathways also restrict the accumulation of oxidized lesions in DNA by removing the mis-inserted 8-oxo-7,8-dihydro-2'-deoxyguanosines (8-oxodGs). In this study, we aimed to investigate the interplay between hMYH DNA glycosylase and hMTH1 for cancer cell survival by using mismatch repair defective T-cell acute lymphoblastic leukemia (T-ALL) cells. To this end, MYH and MTH1 were silenced individually or simultaneously using small hairpin RNAs. Increased sub-G1 population and apoptotic cells were observed upon concurrent depletion of both enzymes. Elevated cell death was consistent with cleaved caspase 3 accumulation in double knockdown cells. Importantly, overexpression of the nuclear isoform of hMYH could remove the G1 arrest and partially rescue the toxicity observed in hMTH1-depleted cells. In addition, expression profiles of human DNA glycosylases were generated using quantitative reverse transcriptase-PCR in MTH1 and/or MYH knockdown cells. NEIL1 DNA glycosylase, involved in repair of oxidized nucleosides, was found to be significantly downregulated as a cellular response to MTH1-MYH co-suppression. Overall, the results suggest that hMYH and hMTH1 functionally cooperate for effective repair and survival in mismatch repair defective T-ALL Jurkat A3 cells.

Sex differences in the diabetes-induced modulation of rat hepatic cytochrome P450 proteins.

Sex differences in the diabetes-induced changes in hepatic cytochrome P450 proteins were investigated in rats treated with streptozotocin. Changes in specific cytochrome P450 proteins were monitored using diagnostic substrates and immunologically utilizing specific polyclonal antibodies. When expressed in terms of nmoles of total cytochrome P450, ethoxyresorufin O-deethylase activity was increased by treatment with streptozotocin, the extent of induction being the same in the two sexes. In contrast, lauric acid hydroxylase and ethylmorphine N-demethylase activities were induced only in the male rat. Finally, p-nitrophenol hydroxylase and pentoxyresorufin O-dealkylase were enhanced by the same treatment in both sexes, the effect being more pronounced in the male. These findings indicate that sex-specific changes in certain cytochrome P450 proteins exist in response to insulin-dependent diabetes but these cannot, however, be ascribed to sex differences in the severity of diabetes induced by streptozotocin since the degrees of hyperketonaemia and hyperglycaemia were the same in the two sexes. These are likely to reflect sex-specific differences in growth hormone and triglyceride levels in the diabetic animals.

MRI of polysplenia syndrome.

The polysplenia syndrome is the association of multiple spleens, situs inversus, congenital heart disease, and azygous continuation of the inferior vena cava. Magnetic resonance (MR) is a noninvasive imaging modality which can easily confirm the multiplicity of spleens, situs inversus, and identify complex congenital cardiovascular malformations. The anomalies of the polysplenia syndrome as imaged by MR are presented.

Mediastinal perigraft seroma: CT and MR imaging.

We report a patient who developed a perigraft seroma after a modified Blalock-Taussig procedure in which a polytetrafluoroethylene (Gore-tex) graft was used. Computed tomography and magnetic resonance were instrumental in the evaluation of this seroma.

Clinical utility of bone SPECT scintigraphy in renal metastases from primary osteosarcoma.

Renal metastases from primary osteosarcomas are rather uncommon and rarely diagnosed early because the patients are asymptomatic and frequently die from other metastatic involvement before renal symptoms develop. The authors present a patient with two clinically silent renal metastases from primary osteosarcoma of the right femur 2 years after surgery of the primary lesion that was first detected on radionuclide bone imaging. Subsequently, a CT scan and a CT guided needle biopsy were performed for confirmation. The patient underwent a left nephrectomy and two separate lesions were proven to be metastatic osteosarcoma. The clinical importance of the nuclear bone scan in the initial management, as well as in the subsequent follow-up of patients after surgery, cannot be overemphasized.

Nitrospira-dominated biofilm within a thermal artesian spring: a case for nitrification-driven primary production in a geothermal setting.

Water chemistry, energetic modeling, and molecular analyses were combined to investigate the microbial ecology of a biofilm growing in a thermal artesian spring within Hot Springs National Park, AR. This unique fresh water spring has a low dissolved chemical load and is isolated from both light and direct terrestrial carbon input - resulting in an oligotrophic ecosystem limited for fixed carbon and electron donors. Evaluation of energy yields of lithotrophic reactions putatively linked to autotrophy identified the aerobic oxidation of methane, hydrogen, sulfide, ammonia, and nitrite as the most exergonic. Small subunit (SSU) rRNA gene libraries from biofilm revealed a low-diversity microbial assemblage populated by bacteria and archaea at a gene copy ratio of 45:1. Members of the bacterial family 'Nitrospiraceae', known for their autotrophic nitrite oxidation, dominated the bacterial SSU rRNA gene library (approximately 45%). Members of the Thaumarchaeota ThAOA/HWCGIII (>96%) and Thaumarchaeota Group I.1b (2.5%), which both contain confirmed autotrophic ammonia oxidizers, dominated the archaeal SSU rRNA library. Archaea appear to dominate among the ammonia oxidizers, as only ammonia monooxygenase subunit A (amoA) genes belonging to members of the Thaumarchaeota were detected. The geochemical, phylogenetic, and genetic data support a model that describes a novel thermophilic biofilm built largely by an autotrophic nitrifying microbial assemblage. This is also the first observation of 'Nitrospiraceae' as the dominant organisms within a geothermal environment.

Comparative mapping of DNA sequences in rye (Secale cereale L.) in relation to the rice genome.

The rice genome has proven a valuable resource for comparative approaches to address individual genomic regions in Triticeae species at the molecular level. To exploit this resource for rye genetics and breeding, an inventory was made of EST-derived markers with known genomic positions in rye, which were related with those in rice. As a first inventory set, 92 EST-SSR markers were mapped which had been drawn from a non-redundant rye EST collection representing 5,423 unigenes and 2.2 Mb of DNA. Using a BC1 mapping population which involved an exotic rye accession as donor parent, these EST-SSR markers were arranged in a linkage map together with 25 genomic SSR markers as well as 131 AFLP and four STS markers. This map comprises seven linkage groups corresponding to the seven rye chromosomes and covers 724 cM of the rye genome. For comparative studies, additional inventory sets of EST-based markers were included which originated from the rye-mapping data published by other authors. Altogether, 502 EST-based markers with known chromosomal localizations in rye were used for BlastN search and 334 of them could be in silico mapped in the rice genome. Additionally, 14 markers were included which lacked sequence information but had been genetically mapped in rice. Based on the 348 markers, each of the seven rye chromosomes could be aligned with distinct portions of the rice genome, providing improved insight into the status of the rye-rice genome relationships. Furthermore, the aligned markers provide genomic anchor points between rye and rice, enabling the identification of conserved ortholog set markers for rye. Perspectives of rice as a model for genome analysis in rye are discussed.

Characterization of an 18,166 EST dataset for cassava (Manihot esculenta Crantz) enriched for drought-responsive genes.

Cassava (Manihot esculenta Crantz) is a staple food for over 600 million people in the tropics and subtropics and is increasingly used as an industrial crop for starch production. Cassava has a high growth rate under optimal conditions but also performs well in drought-prone areas and on marginal soils. To increase the tools for understanding and manipulating drought tolerance in cassava, we generated expressed sequence tags (ESTs) from normalized cDNA libraries prepared from dehydration-stressed and control well-watered tissues. Analysis of a total of 18,166 ESTs resulted in the identification of 8,577 unique gene clusters (5,383 singletons and 3,194 clusters). Functional categories could be assigned to 63% of the unigenes, while another approximately 11% were homologous to hypothetical genes with unclear functions. The remaining approximately 26% were not significantly homologous to sequences in public databases suggesting that some may be novel and putatively specific to cassava. The dehydration-stressed library uncovered numerous ESTs with recognized roles in drought-responses, including those that encode late-embryogenesis-abundant proteins thought to confer osmoprotective functions during water stress, transcription factors, heat-shock proteins as well as proteins involved in signal transduction and oxidative stress. The unigene clusters were screened for short tandem repeats for further development as microsatellite markers. A total of 592 clusters contained 646 repeats, representing 3.3% of the ESTs queried. The ESTs presented here are the first dehydration stress transcriptome of cassava and can be utilized for the development of microarrays and gene-derived molecular markers to further dissect the molecular basis of drought tolerance in cassava.

Prospecting for pig single nucleotide polymorphisms in the human genome: have we struck gold?

Gene-to-gene variation in the frequency of single nucleotide polymorphisms (SNPs) has been observed in humans, mice, rats, primates and pigs, but a relationship across species in this variation has not been described. Here, the frequency of porcine coding SNPs (cSNPs) identified by in silico methods, and the frequency of murine cSNPs, were compared with the frequency of human cSNPs across homologous genes. From 150,000 porcine expressed sequence tag (EST) sequences, a total of 452 SNP-containing sequence clusters were found, totalling 1394 putative SNPs. All the clustered porcine EST annotations and SNP data have been made publicly available at http://sputnik.btk.fi/project?name=swine. Human and murine cSNPs were identified from dbSNP and were characterized as either validated or total number of cSNPs (validated plus non-validated) for comparison purposes. The correlation between in silico pig cSNP and validated human cSNP densities was found to be 0.77 (p < 0.00001) for a set of 25 homologous genes, while a correlation of 0.48 (p < 0.0005) was found for a primarily random sample of 50 homologous human and mouse genes. This is the first evidence of conserved gene-to-gene variability in cSNP frequency across species and indicates that site-directed screening of porcine genes that are homologous to cSNP-rich human genes may rapidly advance cSNP discovery in pigs.

Identifying adolescents at risk for noncompliance with contraceptive therapy.

The sociomedical characteristics of 96 sexually active adolescent females have been studied in order to understand noncompliance with the contraceptive prescription, an important antecendent of adolescent pregnancy. Compliance was found to correlate positively with postmenarchal age, frequency of intercourse, autonomy in making and paying for a clinic appointment specifically for the purpose of obtaining contraception, and acceptance of a method at the time of the initial clinic visit. These findings serve to alert the pediatrician to the adolescent at risk for noncompliance before the experience of failure, and in so doing to identify those in need of special assistance to ensure the success of contraceptive measures.

PIP: 96 sexually active adolescent females attending the Stanford University Adolescent Clinic of the Dept. of Pediatrics between 9/76 and 11/78 were studied to determine factors which contribute to noncompliance with contraceptive prescription, an important antecedent of adolescent pregnancy. 15 variables were selected for the study and included information on demographic characteristics; medical history; patterns of sexual activity; physical development; and motivation for clinic attendance. Chi-square and stepwise discriminant analyses were the analytical methods used. Mean age of study group was 15.9 years; slightly more than half were blacks. One-third of the group had parental knowledge of their sexual activity. Almost 2/3 had a single sex partner, and mean duration of sexual relationship was nearly 1 year. More than 1/3 had intercourse less often than once a week. Half of the patients had initiated their own clinic appointment, while the remaining were influenced by parents, physicians and others. 44% of the subjects complied with the contraceptive regimen during the study period. Adolescents who paid for their own medical care; who had a single sex partner; who were sexually active more often than weekly; who made their own clinic appointments; and who came to the clinic themselves desiring birth control were more likely to be compliant than the others. Using standardized discriminant function coefficient for 6 variables (computed compliance values; method of payment; method of birth control; frequency of intercourse; appointment source; agenda source), patients were correctly assigned to the compliant or noncompliant group in 72% of the cases. Implications of these findings, and the role of the primary care physician in the prevention of adolescent pregnancy, are discussed.

Arabidopsis genome analysis as exemplified by analysis of chromosome 4.

During the last decade the small cruciferous plant Arabidopsis thaliana has become a model organism for flowering plants. Sequencing and analysis of the Arabidopsis genome is nearing completion. Beside an overview on methods and strategies for Arabidopsis genome analysis, a summary of the results from the first analysis is presented. This includes an overview on chromosomal organisation and topological features as well as a first comparison with other genomes.

An RNA-dependent RNA polymerase gene in Arabidopsis is required for posttranscriptional gene silencing mediated by a transgene but not by a virus.

Posttranscriptional gene silencing is a defense mechanism in plants that is similar to quelling in fungi and RNA interference in animals. Here, we describe four genetic loci that are required for posttranscriptional gene silencing in Arabidopsis. One of these, SDE1, is a plant homolog of QDE-1 in Neurospora crassa that encodes an RNA-dependent RNA polymerase. The sde1 mutation was specific for posttranscriptional gene silencing induced by transgenes rather than by viruses. We propose that the role of SDE1 is to synthesize a double-stranded RNA initiator of posttranscriptional gene silencing. According to this idea, when a virus induces posttranscriptional gene silencing, the virus-encoded RNA polymerase would produce the double-stranded RNA and SDE1 would be redundant.

Snipping polymorphisms from large EST collections in barley (Hordeum vulgare L.).

The public EST (expressed sequence tag) databases represent an enormous but heterogeneous repository of sequences, including many from a broad selection of plant species and a wide range of distinct varieties. The significant redundancy within large EST collections makes them an attractive resource for rapid pre-selection of candidate sequence polymorphisms. Here we present a strategy that allows rapid identification of candidate SNPs in barley (Hordeum vulgare L.) using publicly available EST databases. Analysis of 271,630 EST sequences from different cDNA libraries, representing 23 different barley varieties, resulted in the generation of 56,302 tentative consensus sequences. In all, 8171 of these unigene sequences are members of clusters with six or more ESTs. By applying a novel SNP detection algorithm (SNiPpER) to these sequences, we identified 3069 candidate inter-varietal SNPs. In order to verify these candidate SNPs, we selected a small subset of 63 present in 36 ESTs. Of the 63 SNPs selected, we were able to validate 54 (86%) using a direct sequencing approach. For further verification, 28 ESTs were mapped to distinct loci within the barley genome. The polymorphism information content (PIC) and nucleotide diversity (pi) values of the SNPs identified by the SNiPpER algorithm are significantly higher than those that were obtained by random sequencing. This demonstrates the efficiency of our strategy for SNP identification and the cost-efficient development of EST-based SNP-markers.