Simultaneous Detection of FISH Signals and Bromo-Deoxyuridine Incorporation in Fixed Tissue Cultured Cells

FISH (Fluorescence in situ hybridization) is a powerful technique that detects and localises specific DNA sequences on metaphase chromosomes, interphase nuclei or chromatin fibres. When coupled to BrdU (5-Bromo 2-deoxy-uridine) labeling of newly replicated DNA, the replication properties of different DNA sequences can be analysed. However, the technique for the detection of BrdU incorporation is time consuming, and relies on acidic pH buffer treatments, that prevent use of pH sensitive fluorochromes such as FITC (Fluoro-isothiocianate) during FISH. In this work, we describe a simplified protocol that allows the simultaneous detection of FISH signals and BrdU incorporation. Since the technique does not involve paraformaldehyde for cell fixation, or formamide for denaturation of the target DNA and in post-hybridisation washes, it represents a safer alternative to classical FISH techniques

BASS. XXXV. The MBH-σ* Relation of 105 Month Swift-BAT Type 1 AGNs

We present two independent measurements of stellar velocity dispersions (sigma(star)) from the Ca II H+K lambda 3969, 3934 and Mg I b lambda 5183, 5172, 5167 region (3880-5550 angstrom) and the calcium triplet region (8350-8750 angstrom) for 173 hard X-ray-selected Type 1 active galactic nuclei (AGNs; z <= 0.08) from the 105 month Swift-BAT catalog. We construct one of the largest samples of local Type 1 AGNs that have both single-epoch virial black hole mass (M-BH) estimates and sigma(star) measurements obtained from high spectral resolution data, allowing us to test the usage of such methods for supermassive black hole studies. We find that the two independent sigma(star) measurements are highly consistent with each other, with an average offset of only 0.002 +/- 0.001 dex. Comparing M-BH estimates based on broad emission lines and stellar velocity dispersion measurements, we find that the former is systematically lower by approximate to 0.12 dex. Consequently, Eddington ratios estimated through broad-line MBH determinations are similarly biased (but in the opposite way). We argue that the discrepancy is driven by extinction in the broad-line region. We also find an anticorrelation between the offset from the M-BH-sigma(star) relation and the Eddington ratio. Our sample of Type 1 AGNs shows a shallower M-BH-sigma(star) relation (with a power-law exponent of approximate to 3.5) compared with that of inactive galaxies (with a power-law exponent of approximate to 4.5), confirming earlier results obtained from smaller samples

Drug resistance associated genetic polymorphisms in Plasmodium falciparum and Plasmodium vivax collected in Honduras, Central America

Background: In Honduras, chloroquine and primaquine are recommended and still appear to be effective for treatment of Plasmodium falciparum and Plasmodium vivax malaria. The aim of this study was to determine the proportion of resistance associated genetic polymorphisms in P. falciparum and P. vivax collected in Honduras. Methods: Blood samples were collected from patients seeking medical attention at the Hospital Escuela in Tegucigalpa from 2004 to 2006 as well as three regional hospitals, two health centres and one regional laboratory during 2009. Single nucleotide polymorphisms in P. falciparum chloroquine resistance transporter (pfcrt), multidrug resistance 1 (pfmdr1), dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes and in P. vivax multidrug resistance 1 (pvmdr1) and dihydrofolate reductase (pvdhfr) genes were detected using PCR based methods. Results: Thirty seven P. falciparum and 64 P. vivax samples were collected. All P. falciparum infections acquired in Honduras carried pfcrt, pfmdr1, pfdhps and pfdhfr alleles associated with chloroquine, amodiaquine and sulphadoxine-pyrimethamine sensitivity only. One patient with parasites acquired on a Pacific Island had pfcrt 76 T and pfmdr1 86Y alleles. That patient and a patient infected in West Africa had pfdhfr 51I, 59 R and 108 N alleles. Pvmdr1 976 F was found in 7/37 and two copies of pvmdr1 were found in 1/37 samples. Pvdhfr 57 L + 58 R was observed in 2/57 samples. Conclusion: The results indicate that P. falciparum from Honduras remain sensitive to chloroquine and sulphadoxine-pyrimethamine. This suggests that chloroquine and sulphadoxine-pyrimethamine should be efficacious for treatment of uncomplicated P. falciparum malaria, supporting current national treatment guidelines. However, genetic polymorphisms associated with chloroquine and sulphadoxine-pyrimethamine tolerance were detected in local P. vivax and imported P. falciparum infections. Continuous monitoring of the prevalence of drug resistant/tolerant P. falciparum and P. vivax is therefore essential also in Honduras.Swedish International Development Cooperation Agency, Department for research Cooperation (Sida-SAREC) [75007082/03]info:eu-repo/semantics/publishedVersio

Epidemiological and some clinical characteristics of neuroblastoma in Mexican children (1996–2005)

<p>Abstract</p> <p>Background</p> <p>Neuroblastoma (NB) is the principal tumor of the sympathetic nervous system in children under one year of age. The incidence in developed countries is greater than that in developing countries. The aim of this article is to present the epidemiological and some clinical characteristics of Mexican children with NB.</p> <p>Methods</p> <p>A population-based, prolective study, with data obtained from the Childhood Cancer Registry of the Instituto Mexicano de Seguro Social. Statistical analysis: The simple frequencies of the variables of the study and the annual average incidence (per 1,000,000 children/years) by age and sex were obtained. The trend was evaluated by calculating the annual percentage of change. The curves of Kaplan-Meyer were employed for the survival rate and the log-rank test was used to compare the curves.</p> <p>Results</p> <p>Of a total of 2,758 children with cancer registered during the period from 1996–2005, 72 (2.6%) were identified as having Group IV, defined according to the International Classification of Childhood Cancer. The incidence for NB was 3.8 per 1,000,000 children/year; NB was highest in the group of children under one year of age, followed by the group of children between the ages 1–4 years (18.5 and 5.4 per 1,000,000 children/years, respectively). The male/female ratio was 1.1 and there was no trend toward an increase. The time of diagnosis was 26 days (median), but varied according to the stage at diagnosis. Stages III and IV were presented in 88% of the cases. There was no association between the stage, the age at time of diagnosis, or the histological pattern. The overall five-year survival rate was 64%; the patients with stage I, II, III, or IVs did not die; and the five-year survival rate of cases in Stage IV was 40%.</p> <p>Conclusion</p> <p>It is possible that the low incidence of neuroblastoma in Mexican children is due to the difficulty in diagnosing the cases with the best prognosis, some of which could have had spontaneous regression. There was no trend to an increase; the majority of the cases were diagnosed in the advanced stages; and the overall five-years survival rate was similar to that for developed countries.</p

Ancestral diversity improves discovery and fine-mapping of genetic loci for anthropometric traits — The Hispanic/Latino Anthropometry Consortium

Hispanic/Latinos have been underrepresented in genome-wide association studies (GWAS) for anthropometric traits despite their notable anthropometric variability, ancestry proportions, and high burden of growth stunting and overweight/obesity. To address this knowledge gap, we analyzed densely imputed genetic data in a sample of Hispanic/Latino adults to identify and fine-map genetic variants associated with body mass index (BMI), height, and BMI-adjusted waist-to-hip ratio (WHRadjBMI). We conducted a GWAS of 18 studies/consortia as part of the Hispanic/Latino Anthropometry (HISLA) Consortium (stage 1, n = 59,771) and generalized our findings in 9 additional studies (stage 2, n = 10,538). We conducted a trans-ancestral GWAS with summary statistics from HISLA stage 1 and existing consortia of European and African ancestries. In our HISLA stage 1 + 2 analyses, we discovered one BMI locus, as well as two BMI signals and another height signal each within established anthropometric loci. In our trans-ancestral meta-analysis, we discovered three BMI loci, one height locus, and one WHRadjBMI locus. We also identified 3 secondary signals for BMI, 28 for height, and 2 for WHRadjBMI in established loci. We show that 336 known BMI, 1,177 known height, and 143 known WHRadjBMI (combined) SNPs demonstrated suggestive transferability (nominal significance and effect estimate directional consistency) in Hispanic/Latino adults. Of these, 36 BMI, 124 height, and 11 WHRadjBMI SNPs were significant after trait-specific Bonferroni correction. Trans-ancestral meta-analysis of the three ancestries showed a small-to-moderate impact of uncorrected population stratification on the resulting effect size estimates. Our findings demonstrate that future studies may also benefit from leveraging diverse ancestries and differences in linkage disequilibrium patterns to discover novel loci and additional signals with less residual population stratification

Ribosome-Dependent ATPase Interacts with Conserved Membrane Protein in Escherichia coli to Modulate Protein Synthesis and Oxidative Phosphorylation

Elongation factor RbbA is required for ATP-dependent deacyl-tRNA release presumably after each peptide bond formation; however, there is no information about the cellular role. Proteomic analysis in Escherichia coli revealed that RbbA reciprocally co-purified with a conserved inner membrane protein of unknown function, YhjD. Both proteins are also physically associated with the 30S ribosome and with members of the lipopolysaccharide transport machinery. Genome-wide genetic screens of rbbA and yhjD deletion mutants revealed aggravating genetic interactions with mutants deficient in the electron transport chain. Cells lacking both rbbA and yhjD exhibited reduced cell division, respiration and global protein synthesis as well as increased sensitivity to antibiotics targeting the ETC and the accuracy of protein synthesis. Our results suggest that RbbA appears to function together with YhjD as part of a regulatory network that impacts bacterial oxidative phosphorylation and translation efficiency

Possible involvement of caveolin in attenuation of cardioprotective effect of ischemic preconditioning in diabetic rat heart

<p>Abstract</p> <p>Background</p> <p>Nitric oxide (NO) has been noted to produce ischemic preconditioning (IPC)-mediated cardioprotection. Caveolin is a negative regulator of NO, which inhibits endothelial nitric oxide synthase (eNOS) by making caveolin-eNOS complex. The expression of caveolin is increased during diabetes mellitus (DM). The present study was designed to investigate the involvement of caveolin in attenuation of the cardioprotective effect of IPC during DM in rat.</p> <p>Methods</p> <p>Experimental DM was induced by single dose of streptozotocin (50 mg/Kg, <it>i.p</it>,) and animals were used for experiments four weeks later. Isolated heart was mounted on Langendorff's apparatus, and was subjected to 30 min of global ischemia and 120 min of reperfusion. IPC was given by four cycles of 5 min of ischemia and 5 min of reperfusion with Kreb's-Henseleit solution (K-H). Extent of injury was measured in terms of infarct size by triphenyltetrazolium chloride (TTC) staining, and release of lactate dehydrogenase (LDH) and creatin kinase-MB (CK-MB) in coronary effluent. The cardiac release of NO was noted by measuring the level of nitrite in coronary effluent.</p> <p>Results</p> <p>IPC- induced cardioprotection and release of NO was significantly decreased in diabetic rat heart. Pre-treatment of diabetic rat with daidzein (DDZ) a caveolin inhibitor (0.2 mg/Kg/s.c), for one week, significantly increased the release of NO and restored the attenuated cardioprotective effect of IPC. Also perfusion of sodium nitrite (10 μM/L), a precursor of NO, significantly restored the lost effect of IPC, similar to daidzein in diabetic rat. Administration of 5-hydroxy deaconate (5-HD), a mito K_ATPchannel blocker, significantly abolished the observed IPC-induced cardioprotection in normal rat or daidzein and sodium nitrite perfused diabetic rat heart alone or in combination.</p> <p>Conclusions</p> <p>Thus, it is suggested that attenuation of the cardioprotection in diabetic heart may be due to decrease the IPC mediated release of NO in the diabetic myocardium, which may be due to up -regulation of caveolin and subsequently decreased activity of eNOS.</p