Chromatographic resolution of nucleic acids extracted from Penicillium chrysogenum

Summary High molecular weight DNA extracted from Penicillium chrysogenum has been fractionated using RPC-5 Analog, into three distinct types designated 1, 2 and 3. Types 1 and 2 have the same buoyant density of 1.710 g/cm³ and together appear to comprise the nuclear DNA. Type 1 is enriched for repeated sequences which are normally observed in restriction digests of P. chrysogenum total DNA. Conversely, type 2 appears to be composed entirely of non-repetitive sequences. Type 3 has been identified as mitochondrial DNA, having a buoyant density of 1.695 g/cm³ and an estimated molecular weight of 31.6×10⁶ Daltons.     

Heterologous expression of endo-beta-1,4-D-glucanase from Clostridium cellulovorans in Clostridium acetobutylicum ATCC 824 following transformation of the engB gene.

Heterologous expression of the Clostridium cellulovorans engB gene by Clostridium acetobutylicum BKW-1 was detected as zones of hydrolysis on carboxymethyl cellulose (CMC) Trypticase glucose yeast plates stained with Congo red. The extracellular cellulase preparation from C. acetobutylicum BKW-1 has a specific activity towards CMC which is more than fourfold that present in C. acetobutylicum ATCC 824. Western blot (immunoblot) analysis using the C. cellulovorans anti-EngB primary antibody demonstrated that an additional 44-kDa protein band was present in the supernatant derived from C. acetobutylicum BKW-1 but was not present in ATCC 824 or ATCC 824(pMTL500E).     

“Unblocking” Serum Activity <Emphasis Type="Italic">in vitro</Emphasis> in the Polyoma System may Correlate with Antitumour Effects of Antiserum <Emphasis Type="Italic">in vivo</Emphasis>

In a variety of tumour systems, individuals carrying progressively growing neoplasms have lymphoid cells with a specific cytotoxic effect on cultured tumour cells from the same individual^1–4. Since the sera of tumour-bearing individuals have been shown to prevent tumour cell destruction by immune lymphocytes in vitro^2,5–8 and since this serum blocking activity appears early in primary and transplant tumour development^5,7, it has been suggested that the appearance of this serum blocking activity might be responsible for the progressive growth of tumours in individuals having cytotoxic lymphocytes. Counteraction of this blocking activity would thus be of primary importance in facilitating the function of an already existing or bolstered cell-mediated immunity. The serum blocking activity might be inhibited in various ways, by preventing the formation of blocking antibody or by interfering with its action (“unblocking”), as demonstrated in Moloney sarcoma regressor sera⁹. This type of serum also has a therapeutic effect on Moloney sarcomas in vivo^10,11, which has been tentatively attributed to its unblocking activity^8,9 or, possibly, to a complement-dependent cytotoxicity¹⁰. Tumour growth in the Moloney sarcoma system, however, might be due in part to continuous recruitment of neoplastic cells by virus-induced transformation and so the therapeutic effect could be due to a virus-neutralizing serum activity^9,10.     

Microbial transformation of 1,6-dichloro-1,6-dideoxy-β,D-fructofuranosyl-4-chloro-4-deoxy-α,D-galactopyranoside (TGS) by soil populations

Summary Soil microorganisms from one site were shown to be consistently capable of the transformation of 1,6-dichloro-1,6-dideoxy-,d-fructofuranosyl-4-chloro-4-deoxy-,d-galactopyranoside (TGS) in laboratory batch cultures. With fresh soils, all of the available chloride ions were released from the molecule. Subcultures of a TGS-dehalogenating bacterial community produced a progressive decline in the dehalogenating capabilities towards the substrate. The soil organisms did not utilise TGS as a carbon source. The transformation was achieved by co-metabolism and was probably supported by an unknown component in the soil. Four bacterial species were isolated from the TGS-dehalogenating soil community: twoBacillus species, anAcinetobacter group isolate and aMicrococcus group isolate. Thin-layer chromatography confirmed the disappearance of the chlorosugar and high-performance liquid chromatography demonstrated that neither of the constituent monosaccharides—1,6-dichlorofructose nor 4-chlorogalactosucrose was accumulated as an intermediate.

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Resumen Microorganismos de suelo de cierto lugar demostraron consistemente ser capaces de realizar la transformación de 1,6-dicloro-1,6 dideoxi--D-fructofuranosil-4-cloro-4-deoxi-,D-alactopiranosa (TGS) in culturas de laboratorio de tipo discontinuo. Con muestras frescas de suelo, todos los iones cloruro fueron liberados de la molecula. Subculturas de una comunidad bacterial capaz de dehalogenizar TGS produjeron una declinación progresiva de la capacidad de dehalogenizar el substrato. Los microorganismos no utilizaron el TGS como fuente de carbono. La transformación se realiza por co-metabolismo y probablemente se base en un componente del suelo, no determinado. Cuatro especies bacteriales fueron aisladas de la comunidad de bacterias de suelo con capacidad de dehalogenar el TGS: dos especies deBacilo, unaAcinelobacteria y unMicrococo. Estudios de cromatografía de capa delgada confirmaron la desaparición del clorosacárido, y estadios de cromatografía liquida demostraron que ninguno de los componentes monoscáridos — 1,6-diclorofructuosa y 4-clorogalactosucrosa — eran acumulados como productors intermedios.

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Résumé Les microorganismes du sol d'un certain endroit ont été démontrés être capable, sans exception, de la transformation de 1,6-dichloro-1,6-dideoxy-,D-fructofuranosyl-4-chloro-4-deoxy-,D-galactopyranoside (TGS) en cultures de laboratoire du type discontinu. Avec des prélèvements frais du sol, tous les ions disponibles de chlorure ont été libérés de la molécule. Des souscultures d'une communauté bactérienne capable de déhalogeniser le TGS ont produit un déclin progressif de la capacité de déhalogeniser le substrat. Les microorganismes du sol n'ont pas utilisé le TGS comme source de carbone. La transformation s'est accomplie par cometabolisme et, probablement, s'est basée sur un component indéterminé du sol. Quatre espèces bactériennes ont été isolées de la communauté de bactéries du sol capable de déhalogeniser le TGS: deux espèces deBacillus, unAcinetobacter et unMicrococcus. Des études de chromatographie de couches fines ont confirmées la disparition du chlorosaccharide, tandis que des études de chromatographie liquide de haut rendement ont démontrées que, des monosaccharides constituants, ni 1,6-dichlorofructose ni 4-chlorogalactosucrose, n'ont été accumulés comme produits intermédiaires.

     

In situ quantification of aberrant p53 in colorectal neoplasia

Aberrant p53 protein accumulation was measured immunohistologically in 342 colorectal paraffin-embedded tissue sections from 115 patients (24 with adenocarcinoma, 59 with adenoma and 32 'hospital controls'). Subjective scoring was compared with quantitative cell imaging, including dichotomous (p53⁺/p53^-) status, ng p53^mut mg^-1 enterocyte protein, and tumour burden and patient body 'burden' of aberrant p53. A total of 62.5% cancer patients, 23.7% adenoma patients and 3.1% hospital controls were accorded p53⁺ status on the basis of p53 quantification. Quantitative p53⁺/p53^- assignment had a stronger inverse association with survival (χ²=6.17, p=0.013, Kaplan-Meier test) than subjective 'visual estimation' (χ²=0.57, p=0.449). There was a strong inverse relationship between the p53 'body burden' and the months of post-diagnosis survival (hazard ratio=1.42, p=0.0004, Cox proportional hazards). Absolute quantification for inactivated p53 permits objective and reproducible scoring, adjusts for intra-laboratory immunostaining 'batch effects', corrects for fixation artefacts, and standardizes for inter-laboratory differences in fixation, antibody selection and staining method. Clinically, in situ quantification of p53 will permit more accurate survival prognoses and will inform therapy selection and dose. Ultimately, accurate quantitative tissue/blood p53 correlations may provide a minimally invasive and systemic surrogate measure for these same clinical purposes.     

Do energetic demands constrain incubation scheduling in a biparental species?

The high energetic demands of incubation in birds may be animportant ecological factor limiting the evolution of life-historytraits, such as clutch size. In biparental species, however,the demands of incubation may not be a major constraint becausethere may always be sufficient feeding time available for theoff-duty bird to regain energy used during an incubation bout.We investigated whether the energetic demands of incubationconstrain optimum incubation bout length in a biparental incubatorby decreasing the energetic demands of incubation. We put aninsulated cup around the lining of semipalmated sandpiper nestsso that the rate of cooling of eggs was reduced by 21%. Semipalmatedsandpipers responded by increasing their mean incubation boutlength of around 11.1 h by about 10%. Bout lengths in unmanipulatednatural nests became longer as hatch approached (incubationstage), and this was independent of weather. Bout lengths mayhave decreased with increasing rainfall and were independentof time of day. The results suggest that bout length in semipalmatedsandpipers is constrained by their cumulative energetic expenditureduring an incubation bout, and this is determined partly bythe high costs of steady-state incubation. The results alsosuggest that the incubating bird determines the bout lengthrather than the returning bird. Semipalmated sandpipersmay havemaximized incubation bout length to minimize changeovers duringincubation because these probably increasepredation risk. Selectionto minimize the frequency of changeover may then be a factorcontributing to the evolution ofbiparental care and life-historytraits in semipalmated sandpipers.     

Human bronchial epithelium controls TH2 responses by TH1-induced, nitric oxide-mediated STAT5 dephosphorylation: implications for the pathogenesis of asthma

Increased levels of NO in exhaled air in association with increased NO synthetase (NOS)2 expression in bronchial epithelial are hallmark features of asthma. It has been suggested that NO contributes to asthma pathogenesis by selective down-regulation of TH1 responses. We demonstrate, however, that NO can reversibly limit in vitro expansion of both human TH1 and TH2 CD4+ T cells. Mechanistically, NO induces cGMP-mediated reversible STAT5 dephosphorylation and therefore interferes with the IL-2R activation cascade. Human bronchial epithelial cells (HBEC) up-regulate NOS2 after stimulation with IFN-gamma secreted by TH1 CD4+ T cells and release NO, which inhibits both TH1 and TH2 cell proliferation. This reversible T cell growth arrest depends on NO because T cell proliferation is completely restored after in vitro blocking of NOS2 on HBEC. HBEC thus drive the effector end of a TH1-controlled feedback loop, which protects airway mucosal tissues at the potential lesional site in asthma from overwhelming CD4+ TH2 (and potentially TH1) responses following allergen exposure. Variations in the efficiency of this feedback loop provides a plausible mechanism to explain why only a subset of atopics sensitized to ubiquitous aeroallergens progress to expression of clinically relevant levels of airways inflammation.     

Androgen metabolism and JAK/STAT pathway genes and prostate cancer risk

Background: Prostate cancer (PC) is the most frequently diagnosed solid tumor in U.S. men. Genome-wide association studies (GWAS) have identified over 40 risk-associated single nucleotide polymorphisms (SNPs), including variants in androgen pathway genes (e.g., KLK3 and AR). Androgens are important in PC and genes involved in this pathway are therefore candidates for conferring susceptibility to PC. Methods: In this hypothesis-testing study, we evaluated PC risk in association with SNPs in 22 candidate genes involved in androgen metabolism or interactions with the androgen receptor (AR). A total of 187 SNPs were genotyped in 1458 cases and 1351 age-matched controls from a population-based study. PC risk was estimated using adjusted unconditional logistic regression and multinomial regression models. Results: Single SNP analyses showed evidence (p < 0.05) for associations with 14 SNPs in 9 genes: NKX3.1, HSD17B3, AKR1C3, SULT2A1, CYP17A1, KLK3, JAK2, NCOA4 and STAT3. The most significant result was observed for rs2253502 in HSD17B3 (odds ratio, OR = 0.57, 95% CI: 0.39–0.84). In addition, five SNPs in four genes (CYP17A1, HSD17B4, NCOA4, and SULT2A1) were associated with more aggressive disease (p < 0.01). Conclusions: Our results replicate previously reported associations for SNPs in CYP17A1, HSD17B3, ARK1C3, NKX3.1, NCOA4 and KLK3. In addition, novel associations were observed for SNPs in JAK2, HSD17B4, and SULT2A1. These results will require replication in larger studies.     

Carbonized Chicken Eggshell Membranes with 3D Architectures as High‐Performance Electrode Materials for Supercapacitors

Supercapacitor electrode materials are synthesized by carbonizing a common livestock biowaste in the form of chicken eggshell membranes. The carbonized eggshell membrane (CESM) is a three‐dimensional macroporous carbon film composed of interwoven connected carbon fibers containing around 10 wt% oxygen and 8 wt% nitrogen. Despite a relatively low surface area of 221 m² g^?1, exceptional specific capacitances of 297 F g^?1 and 284 F g^?1 are achieved in basic and acidic electrolytes, respectively, in a 3‐electrode system. Furthermore, the electrodes demonstrate excellent cycling stability: only 3% capacitance fading is observed after 10 000 cycles at a current density of 4 A g^?1. These very attractive electrochemical properties are discussed in the context of the unique structure and chemistry of the material.