Formation and electronic structure of an atypical Cu A site

PmoD, a recently discovered protein from methane-oxidizing bacteria, forms a homodimer with a dicopper CuA center at the dimer interface. Although the optical and electron paramagnetic resonance (EPR) spectroscopic signatures of the PmoD CuA bear similarities to those of canonical CuA sites, there are also some puzzling differences. Here we have characterized the rapid formation (seconds) and slow decay (hours) of this homodimeric CuA site to two mononuclear Cu2+ sites, as well as its electronic and geometric structure, using stopped-flow optical and advanced paramagnetic resonance spectroscopies. PmoD CuA formation occurs rapidly and involves a short-lived intermediate with a max of 360 nm. Unlike other CuA sites, the PmoD CuA is unstable, decaying to two type 2 Cu2+ centers. Surprisingly, NMR data indicate that the PmoD CuA has a pure σu∗ ground state rather than the typical equilibrium between σu∗ and πu of all other CuA proteins. EPR, ENDOR, ESEEM, and HYSCORE data indicate the presence of two histidine and two cysteine ligands coordinating the CuA core in a highly symmetrical fashion. This report significantly expands the diversity and understanding of known CuA sites.Fil: Ross, Matthew O.. Northwestern University; Estados UnidosFil: Fisher, Oriana S.. Northwestern University; Estados UnidosFil: Morgada, Marcos Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Krzyaniak, Matthew D.. Northwestern University; Estados UnidosFil: Wasielewski, Michael R.. Northwestern University; Estados UnidosFil: Vila, Alejandro Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Hoffman, Brian M.. Northwestern University; Estados UnidosFil: Rosenzweig, Amy C.. Northwestern University; Estados Unido

Characterisation of new animal cell cultures’ sensitivity to <i>Coxsackievirus B5</i> and <i>Herpes simplex virus‑1</i>

The increase in the number of cell cultures for virology and biotechnology enhances the chances of a successful response to threats related to outbreaks of well-known and new human infectious diseases. It is a vital task to search for cell cultures sensitive to a wide spectrum of viruses.The aim of the study was to investigate the sensitivity of new diploid animal cell cultures (fibroblasts of a foetal pig’s kidneys and larynx) to Coxsackievirus B5 (CVB5) and Herpes simplex virus-1 (HSV-1).Materials and methods. The cultures of porcine foetal kidney fibroblasts (PFKF) and porcine foetal larynx fibroblasts (PFLF) were derived from a foetus of a healthy pig by mild trypsinisation. The study determined the sensitivity of these new PFKF and PFLF cultures to the above-mentioned viruses by the cytopathic effect (CPE) expressed as a percentage. The infectious activity of CVB5 was studied using real-time polymerase chain reaction (PCR) with the determination of amplification cycle threshold values (Ct); that of HSV-1 was studied using quantitative titration of the virus-containing liquid (VCL). Infectious activity values were expressed as tissue culture 50% infective doses (TCID50).Results. The authors developed diploid PFKF and PFLF cell cultures. PFKF cells demonstrated high sensitivity to CVB5, with a CPE of 87.5±3.3% after passage 3 and a satisfactory concentration of enterovirus RNA in the VCL of 22–24 Ct . The sensitivity of PFKF cells to HSV-1 corresponded to a CPE of 92.1±5.5%. In these cells, the infectious activity of HSV-1 corresponded to 104.25 TCID50/0.2 mL. The experiments with PFLF cells showed low CPE and infectious activity values for both viruses.Conclusions. The study demonstrated high CPE values with the CVB5 (CB5-8100) and HSV-1 (HSV-1/L-2) strains as examples and confirmed the sensitivity of the new diploid PFKF cell culture to these test viruses. Thus, the PFKF cell culture offers potential applications in virology and biotechnology and may be a candidate for testing other strains of CVB5 and HSV-1

Charge Delocalization in Self-Assembled Mixed-Valence Aromatic Cation Radicals

The spontaneous assembly of aromatic cation radicals (D+•) with their neutral counterpart (D) affords dimer cation radicals (D2+•). The intermolecular dimeric cation radicals are readily characterized by the appearance of an intervalence charge-resonance transition in the NIR region of their electronic spectra and by ESR spectroscopy. The X-ray crystal structure analysis and DFT calculations of a representative dimer cation radical (i.e., the octamethylbiphenylene dimer cation radical) have established that a hole (or single positive charge) is completely delocalized over both aromatic moieties. The energetics and the geometrical considerations for the formation of dimer cation radicals is deliberated with the aid of a series of cyclophane-like bichromophoric donors with drastically varied interplanar angles between the cofacially arranged aryl moieties. X-ray crystallography of a number of mixed-valence cation radicals derived from monochromophoric benzenoid donors established that they generally assemble in 1D stacks in the solid state. However, the use of polychromophoric intervalence cation radicals, where a single charge is effectively delocalized among all of the chromophores, can lead to higher-order assemblies with potential applications in long-range charge transport. As a proof of concept, we show that a single charge in the cation radical of a triptycene derivative is evenly distributed on all three benzenoid rings and this triptycene cation radical forms a 2D electronically coupled assembly, as established by X-ray crystallography

Review of a series of clinical observations of the use of "Cell culture for substitution therapy"

The aim of the study was to summarize scientific literature data on the clinical effectiveness of cell-based medical biotechnologies for regenerative medicine.Цель исследования – обобщить данные научной литературы о клинической эффективности клеточных медицинских биотехнологий для регенеративной медицины

Характеристика чувствительности новых клеточных культур животного происхождения к вирусам Coxsackievirus B5 и Herpes simplex virus‑1

The increase in the number of cell cultures for virology and biotechnology enhances the chances of a successful response to threats related to outbreaks of well-known and new human infectious diseases. It is a vital task to search for cell cultures sensitive to a wide spectrum of viruses.The aim of the study was to investigate the sensitivity of new diploid animal cell cultures (fibroblasts of a foetal pig’s kidneys and larynx) to Coxsackievirus B5 (CVB5) and Herpes simplex virus-1 (HSV-1).Materials and methods. The cultures of porcine foetal kidney fibroblasts (PFKF) and porcine foetal larynx fibroblasts (PFLF) were derived from a foetus of a healthy pig by mild trypsinisation. The study determined the sensitivity of these new PFKF and PFLF cultures to the above-mentioned viruses by the cytopathic effect (CPE) expressed as a percentage. The infectious activity of CVB5 was studied using real-time polymerase chain reaction (PCR) with the determination of amplification cycle threshold values (Ct); that of HSV-1 was studied using quantitative titration of the virus-containing liquid (VCL). Infectious activity values were expressed as tissue culture 50% infective doses (TCID50).Results. The authors developed diploid PFKF and PFLF cell cultures. PFKF cells demonstrated high sensitivity to CVB5, with a CPE of 87.5±3.3% after passage 3 and a satisfactory concentration of enterovirus RNA in the VCL of 22–24 Ct . The sensitivity of PFKF cells to HSV-1 corresponded to a CPE of 92.1±5.5%. In these cells, the infectious activity of HSV-1 corresponded to 104.25 TCID50/0.2 mL. The experiments with PFLF cells showed low CPE and infectious activity values for both viruses.Conclusions. The study demonstrated high CPE values with the CVB5 (CB5-8100) and HSV-1 (HSV-1/L-2) strains as examples and confirmed the sensitivity of the new diploid PFKF cell culture to these test viruses. Thus, the PFKF cell culture offers potential applications in virology and biotechnology and may be a candidate for testing other strains of CVB5 and HSV-1.Расширение номенклатуры клеточных культур для вирусологии и биотехнологии повышает вероятность успешного реагирования на угрозы, связанные со вспышками известных и новых инфекционных заболеваний человека. Поиск восприимчивых к широкому спектру вирусов клеточных культур является актуальной задачей.Цель работы: изучить чувствительность новых диплоидных клеточных культур животного происхождения (фибробласты почки и гортани плода свиньи) к вирусам Coxsackievirus B5 (CVB5) и Herpes simplex virus-1 (HSV-1).Материалы и методы: клеточные культуры фибробластов почки и гортани плода здоровой свиноматки получены методом щадящей трипсинизации. Чувствительность новых клеточных культур фибробластов почки и гортани плода свиньи (ФППС и ФГПС) к указанным вирусам определяли по степени цитопатического действия (ЦПД), выраженной в процентном соотношении. Изучение инфекционной активности вируса CVB5 проводили методом ПЦР в режиме реального времени с оценкой относительной величины порогового цикла амплификации (Ct); HSV-1 — количественным титрованием вируссодержащей жидкости (ВСЖ), значение показателя выражали в 50% тканевой цитопатической дозе (ТЦД50).Результаты: получены диплоидные клеточные культуры ФППС и ФГПС. Выявлены высокая чувствительность клеток ФППС к вирусу CVB5 с ЦПД 87,5±3,3% на 3 пассаже и удовлетворительная концентрация энтеровирусной РНК в ВСЖ, характеризующаяся значением порогового цикла на уровне 22–24 Ct. Чувствительность клеточной культуры ФППС к HSV-1 соответствовала 92,1±5,5% ЦПД, инфекционная активность — 104,25 ТЦД50/0,2 мл. У клеток ФГПС к изучаемым вирусам определены низкие показатели ЦПД и инфекционной активности.Выводы: новая диплоидная клеточная культура ФППС с подтвержденной чувствительностью к тестируемым вирусам (CVB5 штамма CB5-8100 и HSV-1 штамма HSV-1/L-2) с высоким уровнем ЦПД имеет перспективы использования в вирусологии и биотехнологии. Клеточная культура ФГПС может быть кандидатом для тестирования других представителей CVB5 и HSV-1

Mesomorphism and Photophysics of Some Metallomesogens Based on Hexasubstituted 2,2':6', 2''-Terpyridines : 6', 2''-Terpyridines

The luminescent and mesomorphic properties of a series of metal complexes based on hexacatenar 2,2':6',2''-terpyridines are investigated using experimental methods and density functional theory (DFT). Two types of ligand are examined, namely 5,5''-di(3,4,5-trialkoxyphenyl)terpyridine with or without a fused cyclopentene ring on each pyridine and their complexes were prepared with the following transition metals: ZnII, CoIII, RhIII, IrIII, EuIII and DyIII. The exact geometry of some of these complexes was determined by single X-ray diffraction. All complexes with long alkyl chains were found to be liquid crystalline, which property was induced on complexation. The liquid-crystalline behaviour of the complexes was studied by polarising optical microscopy and small-angle X-ray diffraction. Some of the transition metal complexes (for example, those with ZnII and IrIII) are luminescent in solution, the solid state and the mesophase; their photophysical properties were studied both experimentally and using DFT methods (M06-2X and B3LYP)

Identification of differentially expressed microRNAs in human male breast cancer

<p>Abstract</p> <p>Background</p> <p>The discovery of small non-coding RNAs and the subsequent analysis of microRNA expression patterns in human cancer specimens have provided completely new insights into cancer biology. Genetic and epigenetic data indicate oncogenic or tumor suppressor function of these pleiotropic regulators. Therefore, many studies analyzed the expression and function of microRNA in human breast cancer, the most frequent malignancy in females. However, nothing is known so far about microRNA expression in male breast cancer, accounting for approximately 1% of all breast cancer cases.</p> <p>Methods</p> <p>The expression of 319 microRNAs was analyzed in 9 primary human male breast tumors and in epithelial cells from 15 male gynecomastia specimens using fluorescence-labeled bead technology. For identification of differentially expressed microRNAs data were analyzed by cluster analysis and selected statistical methods.</p> <p>Expression levels were validated for the most up- or down-regulated microRNAs in this training cohort using real-time PCR methodology as well as in an independent test cohort comprising 12 cases of human male breast cancer.</p> <p>Results</p> <p>Unsupervised cluster analysis separated very well male breast cancer samples and control specimens according to their microRNA expression pattern indicating cancer-specific alterations of microRNA expression in human male breast cancer. miR-21, miR519d, miR-183, miR-197, and miR-493-5p were identified as most prominently up-regulated, miR-145 and miR-497 as most prominently down-regulated in male breast cancer.</p> <p>Conclusions</p> <p>Male breast cancer displays several differentially expressed microRNAs. Not all of them are shared with breast cancer biopsies from female patients indicating male breast cancer specific alterations of microRNA expression.</p