Scalable production in human cells and biochemical characterization of full-length normal and mutant huntingtin.

Directory of Open Access Journals (Sweden)

Bin Huang

Full Text Available Huntingtin (Htt is a 350 kD intracellular protein, ubiquitously expressed and mainly localized in the cytoplasm. Huntington's disease (HD is caused by a CAG triplet amplification in exon 1 of the corresponding gene resulting in a polyglutamine (polyQ expansion at the N-terminus of Htt. Production of full-length Htt has been difficult in the past and so far a scalable system or process has not been established for recombinant production of Htt in human cells. The ability to produce Htt in milligram quantities would be a prerequisite for many biochemical and biophysical studies aiming in a better understanding of Htt function under physiological conditions and in case of mutation and disease. For scalable production of full-length normal (17Q and mutant (46Q and 128Q Htt we have established two different systems, the first based on doxycycline-inducible Htt expression in stable cell lines, the second on "gutless" adenovirus mediated gene transfer. Purified material has then been used for biochemical characterization of full-length Htt. Posttranslational modifications (PTMs were determined and several new phosphorylation sites were identified. Nearly all PTMs in full-length Htt localized to areas outside of predicted alpha-solenoid protein regions. In all detected N-terminal peptides methionine as the first amino acid was missing and the second, alanine, was found to be acetylated. Differences in secondary structure between normal and mutant Htt, a helix-rich protein, were not observed in our study. Purified Htt tends to form dimers and higher order oligomers, thus resembling the situation observed with N-terminal fragments, although the mechanism of oligomer formation may be different.

Biochemical Characterization of an In-House Coccidioides Antigen: Perspectives for the Immunodiagnosis of Coccidioidomycosis

OpenAIRE

Renato Evando Moreira Filho; Silviane Praciano Bandeira; Raimunda Sâmia Nogueira Brillhante; Marcos Fábio Gadelha Rocha; Ilka Maria Vasconcelos; Mirella Leite Pereira; Débora de Souza Collares Maia Castelo-Branco; Francisco Airton Castro Rocha; Zoilo Pires de Camargo; Marcio Viana Ramos; Rossana de Aguiar Cordeiro; José Júlio Costa Sidrim

2012-01-01

The objective of this study was to evaluate the reactivity of an in-house antigen, extracted from a strain of C. posadasii isolated in northeastern Brazil, by radial immunodiffusion and Western blotting, as well as to establish its biochemical characterization. The protein antigen was initially extracted with the use of solid ammonium sulfate and characterized by 1-D electrophoresis. Subsequently, it was tested by means of double radial immunodiffusion and Western blottin...

Morphological and biochemical characterization of xanthomonas campestris (pammel) dawson pv. Sesame and it's management by bacterial antagonists

International Nuclear Information System (INIS)

Naqvi, S.F.; Haq, M.I.U.; Tahir, M.I.; Khan, M.A.; Ali, Z.

2013-01-01

Isolates of Xanthomonas campestris (Pammel) Dawson pv. sesami (Xcs) were collected from various locations in Pakistan and were characterized using several biochemical tests including; (KOH) string test, gram staining, H/sub 2/S production, catalase activity, oxidation/fermentation of glucose, oxidase activity and nitrate reduction. The isolates were positive in tests for KOH string assay, H/sub 2/S, catalase and showed negative reactions in gram staining, oxidase test and nitrate reduction while oxidative utilization of glucose was observed. To develop biological control for the pathogen we isolated numerous bacteria from the rhizosphere of sesame plants and screened them for antagonism against the pathogen using the zone inhibition method. Out of 87 isolates, 9 isolates were inhibitory to the pathogen in vitro. Pseudomonas fluorescens ID-3 exhibited the highest zone of inhibition i.e. 7.97mm. All antagonistic isolates were tested for disease control under greenhouse conditions. Four sesame lines that were found to be moderately resistant in previous studies (i.e. 95001, 96007, 96019, and 20003) were selected for evaluating biocontrol efficacy. The isolates were found to control the disease and reduced severity to 3% compared to untreated plants. The isolate P. fluorescens ID-3 was considered to provide the best disease control with each cultivar. Average biocontrol efficiency (BE) of ID-3 was 78.25% while the best BE was obtained in line 96019, i.e. 81%. (author)

Isolation, molecular and biochemical characterization of oil degrading bacteria from contaminated soil at an oil refinery

International Nuclear Information System (INIS)

AL-Deeb, T.M.; Malkawi, H.I.

2009-01-01

Biodegradation using microorganisms is considered to be cost-effective and environmentally friendly treatment of oil-contaminated sites. Oil-biodegrading bacterial strains were isolated, identified and characterized from oil contaminated soil samples at oil refinery in Zarqa (Jordan). Thirty four bacterial isolates were grown on mineral salt media supplemented with crude oil, but 16 showed positive biodegradation of diesel. All the 34 bacterial isolates were characterized at the molecular and bio-chemical levels, and showed positive polymerase chain reaction (PCR) amplification product size of 1500 bp when 16s rDNA bacterial universal primers were used. Eighteen bacterial isolates showed positive PCR amplification product size of 150 bp specific for the genus Pseudomonas and 3 bacterial isolates showed positive amplification product size of 1500 bp specific for the genus Acinetobacter. Biochemical and physiological characterization performed on the 34 bacterial isolates revealed the presence of oil biodegrading bacterial genera and species of Pseudomonas Acidovorans, P. aeruginosa, P. vesicularis, Acinetobacter calcoaceticus, Ac. lowffii, Micro-ococcus luteus, M. varians, M. lylae, M. roseus, Alcaligenes denitrificians, Bacillus megaterium, Comamonas sp., Moralxella sp., Bordetella sp., P. putida, P. stutzeri and P. mallei. (au)

Biochemical characterization of Tunisian grapevine varieties

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Ferjani Ben Abdallah

1998-03-01

The study of GPI, PGM, AAT and peroxydase isozyme banding patterns in combination with berry colour has led to establish a classification of the 61 autochton varieties into 37 groups including 26 varieties definitely differentiated through the results of this biochemical study.

Characterization Plan for Soils Around Drain Line PLA-100115

Energy Technology Data Exchange (ETDEWEB)

D. Shanklin

2006-05-24

This Characterization Plan supports the Hazardous Waste Management Act/Resource Conservation and Recovery Act (HWMA/RCRA) closure of soils that may have been contaminated by releases from drain line PLA-100115, located within the Idaho Nuclear Technology and Engineering Center at the Idaho National Laboratory Site. The requirements to address the closure of soils contaminated by a potential release from this line in a characterization plan was identified in the "HWMA/RCRA Less Than 90-day Generator Closure Report for the VES-SFE-126."

Identification and characterization of mobile genetic elements LINEs from Brassica genome.

Science.gov (United States)

Nouroz, Faisal; Noreen, Shumaila; Khan, Muhammad Fiaz; Ahmed, Shehzad; Heslop-Harrison, J S Pat

2017-09-05

Among transposable elements (TEs), the LTR retrotransposons are abundant followed by non-LTR retrotransposons in plant genomes, the lateral being represented by LINEs and SINEs. Computational and molecular approaches were used for the characterization of Brassica LINEs, their diversity and phylogenetic relationships. Four autonomous and four non-autonomous LINE families were identified and characterized from Brassica. Most of the autonomous LINEs displayed two open reading frames, ORF1 and ORF2, where ORF1 is a gag protein domain, while ORF2 encodes endonuclease (EN) and a reverse transcriptase (RT). Three of four families encoded an additional RNase H (RH) domain in pol gene common to 'R' and 'I' type of LINEs. The PCR analyses based on LINEs RT fragments indicate their high diversity and widespread occurrence in tested 40 Brassica cultivars. Database searches revealed the homology in LINE sequences in closely related genera Arabidopsis indicating their origin from common ancestors predating their separation. The alignment of 58 LINEs RT sequences from Brassica, Arabidopsis and other plants depicted 4 conserved domains (domain II-V) showing similarity to previously detected domains. Based on RT alignment of Brassica and 3 known LINEs from monocots, Brassicaceae LINEs clustered in separate clade, further resolving 4 Brassica-Arabidopsis specific families in 2 sub-clades. High similarities were observed in RT sequences in the members of same family, while low homology was detected in members across the families. The investigation led to the characterization of Brassica specific LINE families and their diversity across Brassica species and their cultivars. Copyright © 2017 Elsevier B.V. All rights reserved.

Biochemical and histological characterization of Agave fourcroydes Lem. plants in vitro acclimatized

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Yunel Pérez Hernández

2016-10-01

Full Text Available Henequen (Agave fourcroydes Lem. is a representative crop of the province of Matanzas, Cuba, which has been exploited extensively for the quality of its fibers. The present work aimed to characterize from the biochemical and histological point of view henequen plants cv. 'Sac Ki' in vitro acclimatized. The biochemical indicators analyzed were concentration of soluble phenols, total soluble carbohydrates, reducing sugars, soluble proteins and total chlorophylls, as well as the relative contents of flavonoids, terpins and anthocyanins. In addition, a histological analysis of the epidermis was performed and the results were compared with plants of one month in rooting culture medium. In vitro acclimatized plants were subsequently transferred to the ex vitro conditions. The contents of total soluble carbohydrates, reducing sugars and chlorophylls decreased, whereas the levels of soluble phenols, proteins and secondary metabolites were higher in relation to in vitro plants rooted for 30 days. The epidermis of the in vitro acclimatized plants showed a greater development of the stomatal apparatus that could positively affect the ex vitro acclimatization, where a survival rate of 87% and a reduction of the acclimatization time were obtained.   Keywords: chlorophyll, henequen, phenol, reducing sugar

Molecular cloning and biochemical characterization of an α-amylase family from Aspergillus niger

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Junying Wang

2018-03-01

Full Text Available Background: α-Amylase is widely used in the starch processing, food and paper industries, hydrolyzing starch, glycogen and other polysaccharides into glucose, maltose and oligosaccharides. An α-amylase gene family from Aspergillus niger CBS513.88 encode eight putative α-amylases. The differences and similarities, biochemical properties and functional diversity among these eight α-amylases remain unknown. Results: The eight genes were cloned and expressed in Pichia pastoris GS115 by shaking-flask fermentation under the induction of methanol. The sequence alignment, biochemical characterizations and product analysis of starch hydrolysis by these α-amylases were investigated. It is found that the eight α-amylases belonged to three different groups with the typical structure of fungal α-amylase. They exhibited maximal activities at 30–40°C except AmyG and were all stable at acidic pH. Ca2+ and EDTA had no effects on the activities of α-amylases except AmyF and AmyH, indicating that the six amylases were Ca2+ independent. Two novel α-amylases of AmyE and AmyF were found. AmyE hydrolyzed starch into maltose, maltotriose and a small amount of glucose, while AmyF hydrolyzed starch into mainly glucose. The excellent physical and chemical properties including high acidic stability, Ca2+-independent and high maltotriose-forming capacity make AmyE suitable in food and sugar syrup industries. Conclusions: This study illustrates that a gene family can encode multiple enzymes members having remarkable differences in biochemical properties. It provides not only new insights into evolution and functional divergence among different members of an α-amylase family, but the development of new enzymes for industrial application. Keywords: Biochemical properties, Food industry, Fungal α-amylase, Glycosyl hydrolase family, Glycosyl hydrolase family, Industrial application, Paper industry, Recombinant Pichia pastoris, Starch processing, α-amylase cloning

The application of HPLC with on-line coupled UV/MS-biochemical detection for isolation of an acetylcholinesterase inhibitor from Narcissus 'Sir Winston Churchill'

NARCIS (Netherlands)

Ingkaninan, K.; Hazekamp, A.; de Best, C.M.; Irth, H.; Tjaden, U.R.; van der Heijden, R.; van der Greef, J.; Verpoorte, R.

2000-01-01

An HPLC with on-line coupled UV/MS-biochemical detection method for acetylcholinesterase (AChE) inhibitors in natural sources has been developed. The potential of this method is shown by the isolation of a new AChE inhibitor from the alcoholic extract of Narcissus 'Sir Winston Churchill'. Combining

Biochemical and biological characterization of Bothriechis schlegelii snake venoms from Colombia and Costa Rica

OpenAIRE

Prezotto-Neto, Jos�� P; Kimura, Louise F; Alves, Andr�� F; Guti��rrez, Jos�� Mar��a; Otero, Rafael; Su��rez, Ana M; Santoro, Marcelo L; Barbaro, Katia C

2016-01-01

Snakebites inflicted by the arboreal viperid snake Bothriechis schlegelii in humans are characterized by pain, edema, and ecchymosis at the site of the bite, rarely with blisters, local necrosis, or defibrination. Herein, a comparative study of Bothriechis schlegelii snake venoms from Colombia (BsCo) and Costa Rica (BsCR) was carried out in order to compare their main activities and to verify the efficacy of Bothrops antivenom produced in Brazil to neutralize them. Biochemical (SDS-PAGE and z...

Radioisotope Characterization of HB Line Low Activity Waste

International Nuclear Information System (INIS)

Snyder, S.J.

1999-01-01

The purpose of this document is to provide a physical, chemical, hazardous and radiological characterization of Low-Level Waste (LLW) generated in HB-Line as required by the 1S Manual, Savannah River Site Waste Acceptance Criteria Manual

Characterizing chemical systems with on-line computers and graphics

International Nuclear Information System (INIS)

Frazer, J.W.; Rigdon, L.P.; Brand, H.R.; Pomernacki, C.L.

1979-01-01

Incorporating computers and graphics on-line to chemical experiments and processes opens up new opportunities for the study and control of complex systems. Systems having many variables can be characterized even when the variable interactions are nonlinear, and the system cannot a priori be represented by numerical methods and models. That is, large sets of accurate data can be rapidly acquired, then modeling and graphic techniques can be used to obtain partial interpretation plus design of further experimentation. The experimenter can thus comparatively quickly iterate between experimentation and modeling to obtain a final solution. We have designed and characterized a versatile computer-controlled apparatus for chemical research, which incorporates on-line instrumentation and graphics. It can be used to determine the mechanism of enzyme-induced reactions or to optimize analytical methods. The apparatus can also be operated as a pilot plant to design control strategies. On-line graphics were used to display conventional plots used by biochemists and three-dimensional response-surface plots

Characterization of a novel telomerase-immortalized human endometrial stromal cell line, St-T1b

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Brosens Jan J

2009-07-01

Full Text Available Abstract Background Coordinated differentiation of the endometrial compartments in the second half of the menstrual cycle is a prerequisite for the establishment of pregnancy. Endometrial stromal cells (ESC decidualize under the influence of ovarian progesterone to accommodate implantation of the blastocyst and support establishment of the placenta. Studies into the mechanisms of decidualization are often hampered by the lack of primary ESC. Here we describe a novel immortalized human ESC line. Methods Primary ESC were immortalized by the transduction of telomerase. The resultant cell line, termed St-T1b, was characterized for its morphological and biochemical properties by immunocytochemistry, RT-PCR and immunoblotting. Its progestational response was tested using progesterone and medroxyprogesterone acetate with and without 8-Br-cAMP, an established inducer of decidualization in vitro. Results St-T1b were positive for the fibroblast markers vimentin and CD90 and negative for the epithelial marker cytokeratin-7. They acquired a decidual phenotype indistinguishable from primary ESC in response to cAMP stimulation. The decidual response was characterized by transcriptional activation of marker genes, such as PRL, IGFBP1, and FOXO1, and enhanced protein levels of the tumor suppressor p53 and the metastasis suppressor KAI1 (CD82. Progestins alone had no effect on St-T1b cells, but medroxyprogesterone acetate greatly enhanced the cAMP-stimulated expression of IGFBP-1 after 3 and 7 days. Progesterone, albeit more weakly, also augmented the cAMP-induced IGFBP-1 production but only after 7 days of treatment. The cell line remained stable in continuous culture for more than 150 passages. Conclusion St-T1b express the appropriate phenotypic ESC markers and their decidual response closely mimics that of primary cultures. Decidualization is efficiently induced by cAMP analog and enhanced by medroxyprogesterone acetate, and, to a lesser extent, by natural

Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

Science.gov (United States)

Malherbe, Christiaan J.; de Beer, Dalene; Joubert, Elizabeth

2012-01-01

Biochemical detection (BCD) methods are commonly used to screen plant extracts for specific biological activities in batch assays. Traditionally, bioactives in the most active extracts were identified through time-consuming bio-assay guided fractionation until single active compounds could be isolated. Not only are isolation procedures often tedious, but they could also lead to artifact formation. On-line coupling of BCD assays to high performance liquid chromatography (HPLC) is gaining ground as a high resolution screening technique to overcome problems associated with pre-isolation by measuring the effects of compounds post-column directly after separation. To date, several on-line HPLC-BCD assays, applied to whole plant extracts and mixtures, have been published. In this review the focus will fall on enzyme-based, receptor-based and antioxidant assays. PMID:22489144

Characterization of Biochemical Composition for Different Types of Spent Mushroom Substrate in Malaysia

International Nuclear Information System (INIS)

Nur Aziera Abd Rasib; Zarina Zakaria; Mohammad Fahrurrazi Tompang; Ridzwan Abdul Rahman; Hakimah Othman

2015-01-01

A preliminary study was conducted to identify the amount and changes of biochemical composition of different types of Malaysian spent mushroom substrate (SMS) before and after several cycle of mushroom cultivation. The characterization of SMS involved the analysis of crude protein, carbohydrate, fat, lignin and ash for selected mushrooms namely as white oyster (Pleuratos oestrous), grey oyster (Pleuratos sajor-caju), abalone (Pleuratos cystidiosus), ganoderma (Ganoderma lucidium) and black jelly (Auricularia polytricha). Overall trend showed that there were increment for crude protein and fat, whereas carbohydrate and lignin showed reduction in the content. Significant results were showed on protein increment where ganoderma attained the highest value, 36.6 g, followed by black jelly, white oyster, grey oyster and abalone. Contradictory, lowest carbohydrate reduction was observed in ganoderma at 70.42 g and the most was in black jelly. Increment in fat and reduction in lignin was almost similar for each SMS. There was an increment in the ash percentage resulted from sterilization process. Clearly cultivation by mushroom had changed biochemical value especially in increasing the protein content that might be useful in protein required industry such as animal feeding. (author)

Purification and biochemical characterization of NpABCG5/NpPDR5, a plant pleiotropic drug resistance transporter expressed in Nicotiana tabacum BY-2 suspension cells.

Science.gov (United States)

Toussaint, Frédéric; Pierman, Baptiste; Bertin, Aurélie; Lévy, Daniel; Boutry, Marc

2017-05-04

Pleiotropic drug resistance (PDR) transporters belong to the ABCG subfamily of ATP-binding cassette (ABC) transporters and are involved in the transport of various molecules across plasma membranes. During evolution, PDR genes appeared independently in fungi and in plants from a duplication of a half-size ABC gene. The enzymatic properties of purified PDR transporters from yeast have been characterized. This is not the case for any plant PDR transporter, or, incidentally, for any purified plant ABC transporter. Yet, plant PDR transporters play important roles in plant physiology such as hormone signaling or resistance to pathogens or herbivores. Here, we describe the expression, purification, enzymatic characterization and 2D analysis by electron microscopy of NpABCG5/NpPDR5 from Nicotiana plumbaginifolia , which has been shown to be involved in the plant defense against herbivores. We constitutively expressed NpABCG5/NpPDR5, provided with a His-tag in a homologous system: suspension cells from Nicotiana tabacum (Bright Yellow 2 line). NpABCG5/NpPDR5 was targeted to the plasma membrane and was solubilized by dodecyl maltoside and purified by Ni-affinity chromatography. The ATP-hydrolyzing specific activity (27 nmol min -1  mg -1 ) was stimulated seven-fold in the presence of 0.1% asolectin. Electron microscopy analysis indicated that NpABCG5/NpPDR5 is monomeric and with dimensions shorter than those of known ABC transporters. Enzymatic data (optimal pH and sensitivity to inhibitors) confirmed that plant and fungal PDR transporters have different properties. These data also show that N. tabacum suspension cells are a convenient host for the purification and biochemical characterization of ABC transporters. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

Biochemical characterization of a recombinant Japanese encephalitis virus RNA-dependent RNA polymerase

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Kim Chan-Mi

2007-07-01

Full Text Available Abstract Background Japanese encephalitis virus (JEV NS5 is a viral nonstructural protein that carries both methyltransferase and RNA-dependent RNA polymerase (RdRp domains. It is a key component of the viral RNA replicase complex that presumably includes other viral nonstructural and cellular proteins. The biochemical properties of JEV NS5 have not been characterized due to the lack of a robust in vitro RdRp assay system, and the molecular mechanisms for the initiation of RNA synthesis by JEV NS5 remain to be elucidated. Results To characterize the biochemical properties of JEV RdRp, we expressed in Escherichia coli and purified an enzymatically active full-length recombinant JEV NS5 protein with a hexahistidine tag at the N-terminus. The purified NS5 protein, but not the mutant NS5 protein with an Ala substitution at the first Asp of the RdRp-conserved GDD motif, exhibited template- and primer-dependent RNA synthesis activity using a poly(A RNA template. The NS5 protein was able to use both plus- and minus-strand 3'-untranslated regions of the JEV genome as templates in the absence of a primer, with the latter RNA being a better template. Analysis of the RNA synthesis initiation site using the 3'-end 83 nucleotides of the JEV genome as a minimal RNA template revealed that the NS5 protein specifically initiates RNA synthesis from an internal site, U81, at the two nucleotides upstream of the 3'-end of the template. Conclusion As a first step toward the understanding of the molecular mechanisms for JEV RNA replication and ultimately for the in vitro reconstitution of viral RNA replicase complex, we for the first time established an in vitro JEV RdRp assay system with a functional full-length recombinant JEV NS5 protein and characterized the mechanisms of RNA synthesis from nonviral and viral RNA templates. The full-length recombinant JEV NS5 will be useful for the elucidation of the structure-function relationship of this enzyme and for the

Biochemical and Kinetic Characterization of Geranylgeraniol 18 ...

African Journals Online (AJOL)

This enzyme and its gene are an attractive target for development of plaunotol production and its detailed biochemical properties need to be understood. Recently, even though the gene (CYP97C27) coding for GGOH 18-hydroxylase has been identified, cloned, and expressed in Escherichia coli system, the enzyme activity ...

Establishment and characterization of rat portal myofibroblast cell lines.

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Michel Fausther

Full Text Available The major sources of scar-forming myofibroblasts during liver fibrosis are activated hepatic stellate cells (HSC and portal fibroblasts (PF. In contrast to well-characterized HSC, PF remain understudied and poorly defined. This is largely due to the facts that isolation of rodent PF for functional studies is technically challenging and that PF cell lines had not been established. To address this, we have generated two polyclonal portal myofibroblast cell lines, RGF and RGF-N2. RGF and RGF-N2 were established from primary PF isolated from adult rat livers that underwent culture activation and subsequent SV40-mediated immortalization. Specifically, Ntpdase2/Cd39l1-sorted primary PF were used to generate the RGF-N2 cell line. Both cell lines were functionally characterized by RT-PCR, immunofluorescence, immunoblot and bromodeoxyuridine-based proliferation assay. First, immortalized RGF and RGF-N2 cells are positive for phenotypic myofibroblast markers alpha smooth muscle actin, type I collagen alpha-1, tissue inhibitor of metalloproteinases-1, PF-specific markers elastin, type XV collagen alpha-1 and Ntpdase2/Cd39l1, and mesenchymal cell marker ecto-5'-nucleotidase/Cd73, while negative for HSC-specific markers desmin and lecithin retinol acyltransferase. Second, both RGF and RGF-N2 cell lines are readily transfectable using standard methods. Finally, RGF and RGF-N2 cells attenuate the growth of Mz-ChA-1 cholangiocarcinoma cells in co-culture, as previously demonstrated for primary PF. Immortalized rat portal myofibroblast RGF and RGF-N2 cell lines express typical markers of activated PF-derived myofibroblasts, are suitable for DNA transfection, and can effectively inhibit cholangiocyte proliferation. Both RGF and RGF-N2 cell lines represent novel in vitro cellular models for the functional studies of portal (myofibroblasts and their contribution to the progression of liver fibrosis.

Biochemical and serological characterization of Escherichia coli ...

African Journals Online (AJOL)

This study was designed to determine the isolation rate, serotypes and biochemical profiles of E. coli from colibacillosis and dead-in-shell embryos in Zaria, Northern-Nigeria. The isolation rate of E. coli from hatcheries studied were 4.67% and 7.50% from farms of Simtu Agricultural Company and National Animal Production ...

Size characterization of airborne SiO2 nanoparticles with on-line and off-line measurement techniques: an interlaboratory comparison study

Science.gov (United States)

Motzkus, C.; Macé, T.; Gaie-Levrel, F.; Ducourtieux, S.; Delvallee, A.; Dirscherl, K.; Hodoroaba, V.-D.; Popov, I.; Popov, O.; Kuselman, I.; Takahata, K.; Ehara, K.; Ausset, P.; Maillé, M.; Michielsen, N.; Bondiguel, S.; Gensdarmes, F.; Morawska, L.; Johnson, G. R.; Faghihi, E. M.; Kim, C. S.; Kim, Y. H.; Chu, M. C.; Guardado, J. A.; Salas, A.; Capannelli, G.; Costa, C.; Bostrom, T.; Jämting, Å. K.; Lawn, M. A.; Adlem, L.; Vaslin-Reimann, S.

2013-10-01

Results of an interlaboratory comparison on size characterization of SiO2 airborne nanoparticles using on-line and off-line measurement techniques are discussed. This study was performed in the framework of Technical Working Area (TWA) 34—"Properties of Nanoparticle Populations" of the Versailles Project on Advanced Materials and Standards (VAMAS) in the project no. 3 "Techniques for characterizing size distribution of airborne nanoparticles". Two types of nano-aerosols, consisting of (1) one population of nanoparticles with a mean diameter between 30.3 and 39.0 nm and (2) two populations of non-agglomerated nanoparticles with mean diameters between, respectively, 36.2-46.6 nm and 80.2-89.8 nm, were generated for characterization measurements. Scanning mobility particle size spectrometers (SMPS) were used for on-line measurements of size distributions of the produced nano-aerosols. Transmission electron microscopy, scanning electron microscopy, and atomic force microscopy were used as off-line measurement techniques for nanoparticles characterization. Samples were deposited on appropriate supports such as grids, filters, and mica plates by electrostatic precipitation and a filtration technique using SMPS controlled generation upstream. The results of the main size distribution parameters (mean and mode diameters), obtained from several laboratories, were compared based on metrological approaches including metrological traceability, calibration, and evaluation of the measurement uncertainty. Internationally harmonized measurement procedures for airborne SiO2 nanoparticles characterization are proposed.

Size characterization of airborne SiO2 nanoparticles with on-line and off-line measurement techniques: an interlaboratory comparison study

International Nuclear Information System (INIS)

Motzkus, C.; Macé, T.; Gaie-Levrel, F.; Ducourtieux, S.; Delvallee, A.; Dirscherl, K.; Hodoroaba, V.-D.; Popov, I.; Popov, O.; Kuselman, I.; Takahata, K.; Ehara, K.; Ausset, P.; Maillé, M.; Michielsen, N.; Bondiguel, S.; Gensdarmes, F.; Morawska, L.; Johnson, G. R.; Faghihi, E. M.

2013-01-01

Results of an interlaboratory comparison on size characterization of SiO 2 airborne nanoparticles using on-line and off-line measurement techniques are discussed. This study was performed in the framework of Technical Working Area (TWA) 34—“Properties of Nanoparticle Populations” of the Versailles Project on Advanced Materials and Standards (VAMAS) in the project no. 3 “Techniques for characterizing size distribution of airborne nanoparticles”. Two types of nano-aerosols, consisting of (1) one population of nanoparticles with a mean diameter between 30.3 and 39.0 nm and (2) two populations of non-agglomerated nanoparticles with mean diameters between, respectively, 36.2–46.6 nm and 80.2–89.8 nm, were generated for characterization measurements. Scanning mobility particle size spectrometers (SMPS) were used for on-line measurements of size distributions of the produced nano-aerosols. Transmission electron microscopy, scanning electron microscopy, and atomic force microscopy were used as off-line measurement techniques for nanoparticles characterization. Samples were deposited on appropriate supports such as grids, filters, and mica plates by electrostatic precipitation and a filtration technique using SMPS controlled generation upstream. The results of the main size distribution parameters (mean and mode diameters), obtained from several laboratories, were compared based on metrological approaches including metrological traceability, calibration, and evaluation of the measurement uncertainty. Internationally harmonized measurement procedures for airborne SiO 2 nanoparticles characterization are proposed

Recombinant expression, purification and biochemical characterization of kievitone hydratase from Nectria haematococca.

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Matthias Engleder

Full Text Available Kievitone hydratase catalyzes the addition of water to the double bond of the prenyl moiety of plant isoflavonoid kievitone and, thereby, forms the tertiary alcohol hydroxy-kievitone. In nature, this conversion is associated with a defense mechanism of fungal pathogens against phytoalexins generated by host plants after infection. As of today, a gene sequence coding for kievitone hydratase activity has only been identified and characterized in Fusarium solani f. sp. phaseoli. Here, we report on the identification of a putative kievitone hydratase sequence in Nectria haematococca (NhKHS, the teleomorph state of F. solani, based on in silico sequence analyses. After heterologous expression of the enzyme in the methylotrophic yeast Pichia pastoris, we have confirmed its kievitone hydration activity and have assessed its biochemical properties and substrate specificity. Purified recombinant NhKHS is obviously a homodimeric glycoprotein. Due to its good activity for the readily available chalcone derivative xanthohumol (XN, this compound was selected as a model substrate for biochemical studies. The optimal pH and temperature for hydratase activity were 6.0 and 35°C, respectively, and apparent Vmax and Km values for hydration of XN were 7.16 μmol min-1 mg-1 and 0.98 ± 0.13 mM, respectively. Due to its catalytic properties and apparent substrate promiscuity, NhKHS is a promising enzyme for the biocatalytic production of tertiary alcohols.

Design and Fabrication of Piezoresistive Based Encapsulated Poly-Si Cantilevers for Bio/chemical Sensing

Science.gov (United States)

Krishna, N. P. Vamsi; Murthy, T. R. Srinivasa; Reddy, K. Jayaprakash; Sangeeth, K.; Hegde, G. M.

Cantilever-based sensing is a growing research field not only within micro regime but also in nano technology. The technology offers a method for rapid, on-line and in-situ monitoring of specific bio/chemical substances by detecting the nanomechanical responses of a cantilever sensor. Cantilever with piezoresistive based detection scheme is more attractive because of its electronics compatibility. Majority of commercially available micromachined piezoresistive sensors are bulk micromachined devices and are fabricated using single crystal silicon wafers. As substrate properties are not important in surface micromachining, the expensive silicon wafers can be replaced by cheaper substrates, such as poly-silicon, glass or plastic. Here we have designed SU-8 based bio/chemical compatible micro electro mechanical device that includes an encapsulated polysilicon piezoresistor for bio/chemical sensing. In this paper we report the design, fabrication and analysis of the encapsulated poly-Si cantilevers. Design and theoretical analysis are carried out using Finite Element Analysis software. For fabrication of poly-silicon piezoresistive cantilevers we followed the surface micromachining process steps. Preliminary characterization of the cantilevers is presented.

Characterization and drug resistance patterns of Ewing's sarcoma family tumor cell lines.

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William A May

Full Text Available Despite intensive treatment with chemotherapy, radiotherapy and surgery, over 70% of patients with metastatic Ewing's Sarcoma Family of Tumors (EFT will die of their disease. We hypothesize that properly characterized laboratory models reflecting the drug resistance of clinical tumors will facilitate the application of new therapeutic agents to EFT. To determine resistance patterns, we studied newly established EFT cell lines derived from different points in therapy: two established at diagnosis (CHLA-9, CHLA-32, two after chemotherapy and progressive disease (CHLA-10, CHLA-25, and two at relapse after myeloablative therapy and autologous bone marrow transplantation (post-ABMT (CHLA-258, COG-E-352. The new lines were compared to widely studied EFT lines TC-71, TC-32, SK-N-MC, and A-673. These lines were extensively characterized with regard to identity (short tandem repeat (STR analysis, p53, p16/14 status, and EWS/ETS breakpoint and target gene expression profile. The DIMSCAN cytotoxicity assay was used to assess in vitro drug sensitivity to standard chemotherapy agents. No association was found between drug resistance and the expression of EWS/ETS regulated genes in the EFT cell lines. No consistent association was observed between drug sensitivity and p53 functionality or between drug sensitivity and p16/14 functionality across the cell lines. Exposure to chemotherapy prior to cell line initiation correlated with drug resistance of EFT cell lines in 5/8 tested agents at clinically achievable concentrations (CAC or the lower tested concentration (LTC: (cyclophosphamide (as 4-HC and doxorubicin at CAC, etoposide, irinotecan (as SN-38 and melphalan at LTC; P<0.1 for one agent, and P<0.05 for four agents. This panel of well-characterized drug-sensitive and drug-resistant cell lines will facilitate in vitro preclinical testing of new agents for EFT.

Biochemical characterization of the triticale TsPAP1, a new type of plant prolyl aminopeptidase, and its impact on proline content and flowering time in transgenic Arabidopsis plants.

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Zdunek-Zastocka, Edyta; Grabowska, Agnieszka; Branicki, Tomasz; Michniewska, Beata

2017-07-01

Proline aminopeptidase (PAP, EC 3.4.11.5) is the only enzyme that effectively releases proline from the N-termini of peptides. The amino acid sequence of the PAP from Triticosecale, TsPAP1, comprises conserved regions, characteristic of the monomeric forms of PAP found in bacteria but not yet identified in plants. Therefore, we aimed to obtain and biochemically characterize the TsPAP1 protein. The recombinant TsPAP1 protein was received through heterologous expression of the TsPAP1 coding sequence in a bacterial expression system and purified with affinity chromatography. Gel filtration chromatography and SDS electrophoresis revealed that TsPAP1 is a monomer with a molecular mass of 37.5 kDa. TsPAP1 prefers substrates with proline at the N-terminus but is also capable of hydrolyzing β-naphthylamides of hydroxyproline and alanine. Among the peptides tested, the most preferred were di- and tripeptides, especially those with glycine in the Y position. The use of diagnostic inhibitors indicated that TsPAP1 is a serine peptidase; however, further characterization revealed that the SH residues are also important for maintaining its activity. To examine the role of TsPAP1 under physiological conditions, we developed transgenic Arabidopsis plants overexpressing TsPAP1. Compared with wild-type plants, the transgenic lines accumulated more proline, flowered an average of 3.5 days earlier, and developed more siliques than did untransformed controls. Our paper is the first to describe the biochemical properties of a novel monomeric plant PAP and contributes to the functional characterization of PAP proteins in plants. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Biochemical characterization of an in-house Coccidioides antigen: perspectives for the immunodiagnosis of coccidioidomycosis.

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Filho, Renato Evando Moreira; Bandeira, Silviane Praciano; Brillhante, Raimunda Sâmia Nogueira; Rocha, Marcos Fábio Gadelha; Vasconcelos, Ilka Maria; Pereira, Mirella Leite; Castelo-Branco, Débora de Souza Collares Maia; Rocha, Francisco Airton Castro; Camargo, Zoilo Pires de; Ramos, Marcio Viana; Cordeiro, Rossana de Aguiar; Sidrim, José Júlio Costa

2012-06-28

The objective of this study was to evaluate the reactivity of an in-house antigen, extracted from a strain of C. posadasii isolated in northeastern Brazil, by radial immunodiffusion and Western blotting, as well as to establish its biochemical characterization. The protein antigen was initially extracted with the use of solid ammonium sulfate and characterized by 1-D electrophoresis. Subsequently, it was tested by means of double radial immunodiffusion and Western blotting. A positive reaction was observed against the antigen by both immunodiagnostic techniques tested on sera from patients suffering from coccidioidomycosis. Besides this, two immunoreactive protein bands were observed and were revealed to be a β-glucosidase and a glutamine synthetase after sequencing of the respective N-terminal regions. Our in-house Coccidioides antigen can be promising as a quick and low-cost diagnostic tool without the risk of direct manipulation of the microorganism.

Isolation and biochemical and molecular characterization of Listeria monocytogenes in food

International Nuclear Information System (INIS)

Helel, Salma

2008-01-01

monocytogenes is a Gram-positive bacteria, saprophytic, non-spore. This is an extremely resistant seeds to environmental conditions outside, especially since the cold psychrotrophic. It can contaminate raw vegetables, cooked meals ready for consumption or foods to be stored in the refrigerator, such as cheese or meat. It is the bacteria responsible for listeriosis. It threatens first unborn children, infants, pregnant women, the elderly and people whose immune system is weakened. Strains of Listeria spp isolated from foods (seafood, meat, meat) were first identified at the stage of the genus by classical tests (Gram staining, catalase test, oxidase test and mobility) and stage of the test case by hemolysis, CAMP test and the gallery Api Listeria. Biochemical characterization allowed after a numerical analysis, to assign 100% of isolates to the genus Listeria. Molecular characterization was performed by PCR amplification of genes coding for protein p60 (iap), the listeriolysine O (hly), the Phosphatidylinositol Phospholipase C (PI-PLC plca) Phosphatidylcholine Phospholipase C (plcB). The result showed an amplification of the iap gene of 100% of the hly gene, plca, plcB of 31.81%. This characterization represents an identification of the collection on the genetic level and shows that 31.81% of isolates, is likely to express the genes responsible for virulence factors of L. monocytogenes, to produce listeriolysine O, phospholipase C and Lecithinase. The molecular identification was performed by microarray technique and identified isolates L. September monocytogenes (five original clinical isolates and two food-borne), fourteen L. innocua (of food) and a strain not identified by DNA chip.. (Author)

Development and characterization of a cell line WAF from freshwater shark Wallago attu.

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Dubey, Akhilesh; Goswami, Mukunda; Yadav, Kamalendra; Sharma, Bhagwati S

2014-02-01

A new epithelial cell line, WAF was developed from caudal fin of freshwater shark, Wallago attu. The cell line was optimally maintained at 28 °C in Leibovitz-15 (L-15) medium supplemented with 20 % fetal bovine serum. The cell line was characterized by various cytogenetic and molecular markers. The cytogenetic analysis revealed a diploid count of 86 chromosomes at different passages. The origin of the cell lines was confirmed by the amplification of 547 and 654 bp sequences of 16S rRNA and cytochrome oxidase subunit I genes of mitochondrial DNA, respectively. WAF cells were characterized for their growth characteristics at different temperature and serum concentration. Epithelial morphology of the cell line was confirmed using immunocytochemistry. Further cell plating efficiency, transfection efficiency and viability of cryopreserved WAF cells was also determined. Cytotoxicity and genotoxicity assessment of cadmium salts on WAF cells by MTT, NR and comet assay illustrated the utility of this cell line as an in vitro model for aquatic toxicological studies. The cell line will be further useful for studying oxidative stress markers against aquatic pollutants.

Size characterization of airborne SiO{sub 2} nanoparticles with on-line and off-line measurement techniques: an interlaboratory comparison study

Energy Technology Data Exchange (ETDEWEB)

Motzkus, C., E-mail: charles.motzkus@lne.fr; Mace, T.; Gaie-Levrel, F.; Ducourtieux, S.; Delvallee, A. [Laboratoire National de Metrologie et d' Essais (LNE) (France); Dirscherl, K. [Danish Fundamental Metrology (DFM) (Denmark); Hodoroaba, V.-D. [BAM Federal Institute for Materials Research and Testing (Germany); Popov, I. [The Hebrew University of Jerusalem, Unit for Nanocharacterization (Israel); Popov, O.; Kuselman, I. [National Physical Laboratory of Israel (INPL) (Israel); Takahata, K.; Ehara, K. [National Institute of Advanced Industrial Science and Technology (AIST), National Metrology Institute of Japan (NMIJ) (Japan); Ausset, P.; Maille, M. [Universite Paris-Est Creteil et Universite Paris-Diderot, Laboratoire Interuniversitaire des Systemes Atmospheriques (LISA), UMR CNRS 7583 (France); Michielsen, N.; Bondiguel, S.; Gensdarmes, F. [Institut de Radioprotection et de Surete Nucleaire (IRSN), PSN-RES, SCA, LPMA (France); Morawska, L.; Johnson, G. R.; Faghihi, E. M. [Queensland University of Technology (QUT), International Laboratory for Air Quality and Health (ILAQH) (Australia); and others

2013-10-15

Results of an interlaboratory comparison on size characterization of SiO{sub 2} airborne nanoparticles using on-line and off-line measurement techniques are discussed. This study was performed in the framework of Technical Working Area (TWA) 34-'Properties of Nanoparticle Populations' of the Versailles Project on Advanced Materials and Standards (VAMAS) in the project no. 3 'Techniques for characterizing size distribution of airborne nanoparticles'. Two types of nano-aerosols, consisting of (1) one population of nanoparticles with a mean diameter between 30.3 and 39.0 nm and (2) two populations of non-agglomerated nanoparticles with mean diameters between, respectively, 36.2-46.6 nm and 80.2-89.8 nm, were generated for characterization measurements. Scanning mobility particle size spectrometers (SMPS) were used for on-line measurements of size distributions of the produced nano-aerosols. Transmission electron microscopy, scanning electron microscopy, and atomic force microscopy were used as off-line measurement techniques for nanoparticles characterization. Samples were deposited on appropriate supports such as grids, filters, and mica plates by electrostatic precipitation and a filtration technique using SMPS controlled generation upstream. The results of the main size distribution parameters (mean and mode diameters), obtained from several laboratories, were compared based on metrological approaches including metrological traceability, calibration, and evaluation of the measurement uncertainty. Internationally harmonized measurement procedures for airborne SiO{sub 2} nanoparticles characterization are proposed.

Biochemical and physiological characterization of oil palm interspecific hybrids (elaeis oleifera x elaeis guineensis) grown in hydroponics

International Nuclear Information System (INIS)

Rivera Mendez, Yurany Dayanna; Moreno Chacon, Andres Leonardo; Romero, Hernan Mauricio

2013-01-01

The interspecific hybrid, Elaeis oleifera x Elaeis guineensis (OxG) is an alternative for improving the competitiveness and sustainability of the Latin American oil palm agro-industry, because of its partial resistance to some lethal diseases and also because of the high quality of its oil. A comparative characterization was conducted of the physiological and biochemical performance of seedlings of six OxG hybrids grown in hydroponics. Gas exchange, vegetative growth, protein, sugar and photosynthetic pigment content, and antioxidant system activity were determined. With the exception of gas exchange, the other variables showed significant differences between materials. The U1273 and U1737 materials showed greater vegetative growth with no expression of biochemical traits, while the U1914 and U1990 materials showed high levels of reducing and total sugars, photosynthetic pigments, and antioxidant system activities, characteristics that could confer them adaptation to stress conditions. With the standardized hydroponics technique, the optimal conditions for the growth of seedlings were ensured, the differences between materials were established, and so those with promising features from the physiological and biochemical standpoint were identified. Finally, it could be used to study in a simple, fast, clean and inexpensive way, the effect of levels and sources of mineral nutrients on the growth and development of oil palm.

Biochemical characterization of blood plasma of coronary artery ...

Indian Academy of Sciences (India)

This study aimed to investigate the biochemical profile of blood plasma of patients with coronary artery disease (CAD) and angiographically normal subjects (controls) to determine biomarkers for their differentiation. In this double blind study, 5 mL venous blood was drawn before angiography from CAD patients (n=60) and ...

Characterizations of p-Wavelets on Positive Half Line Using the Walsh-Fourier Transform

Directory of Open Access Journals (Sweden)

Abdullah Abdullah

2016-03-01

Full Text Available In this paper, we study the characterization of wavelets on positive half line by means of two basic equations in the Fourier domain. We also give another characterization of wavelets.

Molecular characterization of breast cancer cell lines through multiple omic approaches.

Science.gov (United States)

Smith, Shari E; Mellor, Paul; Ward, Alison K; Kendall, Stephanie; McDonald, Megan; Vizeacoumar, Frederick S; Vizeacoumar, Franco J; Napper, Scott; Anderson, Deborah H

2017-06-05

Breast cancer cell lines are frequently used as model systems to study the cellular properties and biology of breast cancer. Our objective was to characterize a large, commonly employed panel of breast cancer cell lines obtained from the American Type Culture Collection (ATCC 30-4500 K) to enable researchers to make more informed decisions in selecting cell lines for specific studies. Information about these cell lines was obtained from a wide variety of sources. In addition, new information about cellular pathways that are activated within each cell line was generated. We determined key protein expression data using immunoblot analyses. In addition, two analyses on serum-starved cells were carried out to identify cellular proteins and pathways that are activated in these cells. These analyses were performed using a commercial PathScan array and a novel and more extensive phosphopeptide-based kinome analysis that queries 1290 phosphorylation events in major signaling pathways. Data about this panel of breast cancer cell lines was also accessed from several online sources, compiled and summarized for the following areas: molecular classification, mRNA expression, mutational status of key proteins and other possible cancer-associated mutations, and the tumorigenic and metastatic capacity in mouse xenograft models of breast cancer. The cell lines that were characterized included 10 estrogen receptor (ER)-positive, 12 human epidermal growth factor receptor 2 (HER2)-amplified and 18 triple negative breast cancer cell lines, in addition to 4 non-tumorigenic breast cell lines. Within each subtype, there was significant genetic heterogeneity that could impact both the selection of model cell lines and the interpretation of the results obtained. To capture the net activation of key signaling pathways as a result of these mutational combinations, profiled pathway activation status was examined. This provided further clarity for which cell lines were particularly deregulated

Metrological characterization methods for confocal chromatic line sensors and optical topography sensors

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Seppä, Jeremias; Niemelä, Karri; Lassila, Antti

2018-05-01

The increasing use of chromatic confocal technology for, e.g. fast, in-line optical topography, and measuring thickness, roughness and profiles implies a need for the characterization of various aspects of the sensors. Single-point, line and matrix versions of chromatic confocal technology, encoding depth information into wavelength, have been developed. Of these, line sensors are particularly suitable for in-line process measurement. Metrological characterization and development of practical methods for calibration and checking is needed for new optical methods and devices. Compared to, e.g. tactile methods, optical topography measurement techniques have limitations related to light wavelength and coherence, optical properties of the sample including reflectivity, specularity, roughness and colour, and definition of optical versus mechanical surfaces. In this work, metrological characterization methods for optical line sensors were developed for scale magnification and linearity, sensitivity to sample properties, and dynamic characteristics. An accurate depth scale calibration method using a single prototype groove depth sample was developed for a line sensor and validated with laser-interferometric sample tracking, attaining (sub)micrometre level or better than 0.1% scale accuracy. Furthermore, the effect of different surfaces and materials on the measurement and depth scale was studied, in particular slope angle, specularity and colour. In addition, dynamic performance, noise, lateral scale and resolution were measured using the developed methods. In the case of the LCI1200 sensor used in this study, which has a 11.3 mm  ×  2.8 mm measurement range, the instrument depth scale was found to depend only minimally on sample colour, whereas measuring steeply sloped specular surfaces in the peripheral measurement area, in the worst case, caused a somewhat larger relative sample-dependent change (1%) in scale.

K basin sandfilter backwash line characterization project, analytical results for campaign 15

International Nuclear Information System (INIS)

Steen, F.H.

1999-01-01

Sample 183KWBMF was taken from the K West Sandfilter Backwash Pit on August 28, 1998 and received by 222-S Laboratory on August 28, 1998. Analyses were performed in accordance with ''Letter of Instruction for K Basins Sandfilter Backwash Line Samples'' (LOI) in support of the K Basin Sandfilter Backwash Line Characterization Project

Grain characterization and milling behaviour of near-isogenic lines differing by hardness.

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Greffeuille, V; Abecassis, J; Rousset, M; Oury, F-X; Faye, A; L'Helgouac'h, C Bar; Lullien-Pellerin, V

2006-12-01

Wheat grain hardness is a major factor affecting the milling behaviour and end-product quality although its exact structural and biochemical basis is still not understood. This study describes the development of new near-isogenic lines selected on hardness. Hard and soft sister lines were characterised by near infrared reflectance (NIR) and particle size index (PSI) hardness index, grain protein content, thousand kernel weight and vitreousness. The milling behaviour of these wheat lines was evaluated on an instrumented micromill which also measures the grinding energy and flour particle size distribution was investigated by laser diffraction. Endosperm mechanical properties were measured using compression tests. Results pointed out the respective effect of hardness and vitreousness on those characteristics. Hardness was shown to influence both the mode of fracture and the mechanical properties of the whole grain and endosperm. Thus, this parameter also acts on milling behaviour. On the other hand, vitreousness was found to mainly play a role on the energy required to break the grain. This study allows us to distinguish between consequences of hardness and vitreousness. Hardness is suggested to influence the adhesion forces between starch granules and protein matrix whereas vitreousness would rather be related to the endosperm microstructure.

K Basin sandfilter backwash line characterization project, analytical results for Campaign 20

International Nuclear Information System (INIS)

STEEN, F.H.

1999-01-01

Sample 112KWBMF was taken from the K West Sandfilter Backwash Pit on June 1 , 1999, and received by 222-S Laboratory on June 2,1999. Analyses were performed on sample 112KWBMF in accordance with Letter of Instruction for K Basins Sandfilter Backwash Line Samples (LOI) in support of the K Basin Sandfilter Backwash Line Characterization Project

Selective enrichment and biochemical characterization of seven human skin fibroblasts cell types in vitro

International Nuclear Information System (INIS)

Rodemann, H.P.; Bayreuther, K.; Francz, P.I.; Dittmann, K.; Albiez, M.

1989-01-01

The mitotic and postmitotic populations of the human skin fibroblast cell line HH-8 are heterogeneous when studied in vitro. There are reproducible changes in the frequencies of the mitotic fibroblasts (MF), MF I, MF II, MF III, and the postmitotic fibroblasts (PMF), PMF IV, PMF V, PMF VI, and PMF VII. For biochemical characterization, methods for selective enrichment of homogeneous populations of these seven fibroblast cell types have been established. Clonal populations with 95% purity for the mitotic fibroblasts MF I, MF II, and MF III can be raised in uniform clone types of fibroblasts (CTF) CTF I, CTF II, and CTF III. Pure clonal subpopulations of MF I type cells are present in mass populations in the range of 1-20 cumulative population doublings (CPD). Populations of mitotic fibroblasts represent nearly homogeneous populations of MF II (75-85% purity) in the range of 28-34 CPD and MF III (73-86% purity) in the range of 48-53 CPD. These populations can be easily expanded to up to 10(7)-10(8) cells. The spontaneous transition of MF III to PMF VI takes 140-180 days. In order to shorten this period and increase the proportion of distinct postmitotic types, mitotic fibroblast mass populations (CPD 30-32, MF II: 75-85% purity) have been induced by uv-irradiation to differentiate to nearly homogeneous populations of PMF IV, PMF V, PMF VI, and PMF VII within 4 to 36 days of culture. Using this method, 10(7) cells of one differentiation stage can be obtained. Spontaneously arising and experimentally selected or induced homogeneous clonal and mass populations of MF I, MF II, MF III, PMF IV, PMF V, PMF VI, and PMF VII express an identical differentiation-dependent and cell-type-specific [35S]methionine-labeled polypeptide pattern

Morphological and biochemical characterization of the membranous hepatitis C virus replication compartment.

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Paul, David; Hoppe, Simone; Saher, Gesine; Krijnse-Locker, Jacomine; Bartenschlager, Ralf

2013-10-01

Like all other positive-strand RNA viruses, hepatitis C virus (HCV) induces rearrangements of intracellular membranes that are thought to serve as a scaffold for the assembly of the viral replicase machinery. The most prominent membranous structures present in HCV-infected cells are double-membrane vesicles (DMVs). However, their composition and role in the HCV replication cycle are poorly understood. To gain further insights into the biochemcial properties of HCV-induced membrane alterations, we generated a functional replicon containing a hemagglutinin (HA) affinity tag in nonstructural protein 4B (NS4B), the supposed scaffold protein of the viral replication complex. By using HA-specific affinity purification we isolated NS4B-containing membranes from stable replicon cells. Complementing biochemical and electron microscopy analyses of purified membranes revealed predominantly DMVs, which contained viral proteins NS3 and NS5A as well as enzymatically active viral replicase capable of de novo synthesis of HCV RNA. In addition to viral factors, co-opted cellular proteins, such as vesicle-associated membrane protein-associated protein A (VAP-A) and VAP-B, that are crucial for viral RNA replication, as well as cholesterol, a major structural lipid of detergent-resistant membranes, are highly enriched in DMVs. Here we describe the first isolation and biochemical characterization of HCV-induced DMVs. The results obtained underline their central role in the HCV replication cycle and suggest that DMVs are sites of viral RNA replication. The experimental approach described here is a powerful tool to more precisely define the molecular composition of membranous replication factories induced by other positive-strand RNA viruses, such as picorna-, arteri- and coronaviruses.

Biochemical characterization of propylglyoxal bis(guanylhydrazone). Facile synthesis of monoalkylglyoxal bis(guanylhydrazones).

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Elo, H; Laine, R; Alhonen-Hongisto, L; Jänne, J; Mutikainen, I; Lumme, P

1985-01-01

Propylglyoxal bis(guanylhydrazone) sulfate, a novel analog of the well-known antileukemic drug methylglyoxal bis(guanylhydrazone), has been prepared from 2,2-dibromopentanal, and the compound has been characterized biochemically. Although it is a powerful inhibitor of S-adenosylmethionine decarboxylase, its Ki value (0.2 microM) is considerably higher than that of ethylglyoxal bis(guanylhydrazone) (0.06 microM). The compound is only poorly taken up by tumor cells, and its accumulation is not stimulated by a prior exposure of the tumor cells to difluoromethylornithine, a compound that causes polyamine depletion. Thus, the uptake characteristics of the compound are similar to those of ethylglyoxal bis(guanylhydrazone), but in striking contrast to those of methylglyoxal and glyoxal bis(guanylhydrazones). Since the configuration of the double bonds in glyoxal, methylglyoxal and propylglyoxal bis(guanylhydrazones) has been shown to be identical, the different uptake characteristics are probably only due to differences in side chain size and/or hydrophobicity.

Derivation and characterization of a pig embryonic stem cell-derived exocrine pancreatic cell line

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The establishment and initial characterization of a pig embryonic stem cell-derived pancreatic cell line, PICM-31, and a colony-cloned derivative cell line, PICM-31A, is described. The cell lines were propagated for several months at split ratios of 1:3 or 1:5 at each passage on STO feeder cells af...

Establishment and conventional cytogenetic characterization of three gastric cancer cell lines.

Science.gov (United States)

Leal, Mariana Ferreira; Martins do Nascimento, José Luiz; da Silva, Carla Elvira Araújo; Vita Lamarão, Maria Fernanda; Calcagno, Danielle Queiroz; Khayat, André Salim; Assumpção, Paulo Pimentel; Cabral, Isabel Rosa; de Arruda Cardoso Smith, Marília; Burbano, Rommel Rodríguez

2009-11-01

Gastric cancer is the fourth most frequent type of cancer and the second most frequent cause of cancer mortality worldwide. Only a modest number of gastric carcinoma cell lines have been isolated thus far. Here we describe the establishment and cytogenetic characterization of three new gastric cancer cell lines obtained from primary gastric adenocarcinoma (ACP02 and ACP03) and cancerous ascitic fluid (AGP01) of individuals from northern Brazil. ACP02, ACP03, and AGP01 cell lines are presently in the 60th passage. The cell lines grew in a disorganized single layer with some agglomerations and heterogeneous divisions (bipolar and multipolar). All cell lines exhibited a composite karyotype with several clonal chromosome alterations. Trisomy 8 was the most frequent alteration. Chromosome 8 aneusomy was confirmed by fluorescence in situ hybridization. All cell lines also exhibited trisomy 7 and deletion of chromosome arm 17p. These results suggest that, although frequent chromosome alterations are commonly observed due to culture process, the ACP02, ACP03, and AGP01 cell lines and primary gastric cancer from individuals of northern Brazil share genetic alterations, supporting use of these cell lines as a model of gastric carcinogenesis in this population.

Cellular and Phenotypic Characterization of Canine Osteosarcoma Cell Lines

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Marie E. Legare, Jamie Bush, Amanda K. Ashley, Taka Kato, William H. Hanneman

2011-01-01

Full Text Available Canine and human osteosarcoma (OSA have many similarities, with the majority of reported cases occurring in the appendicular skeleton, gender predominance noted, high rate of metastasis at the time of presentation, and a lack of known etiology for this devastating disease. Due to poor understanding of the molecular mechanisms underlying OSA, we have characterized seven different OSA canine cell lines: Abrams, D17, Grey, Hughes, Ingles, Jarques, and Marisco and compared them to U2, a human OSA cell line, for the following parameters: morphology, growth, contact inhibition, migrational tendencies, alkaline phosphatase staining, heterologous tumor growth, double-strand DNA breaks, and oxidative damage. All results demonstrated the positive characteristics of the Abrams cell line for use in future studies of OSA. Of particular interest, the robust growth of a subcutaneous tumor and rapid pulmonary metastasis of the Abrams cell line in an immunocompromised mouse shows incredible potential for the future use of Abrams as a canine OSA model. Further investigations utilizing a canine cell model of OSA, such as Abrams, will be invaluable to understanding the molecular events underlying OSA, pharmaceutical inhibition of metastasis, and eventual prevention of this devastating disease.

Characterization and evaluation of rice blast resistance of Chinese indica hybrid rice parental lines

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Yunyu Wu

2017-12-01

Full Text Available The development of resistant varieties and hybrid combinations has been the most effective and economical strategy to control blast disease caused by Magnaporthe oryzae. However, the distribution of major R genes and blast resistance characterization in hybrid rice parents has not been well investigated, resulting in their limited use in hybrid rice blast-resistance breeding. In the present study, 88 elite indica hybrid rice parental lines were evaluated with 30 isolates of M. oryzae collected from the main planting area of indica hybrid rice in China and were characterized for the presence of 11 major resistance genes using molecular markers. The pathogenicity assays showed that four types of hybrid rice parent line showed some resistance to M. oryzae. However, the proportions of highly resistant lines and the mean resistance frequency (RF varied among the four types, with resistance in decreasing order shown by three-line restorer lines, three-line maintainer lines, two-line sterile lines, and two-line restorer lines. All 88 hybrid rice parental lines carried more than one R gene, but none carried the R genes Pi1 and Pi2. Although Pid3 and Pi9 were present only in three-line restorer lines and Pigm only in three-line maintainer lines, the remaining six R genes (Pib, Pid2, Pi5, Pia, Pi54, and Pita were present in the four types of hybrid rice parent with significantly different distribution frequencies. The correlation between R genes and resistance reactions was investigated. The results are expected to provide useful information for rational utilization of major R genes in hybrid rice breeding programs. Keywords: Hybrid rice parental lines, Magnaporthe oryzae, Pi genes, Resistance evaluation, Molecular markers

Biochemical characterization of amandin, the major storage protein in almond (Prunus dulcis L.).

Science.gov (United States)

Sathe, Shridhar K; Wolf, Walter J; Roux, Kenneth H; Teuber, Suzanne S; Venkatachalam, Mahesh; Sze-Tao, Kar Wai Clara

2002-07-17

The almond major storage protein, amandin, was prepared by column chromatography (amandin-1), cryoprecipitation (amandin-2), and isoelectric precipitation (amandin-3) methods. Amandin is a legumin type protein characterized by a sedimentation value of 14S. Amandin is composed of two major types of polypeptides with estimated molecular weights of 42-46 and 20-22 kDa linked via disulfide bonds. Several additional minor polypeptides were also present in amandin. Amandin is a storage protein with an estimated molecular weight of 427,300 +/- 47,600 Da (n = 7) and a Stokes radius of 65.88 +/- 3.21 A (n = 7). Amandin is not a glycoprotein. Amandin-1, amandin-2, and amandin-3 are antigenically related and have similar biochemical properties. Amandin-3 is more negatively charged than either amandin-1 or amandin-2. Methionine is the first essential limiting amino acid in amandin followed by lysine and threonine.

Improving Marine Ecosystem Models with Biochemical Tracers

Science.gov (United States)

Pethybridge, Heidi R.; Choy, C. Anela; Polovina, Jeffrey J.; Fulton, Elizabeth A.

2018-01-01

Empirical data on food web dynamics and predator-prey interactions underpin ecosystem models, which are increasingly used to support strategic management of marine resources. These data have traditionally derived from stomach content analysis, but new and complementary forms of ecological data are increasingly available from biochemical tracer techniques. Extensive opportunities exist to improve the empirical robustness of ecosystem models through the incorporation of biochemical tracer data and derived indices, an area that is rapidly expanding because of advances in analytical developments and sophisticated statistical techniques. Here, we explore the trophic information required by ecosystem model frameworks (species, individual, and size based) and match them to the most commonly used biochemical tracers (bulk tissue and compound-specific stable isotopes, fatty acids, and trace elements). Key quantitative parameters derived from biochemical tracers include estimates of diet composition, niche width, and trophic position. Biochemical tracers also provide powerful insight into the spatial and temporal variability of food web structure and the characterization of dominant basal and microbial food web groups. A major challenge in incorporating biochemical tracer data into ecosystem models is scale and data type mismatches, which can be overcome with greater knowledge exchange and numerical approaches that transform, integrate, and visualize data.

K Basin Sandfilter Backwash Line Characterization Project and Analytical Results for Campaign 24

International Nuclear Information System (INIS)

STEEN, F.H.

1999-01-01

Sample 203KWBMF was taken from the K West Sandfilter Backwash Pit on November 4, 1999 and received by 2224 Laboratory on November 4, 1999. Analyses were performed on sample 203KWBMF in accordance with ''Letter of Instruction for K Basins sandfilter Backwash Line Samples'' (LOI) in support of the K Basin Sandfilter Backwash Line Characterization Project

Biochemical characterization of systemic bacteria in bananas, sensitivity to antibiotics and plant phytotoxicity during shoot proliferation

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Janiffe Peres de Oliveira

2016-04-01

Full Text Available The objective of this work was to characterize the biochemically systemic bacterial isolated from banana plants, to evaluate the bacterial sensitivity to antibiotics, and to determine the phytotoxicity of banana shoots during in vitro proliferation. Systemic bacteria belonging to the Klebsiella and Aeromonas genera were isolated from the “Maravilha” (FHIA 01 AAAB, “Preciosa” (PV 4285 AAAB and “Thap Maeo” (AAB varieties and were then characterized. Tests of shoot sensitivity to antibiotics were performed, and the minimum inhibitory concentration (MIC and phytotoxic effects of selected antibiotics to plants were determined. Among the 20 antibiotics evaluated, the strains showed sensitivity to cefaclor, cefalexin, cefalotin, nalidixic acid, chloramphenicol, and vancomycin. However, during MIC determination, the best results were obtained with cefaclor, vancomycin or nalidixic acid alone in concentrations ranging from 512 to 1,024 mg L-1. In culture medium, cefaclor at 1,024 mg L-1 was the only antibiotic to affect the multiplication and the shoot survival in culture.

Biochemical Characterization of an In-House Coccidioides Antigen: Perspectives for the Immunodiagnosis of Coccidioidomycosis

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Renato Evando Moreira Filho

2012-06-01

Full Text Available The objective of this study was to evaluate the reactivity of an in-house antigen, extracted from a strain of C. posadasii isolated in northeastern Brazil, by radial immunodiffusion and Western blotting, as well as to establish its biochemical characterization. The protein antigen was initially extracted with the use of solid ammonium sulfate and characterized by 1-D electrophoresis. Subsequently, it was tested by means of double radial immunodiffusion and Western blotting. A positive reaction was observed against the antigen by both immunodiagnostic techniques tested on sera from patients suffering from coccidioidomycosis. Besides this, two immunoreactive protein bands were observed and were revealed to be a β-glucosidase and a glutamine synthetase after sequencing of the respective N-terminal regions. Our in-house Coccidioides antigen can be promising as a quick and low-cost diagnostic tool without the risk of direct manipulation of the microorganism.

Molecular and biochemical characterizations of human oral lactobacilli as putative probiotic candidates.

Science.gov (United States)

Strahinic, I; Busarcevic, M; Pavlica, D; Milasin, J; Golic, N; Topisirovic, L

2007-04-01

The objective of this study was to characterize the lactobacilli from the human oral cavity as a potential source of probiotic strains. Samples were collected from four different locations within the oral cavity: surface of healthy tooth, oral mucous membrane, surface of tooth decay and deep tooth decay. On the basis of morphological and biochemical properties eight categories were formed and 26 isolates were selected for further characterization. The isolates were determined as Lactobacillus sp. using primers specific for 16S rDNA. Sequencing of 16S rDNA genes and repetitive sequence-based polymerase chain reactions were used for determination to species and subspecies levels. Predominant species were Lactobacillus fermentum, Lactobacillus plantarum, Lactobacillus salivarius and Lactobacillus paracasei subsp. paracasei, while Lactobacillus acidophilus, Lactobacillus cellobiosus, Lactobacillus delbrueckii subsp. lactis and Lactobacillus gasseri were also present. The isolates Lactobacillus salivarius BGHO1, Lactobacillus fermentum BGHO36 and BGHO64, Lactobacillus gasseri BGHO89 and Lactobacillus delbrueckii subsp. lactis BGHO99 exhibited antagonistic action on the growth of Staphylococcus aureus, Enterococcus faecalis, Micrococcus flavus, Salmonella enteritidis, Streptococcus pneumoniae and Streptococcus mutans, but not on growth of Candida albicans. Moreover, the isolates L. salivarius BGHO1 and L. gasseri BGHO89 were tolerant to low pH and high concentration of bile salts. Taken together, these findings imply that L. salivarius BGHO1 and L. gasseri BGHO89 might be subjects for additional investigation as potential probiotic strains.

Establishment and characterization of a novel head and neck squamous cell carcinoma cell line USC-HN1.

Science.gov (United States)

Liebertz, Daniel J; Lechner, Melissa G; Masood, Rizwan; Sinha, Uttam K; Han, Jing; Puri, Raj K; Correa, Adrian J; Epstein, Alan L

2010-02-22

Head and neck squamous cell carcinoma (HNSCC) is an aggressive and lethal malignancy. Publically available cell lines are mostly of lingual origin, or have not been carefully characterized. Detailed characterization of novel HNSCC cell lines is needed in order to provide researchers a concrete keystone on which to build their investigations. The USC-HN1 cell line was established from a primary maxillary HNSCC biopsy explant in tissue culture. The immortalized cells were then further characterized by heterotransplantation in Nude mice; immunohistochemical staining for relevant HNSCC biomarkers; flow cytometry for surface markers; cytogenetic karyotypic analysis; human papillomavirus and Epstein-Barr virus screening; qRT-PCR for oncogene and cytokine analysis; investigation of activated, cleaved Notch1 levels; and detailed 35,000 gene microarray analysis. Characterization experiments confirmed the human HNSCC origin of USC-HN1, including a phenotype similar to the original tumor. Viral screening revealed no HPV or EBV infection, while western blotting displayed significant upregulation of activated, cleaved Notch1. USC-HN1, a novel immortalized cell line has been derived from a maxillary HNSCC. Characterization studies have shown that the cell line is of HNSCC origin and displays many of the same markers previously reported in the literature. USC-HN1 is available for public research and will further the investigation of HNSCC and the development of new therapeutic modalities.

Fragrance analysis using molecular and biochemical methods in ...

African Journals Online (AJOL)

For molecular and biochemical analysis of aroma, a mapping population comprising 208 recombinant inbred lines (RILs) derived from a diverse cross between CSR10 and Taraori Basmati through Single seed descent (SSD) method was used. RILs are among the best mapping populations, which provide a novel material ...

Isolation and characterization of active LINE and SINEs from the eel.

Science.gov (United States)

Kajikawa, Masaki; Ichiyanagi, Kenji; Tanaka, Nozomu; Okada, Norihiro

2005-03-01

Long interspersed elements (LINEs) and short interspersed elements (SINEs) are retrotransposons. These elements can mobilize by the "copy-and-paste" mechanism, in which their own RNA is reverse-transcribed into complementary DNA (cDNA). LINEs and SINEs not only are components of eukaryotic genomes but also drivers of genomic evolution. Thus, studies of the amplification mechanism of LINEs and SINEs are important for understanding eukaryotic genome evolution. Here we report the characterization of one LINE family (UnaL2) and two SINE families (UnaSINE1 and UnaSINE2) from the eel (Anguilla japonica) genome. UnaL2 is approximately 3.6 kilobases (kb) and encodes only one open reading frame (ORF). UnaL2 belongs to the stringent type--thought to be a major group of LINEs--and can mobilize in HeLa cells. We also show that UnaL2 and the two UnaSINEs have similar 3' tails, and that both UnaSINE1 and UnaSINE2 can be mobilized by UnaL2 in HeLa cells. These elements are thus useful for delineating the amplification mechanism of stringent type LINEs as well as that of SINEs.

Establishment and Biological Characterization of a Panel of Glioblastoma Multiforme (GBM) and GBM Variant Oncosphere Cell Lines.

Science.gov (United States)

Binder, Zev A; Wilson, Kelli M; Salmasi, Vafi; Orr, Brent A; Eberhart, Charles G; Siu, I-Mei; Lim, Michael; Weingart, Jon D; Quinones-Hinojosa, Alfredo; Bettegowda, Chetan; Kassam, Amin B; Olivi, Alessandro; Brem, Henry; Riggins, Gregory J; Gallia, Gary L

2016-01-01

Human tumor cell lines form the basis of the majority of present day laboratory cancer research. These models are vital to studying the molecular biology of tumors and preclinical testing of new therapies. When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity. Using a modified neural stem cell culture technique, here we report the characterization of GBM cell lines including GBM variants. Tumor tissue samples were obtained intra-operatively and cultured in neural stem cell conditions containing growth factors. Tumor lines were characterized in vitro using differentiation assays followed by immunostaining for lineage-specific markers. In vivo tumor formation was assayed by orthotopic injection in nude mice. Genetic uniqueness was confirmed via short tandem repeat (STR) DNA profiling. Thirteen oncosphere lines derived from GBM and GBM variants, including a GBM with PNET features and a GBM with oligodendroglioma component, were established. All unique lines showed distinct genetic profiles by STR profiling. The lines assayed demonstrated a range of in vitro growth rates. Multipotency was confirmed using in vitro differentiation. Tumor formation demonstrated histologic features consistent with high grade gliomas, including invasion, necrosis, abnormal vascularization, and high mitotic rate. Xenografts derived from the GBM variants maintained histopathological features of the primary tumors. We have generated and characterized GBM suspension lines derived from patients with GBMs and GBM variants. These oncosphere cell lines will expand the resources available for preclinical study.

Physicochemical and biochemical characterization of biosurfactants released by Lactobacillus strains

NARCIS (Netherlands)

Velraeds, MMC; vanderMei, HC; Reid, G; Busscher, HJ

1996-01-01

Biosurfactants from Lactobacillus casei subsp. rhamnosus 36 and ATCC 7469, Lactobacillus fermentum B54 and Lactobacillus acidophilus RC14 were isolated from bacteria in their mid-exponential (4-5 h) and stationary growth phases (18 h) and physicochemical and biochemical properties of the

Simulation studies in biochemical signaling and enzyme reactions

Science.gov (United States)

Nelatury, Sudarshan R.; Vagula, Mary C.

2014-06-01

Biochemical pathways characterize various biochemical reaction schemes that involve a set of species and the manner in which they are connected. Determination of schematics that represent these pathways is an important task in understanding metabolism and signal transduction. Examples of these Pathways are: DNA and protein synthesis, and production of several macro-molecules essential for cell survival. A sustained feedback mechanism arises in gene expression and production of mRNA that lead to protein synthesis if the protein so synthesized serves as a transcription factor and becomes a repressor of the gene expression. The cellular regulations are carried out through biochemical networks consisting of reactions and regulatory proteins. Systems biology is a relatively new area that attempts to describe the biochemical pathways analytically and develop reliable mathematical models for the pathways. A complete understanding of chemical reaction kinetics is prohibitively hard thanks to the nonlinear and highly complex mechanisms that regulate protein formation, but attempting to numerically solve some of the governing differential equations seems to offer significant insight about their biochemical picture. To validate these models, one can perform simple experiments in the lab. This paper introduces fundamental ideas in biochemical signaling and attempts to take first steps into the understanding of biochemical oscillations. Initially, the two-pool model of calcium is used to describe the dynamics behind the oscillations. Later we present some elementary results showing biochemical oscillations arising from solving differential equations of Elowitz and Leibler using MATLAB software.

Biochem-Env, a plateform of environmental biochemistry for research

OpenAIRE

GRONDIN, VIRGINIE; Nelieu, Sylvie; Crouzet, Olivier; Hedde, Mickaël; Mougin, Christian

2016-01-01

As a service of the research infrastructure AnaEE-France (http://www.anaee-france.fr/fr/), the platform Biochem-Env (http://www.biochemenv.fr) offers skills and innovative analytical tools for biochemical characterizations of soils, sediments, and micro-macro-organisms living in terrestrial and aquatic ecosystems. The platform provides methods validated according to Quality Guidelines, i.e. to measure global soil enzymatic activities. Our robot-supported protocols allow great number of enzyme...

Extracellular vesicles secreted from cancer cell lines stimulate secretion of MMP-9, IL-6, TGF-β1 and EMMPRIN.

Directory of Open Access Journals (Sweden)

Jasmina S Redzic

Full Text Available Extracellular vesicles (EVs are key contributors to cancer where they play an integral role in cell-cell communication and transfer pro-oncogenic molecules to recipient cells thereby conferring a cancerous phenotype. Here, we purified EVs using straightforward biochemical approaches from multiple cancer cell lines and subsequently characterized these EVs via multiple biochemical and biophysical methods. In addition, we used fluorescence microscopy to directly show internalization of EVs into the recipient cells within a few minutes upon addition of EVs to recipient cells. We confirmed that the transmembrane protein EMMPRIN, postulated to be a marker of EVs, was indeed secreted from all cell lines studied here. We evaluated the response to EV stimulation in several different types of recipient cells lines and measured the ability of these purified EVs to induce secretion of several factors highly upregulated in human cancers. Our data indicate that purified EVs preferentially stimulate secretion of several proteins implicated in driving cancer in monocytic cells but only harbor limited activity in epithelial cells. Specifically, we show that EVs are potent stimulators of MMP-9, IL-6, TGF-β1 and induce the secretion of extracellular EMMPRIN, which all play a role in driving immune evasion, invasion and inflammation in the tumor microenvironment. Thus, by using a comprehensive approach that includes biochemical, biological, and spectroscopic methods, we have begun to elucidate the stimulatory roles.

Molecular and Functional Characterization of GR2-R1 Event Based Backcross Derived Lines of Golden Rice in the Genetic Background of a Mega Rice Variety Swarna.

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Haritha Bollinedi

Full Text Available Homozygous Golden Rice lines developed in the background of Swarna through marker assisted backcross breeding (MABB using transgenic GR2-R1 event as a donor for the provitamin A trait have high levels of provitamin A (up to 20 ppm but are dwarf with pale green leaves and drastically reduced panicle size, grain number and yield as compared to the recurrent parent, Swarna. In this study, we carried out detailed morphological, biochemical and molecular characterization of these lines in a quest to identify the probable reasons for their abnormal phenotype. Nucleotide blast analysis with the primer sequences used to amplify the transgene revealed that the integration of transgene disrupted the native OsAux1 gene, which codes for an auxin transmembrane transporter protein. Real time expression analysis of the transgenes (ZmPsy and CrtI driven by endosperm-specific promoter revealed the leaky expression of the transgene in the vegetative tissues. We propose that the disruption of OsAux1 disturbed the fine balance of plant growth regulators viz., auxins, gibberellic acid and abscisic acid, leading to the abnormalities in the growth and development of the lines homozygous for the transgene. The study demonstrates the conserved roles of OsAux1 gene in rice and Arabidopsis.

Characterization of the RAPD for 6 Durum wheat lines (Triticum durum desf.) selected from M4-irradiated population under drought conditions

International Nuclear Information System (INIS)

Kalil, M.K.; Nesiem, M.R.A.; Kassem, M.K.M.; Basyouny, M.A.E.

2012-01-01

Grains of two durum wheat cultivars Sohag 3 and Beni Suef 3 were irradiated with different doses of gamma ray 0, 150, 250 and 350 Gy to obtain new durum wheat lines characterized by high yielding and drought tolerance. Irradiated grins were cultivated in the field under normal and drought conditions during 2005-2009 Results of field experiments showed that there were new six putative lines S1, S2, S3, S4, B1 and B2. Each putative line had superiority than its parent in grain yield / plant. The putative lines S1 and S2 had superiority over their parent Sohag 3 under normal conditions in grain yield per plant this increase equal 52 and 60% respectively. The putative lines S3 and S4 had superiority in grain yield per plan as compared to Sohag 3 under drought conditions this increase equal 75 to 58% respectively. The putative lines B1 and B2 had superiority in grain yield per plant than their parent Beni Suef 3 under normal condition this increase equal 46 and 12% respectively. Results for RAPD markers showed that each putative line was characterized by positive and negative unique marker. The putative line S1: characterized by four negative marker amplified by OPM-05, OPN-04, OPA-18 and OPB-12 primers. The putative line S2: characterized by one negative unique marker amplified by OPQ-14 marker. The putative line S3: characterized by two positive markers amplified by OPB-07 and OPG-12 markers and one negative unique marker amplified by OPA-10 marker. The putative line S4: characterized by four negative markers amplified by OPM-05, OPN-13, OPQ-12 and OPQ-14 markers and one positive unique marker amplified by OPC-05 marker. The putative line B1: characterized by four positive markers amplified by OPA-10, OPG-12, OPB-07 and OPA-18 markers and three negative markers amplified with OPM-05, OPC-05 and OPB-12 markers. The putative line B2: characterized by three positive unique marker amplified by OPB-07, OPN-04, OPN-10 markers and four negative markers amplified with OPA-10

Biochemical and biophysical characterization of the transmissible gastroenteritis coronavirus fusion core

International Nuclear Information System (INIS)

Ma Guangpeng; Feng Youjun; Gao Feng; Wang Jinzi; Liu Cheng; Li Yijing

2005-01-01

Transmissible gastroenteritis coronavirus (TGEV) is one of the most destructive agents, responsible for the enteric infections that are lethal for suckling piglets, causing enormous economic loss to the porcine fostering industry every year. Although it has been known that TGEV spiker protein is essential for the viral entry for many years, the detail knowledge of the TGEV fusion protein core is still very limited. Here, we report that TGEV fusion core (HR1-SGGRGG-HR2), in vitro expressed in GST prokaryotic expression system, shares the typical properties of the trimer of coiled-coil heterodimer (six α-helix bundle), which has been confirmed by a combined series of biochemical and biophysical evidences including size exclusion chromatography (gel-filtration), chemical crossing, and circular diagram. The 3D homologous structure model presents its most likely structure, extremely similar to those of the coronaviruses documented. Taken together, TGEV spiker protein belongs to the class I fusion protein, characterized by the existence of two heptad-repeat (HR) regions, HR1 and HR2, and the present knowledge about the truncated TGEV fusion protein core may facilitate in the design of the small molecule or polypeptide drugs targeting the membrane fusion between TGEV and its host

Biochemical Characterization of CPS-1, a Subclass B3 Metallo-β-Lactamase from a Chryseobacterium piscium Soil Isolate

DEFF Research Database (Denmark)

Gudeta, Dereje Dadi; Pollini, Simona; Docquier, Jean-Denis

2016-01-01

CPS-1 is a subclass B3 metallo-β-lactamase from a Chryseobacterium piscium isolated from soil, showing 68 % amino acid identity to GOB-1 enzyme. CPS-1 was overproduced in Escherichia coli Rosetta (DE3), purified by chromatography and biochemically characterized. This enzyme exhibits a broad spect...... spectrum substrate profile including penicillins, cephalosporins and carbapenems, which overall resembles those of L1, GOB-1 and acquired subclass B3 enzymes AIM-1 and SMB-1....

Purification and Biochemical Characterization of Three Myotoxins from Bothrops mattogrossensis Snake Venom with Toxicity against Leishmania and Tumor Cells

Directory of Open Access Journals (Sweden)

Andréa A. de Moura

2014-01-01

Full Text Available Bothrops mattogrossensis snake is widely distributed throughout eastern South America and is responsible for snakebites in this region. This paper reports the purification and biochemical characterization of three new phospholipases A2 (PLA2s, one of which is presumably an enzymatically active Asp49 and two are very likely enzymatically inactive Lys49 PLA2 homologues. The purification was obtained after two chromatographic steps on ion exchange and reverse phase column. The 2D SDS-PAGE analysis revealed that the proteins have pI values around 10, are each made of a single chain, and have molecular masses near 13 kDa, which was confirmed by MALDI-TOF mass spectrometry. The N-terminal similarity analysis of the sequences showed that the proteins are highly homologous with other Lys49 and Asp49 PLA2s from Bothrops species. The PLA2s isolated were named BmatTX-I (Lys49 PLA2-like, BmatTX-II (Lys49 PLA2-like, and BmatTX-III (Asp49 PLA2. The PLA2s induced cytokine release from mouse neutrophils and showed cytotoxicity towards JURKAT (leukemia T and SK-BR-3 (breast adenocarcinoma cell lines and promastigote forms of Leishmania amazonensis. The structural and functional elucidation of snake venoms components may contribute to a better understanding of the mechanism of action of these proteins during envenomation and their potential pharmacological and therapeutic applications.

Generation and Characterization of an Immortalized Human Esophageal Myofibroblast Line.

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Chao Niu

Full Text Available Stromal cells with a myofibroblast phenotype present in the normal human esophagus are increased in individuals with gastro-esophageal reflux disease (GERD. We have previously demonstrated that myofibroblasts stimulated with acid and TLR4 agonists increase IL-6 and IL-8 secretion using primary cultures of myofibroblasts established from normal human esophagus. While primary cultures have the advantage of reflecting the in vivo environment, a short life span and unavoidable heterogeneity limits the usefulness of this model in larger scale in vitro cellular signaling studies. The major aim of this paper therefore was to generate a human esophageal myofibroblast line with an extended lifespan. In the work presented here we have generated and characterized an immortalized human esophageal myofibroblast line by transfection with a commercially available GFP-hTERT lentivirus. Immortalized human esophageal myofibroblasts demonstrate phenotypic, genotypic and functional similarity to primary cultures of esophageal myofibroblasts we have previously described. We found that immortalized esophageal myofibroblasts retain myofibroblast spindle-shaped morphology at low and high confluence beyond passage 80, and express α-SMA, vimentin, and CD90 myofibroblast markers. Immortalized human esophageal myofibroblasts also express the putative acid receptor TRPV1 and TLR4 and retain the functional capacity to respond to stimuli encountered in GERD with secretion of IL-6. Finally, immortalized human esophageal myofibroblasts also support the stratified growth of squamous esophageal epithelial cells in 3D organotypic cultures. This newly characterized immortalized human esophageal myofibroblast cell line can be used in future cellular signaling and co-culture studies.

Biochemical and functional characterization of Bothropoidin: the first haemorrhagic metalloproteinase from Bothrops pauloensis snake venom.

Science.gov (United States)

Gomes, Mário Sérgio R; Naves de Souza, Dayane L; Guimarães, Denise O; Lopes, Daiana S; Mamede, Carla C N; Gimenes, Sarah Natalie C; Achê, David C; Rodrigues, Renata S; Yoneyama, Kelly A G; Borges, Márcia H; de Oliveira, Fábio; Rodrigues, Veridiana M

2015-03-01

We present the biochemical and functional characterization of Bothropoidin, the first haemorrhagic metalloproteinase isolated from Bothrops pauloensis snake venom. This protein was purified after three chromatographic steps on cation exchange CM-Sepharose fast flow, size-exclusion column Sephacryl S-300 and anion exchange Capto Q. Bothropoidin was homogeneous by SDS-PAGE under reducing and non-reducing conditions, and comprised a single chain of 49,558 Da according to MALDI TOF analysis. The protein presented an isoelectric point of 3.76, and the sequence of six fragments obtained by MS (MALDI TOF\\TOF) showed a significant score when compared with other PIII Snake venom metalloproteinases (SVMPs). Bothropoidin showed proteolytic activity on azocasein, Aα-chain of fibrinogen, fibrin, collagen and fibronectin. The enzyme was stable at pH 6-9 and at lower temperatures when assayed on azocasein. Moreover, its activity was inhibited by EDTA, 1.10-phenanthroline and β-mercaptoethanol. Bothropoidin induced haemorrhage [minimum haemorrhagic dose (MHD) = 0.75 µg], inhibited platelet aggregation induced by collagen and ADP, and interfered with viability and cell adhesion when incubated with endothelial cells in a dose and time-dependent manner. Our results showed that Bothropoidin is a haemorrhagic metalloproteinase that can play an important role in the toxicity of B. pauloensis envenomation and might be used as a tool for studying the effects of SVMPs on haemostatic disorders and tumour metastasis. © The Authors 2014. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Cohnella amylopullulanases: Biochemical characterization of two recombinant thermophilic enzymes.

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Fatemeh Zebardast Roodi

Full Text Available Some industries require newer, more efficient recombinant enzymes to accelerate their ongoing biochemical reactions in harsh environments with less replenishment. Thus, the search for native enzymes from extremophiles that are suitable for use under industrial conditions is a permanent challenge for R & D departments. Here and toward such discoveries, two sequences homologous to amylopullulanases (EC 3.2.1.41, GH57 from an endogenous Cohnella sp., [Coh00831 (KP335161; 1998 bp and Coh01133 (KP335160: 3678 bp] were identified. The genes were heterologously expressed in E. coli to both determine their type and further characterize their properties. The isolated DNA was PCR amplified with gene specific primers and cloned in pET28a, and the recombinant proteins were expressed in E. coli BL21 (DE3. The temperatures and pH optima of purified recombinants Coh 01133 and Coh 00831 enzymes were 70°C and 8, and 60°C and 6, respectively. These enzymes are stable more than 90% in 60°C and 50°C for 90 min respectively. The major reactions released sugars which could be fractionated by HPLC analysis, from soluble starch were mainly maltose (G2, maltotriose (G3 and maltotetraose (G4. The enzymes hydrolyzed pullulan to maltotriose (G3 only. Enzyme activities for both proteins were improved in the availability of Mn2+, Ba2+, Ca2+, and Mg2+ and reduced in the presence of Fe2+, Li2+, Na2+, Triton X100 and urea. Moreover, Co2+, K+, and Cu2+ had a negative effect only on Coh 01133 enzyme.

Cohnella amylopullulanases: Biochemical characterization of two recombinant thermophilic enzymes.

Science.gov (United States)

Zebardast Roodi, Fatemeh; Aminzadeh, Saeed; Farrokhi, Naser; Karkhane, AliAsghar; Haghbeen, Kamahldin

2017-01-01

Some industries require newer, more efficient recombinant enzymes to accelerate their ongoing biochemical reactions in harsh environments with less replenishment. Thus, the search for native enzymes from extremophiles that are suitable for use under industrial conditions is a permanent challenge for R & D departments. Here and toward such discoveries, two sequences homologous to amylopullulanases (EC 3.2.1.41, GH57) from an endogenous Cohnella sp., [Coh00831 (KP335161; 1998 bp) and Coh01133 (KP335160: 3678 bp)] were identified. The genes were heterologously expressed in E. coli to both determine their type and further characterize their properties. The isolated DNA was PCR amplified with gene specific primers and cloned in pET28a, and the recombinant proteins were expressed in E. coli BL21 (DE3). The temperatures and pH optima of purified recombinants Coh 01133 and Coh 00831 enzymes were 70°C and 8, and 60°C and 6, respectively. These enzymes are stable more than 90% in 60°C and 50°C for 90 min respectively. The major reactions released sugars which could be fractionated by HPLC analysis, from soluble starch were mainly maltose (G2), maltotriose (G3) and maltotetraose (G4). The enzymes hydrolyzed pullulan to maltotriose (G3) only. Enzyme activities for both proteins were improved in the availability of Mn2+, Ba2+, Ca2+, and Mg2+ and reduced in the presence of Fe2+, Li2+, Na2+, Triton X100 and urea. Moreover, Co2+, K+, and Cu2+ had a negative effect only on Coh 01133 enzyme.

Characterization of inorganic phosphate transport in the triple-negative breast cancer cell line, MDA-MB-231.

Science.gov (United States)

Russo-Abrahão, Thais; Lacerda-Abreu, Marco Antônio; Gomes, Tainá; Cosentino-Gomes, Daniela; Carvalho-de-Araújo, Ayra Diandra; Rodrigues, Mariana Figueiredo; Oliveira, Ana Carolina Leal de; Rumjanek, Franklin David; Monteiro, Robson de Queiroz; Meyer-Fernandes, José Roberto

2018-01-01

Recent studies demonstrate that interstitial inorganic phosphate is significantly elevated in the breast cancer microenvironment as compared to normal tissue. In addition it has been shown that breast cancer cells express high levels of the NaPi-IIb carrier (SLC34A2), suggesting that this carrier may play a role in breast cancer progression. However, the biochemical behavior of inorganic phosphate (Pi) transporter in this cancer type remains elusive. In this work, we characterize the kinetic parameters of Pi transport in the aggressive human breast cancer cell line, MDA-MB-231, and correlated Pi transport with cell migration and adhesion. We determined the influence of sodium concentration, pH, metabolic inhibitors, as well as the affinity for inorganic phosphate in Pi transport. We observed that the inorganic phosphate is dependent on sodium transport (K0,5 value = 21.98 mM for NaCl). Furthermore, the transport is modulated by different pH values and increasing concentrations of Pi, following the Michaelis-Menten kinetics (K0,5 = 0.08 mM Pi). PFA, monensin, furosemide and ouabain inhibited Pi transport, cell migration and adhesion. Taken together, these results showed that the uptake of Pi in MDA-MB-231 cells is modulated by sodium and by regulatory mechanisms of intracellular sodium gradient. General Significance: Pi transport might be regarded as a potential target for therapy against tumor progression.

Characterizing performances of solder paste printing process at flexible manufacturing lines

International Nuclear Information System (INIS)

Siew, Jit Ping; Low, Heng Chin; Teoh, Ping Chow

2015-01-01

Solder paste printing (SPP) has been a challenge on printed circuit board (PCB) manufacturing, evident by the proliferation of solder paste inspection equipment, or substituted by rigorous non-value added activity of manual inspections. The objective of this study is to characterize the SPP performance of various products manufactured in flexible production lines with different equipment configurations, and determine areas for process improvement. The study began by collecting information on SPP performance relative to component placement (CP) process, and to the proportion of mixed products. Using a clustering algorithm to group similar elements together, SPP performance across all product-production line pairs are statistically modeled to discover the trend and the influential factors. The main findings are: (a) Ratio of overall dpku for CP and SPP processes are 2:1; (b) logistic regression models of SPP performance indicated that only effects of product-production line and solder paste printer configuration are significant; (c) PCB circuitry design with BGA components and single solder paste printer line configurations generated the highest monthly defects, with the highest variation in the latter

Characterizing performances of solder paste printing process at flexible manufacturing lines

Energy Technology Data Exchange (ETDEWEB)

Siew, Jit Ping; Low, Heng Chin [University of Science Malaysia, 11800 Minden, Penang (Malaysia); Teoh, Ping Chow [Wawasan Open University, 54 Jalan Sultan Ahmad Shah, 10050 Penang (Malaysia)

2015-02-03

Solder paste printing (SPP) has been a challenge on printed circuit board (PCB) manufacturing, evident by the proliferation of solder paste inspection equipment, or substituted by rigorous non-value added activity of manual inspections. The objective of this study is to characterize the SPP performance of various products manufactured in flexible production lines with different equipment configurations, and determine areas for process improvement. The study began by collecting information on SPP performance relative to component placement (CP) process, and to the proportion of mixed products. Using a clustering algorithm to group similar elements together, SPP performance across all product-production line pairs are statistically modeled to discover the trend and the influential factors. The main findings are: (a) Ratio of overall dpku for CP and SPP processes are 2:1; (b) logistic regression models of SPP performance indicated that only effects of product-production line and solder paste printer configuration are significant; (c) PCB circuitry design with BGA components and single solder paste printer line configurations generated the highest monthly defects, with the highest variation in the latter.

Biochemical Characterization of Bovine Brain Myristoyl-CoA:Protein N-Myristoyltransferase Type 2

Directory of Open Access Journals (Sweden)

Ponniah Selvakumar

2009-01-01

Full Text Available Protein N-myristoylation is a lipidic modification which refers to the covalent attachment of myristate, a 14-carbon saturated fatty acid, to the N-terminal glycine residue of a number of mammalian, viral, and fungal proteins. In this paper, we have cloned the gene coding for myristoyl-CoA:protein N-myristoyltransferase (NMT from Bos tarus brain. The open reading frame codes for a 410-amino-acid protein and overexpressed in Escherichia coli. Kinetic studies suggested that bovine brain NMT2 and human NMT1 show significant differences in their peptide substrate specificities. The metal ion Ca2+ had stimulatory effects on NMT2 activity while Mn2+ and Zn2+ inhibited the enzyme activity. In addition, NMT2 activity was inhibited by various organic solvents and other detergents while NMT1 had a stimulatory effect. Biochemical characterization suggested that both forms of NMT have unique characteristics. Further analysis towards functional role NMT2 will lead the development of therapeutic target for the progression of various diseases such as cancer, cardiovascular diseases, and neurodegenerative diseases.

Derivation and Osmotolerance Characterization of Three Immortalized Tilapia (Oreochromis mossambicus) Cell Lines

Science.gov (United States)

Gardell, Alison M.; Qin, Qin; Rice, Robert H.; Li, Johnathan; Kültz, Dietmar

2014-01-01

Fish cell cultures are becoming more widely used models for investigating molecular mechanisms of physiological response to environmental challenge. In this study, we derived two immortalized Mozambique tilapia (Oreochromis mossambicus) cell lines from brain (OmB) and lip epithelium (OmL), and compared them to a previously immortalized bulbus arteriosus (TmB) cell line. The OmB and OmL cell lines were generated without or with Rho-associated kinase (ROCK) inhibitor/3T3 feeder layer supplementation. Although both approaches were successful, ROCK inhibitor/feeder layer supplementation was found to offer the advantages of selecting for epithelial-like cell type and decreasing time to immortalization. After immortalization (≥ passage 5), we characterized the proteomes of the newly derived cell lines (OmB and OmL) using LCMS and identified several unique cell markers for each line. Subsequently, osmotolerance for each of the three cell lines following acute exposure to elevated sodium chloride was evaluated. The acute maximum osmotolerance of these tilapia cell lines (>700 mOsm/kg) was markedly higher than that of any other known vertebrate cell line, but was significantly higher in the epithelial-like OmL cell line. To validate the physiological relevance of these tilapia cell lines, we quantified the effects of acute hyperosmotic challenge (450 mOsm/kg and 700 mOsm/kg) on the transcriptional regulation of two enzymes involved in biosynthesis of the compatible organic osmolyte, myo-inositol. Both enzymes were found to be robustly upregulated in all three tilapia cell lines. Therefore, the newly established tilapia cells lines represent valuable tools for studying molecular mechanisms involved in the osmotic stress response of euryhaline fish. PMID:24797371

Variation of several agronomic and biochemical traits in γ-ray induced mutant soybeans

International Nuclear Information System (INIS)

Shim, Kyo Moon; Kim, Sun Hyung; Kim, Nam Soo; Son, Hi Sup; Rhee, Hae Ik

1996-01-01

Two soybean cultivars(Paldalkong and Bangsakong) were irradiated with gamma-ray to reduce seed size, which is an important trait for soybean sprout and the derived mutant soybeans were analyzed in several agronomic and biochemical traits. There were high levels of variations in quantitative traits among the mutants. Several mutant lines showed higher yield and smaller seed than their parents. Qualitative traits such as seed coat color or pubescent color were also changed in a few lines. Biochemical variations were also observed among the mutants. In seed storage protein analysis, many mutant lines showed reduced intensities in β-subunits in 7S globulin than their parents and an additional band in the acidic subunit at 31KDa. Two mutant lines derived from Paldalkong showed an additional band and a shifted band of protease inhibitor by electrophoretic analysis. Variations in isozymes and RAPD were also observed among the mutants. Six isozymes(Adh, Est, Gdh, Idh, Mdh and Pgm) among eleven isozymes showed some variations and six out of ten primers showed polymorphic amplified DNA fragments among the mutants. (author)

Establishment, characterization and chemosensitivity of three mismatch repair deficient cell lines from sporadic and inherited colorectal carcinomas.

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Claudia Maletzki

Full Text Available BACKGROUND: Colorectal cancer (CRC represents a morphologic and molecular heterogenic disease. This heterogeneity substantially impairs drug effectiveness and prognosis. The subtype of mismatch repair deficient (MMR-D CRCs, accounting for about 15% of all cases, shows particular differential responses up to resistance towards currently approved cytostatic drugs. Pre-clinical in vitro models representing molecular features of MMR-D tumors are thus mandatory for identifying biomarkers that finally help to predict responses towards new cytostatic drugs. Here, we describe the successful establishment and characterization of three patient-derived MMR-D cell lines (HROC24, HROC87, and HROC113 along with their corresponding xenografts. METHODOLOGY: MMR-D cell lines (HROC24, HROC87, and HROC113 were established from a total of ten clinicopathological well-defined MMR-D cases (120 CRC cases in total. Cells were comprehensively characterized by phenotype, morphology, growth kinetics, invasiveness, and molecular profile. Additionally, response to clinically relevant chemotherapeutics was examined in vitro and in vivo. PRINCIPAL FINDINGS: Two MMR-D lines showing CIMP-H derived from sporadic CRC (HROC24: K-ras(wt, B-raf(mut, HROC87: K-ras(wt, B-raf(mut, whereas the HROC113 cell line (K-ras(mut, B-raf(wt was HNPCC-associated. A diploid DNA-status could be verified by flow cytometry and SNP Array analysis. All cell lines were characterized as epithelial (EpCAM(+ tumor cells, showing surface tumor marker expression (CEACAM(+. MHC-class II was inducible by Interferon-γ stimulation. Growth kinetics as well as invasive potential was quite heterogeneous between individual lines. Besides, MMR-D cell lines exhibited distinct responsiveness towards chemotherapeutics, even when comparing in vitro and in vivo sensitivity. CONCLUSIONS: These newly established and well-characterized, low-passage MMR-D cell lines provide a useful tool for future investigations on the

Biochemical characterization of legume seeds as ingredients in animal feed

Energy Technology Data Exchange (ETDEWEB)

Martín-Pedrosa, M.; Varela, A.; Guillamon, E.; Cabellos, B.; Burbano, C.; Gomez-Fernandez, J.; Mercado, E. de; Gomez-Izquierdo, E.; Cuadrado, C.; Muzquiz, M.

2016-11-01

The current European protein deficit is estimated as high as 70% of present needs. Because of the high protein content of their seeds, grain legumes are attractive candidates for lowering the deficiency in plant protein production. The objective of this work was to identify new sources of vegetable protein that would reduce our high dependence of soy, the main source of protein in the manufacture of feedstuffs. To achieve this goal, we determined the proximate composition, the bioactive components, as well as the antinutritional factors present in the studied seeds. In general, the protein, fat and carbohydrates content of legume seeds studied were within the limits found in the literature. The bioactive compounds detected in all the seeds were α-galactosides, myoinositol phosphates, protease inhibitors and phenols. IP6 (phytic acid) was the main inositol phosphate form in all the samples. The highest protease inhibitors content was detected in both Lathyrus cicera cultivars. Vicia ervilia and L. cicera cultivars showed low haemagglutinating activity (20.4 HU/g). The γ-glutamyl-S-ethenyl-cysteine content in Vicia narbonensis was around 16.0 mg/g. Both L. cicera varieties presented similar β-N-oxalyl-L-α, β-diaminopropionic acid content (0.80 mg/g). The two V. ervilia varieties showed high canavanine concentration (1.93-5.28 mg/g). Vicine was only detected in V. narbonensis cultivars (0.3 mg/g). The biochemical characterization carried out in this study allows us to know the limits of inclusion of these minor crop seeds in feed formulations in order to replace the soybean. (Author)

Biochemical characterization of legume seeds as ingredients in animal feed

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Mercedes Martín-Pedrosa

2016-03-01

Full Text Available The current European protein deficit is estimated as high as 70% of present needs. Because of the high protein content of their seeds, grain legumes are attractive candidates for lowering the deficiency in plant protein production. The objective of this work was to identify new sources of vegetable protein that would reduce our high dependence of soy, the main source of protein in the manufacture of feedstuffs. To achieve this goal, we determined the proximate composition, the bioactive components, as well as the antinutritional factors present in the studied seeds. In general, the protein, fat and carbohydrates content of legume seeds studied were within the limits found in the literature. The bioactive compounds detected in all the seeds were α-galactosides, myoinositol phosphates, protease inhibitors and phenols. IP6 (phytic acid was the main inositol phosphate form in all the samples. The highest protease inhibitors content was detected in both Lathyrus cicera cultivars. Vicia ervilia and L. cicera cultivars showed low haemagglutinating activity (20.4 HU/g. The γ-glutamyl-S-ethenyl-cysteine content in Vicia narbonensis was around 16.0 mg/g. Both L. cicera varieties presented similar β-N-oxalyl-L-α, β-diaminopropionic acid content (0.80 mg/g. The two V. ervilia varieties showed high canavanine concentration (1.93-5.28 mg/g. Vicine was only detected in V. narbonensis cultivars (0.3 mg/g. The biochemical characterization carried out in this study allows us to know the limits of inclusion of these minor crop seeds in feed formulations in order to replace the soybean.

[Establishment and characterization of a cell line derived from human ovarian mucinous cystadenocarcinoma].

Science.gov (United States)

Wan, Q; Xu, D; Li, Z

2001-07-01

To establish a cell line of human ovarian cancer, and study its characterization. The cell line was established by the cultivation of subsides walls, and kept by freezing. The morphology was observed by microscope and electromicroscope. The authors studied its growth and propagation, the agglutination test of phytohemagglutinin (PHA), the chromosome analysis, heterotransplanting, immuno-histochemistry staining, the analysis of hormone, the pollution examination and the test of sensitivity to virus etc. A new human ovarian carcinoma cell line, designated ovarian mucinous cystadenocarcinoma 685 (OMC685), was established from mucinous cystadenocarcinoma. This cell line had subcultured to 91 generations, and some had been frozen for 8 years and revived, still grew well. This cell line possessed the feature of glandular epithelium cancer cell. The cells grew exuberantly, and the agglutinating test of PHA was positive. Karyotype was subtriploid with distortion. Heterotransplantations, alcian blue periobic acid-schiff (AbPAS), mucicarmine, alcian blue stainings, estradiol (E2) and progesterone were all positive. Without being polluted, it was sensitive to polivirus-I, adenovirus 7 and measles virus. OMC685 is a distinct human ovarian tumous cell line.

Genome-wide identification of the regulatory targets of a transcription factor using biochemical characterization and computational genomic analysis

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Jolly Emmitt R

2005-11-01

Full Text Available Abstract Background A major challenge in computational genomics is the development of methodologies that allow accurate genome-wide prediction of the regulatory targets of a transcription factor. We present a method for target identification that combines experimental characterization of binding requirements with computational genomic analysis. Results Our method identified potential target genes of the transcription factor Ndt80, a key transcriptional regulator involved in yeast sporulation, using the combined information of binding affinity, positional distribution, and conservation of the binding sites across multiple species. We have also developed a mathematical approach to compute the false positive rate and the total number of targets in the genome based on the multiple selection criteria. Conclusion We have shown that combining biochemical characterization and computational genomic analysis leads to accurate identification of the genome-wide targets of a transcription factor. The method can be extended to other transcription factors and can complement other genomic approaches to transcriptional regulation.

Molecular cloning and biochemical characterization of three phosphoglycerate kinase isoforms from developing sunflower (Helianthus annuus L.) seeds.

Science.gov (United States)

Troncoso-Ponce, M A; Rivoal, J; Venegas-Calerón, M; Dorion, S; Sánchez, R; Cejudo, F J; Garcés, R; Martínez-Force, E

2012-07-01

Three cDNAs encoding different phosphoglycerate kinase (PGK, EC 2.7.2.3) isoforms, two cytosolic (HacPGK1 and HacPGK2) and one plastidic (HapPGK), were cloned and characterized from developing sunflower (Helianthus annuus L.) seeds. The expression profiles of these genes showed differences in heterotrophic tissues, such as developing seeds and roots, where HacPGK1 was predominant, while HapPGK was highly expressed in photosynthetic tissues. The cDNAs were expressed in Escherichia coli, and the corresponding proteins purified to electrophoretic homogeneity, using immobilized metal ion affinity chromatography, and biochemically characterized. Despite the high level of identity between sequences, the HacPGK1 isoform showed strong differences in terms of specific activity, temperature stability and pH sensitivity in comparison to HacPGK2 and HapPGK. A polyclonal immune serum was raised against the purified HacPGK1 isoform, which showed cross-immunoreactivity with the other PGK isoforms. This serum allowed the localization of high expression levels of PGK isozymes in embryo tissues. Copyright © 2012 Elsevier Ltd. All rights reserved.

Establishment and characterization of a novel osteosarcoma cell line: CHOS.

Science.gov (United States)

Liu, Yunlu; Feng, Xiaobo; Zhang, Yukun; Jiang, Hongyan; Cai, Xianyi; Yan, Xinxin; Huang, Zengfa; Mo, Fengbo; Yang, Wen; Yang, Cao; Yang, Shuhua; Liu, Xianzhe

2016-12-01

Osteosarcoma has a well-recognized bimodal distribution, with the first peak in adolescence and another in the elderly age-group. The elderly patients have different clinical features and a poorer prognosis as compared to adolescents. To better understand the biological features of osteosarcoma in the elderly population, we established a new human osteosarcoma cell line from a 58-year-old man with primary chondroblastic osteosarcoma. After 6 months of continuous culture in vitro for over 50 passages, an immortalized cell line CHOS was established. The cell line was well-characterized by cytogenetic, biomarker, functional, and histological analyses. The CHOS cells exhibited a spindle-shaped morphology and a doubling time of 36 h. Cytogenetic analysis of CHOS cells revealed the loss of chromosome Y and the gain of chromosome 12. Quantitative real-time polymerase chain reaction (RT-PCR), Western blotting and/or immunofluorescence revealed the expression of chondroblastic, mesenchymal and tumor metastasis markers in the CHOS cells. Compared with the osteosarcoma cell line, the CHOS cells were found to be more sensitive to cisplatin and doxorubicin, but were resistant to methotrexate. The cell line was highly tumorigenic and maintained the histological characteristics and invasive nature of the original tumor. Furthermore, on immunohistochemical analysis, the xenografts and metastases were found to co-express collagen II, aggrecan, vimentin and S100A4 that resembled the original tumor cells. Our results indicate, the potential of CHOS cell line to serve as a useful tool for further studies on the molecular biology of osteosarcoma, especially in the elderly patients. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:2116-2125, 2016. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

Establishment and characterization of a new cell line derived from human colorectal laterally spreading tumor

Science.gov (United States)

Wang, Xin-Ying; Lai, Zhou-Sheng; Yeung, Chung-Man; Wang, Ji-De; Deng, Wen; Li, Hoi Yee; Han, Yu-Jing; Kung, Hsiang-Fu; Jiang, Bo; Lin, Marie Chia-mi

2008-01-01

AIM: To study the molecular mechanism of laterally spreading tumor (LST), a cell line [Laterally Spreading Tumor-Rectum 1 (LST-R1)] was derived and the characteristics of this cell line were investigated. METHODS: A new cell line (LST-R1) originated from laterally spreading tumor was established. Properties of the cell line were characterized using scanning and transmission electron microscopy, immunohistochemistry method, cytogenetic analysis and nude mice xenograft experiments. In vitro invasion assay, cDNA microarray and Western blotting were used to compare the difference between the LST-R1 and other colorectal cancer cell lines derived from prudent colon cancer. RESULTS: Our study demonstrated that both epithelial special antigen (ESA) and cytokeratin-20 (CK20) were expressed in LST-R1. The cells presented microvilli and tight junction with large nuclei. The karyotypic analysis showed hyperdiploid features with structural chromosome aberrations. The in vivo tumorigenicity was also demonstrated in nude mice xenograft experiments. The invasion assay suggested this cell line has a higher invasive ability. cDNA microarray and Western blotting show the loss of the expression of E-cadherin in LST-R1 cells. CONCLUSION: We established and characterized a colorectal cancer cell line, LST-R1 and LST-R1 has an obvious malignant tendency, which maybe partially attributed to the changes of the expression of some adhesion molecules, such as E-cadherin. It is also a versatile tool for exploring the original and progressive mechanisms of laterally spreading tumor and the early colon cancer genesis. PMID:18300345

Creating Sunflower Mutant Lines (Helianthus Annuus L.) Using Induced Mutagenesis

International Nuclear Information System (INIS)

Encheva, J.

2009-01-01

Immature sunflower zygotic embryos of sunflower fertility restorer line 374 R were treated with ultrasound and gamma radiation before plating embryos to culture medium. All plants were isolated and self-pollinated for several generations. New sunflower forms with inherited morphological and biochemical changes were obtained. The genetic changes occurring during the mutation procedure included fourteen morphological and biochemical characters. In comparison to the check line 374 R, decreasing of the mean value of the indexes was registered for 33 % of the total number of characters and vise verse, significant increasing was observed for 60 %. Mutation for resistance to the local population of Orobanche cumana race A-E was obtained from the susceptible Bulgarian control line 374 R. Two investigated mutant lines possessed 100 % resistance to Orobanche and stable inheritance in the next generations. Our results showed that induced mutagenesis in sunflower can be successfully used to develop new lines useful for heterosis breeding

Biochemical Characterization of Glutamate Racemase-A New Candidate Drug Target against Burkholderia cenocepacia Infections.

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Aygun Israyilova

Full Text Available The greatest obstacle for the treatment of cystic fibrosis patients infected with the Burkholderia species is their intrinsic antibiotic resistance. For this reason, there is a need to develop new effective compounds. Glutamate racemase, an essential enzyme for the biosynthesis of the bacterial cell wall, is an excellent candidate target for the design of new antibacterial drugs. To this aim, we recombinantly produced and characterized glutamate racemase from Burkholderia cenocepacia J2315. From the screening of an in-house library of compounds, two Zn (II and Mn (III 1,3,5-triazapentadienate complexes were found to efficiently inhibit the glutamate racemase activity with IC50 values of 35.3 and 10.0 μM, respectively. Using multiple biochemical approaches, the metal complexes have been shown to affect the enzyme activity by binding to the enzyme-substrate complex and promoting the formation of an inhibited dimeric form of the enzyme. Our results corroborate the value of glutamate racemase as a good target for the development of novel inhibitors against Burkholderia.

Biochemical composition and methane production correlations

OpenAIRE

Charnier, Cyrille; Latrille, Eric; Moscoviz, Roman; Miroux, Jérémie; Steyer, Jean-Philippe

2016-01-01

Substrates for anaerobic digestion are composed of heterogeneous and complex organic matter. General parameters of the organic matter can be used to describe its composition such as sugar, protein and lipid contents, Chemical Oxygen Demand (COD), Biochemical Methane Potential (BMP) and kinetic of methane production. These parameters are required for the monitoring of digesters but their characterization are time consuming and expensive; thus, these parameters are rarely assessed all together....

Validation of in-line surface characterization by light scattering in Robot Assisted Polishing

DEFF Research Database (Denmark)

Pilny, Lukas; Bissacco, Giuliano; De Chiffre, Leonardo

2014-01-01

The suitability of a commercial scattered light sensor for in-line characterization of fine surfaces in the roughness range Sa 1 – 30 nm generated by the Robot Assisted Polishing (RAP) was investigated and validated. A number of surfaces were generated and directly measured with the scattered light...

Spent Pot Lining Characterization Framework

Science.gov (United States)

Ospina, Gustavo; Hassan, Mohamed I.

2017-09-01

Spent pot lining (SPL) management represents a major concern for aluminum smelters. There are two key elements for spent pot lining management: recycling and safe storage. Spent pot lining waste can potentially have beneficial uses in co-firing in cement plants. Also, safe storage of SPL is of utmost importance. Gas generation of SPL reaction with water and ignition sensitivity must be studied. However, determining the feasibility of SPL co-firing and developing the required procedures for safe storage rely on determining experimentally all the necessary SPL properties along with the appropriate test methods, recognized by emissions standards and fire safety design codes. The applicable regulations and relevant SPL properties for this purpose are presented along with the corresponding test methods.

Proposed particle-beam characterizations for the APS undulator test line

Energy Technology Data Exchange (ETDEWEB)

Lumpkin, A.H.; Borland, M.; Milton, S.

1993-09-01

A research and development effort is underway at the Advanced Photon Source (APS) to use an rf gun as a low-emittance electron source for injection into the 100- to 650-MeV linac subsystem and subsequently to an undulator test area. This configuration would combine the acceleration capability of the 200-MeV S-band electron linac and the in-line 450-MeV positron linac that normally provide positrons to the positron accumulator ring (PAR). A transport line that bypasses the PAR will bring the electrons to the undulator test area. Characterization techniques will be discussed for the electron beam with a normalized, rms emittance of <10 {pi} mm mrad (1{sigma}) at micropulse charges of up to 350 pC and micropulse durations of {approximately}5 ps (FWHM). Tests proposed include measurement of particle beam transport effects (at one-tenth the storage ring beam rigidity) caused by small undulator field errors as well as operations intended to produce coherent, short wavelength radiation (<200 nm).

Characterization of cell lines developed from the glassy-winged sharpshooter, Homalodisca coagulata (Hemiptera: Cicadellidae).

Science.gov (United States)

Kamita, Shizuo G; Do, Zung N; Samra, Aman I; Hagler, James R; Hammock, Bruce D

2005-01-01

Four continuous cell lines were established from the embryos of the glassy-winged sharpshooter, Homalodisca coagulata (Say), an economically important insect vector of bacterial pathogens of grape, almond, citrus, oleander, and other agricultural and ornamental plantings. The cell lines were designated GWSS-Z10, GWSS-Z15, GWSS-G3, and GWSS-LH. The GWSS-Z10, GWSS-Z15, and GWSS-G3 lines were cultured in Ex-Cell 401 medium supplemented with 10% fetal bovine serum (FBS), whereas the GWSS-LH line was cultured in LH medium supplemented with 20% FBS. The cell lines were characterized in terms of their morphology, growth, protein composition, and polymerase chain reaction- amplification patterns of their chromosomal deoxyribonucleic acid. The population doubling times of GWSS-Z10, GWSS-Z15, GWSS-G3, and GWSS-LH were 46.2, 90.9, 100.3, and 60.2 h, respectively. These lines should be useful for the study of insect-pathogenic viruses of leafhoppers, aphids, treehoppers, and other related insects as well as plant-pathogenic viruses that are transmitted by these insects.

Molecular and biochemical characterization of calmodulin from Echinococcus granulosus.

Science.gov (United States)

Wang, Ning; Zhong, Xiuqin; Song, Xingju; Gu, Xiaobin; Lai, Weiming; Xie, Yue; Peng, Xuerong; Yang, Guangyou

2017-12-04

Echinococcus granulosus is a harmful cestode parasite that causes cystic echinococcosis in humans as well as various livestock species and wild animals. Calmodulin (CaM), a Ca 2+ sensor protein, is widely expressed in eukaryotes and mediates a variety of cellular signaling activities. In the present study, the cDNA encoding CaM in Echinococcus granulosus (rEgCaM) was successfully cloned and the molecular and biochemical characterizations carried out. The antigenicity and immunoreactivity of rEgCaM was detected and the preliminary enzyme-linked immunosorbent assay (ELISA)-based serodiagnostic potential of EgCaM was assessed. The locations of this protein in the adult worm and larval stage, and the mRNA expression in different states of E. granulosus protoscoleces (PSCs) were defined clearly. Moreover, the Ca 2+ -binding properties of EgCaM were measured. rEgCaM is a highly conserved calcium-binding protein, consisting of 149 amino acids. Immunoblotting analysis revealed that rEgCaM could be identified using E. granulosus infected sheep serum. The use of rEgCaM as an antigen was evaluated by indirect ELISA which exhibited a high sensitivity (90.3%), but low specificity (47.1%). rEgCaM was ubiquitously expressed in protoscoleces and adults of E. granulosus, as well as in the germinal layer of the cyst wall. The mRNA expression level of rEgCaM was increased from the start of H 2 O 2 exposure and then gradually decreased because of the increased apoptosis of PSCs. In electrophoretic mobility tests and 1-anilinonaphthalene-8-sulfonic acid assays, rEgCaM showed a typical characteristic of a calcium-binding protein. To our knowledge, this is the first report on CaM from E. granulosus and rEgCaM is likely to be involved in some important biological function of E. granulosus as a calcium-binding protein.

Biochemical characterization of the purple form of Marinobacter hydrocarbonoclasticus nitrous oxide reductase

Science.gov (United States)

Dell'Acqua, Simone; Pauleta, Sofia R.; Moura, José J. G.; Moura, Isabel

2012-01-01

Nitrous oxide reductase (N2OR) catalyses the final step of the denitrification pathway—the reduction of nitrous oxide to nitrogen. The catalytic centre (CuZ) is a unique tetranuclear copper centre bridged by inorganic sulphur in a tetrahedron arrangement that can have different oxidation states. Previously, Marinobacter hydrocarbonoclasticus N2OR was isolated with the CuZ centre as CuZ*, in the [1Cu2+ : 3Cu+] redox state, which is redox inert and requires prolonged incubation under reductive conditions to be activated. In this work, we report, for the first time, the isolation of N2OR from M. hydrocarbonoclasticus in the ‘purple’ form, in which the CuZ centre is in the oxidized [2Cu2+ : 2Cu+] redox state and is redox active. This form of the enzyme was isolated in the presence of oxygen from a microaerobic culture in the presence of nitrate and also from a strictly anaerobic culture. The purple form of the enzyme was biochemically characterized and was shown to be a redox active species, although it is still catalytically non-competent, as its specific activity is lower than that of the activated fully reduced enzyme and comparable with that of the enzyme with the CuZ centre in either the [1Cu2+ : 3Cu+] redox state or in the redox inactive CuZ* state. PMID:22451106

Genetic and Genomic Characterization of 462 Melanoma Patient-Derived Xenografts, Tumor Biopsies, and Cell Lines

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Bradley Garman

2017-11-01

Full Text Available Summary: Tumor-sequencing studies have revealed the widespread genetic diversity of melanoma. Sequencing of 108 genes previously implicated in melanomagenesis was performed on 462 patient-derived xenografts (PDXs, cell lines, and tumors to identify mutational and copy number aberrations. Samples came from 371 unique individuals: 263 were naive to treatment, and 108 were previously treated with targeted therapy (34, immunotherapy (54, or both (20. Models of all previously reported major melanoma subtypes (BRAF, NRAS, NF1, KIT, and WT/WT/WT were identified. Multiple minor melanoma subtypes were also recapitulated, including melanomas with multiple activating mutations in the MAPK-signaling pathway and chromatin-remodeling gene mutations. These well-characterized melanoma PDXs and cell lines can be used not only as reagents for a large array of biological studies but also as pre-clinical models to facilitate drug development. : Garman et al. have characterized melanoma PDXs and cell lines described in Krepler et al. (see the related paper in this issue of Cell Reports, identifying major and minor subtypes, some of which were previously not well defined, targeted and immunotherapy resistance, and tumor heterogeneity, creating a set of reagents for future drug discovery and biological studies. Keywords: melanoma, patient-derived xenografts, massively parallel sequencing, cell lines

Characterizing Excavation Damaged Zone and Stability of Pressurized Lined Rock Caverns for Underground Compressed Air Energy Storage

Science.gov (United States)

Kim, Hyung-Mok; Rutqvist, Jonny; Jeong, Ju-Hwan; Choi, Byung-Hee; Ryu, Dong-Woo; Song, Won-Kyong

2013-09-01

In this paper, we investigate the influence of the excavation damaged zone (EDZ) on the geomechanical performance of compressed air energy storage (CAES) in lined rock caverns. We conducted a detailed characterization of the EDZ in rock caverns that have been excavated for a Korean pilot test program on CAES in (concrete) lined rock caverns at shallow depth. The EDZ was characterized by measurements of P- and S-wave velocities and permeability across the EDZ and into undisturbed host rock. Moreover, we constructed an in situ concrete lining model and conducted permeability measurements in boreholes penetrating the concrete, through the EDZ and into the undisturbed host rock. Using the site-specific conditions and the results of the EDZ characterization, we carried out a model simulation to investigate the influence of the EDZ on the CAES performance, in particular related to geomechanical responses and stability. We used a modeling approach including coupled thermodynamic multiphase flow and geomechanics, which was proven to be useful in previous generic CAES studies. Our modeling results showed that the potential for inducing tensile fractures and air leakage through the concrete lining could be substantially reduced if the EDZ around the cavern could be minimized. Moreover, the results showed that the most favorable design for reducing the potential for tensile failure in the lining would be a relatively compliant concrete lining with a tight inner seal, and a relatively stiff (uncompliant) host rock with a minimized EDZ. Because EDZ compliance depends on its compressibility (or modulus) and thickness, care should be taken during drill and blast operations to minimize the damage to the cavern walls.

Biochemical Characterization of Mycobacterium tuberculosis DNA Repair Enzymes – Nfo, XthA and Nei2

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Sailau Abeldenov

2014-01-01

Full Text Available Introduction: Tuberculosis (TB is a human disease caused by Mycobacterium tuberculosis (Mtb. Treatment of TB requires long-term courses of multi-drug therapies to eliminate subpopulations of bacteria, which sometimes persist against antibiotics. Therefore, understanding of the mechanism of Mtb antibiotic-resistance is extremely important. During infection, Mtb overcomes a variety of body defense mechanisms, including treatment with the reactive species of oxygen and nitrogen. The bases in DNA molecule are susceptible to the damages caused by reactive forms of intermediate compounds of oxygen and nitrogen. Most of this damage is repaired by the base excision repair (BER pathway. In this study, we aimed to biochemically characterize three Mtb DNA repair enzymes of BER pathway. Methods: XthA, nfo, and nei genes were identified in mycobacteria by homology search of genomic sequences available in the GenBank database. We used standard methods of genetic engineering  to clone and sequence Mtb genes, which coded Nfo, XthA and Nei2 repair enzymes. The protein products of Mtb genes were expressed and purified in Escherichia coli using affinity tags. The enzymatic activity of purified Nfo, XthA, and Nei2 proteins were measured using radioactively labeled DNA substrates containing various modified residues. Results: The genes end (Rv0670, xthA (Rv0427c, and nei (Rv3297 were PCR amplified using genomic DNA of Mtb H37Rv with primers that contain specific restriction sites. The amplified products were inserted into pET28c(+ expression vector in such a way that the recombinant proteins contain C-terminal histidine tags. The plasmid constructs were verified by sequencing and then transformed into the Escherichia coli BL21 (DE3 strain. Purification of recombinant proteins was performed using Ni2+ ions immobilized affinity column, coupled with the fast performance liquid chromatography machine AKTA. Identification of the isolated proteins was performed by

Craniometaphyseal dysplasia with obvious biochemical abnormality and rickets-like features.

Science.gov (United States)

Wu, Bo; Jiang, Yan; Wang, Ou; Li, Mei; Xing, Xiao-Ping; Xia, Wei-Bo

2016-05-01

Craniometaphyseal dysplasia (CMD) is a rare genetic disorder that is characterized by progressive sclerosis of the craniofacial bones and metaphyseal widening of long bones, and biochemical indexes were mostly normal. To further the understanding of the disease from a biochemical perspective, we reported a CMD case with obviously abnormal biochemical indexes. A 1-year-old boy was referred to our clinic. Biochemical test showed obviously increased alkaline phosphatase (ALP) and parathyroid hormone (PTH), mild hypocalcemia and hypophosphatemia. Moreover, significant elevated receptor activator of nuclear factor kappa-B ligand (RANKL) level, but normal β-C-terminal telopeptide of type I collagen (β-CTX) concentration were revealed. He was initially suspected of rickets, because the radiological examination also showed broadened epiphysis in his long bones. Supplementation with calcium and calcitriol alleviated biochemical abnormality. However, the patient gradually developed osteosclerosis which was inconformity with rickets. Considering that he was also presented with facial paralysis and nasal obstruction symptom, the diagnosis of craniometaphyseal dysplasia was suspected, and then was confirmed by the mutation analysis of ANKH of the proband and his family, which showed a de novo heterozygous mutation (C1124-1126delCCT) on exon 9. Our study revealed that obvious biochemical abnormality and rickets-like features might present as uncommon characteristics in CMD patients, and the calcium and calcitriol supplementation could alleviate biochemical abnormalities. Furthermore, although early osteoclast differentiation factor was excited in CMD patient, activity of osteoclast was still inert. Copyright © 2016. Published by Elsevier B.V.

Biochemical characterization of the GM2 gangliosidosis B1 variant

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Tutor J.C.

2004-01-01

Full Text Available The deficiency of the A isoenzyme of ß-hexosaminidase (Hex produced by different mutations of the gene that codes for the alpha subunit (Tay-Sachs disease has two variants with enzymological differences: the B variant consists of the absence of Hex A isoenzyme and the B1 variant produces an inactive Hex A isoenzyme for the hydrolysis of the GM2 ganglioside and synthetic substrates with negative charge. In contrast to the early childhood form of the B variant, the B1 variant appears at a later clinical stage (3 to 7 years of age with neurodegenerative symptoms leading to the death of the patient in the second decade of life. The most frequent mutation responsible for the GM2 gangliosidosis B1 variant is R178H, which has a widespread geographic and ethnic distribution. The highest incidence has been described in Portugal, which has been suggested as the point of origin of this mutation. Biochemical characterization of this lysosomal disease is carried out using negatively charged synthetic alpha subunit-specific sulfated substrates, since Hex A isoenzyme heat-inactivation assays are not applicable. However, the determination of the apparent activation energy of Hex using the neutral substrate 3,3'-dichlorophenolsulfonphthaleinyl N-acetyl-ß-D-glucosaminide, may offer a valid alternative. The presence of an alpha subunit in the alphaß heterodimer Hex A means that its activation energy (41.8 kJ/mol is significantly lower than that of the ßß homodimer Hex B (75.1 kJ/mol; however, as mutation inactivates the alpha subunit, the Hex A of the B1 variant presents an activation energy that is similar to that of the Hex B isoenzyme.

Caracterização bioquímica de linhagens de soja com alto teor de proteína Biochemical characterization of high protein soybean lines

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Rita Maria Alves de Moraes

2006-05-01

Full Text Available O objetivo deste trabalho foi caracterizar bioquimicamente duas isolinhas de soja com alto teor de proteína. O aumento do teor de proteína nas isolinhas foi acompanhado por redução no teor de óleo e de carboidratos totais. Em relação à composição aminoacídica, o aumento do teor de proteína promoveu acréscimo em todos os aminoácidos, exceto glicina, alanina, metionina, cisteína e tirosina, mantendo a relação enxofre/nitrogênio. A quantificação dos polipeptídios mostrou que o aumento do teor de proteína manteve inalterado o teor das proteínas 7S, promoveu aumento no teor das proteínas 11S e, conseqüentemente, da relação 11S/7S. Pode haver melhoria na qualidade do farelo de soja das isolinhas, uma vez que as proteínas 11S têm melhor qualidade nutricional do que as proteínas 7S.The objective of this work was to characterize high protein soybean near isogenic lines. The increasing of protein was followed by reducing of oil and carbohydrate. In respect to aminoacid composition, increasing of protein promoted a rising in all aminoacids, except for glycine, alanine, methionine, cysteine and tyrosine, although the ratio S/N has been kept. The measure of polypeptides showed that the increasing of protein did not alter the quantity of 7S proteins, provided increasing of 11S proteins and 11S/7S ratio. An improvement of meal quality in these lines can occur once the 11S proteins have a better nutritional quality than 7S proteins.

Biochemical markers predictive for bone marrow involvement in systemic mastocytosis

NARCIS (Netherlands)

Donker, Marjolein L.; van Doormaal, Jasper J.; van Doormaal, Frederiek F.; Kluin, Philip M.; van der Veer, Eveline; de Monchy, Jan G. R.; Kema, Ido P.; Kluin-Nelemans, Hanneke C.

Systemic mastocytosis is characterized by bone marrow involvement, which requires a bone marrow biopsy for diagnostic work-up. We questioned whether bone marrow involvement could be predicted using biochemical markers. We selected patients with various symptoms suggestive of indolent systemic

Isolation and characterization of conditionally immortalized mouse glomerular endothelial cell lines.

Science.gov (United States)

Rops, Angelique L; van der Vlag, Johan; Jacobs, Cor W; Dijkman, Henry B; Lensen, Joost F; Wijnhoven, Tessa J; van den Heuvel, Lambert P; van Kuppevelt, Toin H; Berden, Jo H

2004-12-01

The culture and establishment of glomerular cell lines has proven to be an important tool for the understanding of glomerular cell functions in glomerular physiology and pathology. Especially, the recent establishment of a conditionally immortalized visceral epithelial cell line has greatly boosted the research on podocyte biology. Glomeruli were isolated from H-2Kb-tsA58 transgenic mice that contain a gene encoding a temperature-sensitive variant of the SV40 large tumor antigen, facilitating proliferative growth at 33 degrees C and differentiation at 37 degrees C. Glomerular endothelial cells were isolated from glomerular outgrowth by magnetic beads loaded with CD31, CD105, GSL I-B4, and ULEX. Clonal cell lines were characterized by immunofluorescence staining with antibodies/lectins specific for markers of endothelial cells, podocytes, and mesangial cells. Putative glomerular endothelial cell lines were analyzed for (1) cytokine-induced expression of adhesion molecules; (2) tube formation on Matrigel coating; and (3) the presence of fenestrae. As judged by immunostaining for Wilms tumor-1, smooth muscle actin (SMA), podocalyxin, and von Willebrand factor (vWF), we obtained putative endothelial, podocyte and mesangial cell lines. The mouse glomerular endothelial cell clone #1 (mGEnC-1) was positive for vWF, podocalyxin, CD31, CD105, VE-cadherin, GSL I-B4, and ULEX, internalized acetylated-low-density lipoprotein (LDL), and showed increased expression of adhesion molecules after activation with proinflammatory cytokines. Furthermore, mGEnC-1 formed tubes and contained nondiaphragmed fenestrae. The mGEnC-1 represents a conditionally immortalized cell line with various characteristics of differentiated glomerular endothelial cells when cultured at 37 degrees C. Most important, mGEnC-1 contains nondiaphragmed fenestrae, which is a unique feature of glomerular endothelial cells.

Canine degenerative myelopathy: biochemical characterization of superoxide dismutase 1 in the first naturally occurring non-human amyotrophic lateral sclerosis model.

Science.gov (United States)

Crisp, Matthew J; Beckett, Jeffrey; Coates, Joan R; Miller, Timothy M

2013-10-01

Mutations in canine superoxide dismutase 1 (SOD1) have recently been shown to cause canine degenerative myelopathy, a disabling neurodegenerative disorder affecting specific breeds of dogs characterized by progressive motor neuron loss and paralysis until death, or more common, euthanasia. This discovery makes canine degenerative myelopathy the first and only naturally occurring non-human model of amyotrophic lateral sclerosis (ALS), closely paralleling the clinical, pathological, and genetic presentation of its human counterpart, SOD1-mediated familial ALS. To further understand the biochemical role that canine SOD1 plays in this disease and how it may be similar to human SOD1, we characterized the only two SOD1 mutations described in affected dogs to date, E40K and T18S. We show that a detergent-insoluble species of mutant SOD1 is present in spinal cords of affected dogs that increases with disease progression. Our in vitro results indicate that both canine SOD1 mutants form enzymatically active dimers, arguing against a loss of function in affected homozygous animals. Further studies show that these mutants, like most human SOD1 mutants, have an increased propensity to form aggregates in cell culture, with 10-20% of cells possessing visible aggregates. Creation of the E40K mutation in human SOD1 recapitulates the normal enzymatic activity but not the aggregation propensity seen with the canine mutant. Our findings lend strong biochemical support to the toxic role of SOD1 in canine degenerative myelopathy and establish close parallels for the role mutant SOD1 plays in both canine and human disorders. Copyright © 2013 Elsevier Inc. All rights reserved.

Biochemical and morphological characterization of light and heavy sarcoplasmic reticulum vesicles

Energy Technology Data Exchange (ETDEWEB)

Campbell, Kevin Peter [Univ. of Rochester, NY (United States)

1978-01-01

Light (30 to 32.5% sucrose) and heavy (38.5 to 42% sucrose) sarcoplasmic reticulum vesicles (LSR,HSR) were isolated from rabbit leg muscle using a combination of differential centrifugation and isopycnic zonal ultracentrifugation. Thin-section electron microscopy of LSR vesicles reveals empty vesicles of various sizes and shapes whereas the HSR vesicles appear as rounded vesicles of uniform size filled with electron dense material, similar to that seen in the terminal cisternae of the sarcoplasmic reticulum. The sucrose HSR vesicles have an additional morphological feature which appears as membrane projections that resemble the SR feet. The freeze-fracture morphology of either type of SR reveals an asymmetric distribution of intramembraneous particles in the same orientation and distribution as the sarcoplasmic reticulum in vivo. Biochemical studies were made on the content of Ca, Mg, ATPase, and protein of the vesicles and phosphorylation of the vesicles. The biochemical and morphological data indicate that the LSR is derived from the longitudinal sarcoplasmic reticulum and the HSR is derived from the terminal cisternae of the sarcoplasmic reticulum, contains junctional SR membrane and has three unique proteins (calsequestrin, an intrinsic 30,000 dalton protein and a 9000 dalton proteolipid).

Biochemical characterization of the Arctic char (Salvelinus alpinus ovarian progestin membrane receptor

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Thomas Peter

2005-11-01

Full Text Available Abstract Membrane progestin receptors are involved in oocyte maturation in teleosts. However, the maturation-inducing steroid (MIS does not appear to be conserved among species and several progestins may fulfill this function. So far, complete biochemical characterization has only been performed on a few species. In the present study we have characterized the membrane progestin receptor in Arctic char (Salvelinus alpinus and show that the 17,20beta-dihydroxy-4-pregnen-3-one (17,20beta-P receptor also binds several xenobiotics, thus rendering oocyte maturation sensitive to environmental pollutants. We identified a single class of high affinity (Kd, 13.8 ± 1.1 nM, low capacity (Bmax, 1.6 ± 0.6 pmol/g ovary binding sites by saturation and Scatchard analyses. Receptor binding displayed rapid association and dissociation kinetics typical of steroid membrane receptors, with t1/2 s of less than 1 minute. The 17,20beta-P binding also displayed tissue specificity with high, saturable, and specific 17,20beta-P binding detected in ovaries, heart and gills while no specific binding was observed in muscle, brain or liver. Changes in 17,20beta-P binding during oocyte maturation were consistent with its identity as the oocyte MIS membrane receptor. Incubation of fully-grown ovarian follicles with gonadotropin induced oocyte maturation, which was accompanied by a five-fold increase in 17,20beta-P receptor binding. In addition, competition studies with a variety of steroids revealed that receptor binding is highly specific for 17,20beta-P, the likely maturation-inducing steroid (MIS in Arctic char. The relative-binding affinities of all the other progestogens and steroids tested were less than 5% of that of 17,20beta-P for the receptor. Several ortho, para derivatives of DDT also showed weak binding affinity for the 17,20beta-P receptor supporting the hypothesis that xenobiotics may bind steroid receptors on the oocyte's surface and might thereby interfere

Biochemical and structural characterization of Klebsiella pneumoniae oxamate amidohydrolase in the uric acid degradation pathway

Energy Technology Data Exchange (ETDEWEB)

Hicks, Katherine A.; Ealick, Steven E.

2016-05-25

HpxW from the ubiquitous pathogenKlebsiella pneumoniaeis involved in a novel uric acid degradation pathway downstream from the formation of oxalurate. Specifically, HpxW is an oxamate amidohydrolase which catalyzes the conversion of oxamate to oxalate and is a member of the Ntn-hydrolase superfamily. HpxW is autoprocessed from an inactive precursor to form a heterodimer, resulting in a 35.5 kDa α subunit and a 20 kDa β subunit. Here, the structure of HpxW is presented and the substrate complex is modeled. In addition, the steady-state kinetics of this enzyme and two active-site variants were characterized. These structural and biochemical studies provide further insight into this class of enzymes and allow a mechanism for catalysis consistent with other members of the Ntn-hydrolase superfamily to be proposed.

Purification, biochemical, and structural characterization of a novel fibrinolytic enzyme from Mucor subtilissimus UCP 1262.

Science.gov (United States)

Nascimento, Thiago Pajeú; Sales, Amanda Emmanuelle; Porto, Tatiana Souza; Costa, Romero Marcos Pedrosa Brandão; Breydo, Leonid; Uversky, Vladimir N; Porto, Ana Lúcia Figueiredo; Converti, Attilio

2017-08-01

Fibrinolytic proteases are enzymes that degrade fibrin. They provide a promising alternative to existing drugs for thrombolytic therapy. A protease isolated from the filamentous fungus Mucor subtilissimus UCP 1262 was purified in three steps by ammonium sulfate fractionation, ion exchange, and molecular exclusion chromatographies, and characterized biochemically and structurally. The purified protease exhibited a molecular mass of 20 kDa, an apparent isoelectric point of 4.94 and a secondary structure composed mainly of α-helices. Selectivity for N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide as substrate suggests that this enzyme is a chymotrypsin-like serine protease, whose activity was enhanced by the addition of Cu 2+ , Mg 2+ , and Fe 2+ . The enzyme showed a fibrinolytic activity of 22.53 U/mL at 40 °C and its contact with polyethylene glycol did not lead to any significant alteration of its secondary structure. This protein represents an important example of a novel fibrinolytic enzyme with potential use in the treatment of thromboembolic disorders such as strokes, pulmonary emboli, and deep vein thrombosis.

Genome characterization of the selected long- and short-sleep mouse lines.

Science.gov (United States)

Dowell, Robin; Odell, Aaron; Richmond, Phillip; Malmer, Daniel; Halper-Stromberg, Eitan; Bennett, Beth; Larson, Colin; Leach, Sonia; Radcliffe, Richard A

2016-12-01

The Inbred Long- and Short-Sleep (ILS, ISS) mouse lines were selected for differences in acute ethanol sensitivity using the loss of righting response (LORR) as the selection trait. The lines show an over tenfold difference in LORR and, along with a recombinant inbred panel derived from them (the LXS), have been widely used to dissect the genetic underpinnings of acute ethanol sensitivity. Here we have sequenced the genomes of the ILS and ISS to investigate the DNA variants that contribute to their sensitivity difference. We identified ~2.7 million high-confidence SNPs and small indels and ~7000 structural variants between the lines; variants were found to occur in 6382 annotated genes. Using a hidden Markov model, we were able to reconstruct the genome-wide ancestry patterns of the eight inbred progenitor strains from which the ILS and ISS were derived, and found that quantitative trait loci that have been mapped for LORR were slightly enriched for DNA variants. Finally, by mapping and quantifying RNA-seq reads from the ILS and ISS to their strain-specific genomes rather than to the reference genome, we found a substantial improvement in a differential expression analysis between the lines. This work will help in identifying and characterizing the DNA sequence variants that contribute to the difference in ethanol sensitivity between the ILS and ISS and will also aid in accurate quantification of RNA-seq data generated from the LXS RIs.

Development and characterization of two cell lines from gills of Atlantic salmon

Science.gov (United States)

Gjessing, Mona C.; Aamelfot, Maria; Batts, William N.; Benestad, Sylvie L.; Dale, Ole B.; Thoen, Even; Weli, Simon C.; Winton, James R.

2018-01-01

Gill disease in Atlantic salmon, Salmo salar L., causes big losses in the salmon farming industry. Until now, tools to cultivate microorganisms causing gill disease and models to study the gill responses have been lacking. Here we describe the establishment and characterization of two cell lines from the gills of Atlantic salmon. Atlantic salmon gill cell ASG-10 consisted of cells staining for cytokeratin and e-cadherin and with desmosomes as seen by transmission electron microscopy suggesting the cells to be of epithelial origin. These structures were not seen in ASG-13. The cell lines have been maintained for almost 30 passages and both cell lines are fully susceptible to infection by infectious hematopoietic necrosis virus (IHNV), viral hemorrhagic septicemia virus (VHSV), infectious pancreatic necrosis virus (IPNV), Atlantic salmon reovirus TS (TSRV) and Pacific salmon paramyxovirus (PSPV). While infectious salmon anemia virus (ISAV) did not cause visible CPE, immunofluorescent staining revealed a sub-fraction of cells in both the ASG-10 and ASG-13 lines may be permissive to infection. ASG-10 is able to proliferate and migrate to close scratches in the monolayer within seven days in vitro contrary to ASG-13, which does not appear to do have the same proliferative and migratory ability. These cell lines will be useful in studies of gill diseases in Atlantic salmon and may represent an important contribution for alternatives to experimental animals and studies of epithelial–mesenchymal cell biology.

Derivation and characterization of human embryonic stem cell lines from the Chinese population

Institute of Scientific and Technical Information of China (English)

Zhao Wu; Huimin Dai; Lei Qian; Qing Tian; Lei Xiao; Xiaojun Tan; Hui Li; Lingjun Rao; Lixiazi He; Lei Bao; Jing Liao; Chun Cui; Zhenyu Zuo; Qiao Li

2011-01-01

Human embryonic stem cells (hESCs) can self-renew indefinitely and differentiate into all cell types in the human body. Therefore, they are valuable in regenerative medicine, human developmental biology and drug discovery. A number of hESC lines have been derived from the Chinese population,but limited of them are available for research purposes. Here we report the derivation and characterization of two hESC lines derived from human blastocysts of Chinese origin. These hESCs express alkaline phosphatase and hESC-specific markers, including Oct4, Nanog, SSEA-3, SSEA-4,TRA-1-60 and TRA-1-81. They also have high levels of telomerase activity and normal karyotypes. These cells can form embryoid body in vitro and can be differentiated into all three germ layers in vivo by teratoma formation. The newly established hESCs will be distributed for research purposes.The availability of hESC lines from the Chinese population will facilitate studies on the differences in hESCs from different ethnic groups.

Biochemical techniques for the characterization of G-quadruplex structures: EMSA, DMS footprinting, and DNA polymerase stop assay.

Science.gov (United States)

Sun, Daekyu; Hurley, Laurence H

2010-01-01

The proximal promoter region of many human growth-related genes contains a polypurine/polypyrimidine tract that serves as multiple binding sites for Sp1 or other transcription factors. These tracts often contain a guanine-rich sequence consisting of four runs of three or more contiguous guanines separated by one or more bases, corresponding to a general motif known for the formation of an intramolecular G-quadruplex. Recent results provide strong evidence that specific G-quadruplex structures form naturally within these polypurine/polypyrimidine tracts in many human promoter regions, raising the possibility that the transcriptional control of these genes can be modulated by G-quadruplex-interactive agents. In this chapter, we describe three general biochemical methodologies, electrophoretic mobility shift assay (EMSA), dimethylsulfate (DMS) footprinting, and the DNA polymerase stop assay, which can be useful for initial characterization of G-quadruplex structures formed by G-rich sequences.

Establishment and characterization of a new epithelial cell line, KC-1, from koala (Phascolarctos cinereus) conjunctiva.

Science.gov (United States)

Girjes, Adeeb A; Lee, Kristen E; Carrick, Frank N

2003-01-01

A novel, untransformed koala cell line (KC-1) was established by culturing koala conjunctival tissue in growth medium, which has permitted the study of the cell biology of this unique system. After the establishment of the KC-1 cell line, the cells were characterized by light microscopy, doubling time, and Western blot analysis. Light microscopy revealed that the cells have an epithelial morphology. Doubling times were significantly different (P koala cell line was adapted to grow continuously in Dulbecco modified Eagle medium containing 10% FCS for at least 30 passages. This unique cell line is an ideal tool for further investigation on koala cell biology and cytogenetics and for exploration of the pathophysiological mechanism of eye infections caused by different pathogens in koalas.

The Influence of LINE-1 and SINE Retrotransposons on Mammalian Genomes.

Science.gov (United States)

Richardson, Sandra R; Doucet, Aurélien J; Kopera, Huira C; Moldovan, John B; Garcia-Perez, José Luis; Moran, John V

2015-04-01

Transposable elements have had a profound impact on the structure and function of mammalian genomes. The retrotransposon Long INterspersed Element-1 (LINE-1 or L1), by virtue of its replicative mobilization mechanism, comprises ∼17% of the human genome. Although the vast majority of human LINE-1 sequences are inactive molecular fossils, an estimated 80-100 copies per individual retain the ability to mobilize by a process termed retrotransposition. Indeed, LINE-1 is the only active, autonomous retrotransposon in humans and its retrotransposition continues to generate both intra-individual and inter-individual genetic diversity. Here, we briefly review the types of transposable elements that reside in mammalian genomes. We will focus our discussion on LINE-1 retrotransposons and the non-autonomous Short INterspersed Elements (SINEs) that rely on the proteins encoded by LINE-1 for their mobilization. We review cases where LINE-1-mediated retrotransposition events have resulted in genetic disease and discuss how the characterization of these mutagenic insertions led to the identification of retrotransposition-competent LINE-1s in the human and mouse genomes. We then discuss how the integration of molecular genetic, biochemical, and modern genomic technologies have yielded insight into the mechanism of LINE-1 retrotransposition, the impact of LINE-1-mediated retrotransposition events on mammalian genomes, and the host cellular mechanisms that protect the genome from unabated LINE-1-mediated retrotransposition events. Throughout this review, we highlight unanswered questions in LINE-1 biology that provide exciting opportunities for future research. Clearly, much has been learned about LINE-1 and SINE biology since the publication of Mobile DNA II thirteen years ago. Future studies should continue to yield exciting discoveries about how these retrotransposons contribute to genetic diversity in mammalian genomes.

Establishment and characterization of 7 novel hepatocellular carcinoma cell lines from patient-derived tumor xenografts.

Directory of Open Access Journals (Sweden)

Hong Xin

Full Text Available Hepatocellular carcinoma (HCC is a common cancer with poor prognosis worldwide and the molecular mechanism is not well understood. This study aimed to establish a collection of human HCC cell lines from patient-derived xenograft (PDX models. From the 20 surgical HCC sample collections, 7 tumors were successfully developed in immunodeficient mice and further established 7 novel HCC cell lines (LIXC002, LIXC003, LIXC004, LIXC006, LIXC011, LIXC012 and CPL0903 by primary culture. The characterization of cell lines was defined by morphology, growth kinetics, cell cycle, chromosome analysis, short tandem repeat (STR analysis, molecular profile, and tumorigenicity. Additionally, response to clinical chemotherapeutics was validated both in vitro and in vivo. STR analysis indicated that all cell lines were unique cells different from known cell lines and free of contamination by bacteria or mycoplasma. The other findings were quite heterogeneous between individual lines. Chromosome aberration could be found in all cell lines. Alpha-fetoprotein was overexpressed only in 3 out of 7 cell lines. 4 cell lines expressed high level of vimentin. Ki67 was strongly stained in all cell lines. mRNA level of retinoic acid induced protein 3 (RAI3 was decreased in all cell lines. The 7 novel cell lines showed variable sensitivity to 8 tested compounds. LIXC011 and CPL0903 possessed multiple drug resistance property. Sorafenib inhibited xenograft tumor growth of LIXC006, but not of LIXC012. Our results indicated that the 7 novel cell lines with low passage maintaining their clinical and pathological characters could be good tools for further exploring the molecular mechanism of HCC and anti-cancer drug screening.

BIOCHEMICAL AND BIOPHYSICAL CHARACTERIZATION OF RECOMBINANT YEAST PROTEASOME MATURATION FACTOR UMP1

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Bebiana Sá-Moura

2013-04-01

We show that recombinant Ump1 is purified as a mixture of different oligomeric species and thatoligomerization is mediated by intermolecular disulfide bond formation involving the only cysteine residue present in the protein.Furthermore, a combination of bioinformatic, biochemical and structural analysis revealed that Ump1 shows characteristics of anintrinsically disordered protein, which might become structured only upon interaction with the proteasomesubunits.

Biochemical and functional characterization of an albumin protein belonging to the hemopexin superfamily from Lens culinaris seeds.

Science.gov (United States)

Scarafoni, Alessio; Gualtieri, Elisa; Barbiroli, Alberto; Carpen, Aristodemo; Negri, Armando; Duranti, Marcello

2011-09-14

The present paper reports the purification and biochemical characterization of an albumin identified in mature lentil seeds with high sequence similarity to pea PA2. These proteins are found in many edible seeds and are considered potentially detrimental for human health due to the potential allergenicity and lectin-like activity. Thus, the description of their possible presence in food and the assessment of the molecular properties are relevant. The M(r), pI, and N-terminal sequence of this protein have been determined. The work included the study of (i) the binding properties to hemine to assess the presence of hemopexin structural domains and (ii) the binding properties of the protein to thiamin. In addition, the structural changes induced by heating have been evaluated by means of spectroscopic techniques. Denaturation temperature has also been determined. The present work provides new insights about the structural molecular features and the ligand-binding properties and dynamics of this kind of seed albumin.

Biochemical Characterization of Porphobilinogen Deaminase–Deficient Mice During Phenobarbital Induction of Heme Synthesis and the Effect of Enzyme Replacement

Science.gov (United States)

Johansson, Annika; Möller, Christer; Fogh, Jens; Harper, Pauline

2003-01-01

Acute intermittent porphyria (AIP) is a genetic disorder caused by a deficiency of porphobilinogen deaminase (PBGD), the 3rd enzyme in heme synthesis. It is clinically characterized by acute attacks of neuropsychiatric symptoms and biochemically by increased urinary excretion of the porphyrin precursors porphobilinogen (PBG) and 5-aminolevulinic acid (ALA). A mouse model that is partially deficient in PBGD and biochemically mimics AIP after induction of the hepatic ALA synthase by phenobarbital was used in this study to identify the site of formation of the presumably toxic porphyrin precursors and study the effect of enzyme-replacement therapy by using recombinant human PBGD (rhPBGD). After 4 d of phenobarbital administration, high levels of PBG and ALA were found in liver, kidney, plasma, and urine of the PBGD-deficient mice. The administration of rhPBGD intravenously or subcutaneously after a 4-d phenobarbital induction was shown to lower the PBG level in plasma in a dose-dependent manner with maximal effect seen after 30 min and 2 h, respectively. Injection of rhPBGD subcutaneously twice daily during a 4-d phenobarbital induction reduced urinary PBG excretion to 25% of the levels found in PBGD-deficient mice administered with only phenobarbital. This study points to the liver as the main producer of PBG and ALA in the phenobarbital-induced PBGD-deficient mice and demonstrates efficient removal of accumulated PBG in plasma and urine by enzyme-replacement therapy. PMID:15208740

Cytokeratin characterization of human prostatic carcinoma and its derived cell lines.

Science.gov (United States)

Nagle, R B; Ahmann, F R; McDaniel, K M; Paquin, M L; Clark, V A; Celniker, A

1987-01-01

Two murine monoclonal anti-cytokeratin antibodies with defined specificity were shown to distinguish between basal cells and luminal cells in human prostate tissue. Forty-one biopsies or transurethral resection specimens were characterized using these two antibodies. In cases of benign prostatic hyperplasia, focal loss of the basal cell layer was noted in areas of glandular proliferation. Ten cases of adenocarcinoma of the prostate, varying in Gleason's histological grade from 2 to 4, were also studied. In each case the carcinoma was shown to represent the luminal cell phenotype with no evidence of involvement of the basal cell phenotype. An analysis of three established metastatic prostatic carcinoma cell lines (DU-145, PC-3, and LNCaP) using two-dimensional electrophoresis showed that the cytokeratin complement of each cell line was slightly different but retained the phenotype of the luminal cell. It was concluded that during both hyperplasia and neoplastic transformation of the prostate, the luminal cell phenotype is primarily involved and that the basal cell phenotype does not appear to contribute to either intraluminal proliferation or invasive cell populations.

Sublethal microcystin exposure and biochemical outcomes among hemodialysis patients.

Directory of Open Access Journals (Sweden)

Elizabeth D Hilborn

Full Text Available Cyanobacteria are commonly-occurring contaminants of surface waters worldwide. Microcystins, potent hepatotoxins, are among the best characterized cyanotoxins. During November, 2001, a group of 44 hemodialysis patients were exposed to microcystins via contaminated dialysate. Serum microcystin concentrations were quantified with enzyme-linked immunosorbent assay which measures free serum microcystin LR equivalents (ME. We describe serum ME concentrations and biochemical outcomes among a subset of patients during 8 weeks following exposure. Thirteen patients were included; 6 were males, patients' median age was 45 years (range 16-80, one was seropositive for hepatitis B surface antigen. The median serum ME concentration was 0.33 ng/mL (range: <0.16-0.96. One hundred thirty nine blood samples were collected following exposure. Patients' biochemical outcomes varied, but overall indicated a mixed liver injury. Linear regression evaluated each patient's weekly mean biochemical outcome with their maximum serum ME concentration; a measure of the extrinsic pathway of clotting function, prothrombin time, was negatively and significantly associated with serum ME concentrations. This group of exposed patients' biochemical outcomes display evidence of a mixed liver injury temporally associated with microcystin exposure. Interpretation of biochemical outcomes are complicated by the study population's underlying chronic disease status. It is clear that dialysis patients are a distinct 'at risk' group for cyanotoxin exposures due to direct intravenous exposure to dialysate prepared from surface drinking water supplies. Careful monitoring and treatment of water supplies used to prepare dialysate is required to prevent future cyanotoxin exposure events.

An integrated in-line fluid characterization system for industrial applications(In-situ fluid mechanics experiments)

OpenAIRE

Johan, Wiklund; Reinhardt, Kotze; Beat, Birkhofer; Stefano, Ricci; Valentino, Meacci; Mats, Stading; Rainer, Haldenwang; SP-Technical Research Institute of Sweden; FPRC, Cape Peninsula University of Technology; Sika Services AG; Information Engineering Department - University of Florence; Information Engineering Department - University of Florence; SP-Technical Research Institute of Sweden; FPRC, Cape Peninsula University of Technology

2015-01-01

In this work we have presented the world's first commercially available embedded in-line fluids characterization system, "Flow-Viz". It has been specifically designed for the non-invasive, in-line, continuous, real-time velocity profile and rheological assessment of opaque, non-Newtonian industrial fluids. The Flow-Viz system has been successfully installed in pilot plants of international companies and used also for academic research. The technology has been applied to a wide range of fluids...

In-line characterization and identification of micro-droplets on-chip

Directory of Open Access Journals (Sweden)

Weber Emanuel

2014-01-01

Full Text Available We present an integrated optofluidic sensor system for in-line characterization of micro-droplets. The device provides information about the droplet generation frequency, the droplet volume, and the content of the droplet. Due to its simplicity this principle can easily be implemented with other microfluidic components on one and the same device. The sensor is based on total internal reflection phenomena. Droplets are pushed through a microfluidic channel which is hit by slightly diverging monochromatic light. At the solid-liquid interface parts of the rays experience total internal reflection while another part is transmitted. The ratio of reflected to transmitted light depends on the refractive index of the solution. Both signals are recorded simultaneously and provide a very stable output signal for the droplet characterization. With the proposed system passing droplets were counted up to 320 droplets per second and droplets with different volumes could be discriminated. In a final experiment droplets with different amounts of dissolved CaCl2 were distinguished based on their reflected and transmitted light pattern. This principle can be applied for the detection of any molecules in microdroplets which significantly influence the refractive index of the buffer solution.

Medicinal plants: production and biochemical characterization

International Nuclear Information System (INIS)

Chunzhao Liu; Zobayed, S.M.A; Murch, S.J.; Saxena, P.K.

2002-01-01

Recent advances in the area of biotechnology offer some possibility for the development of new technologies for the conservation, characterization and mass production of medicinal plant species, (i.e. in vitro cell culture techniques for the mass production of sterile, consistent, standardized medicinal plant materials). This paper discussed the following subjects - plant tissue culture, de novo shoot organogenesis, de novo root organogenesis, somatic embryogenesis, large scale propagation in bioreactors and discovery of unique biomolecules

Impact of divergent selection for ultimate pH of pectoralis major muscle on biochemical, histological, and sensorial attributes of broiler meat.

Science.gov (United States)

Alnahhas, N; Le Bihan-Duval, E; Baéza, E; Chabault, M; Chartrin, P; Bordeau, T; Cailleau-Audouin, E; Meteau, K; Berri, C

2015-09-01

The impact of divergent selection based on the ultimate pH (pHu) of pectoralis major (P. major) muscle on the chemical, biochemical, and histological profiles of the muscle and sensorial quality of meat was investigated in broiler chickens. The protein, lipid, DM, glycogen and lactate content, glycolytic potential, proteolysis, lipid and protein oxidation index, muscle fiber cross-sectional area, capillary density, and collagen surface were determined on the breast P. major muscle of 6-wk-old broilers issued from the high-pHu (pHu+) and low-pHu (pHu-) lines. Sensory attributes were also evaluated on the breast (roasted or grilled) and thigh (roasted) meat of the 2 lines. Protein, lipid, and DM content of P. major muscle were not affected by selection ( > 0.05). However, the P. major muscle of the pHu+ line was characterized by lower residual glycogen (-16%; ≤ 0.001) and lactate (-14%; ≤ 0.001) content and lower glycolytic potential (-14%; ≤ 0.001) compared with the pHu- line. Although the average cross-sectional area of muscle fibers and surface occupied by collagen were similar ( > 0.05) in both lines, fewer capillaries per fiber (-15%; ≤ 0.05) were observed in the pHu+ line. The pHu+ line was also characterized by lower lipid oxidation (thiobarbituric acid reactive substance index: -23%; ≤ 0.05) but protein oxidation and proteolysis index were not different ( > 0.05) between the 2 lines. At the sensory level, selection on breast muscle pHu mainly affected the texture of grilled and roast breast meat, which was judged significantly more tender ( ≤ 0.001) in the pHu+ line, and the acid taste, which was less pronounced in the roasted breast meat of the pHu+ line ( ≤ 0.002). This study highlighted that selection based on pHu does not affect the chemical composition and structure of breast meat. However, by modifying muscle blood supply and glycogen turnover, it affects meat acidity and oxidant status, both of which are likely to contribute to the large

Biochemical composition and caloric content in the sand whiting Sillago sihama (Forsskal), from Zuari Estuary, Goa

Digital Repository Service at National Institute of Oceanography (India)

Shamsan, E.F.; Ansari, Z.A.

to prevent a number of coronary heart diseases (Edirisinghe, 1998), is also present in fishes. Although several studies deal with the proximate composition of biochemical components of many commercially important fishes but no work on similar lines has...

Sub-acute toxicity and biochemical effects of extracts of Anaphe ...

African Journals Online (AJOL)

Ataxia syndrome which is characterized by sudden onset of severe muscular tremor and gait ataxia has been shown to be associated with the consumption of the larvae of Anaphe venata in South Western part of Nigeria. In this report, the sub -acute toxicity and biochemical effects of polar and nonpolar extracts of Anaphe ...

Establishment and characterization of a unique 1 microm diameter liver-derived progenitor cell line.

Science.gov (United States)

Aravalli, Rajagopal N; Behnan Sahin, M; Cressman, Erik N K; Steer, Clifford J

2010-01-01

Liver-derived progenitor cells (LDPCs) are recently identified novel stem/progenitor cells from healthy, unmanipulated adult rat livers. They are distinct from other known liver stem/progenitor cells such as the oval cells. In this study, we have generated a LDPC cell line RA1 by overexpressing the simian virus 40 (SV40) large T antigen (TAg) in primary LDPCs. This cell line was propagated continuously for 55 passages in culture, after which it became senescent. Interestingly, following transformation with SV40 TAg, LDPCs decreased in size significantly and the propagating cells measured 1 microm in diameter. RA1 cells proliferated in vitro with a doubling time of 5-7 days, and expressed cell surface markers of LDPCs. In this report, we describe the characterization of this novel progenitor cell line that might serve as a valuable model to study liver cell functions and stem cell origin of liver cancers. Copyright 2009 Elsevier Inc. All rights reserved.

Molecular cytogenetic characterization of a human thyroid cancercell line

Energy Technology Data Exchange (ETDEWEB)

Weier, Heinz-Ulrich G.; Tuton, Tiffany B.; Ito, Yuko; Chu, LisaW.; Lu, Chung-Mei; Baumgartner, Adolf; Zitzelsberger, Horst F.; Weier,Jingly F.

2006-01-04

The incidence of papillary thyroid carcinoma (PTC) increases significantly after exposure of the head and neck region to ionizing radiation, yet we know neither the steps involved in malignant transformation of thyroid epithelium nor the specific carcinogenic mode of action of radiation. Such increased tumor frequency became most evident in children after the 1986 nuclear accident in Chernobyl, Ukraine. In the twelve years following the accident, the average incidence of childhood PTCs (chPTC) increased over one hundred-fold compared to the rate of about 1 tumor incidence per 10{sup 6} children per year prior to 1986. To study the etiology of radiation-induced thyroid cancer, we formed an international consortium to investigate chromosomal changes and altered gene expression in cases of post-Chernobyl chPTC. Our approach is based on karyotyping of primary cultures established from chPTC specimens, establishment of cell lines and studies of genotype-phenotype relationships through high resolution chromosome analysis, DNA/cDNA micro-array studies, and mouse xenografts that test for tumorigenicity. Here, we report the application of fluorescence in situ hybridization (FISH)-based techniques for the molecular cytogenetic characterization of a highly tumorigenic chPTC cell line, S48TK, and its subclones. Using chromosome 9 rearrangements as an example, we describe a new approach termed ''BAC-FISH'' to rapidly delineate chromosomal breakpoints, an important step towards a better understanding of the formation of translocations and their functional consequences.

Characterization of Neptunium Oxide Generated Using the HB-Line Phase II Flowsheet

Energy Technology Data Exchange (ETDEWEB)

Duffey, J

2003-08-29

Approximately 98 grams of neptunium(IV) oxide (NpO{sub 2}) were produced at the Savannah River Technology Center (SRTC) for use in gas generation tests to support the neptunium stabilization program at the Savannah River Site (SRS). The NpO{sub 2} was produced according to the anticipated HB-Line flowsheet consisting of anion exchange, oxalate precipitation, filtration, and calcination. Characterization of the NpO{sub 2} product to be used in gas generation tests included bulk and tap density measurements, X-ray diffraction, particle size distribution, specific surface area measurements, and moisture analysis.

Hydrodynamic behaviour and biochemical characterization of a ...

African Journals Online (AJOL)

STORAGESEVER

2008-12-03

Dec 3, 2008 ... Available online at http://www.academicjournals.org/AJB ... characterization of a simple custom expanded bed ... over the traditional methods of recovery, e.g. circum- ... may be considered to be currently limited by the.

Characterization of a mutant rat kangaroo cell line with alterations in the cell cycle and DNA repair

OpenAIRE

Miyaji, E.N.; Johnson, R.T.; Downes, C.S.; Eveno, E.; Mezzina, M.; Sarasin, A.; Menck, C.F.M.

2000-01-01

Using a positive selection system for isolating DNA replication and repair related mutants, we isolated a clone from a rat kangaroo cell line (PtK2) that has increased sensitivity to UV light. Characterization of this clone indicated normal post-replication repair after UV irradiation, and normal removal rates of cyclobutane pyrimidine dimers and pyrimidine(6-4)pyrimidone photoproducts by excision repair. However, this cell line has decreased ability to make early incisions on damaged DNA, po...

Establishment and characterization of a new cell line of canine inflammatory mammary cancer: IPC-366.

Directory of Open Access Journals (Sweden)

Sara Caceres

Full Text Available Canine inflammatory mammary cancer (IMC shares epidemiologic, histopathological and clinical characteristics with the disease in humans and has been proposed as a natural model for human inflammatory breast cancer (IBC. The aim of this study was to characterize a new cell line from IMC (IPC-366 for the comparative study of both IMC and IBC. Tumors cells from a female dog with clinical IMC were collected. The cells were grown under adherent conditions. The growth, cytological, ultrastructural and immunohistochemical (IHC characteristics of IPC-366 were evaluated. Ten female Balb/SCID mice were inoculated with IPC-366 cells to assess their tumorigenicity and metastatic potential. Chromosome aberration test and Karyotype revealed the presence of structural aberration, numerical and neutral rearrangements, demonstrating a chromosomal instability. Microscopic examination of tumor revealed an epithelial morphology with marked anysocytosis. Cytological and histological examination of smears and ultrathin sections by electron microscopy revealed that IPC-366 is formed by highly malignant large round or polygonal cells characterized by marked atypia and prominent nucleoli and frequent multinucleated cells. Some cells had cytoplasmic empty spaces covered by cytoplasmic membrane resembling capillary endothelial cells, a phenomenon that has been related to s vasculogenic mimicry. IHC characterization of IPC-366 was basal-like: epithelial cells (AE1/AE3+, CK14+, vimentin+, actin-, p63-, ER-, PR-, HER-2, E-cadherin, overexpressed COX-2 and high Ki-67 proliferation index (87.15 %. At 2 weeks after inoculating the IPC-366 cells, a tumor mass was found in 100 % of mice. At 4 weeks metastases in lung and lymph nodes were found. Xenograph tumors maintained the original IHC characteristics of the female dog tumor. In summary, the cell line IPC-366 is a fast growing malignant triple negative cell line model of inflammatory mammary carcinoma that can be used for the

Metabolic characterization of invaded cells of the pancreatic cancer cell line, PANC?1

OpenAIRE

Fujita, Mayumi; Imadome, Kaori; Imai, Takashi

2017-01-01

We previously reported that about 0.4% of cells in the cultured human pancreatic cancer cell line, PANC?1, can invade matrigel during the transwell invasion assay, suggesting that these invaded PANC?1 cells may have specific characteristics to keep their invasive potential. To identify the metabolic characterization specific in the invaded PANC?1 cells, metabolome analysis of the invaded PANC?1 compared with the whole cultured PANC?1 was performed using CE?TOFMS, and concentrations of 110 met...

Study of cancer cell lines with Fourier transform infrared (FTIR)/vibrational absorption (VA) spectroscopy

DEFF Research Database (Denmark)

Uceda Otero, E. P.; Eliel, G. S. N.; Fonseca, E. J. S.

2013-01-01

In this work we have used Fourier transform infrared (FTIR) / vibrational absorption (VA) spectroscopy to study two cancer cell lines: the Henrietta Lacks (HeLa) human cervix carcinoma and 5637 human bladder carcinoma cell lines. Our goal is to experimentally investigate biochemical changes...

Biochemical and Molecular Characterization of Saccharomyces cerevisiae Strains Obtained from Sugar-Cane Juice Fermentations and Their Impact in Cachaça Production▿

Science.gov (United States)

Oliveira, Valdinéia Aparecida; Vicente, Maristela Araújo; Fietto, Luciano Gomes; de Miranda Castro, Ieso; Coutrim, Maurício Xavier; Schüller, Dorit; Alves, Henrique; Casal, Margarida; de Oliveira Santos, Juliana; Araújo, Leandro Dias; da Silva, Paulo Henrique Alves; Brandão, Rogelio Lopes

2008-01-01

Saccharomyces cerevisiae strains from different regions of Minas Gerais, Brazil, were isolated and characterized aiming at the selection of starter yeasts to be used in the production of cachaça, the Brazilian sugar cane spirit. The methodology established took into account the screening for biochemical traits desirable in a yeast cachaça producer, such as no H2S production, high tolerance to ethanol and high temperatures, high fermentative capacity, and the abilities to flocculate and to produce mycocins. Furthermore, the yeasts were exposed to drugs such as 5,5′,5"-trifluor-d,l-leucine and cerulenin to isolate those that potentially overproduce higher alcohols and esters. The utilization of a random amplified polymorphic DNA-PCR method with primers based on intron splicing sites flanking regions of the COX1 gene, as well as microsatellite analysis, was not sufficient to achieve good differentiation among selected strains. In contrast, karyotype analysis allowed a clear distinction among all strains. Two selected strains were experimentally evaluated as cachaça producers. The results suggest that the selection of strains as fermentation starters requires the combined use of biochemical and molecular criteria to ensure the isolation and identification of strains with potential characteristics to produce cachaça with a higher quality standard. PMID:18065624

Synthesis of Biochemical Applications on Digital Microfluidic Biochips with Operation Execution Time Variability

DEFF Research Database (Denmark)

Alistar, Mirela; Pop, Paul

2015-01-01

that each biochemical operation in an application is characterized by a worst-case execution time (wcet). However, during the execution of the application, due to variability and randomness in biochemical reactions, operations may finish earlier than their wcetswcets, resulting in unexploited slack...... in the schedule. In this paper, we first propose an online synthesis strategy that re-synthesizes the application at runtime when operations experience variability in their execution time, exploiting thus the slack to obtain shorter application completion times. We also propose a quasi-static synthesis strategy...... approaches have been proposed for the synthesis of digital microfluidic biochips, which, starting from a biochemical application and a given biochip architecture, determine the allocation, resource binding, scheduling, placement and routing of the operations in the application. Researchers have assumed...

Characterization of blood biochemical markers during aging in the Grey Mouse Lemur (Microcebus murinus: impact of gender and season

Directory of Open Access Journals (Sweden)

Marchal Julia

2012-11-01

Full Text Available Abstract Background Hematologic and biochemical data are needed to characterize the health status of animal populations over time to determine the habitat quality and captivity conditions. Blood components and the chemical entities that they transport change predominantly with sex and age. The aim of this study was to utilize blood chemistry monitoring to establish the reference levels in a small prosimian primate, the Grey Mouse Lemur (Microcebus murinus. Method In the captive colony, mouse lemurs may live 10–12 years, and three age groups for both males and females were studied: young (1–3 years, middle-aged (4–5 years and old (6–10 years. Blood biochemical markers were measured using the VetScan Comprehensive Diagnostic Profile. Because many life history traits of this primate are highly dependent on the photoperiod (body mass and reproduction, the effect of season was also assessed. Results The main effect of age was observed in blood markers of renal functions such as creatinine, which was higher among females. Additionally, blood urea nitrogen significantly increased with age and is potentially linked to chronic renal insufficiency, which has been described in captive mouse lemurs. The results demonstrated significant effects related to season, especially in blood protein levels and glucose rates; these effects were observed regardless of gender or age and were likely due to seasonal variations in food intake, which is very marked in this species. Conclusion These results were highly similar with those obtained in other primate species and can serve as references for future research of the Grey Mouse Lemur.

Biochemical characterization of indigenous Fulani and Yoruba ...

African Journals Online (AJOL)

The study was carried out to characterize two indigenous chickens of Nigeria using protein markers; haemolglobin (HB) and carbonic anhydrase (CA). Separation of the two proteins was achieved by cellulose acetate electrophoresis and direct gene counting method was employed to interpret the result. Palentological ...

Biochemical comparison of osteoarthritic knees with and without effusion

Science.gov (United States)

2011-01-01

Background Several symptom-relieving interventions have been shown to be efficacious among osteoarthritis (OA) patients with knee effusion; however, not every symptomatic knee OA patient has clinical effusion. Results may be over-generalized since it is unclear if effused knees represent a unique pathological condition or subset compared to knees without effusion. The primary purpose of this study was to determine if biochemical differences existed between OA knees with and without effusion. Methods The present cross-sectional study consisted of 22 volunteers (11 with knee effusion, 11 without knee effusion) with confirmed late-stage radiographic knee OA (Kellgren-Lawrence score ≥ 3). Synovial fluid samples were collected and analyzed using a custom multiplex enzyme-linked immunosorbent assay to determine eight specific biomarker concentrations (e.g., catabolic, anabolic). Results Matrix metalloproteinase (MMP)-3, tissue inhibitor of MMPs (TIMP)-1, TIMP-2, and interleukin-10 were significantly higher in the knees with effusion than in the knees without effusion. Conclusions The biochemical differences that existed between knees with and without effusion provide support that OA subsets may exist, characterized by distinct biochemical characteristics and clinical findings (e.g., effusion). PMID:22122951

Establishment of a novel human medulloblastoma cell line characterized by highly aggressive stem-like cells.

Science.gov (United States)

Silva, Patrícia Benites Gonçalves da; Rodini, Carolina Oliveira; Kaid, Carolini; Nakahata, Adriana Miti; Pereira, Márcia Cristina Leite; Matushita, Hamilton; Costa, Silvia Souza da; Okamoto, Oswaldo Keith

2016-08-01

Medulloblastoma is a highly aggressive brain tumor and one of the leading causes of morbidity and mortality related to childhood cancer. These tumors display differential ability to metastasize and respond to treatment, which reflects their high degree of heterogeneity at the genetic and molecular levels. Such heterogeneity of medulloblastoma brings an additional challenge to the understanding of its physiopathology and impacts the development of new therapeutic strategies. This translational effort has been the focus of most pre-clinical studies which invariably employ experimental models using human tumor cell lines. Nonetheless, compared to other cancers, relatively few cell lines of human medulloblastoma are available in central repositories, partly due to the rarity of these tumors and to the intrinsic difficulties in establishing continuous cell lines from pediatric brain tumors. Here, we report the establishment of a new human medulloblastoma cell line which, in comparison with the commonly used and well-established cell line Daoy, is characterized by enhanced proliferation and invasion capabilities, stem cell properties, increased chemoresistance, tumorigenicity in an orthotopic metastatic model, replication of original medulloblastoma behavior in vivo, strong chromosome structural instability and deregulation of genes involved in neural development. These features are advantageous for designing biologically relevant experimental models in clinically oriented studies, making this novel cell line, named USP-13-Med, instrumental for the study of medulloblastoma biology and treatment.

Biochemical and Functional Characterization of Parawixia bistriata Spider Venom with Potential Proteolytic and Larvicidal Activities

Science.gov (United States)

Gimenez, Gizeli S.; Coutinho-Neto, Antonio; Kayano, Anderson M.; Simões-Silva, Rodrigo; Trindade, Frances; de Almeida e Silva, Alexandre; Marcussi, Silvana; da Silva, Saulo L.; Fernandes, Carla F. C.; Zuliani, Juliana P.; Calderon, Leonardo A.; Soares, Andreimar M.; Stábeli, Rodrigo G.

2014-01-01

Toxins purified from the venom of spiders have high potential to be studied pharmacologically and biochemically. These biomolecules may have biotechnological and therapeutic applications. This study aimed to evaluate the protein content of Parawixia bistriata venom and functionally characterize its proteins that have potential for biotechnological applications. The crude venom showed no phospholipase, hemorrhagic, or anti-Leishmania activities attesting to low genotoxicity and discrete antifungal activity for C. albicans. However the following activities were observed: anticoagulation, edema, myotoxicity and proteolysis on casein, azo-collagen, and fibrinogen. The chromatographic and electrophoretic profiles of the proteins revealed a predominance of acidic, neutral, and polar proteins, highlighting the presence of proteins with high molecular masses. Five fractions were collected using cation exchange chromatography, with the P4 fraction standing out as that of the highest purity. All fractions showed proteolytic activity. The crude venom and fractions P1, P2, and P3 showed larvicidal effects on A. aegypti. Fraction P4 showed the presence of a possible metalloprotease (60 kDa) that has high proteolytic activity on azo-collagen and was inhibited by EDTA. The results presented in this study demonstrate the presence of proteins in the venom of P. bistriata with potential for biotechnological applications. PMID:24895632

Initial biochemical and functional characterization of a 5'-nucleotidase from Xylella fastidiosa related to the human cytosolic 5'-nucleotidase I.

Science.gov (United States)

Santos, Clelton A; Saraiva, Antonio M; Toledo, Marcelo A S; Beloti, Lilian L; Crucello, Aline; Favaro, Marianna T P; Horta, Maria A C; Santiago, André S; Mendes, Juliano S; Souza, Alessandra A; Souza, Anete P

2013-01-01

The 5'-nucleotidases constitute a ubiquitous family of enzymes that catalyze either the hydrolysis or the transfer of esterified phosphate at the 5' position of nucleoside monophosphates. These enzymes are responsible for the regulation of nucleotide and nucleoside levels in the cell and can interfere with the phosphorylation-dependent activation of nucleoside analogs used in therapies targeting solid tumors and viral infections. In the present study, we report the initial biochemical and functional characterization of a 5'-nucleotidase from Xylella fastidiosa that is related to the human cytosolic 5'-nucleotidase I. X. fastidiosa is a plant pathogenic bacterium that is responsible for numerous economically important crop diseases. Biochemical assays confirmed the phosphatase activity of the recombinant purified enzyme and revealed metal ion dependence for full enzyme activity. In addition, we investigated the involvement of Xf5'-Nt in the formation of X. fastidiosa biofilms, which are structures that occlude the xylem vessels of susceptible plants and are strictly associated with bacterial pathogenesis. Using polyclonal antibodies against Xf5'-Nt, we observed an overexpression of Xf5'-Nt during the initial phases of X. fastidiosa biofilm formation that was not observed during X. fastidiosa planktonic growth. Our results demonstrate that the de/phosphorylation network catalyzed by 5'-nucleotidases may play an important role in bacterial biofilm formation, thereby contributing novel insights into bacterial nucleotide metabolism and pathogenicity. Copyright © 2013 Elsevier Ltd. All rights reserved.

Establishment and characterization of a unique 1 μm diameter liver-derived progenitor cell line

International Nuclear Information System (INIS)

Aravalli, Rajagopal N.; Behnan Sahin, M.; Cressman, Erik N.K.; Steer, Clifford J.

2010-01-01

Liver-derived progenitor cells (LDPCs) are recently identified novel stem/progenitor cells from healthy, unmanipulated adult rat livers. They are distinct from other known liver stem/progenitor cells such as the oval cells. In this study, we have generated a LDPC cell line RA1 by overexpressing the simian virus 40 (SV40) large T antigen (TAg) in primary LDPCs. This cell line was propagated continuously for 55 passages in culture, after which it became senescent. Interestingly, following transformation with SV40 TAg, LDPCs decreased in size significantly and the propagating cells measured 1 μm in diameter. RA1 cells proliferated in vitro with a doubling time of 5-7 days, and expressed cell surface markers of LDPCs. In this report, we describe the characterization of this novel progenitor cell line that might serve as a valuable model to study liver cell functions and stem cell origin of liver cancers.

Establishment and characterization of a unique 1 {mu}m diameter liver-derived progenitor cell line

Energy Technology Data Exchange (ETDEWEB)

Aravalli, Rajagopal N., E-mail: arava001@umn.edu [Department of Radiology, University of Minnesota Medical School, Minneapolis, MN 55455 (United States); Behnan Sahin, M. [Department of Medicine, University of Minnesota Medical School, Minneapolis, MN 55455 (United States); Cressman, Erik N.K. [Department of Radiology, University of Minnesota Medical School, Minneapolis, MN 55455 (United States); Steer, Clifford J., E-mail: steer001@umn.edu [Department of Medicine, University of Minnesota Medical School, Minneapolis, MN 55455 (United States); Department of Genetics, Cell Biology, and Development, University of Minnesota Medical School, Minneapolis, MN 55455 (United States)

2010-01-01

Liver-derived progenitor cells (LDPCs) are recently identified novel stem/progenitor cells from healthy, unmanipulated adult rat livers. They are distinct from other known liver stem/progenitor cells such as the oval cells. In this study, we have generated a LDPC cell line RA1 by overexpressing the simian virus 40 (SV40) large T antigen (TAg) in primary LDPCs. This cell line was propagated continuously for 55 passages in culture, after which it became senescent. Interestingly, following transformation with SV40 TAg, LDPCs decreased in size significantly and the propagating cells measured 1 {mu}m in diameter. RA1 cells proliferated in vitro with a doubling time of 5-7 days, and expressed cell surface markers of LDPCs. In this report, we describe the characterization of this novel progenitor cell line that might serve as a valuable model to study liver cell functions and stem cell origin of liver cancers.

Molecular Analysis, Biochemical Characterization, Antimicrobial Activity, and Immunological Analysis of Proteus mirabilis Isolated from Broilers.

Science.gov (United States)

Yeh, Hung-Yueh; Line, John E; Hinton, Arthur

2018-03-01

Proteus mirabilis, a Gram-negative bacterium, is ubiquitous in the environment and is considered as the normal microflora in the human gastrointestinal tract. However, this bacterium is an opportunistic pathogen in humans, often causing urinary tract infections. Moreover, Proteus has been frequently isolated from food animals, including poultry. Whether this bacterium contributes to the foodborne illness in humans is unclear. In this report, P. mirabilis isolates recovered from broilers during housing in the units were characterized, their antimicrobial activity was assayed, and broiler immune response to the soluble proteins was determined. Cecal contents and fecal droppings were treated according to the standard protocol for isolation. Speciation based on biochemical reactions and the antimicrobial activity of the isolates were carried out using commercial kits. Immunoblot was assayed to determine immune status of broilers against P. mirabilis. A total of 10 isolates of P. mirabilis were selected for further characterization. These isolates could grow in pH 6.0 and 1% NaCl conditions. They were resistant to sodium lactate, troleandomycin, rifamycin SV, vancomycin, but sensitive to nalidixic acid, cefotaxime and novobiocin. Moreover, the CTX, ACC, CMY-1, BIC, NDM, VEB, qnrB and qnrD genes were detected by PCR amplification in all isolates. Sera from broilers harboring this bacterium reacted to the P. mirabilis soluble proteins, but not from litter- and age-matched P. mirabilis negative and SPF chickens, indicating that this bacterium infected chickens that could have humoral immune response against P. mirabilis. This study provides a rationale for further monitoring P. mirabilis during poultry production to determine whether this bacterium poses potential threats to public health. Published 2018. This article is a U.S. Government work and is in the public domain in the USA.

Characterization of breakpoint cluster region kinase and SH2-binding activities.

Science.gov (United States)

Afar, D E; Witte, O N

1995-01-01

BCR is an interesting signaling protein, whose cellular function is currently unknown. Its biochemical properties include serine kinase activity, SH2-binding activity, and a GTPase-activating activity. The SH2-binding activity is particularly interesting because it may link BCR to signaling pathways involving SH2-containing molecules. Since tyrosine phosphorylation of BCR has been detected in CML-derived cell lines and since tyrosine-phosphorylated BCR shows increased affinity toward certain SH2 domains, it seems particularly important to further characterize this activity. This chapter described a simple purification scheme for partial purification of BCR, which can be used to assess in vitro kinase and SH2-binding activities.

Physiological and biochemical characterization of mango tree with paclobutrazol application via irrigation.

OpenAIRE

SOUZA, M. A. de; MESQUITA, A. C.; SIMOES, W. L.; FERREITA, K. M.; ARAÚJO, E. F. J.

2017-01-01

Paclobutrazol application in mango tree floral induction is followed by changes in the hormonal balance and carbohydrates production. This study aimed at evaluating the physiological and biochemical variables of mango tree (Palmer cultivar) by applying paclobutrazol doses via irrigation. The experimental design was randomized blocks, with five paclobutrazol doses (0.7 g, 1.0 g, 1.3 g, 1.6 g and 1.9 g of a.i. per linear meter of canopy) and one additional treatment, with one dose in the conven...

Biochemical Importance of Glycosylation of Plasminogen Activator Inhibitor-1

DEFF Research Database (Denmark)

Gils, Ann; Pedersen, Katrine Egelund; Skottrup, Peter

2003-01-01

The serpin plasminogen activator inhibitor-1 (PAI-1) is a potential target for anti-thrombotic and anti-cancer therapy. PAI-1 has 3 potential sites for N-linked glycosylation. We demonstrate here that PAI-1 expressed recombinantly or naturally by human cell lines display a heterogeneous glycosyla......The serpin plasminogen activator inhibitor-1 (PAI-1) is a potential target for anti-thrombotic and anti-cancer therapy. PAI-1 has 3 potential sites for N-linked glycosylation. We demonstrate here that PAI-1 expressed recombinantly or naturally by human cell lines display a heterogeneous...... with the glycosylation sites could be excluded as explanation for the differential reactivity. The latency transition of non-glycosylated, but not of glycosylated PAI-1, was strongly accelerated by a non-ionic detergent. The different biochemical properties of glycosylated and non-glycosylated PAI-1 depended...

Characterization of immortalized MARCO and SR-AI/II-deficient murine alveolar macrophage cell lines

Directory of Open Access Journals (Sweden)

Imrich Amy

2008-05-01

Full Text Available Abstract Background Alveolar macrophages (AM avidly bind and ingest unopsonized inhaled particles and bacteria through class A scavenger receptors (SRAs MARCO and SR-AI/II. Studies to characterize the function of these SRAs have used AMs from MARCO or SR-AI/II null mice, but this approach is limited by the relatively low yield of AMs. Moreover, studies using both MARCO and SR-AI/II-deficient (MS-/- mice have not been reported yet. Hence, we sought to develop continuous cell lines from primary alveolar macrophages from MS-/- mice. Results We used in vitro infection of the primary AMs with the J2 retrovirus carrying the v-raf and v-myc oncogenes. Following initial isolation in media supplemented with murine macrophage colony-stimulating factor (M-CSF, we subcloned three AM cell lines, designated ZK-1, ZK-2 and ZK-6. These cell lines grow well in RPMI-1640-10% FBS in the absence of M-CSF. These adherent but trypsin-sensitive cell lines have a doubling time of approximately 14 hours, exhibit typical macrophage morphology, and express macrophage-associated cell surface Mac-1 (CD11b and F4/80 antigens. The cell lines show robust Fc-receptor dependent phagocytosis of opsonized red blood cells. Similar to freshly isolated AMs from MS-/- mice, the cell lines exhibit decreased phagocytosis of unopsonized titanium dioxide (TiO2, fluorescent latex beads and bacteria (Staphylococcus aureus compared with the primary AMs from wild type (WT C57BL/6 mice. Conclusion Our results indicated that three contiguous murine alveolar macrophage cell lines with MS-/- (ZK1, ZK2 and ZK6 were established successfully. These cell lines demonstrated macrophage morphology and functional activity. Interestingly, similar to freshly isolated AMs from MS-/- mice, the cell lines have a reduced, but not absent, ability to bind and ingest particles, with an altered pattern of blockade by scavenger receptor inhibitors. These cell lines will facilitate in vitro studies to further define

Biological and Biochemical Potential of Sea Snake Venom and Characterization of Phospholipase A2 and Anticoagulation Activity.

Science.gov (United States)

Damotharan, Palani; Veeruraj, Anguchamy; Arumugam, Muthuvel; Balasubramanian, Thangavel

2016-03-01

This study is designed to isolate and purify a novel anti-clotting protein component from the venom of Enhydrina schistosa, and explore its biochemical and biological activities. The active protein was purified from the venom of E. schistosa by ion-exchange chromatography using DEAE-cellulose. The venom protein was tested by various parameters such as, proteolytic, haemolytic, phospholipase and anti-coagulant activities. 80 % purity was obtained in the final stage of purification and the purity level of venom was revealed as a single protein band of about 44 kDa in SDS-polyacrylamide electrophoresis under reducing conditions. The results showed that the Potent hemolytic activity was observed against cow, goat, chicken and human (A, B and O positive) erythrocytes. Furthermore, the clotting assays showed that the venom of E. schistosa significantly prolonged in activated partial thromboplastin time, thrombin time, and prothrombin time. Venomous enzymes which hydrolyzed casein and gelatin substrate were found in this venom protein. Gelatinolytic activity was optimal at pH 5-9 and (1)H NMR analysis of purified venom was the base line information for the structural determination. These results suggested that the E. schistosa venom holds good promise for the development of novel lead compounds for pharmacological applications in near future.

Production and characterization of femtosecond laser-written double line waveguides in heavy metal oxide glasses

Science.gov (United States)

da Silva, Diego Silvério; Wetter, Niklaus Ursus; de Rossi, Wagner; Kassab, Luciana Reyes Pires; Samad, Ricardo Elgul

2018-01-01

We report the fabrication and characterization of double line waveguides directly written in tellurite and germanate glasses using a femtosecond laser delivering 30 μJ, 80 fs pulses at 4 kHz repetition rate. The double line waveguides produced presented internal losses inferior to 2.0 dB/cm. The output mode profile and the M2 measurements indicate multimodal guiding behavior. A better beam quality for the GeO2 - PbO waveguide was observed when compared with TeO2 - ZnO glass. Raman spectroscopy of the waveguides showed structural modification of the glassy network and indicates that a negative refractive index modification occurs at the focus of the laser beam, therefore allowing for light guiding in between two closely spaced laser written lines. The refractive index change at 632 nm is around 10-4, and the structural changes in the laser focal region of the writing, evaluated by Raman spectroscopy, corroborated our findings that these materials are potential candidates for optical waveguides and passive components. To the best of our knowledge, the two double line configuration demonstrated in the present work was not reported before for germanate or tellurite glasses.

Characterization and biological activities of extracellular melanin produced by Schizophyllum commune (Fries).

Science.gov (United States)

Arun, G; Eyini, M; Gunasekaran, P

2015-06-01

Melanins are enigmatic pigments produced by a wide variety of microorganisms including bacteria and fungi. Here, we have isolated and characterized extracellular melanin from mushroom fungus, Schizophyllum commune. The extracellular dark pigment produced by the broth culture of S. commune, after 21 days of incubation was recovered by hot acid-alkali treatment. The melanin nature of the pigment was characterized by biochemical tests and further, confirmed by UV, IR, EPR, NMR and MALDI-TOF Mass Spectra. Extracellular melanin, at 100 μg/ml, showed significant antibacterial activity against Escherichia coli, Bacillus subtilis, Klebsiella pneumoniae and Pseudomonas fluorescens and antifungal activity against Trichophyton simii and T. rubrum. At a concentration of 50 μg/ml, melanin showed high free radical scavenging activity of DPPH (2,2-diphenyl-1-picrylhydrazyl) indicating its antioxidant potential. It showed concentration dependent inhibition of cell proliferation of Human Epidermoid Larynx Carcinoma Cell Line (HEP-2). This study has demonstrated characterization of melanin from basidiomycetes mushroom fungus, Schizophyllum commune and its applications.

Biochemical and morphological changes associated with macrophages and osteoclasts when challenged with infection - biomed 2011.

Science.gov (United States)

Wiggers, Erin Callie; Johnson, William; Tucci, Michelle; Benghuzzi, Hamed

2011-01-01

Osteomyelitis is a bacterial infection of the bone that occurs frequently as a complication of open fractures and various kinds of orthopedic surgery. This infection can often lead to more extensive surgeries and even death of the patient. In animal models of osteomyelitis, the site of infection by Staphylococcus aureus was observed to have high numbers of both macrophages and osteoclasts, both of which may contribute to large amounts of osteolysis and tissue damage. In order to evaluate the immune response in both types of cells, two cells lines, a macrophage cell line and a macrophage cell line stimulated to become osteoclasts by the addition of receptor activator of nuclear-factor B (RANKL), were exposed to lipopolysaccharides, opsonized S. aureus, and unopsonized S. aureus. The results showed that both cell types activated a biochemical cascade that included the release of cytokines and nitric oxide associated with cell damage and death in response to infection. However, macrophages and osteoclasts differed in response magnitude, most likely due to differences in cell-membrane receptors. This data supports the growing body of research that links the immune and skeletal systems. Further understanding of biochemical pathways shared by the two systems could lead to significant advances in the treatment of osteomyelitis and the success of prostheses.

Soya bean Gα proteins with distinct biochemical properties exhibit differential ability to complement Saccharomyces cerevisiae gpa1 mutant.

Science.gov (United States)

Roy Choudhury, Swarup; Wang, Yuqi; Pandey, Sona

2014-07-01

Signalling pathways mediated by heterotrimeric G-proteins are common to all eukaryotes. Plants have a limited number of each of the G-protein subunits, with the most elaborate G-protein network discovered so far in soya bean (Glycine max, also known as soybean) which has four Gα, four Gβ and ten Gγ proteins. Biochemical characterization of Gα proteins from plants suggests significant variation in their properties compared with the well-characterized non-plant proteins. Furthermore, the four soya bean Gα (GmGα) proteins exhibit distinct biochemical activities among themselves, but the extent to which such biochemical differences contribute to their in vivo function is also not known. We used the yeast gpa1 mutant which displays constitutive signalling and growth arrest in the pheromone-response pathway as an in vivo model to evaluate the effect of distinct biochemical activities of GmGα proteins. We showed that specific GmGα proteins can be activated during pheromone-dependent receptor-mediated signalling in yeast and they display different strengths towards complementation of yeast gpa1 phenotypes. We also identified amino acids that are responsible for differential complementation abilities of specific Gα proteins. These data establish that specific plant Gα proteins are functional in the receptor-mediated pheromone-response pathway in yeast and that the subtle biochemical differences in their activity are physiologically relevant.

Plasma density characterization at SPARC-LAB through Stark broadening of Hydrogen spectral lines

Energy Technology Data Exchange (ETDEWEB)

Filippi, F., E-mail: francesco.filippi@roma1.infn.it [Dipartimento di Scienze di Base e Applicate per l' Ingegneria (SBAI), ‘Sapienza’ Università di Roma, Via A. Scarpa 14-16, 00161 Roma (Italy); INFN-Roma1, Piazzale Aldo Moro, 2 00161 Roma (Italy); Anania, M.P.; Bellaveglia, M.; Biagioni, A.; Chiadroni, E. [Laboratori Nazionali di Frascati, INFN, Via E. Fermi, Frascati (Italy); Cianchi, A. [Dipartimento di Fisica, Universitá di Roma Tor Vergata, Via della Ricerca Scientifica 1, 00133 Roma (Italy); Di Giovenale, D.; Di Pirro, G.; Ferrario, M. [Laboratori Nazionali di Frascati, INFN, Via E. Fermi, Frascati (Italy); Mostacci, A.; Palumbo, L. [Dipartimento di Scienze di Base e Applicate per l' Ingegneria (SBAI), ‘Sapienza’ Università di Roma, Via A. Scarpa 14-16, 00161 Roma (Italy); INFN-Roma1, Piazzale Aldo Moro, 2 00161 Roma (Italy); Pompili, R.; Shpakov, V.; Vaccarezza, C.; Villa, F. [Laboratori Nazionali di Frascati, INFN, Via E. Fermi, Frascati (Italy); Zigler, A. [Hebrew University of Jerusalem, Jerusalem 91904 (Israel)

2016-09-01

Plasma-based acceleration techniques are of great interest for future, compact accelerators due to their high accelerating gradient. Both particle-driven and laser-driven Plasma Wakefield Acceleration experiments are foreseen at the SPARC-LAB Test Facility (INFN National Laboratories of Frascati, Italy), with the aim to accelerate high-brightness electron beams. In order to optimize the efficiency of the acceleration in the plasma and preserve the quality of the accelerated beam, the knowledge of the plasma electron density is mandatory. The Stark broadening of the Hydrogen spectral lines is one of the candidates used to characterize plasma density. The implementation of this diagnostic for plasma-based experiments at SPARC-LAB is presented. - Highlights: • Stark broadening of Hydrogen lines has been measured to determine plasma density. • Plasma density diagnostic tool for plasma-based experiments at SPARC-LAB is presented. • Plasma density in tapered laser triggered ablative capillary discharge was measured. • Results of plasma density measurements in ablative capillaries are shown.

Plasma density characterization at SPARC-LAB through Stark broadening of Hydrogen spectral lines

International Nuclear Information System (INIS)

Filippi, F.; Anania, M.P.; Bellaveglia, M.; Biagioni, A.; Chiadroni, E.; Cianchi, A.; Di Giovenale, D.; Di Pirro, G.; Ferrario, M.; Mostacci, A.; Palumbo, L.; Pompili, R.; Shpakov, V.; Vaccarezza, C.; Villa, F.; Zigler, A.

2016-01-01

Plasma-based acceleration techniques are of great interest for future, compact accelerators due to their high accelerating gradient. Both particle-driven and laser-driven Plasma Wakefield Acceleration experiments are foreseen at the SPARC-LAB Test Facility (INFN National Laboratories of Frascati, Italy), with the aim to accelerate high-brightness electron beams. In order to optimize the efficiency of the acceleration in the plasma and preserve the quality of the accelerated beam, the knowledge of the plasma electron density is mandatory. The Stark broadening of the Hydrogen spectral lines is one of the candidates used to characterize plasma density. The implementation of this diagnostic for plasma-based experiments at SPARC-LAB is presented. - Highlights: • Stark broadening of Hydrogen lines has been measured to determine plasma density. • Plasma density diagnostic tool for plasma-based experiments at SPARC-LAB is presented. • Plasma density in tapered laser triggered ablative capillary discharge was measured. • Results of plasma density measurements in ablative capillaries are shown.

In vitro biochemical characterization of all barley endosperm starch synthases

DEFF Research Database (Denmark)

Cuesta-Seijo, Jose A.; Nielsen, Morten M.; Ruzanski, Christian

2016-01-01

Starch is the main storage polysaccharide in cereals and the major source of calories in the human diet. It is synthesized by a panel of enzymes including five classes of starch synthases (SSs). While the overall starch synthase (SS) reaction is known, the functional differences between the five SS....... Here we provide a detailed biochemical study of the activity of all five classes of SSs in barley endosperm. Each enzyme was produced recombinantly in E. coli and the properties and modes of action in vitro were studied in isolation from other SSs and other substrate modifying activities. Our results...... define the mode of action of each SS class in unprecedented detail; we analyze their substrate selection, temperature dependence and stability, substrate affinity and temporal abundance during barley development. Our results are at variance with some generally accepted ideas about starch biosynthesis...

Establishment, characterization, virus susceptibility and transfection of cell lines from cobia, Rachycentron canadum (L.), brain and fin.

Science.gov (United States)

Cheng, T-C; Lai, Y-S; Lin, I-Y; Wu, C-P; Chang, S-L; Chen, T-I; Su, M-S

2010-02-01

Establishment and characterization of two cobia, Rachycentron canadum, cell lines derived from cobia brain (CB) and cobia fin (CF) are described. Caudal fin and brain from juvenile cobia were dissociated for 30 and 10 min, respectively, in phosphate-buffered saline containing 0.25% trypsin at 25 degrees C. The optimal culture condition for both dissociated cells (primary cell culture) was at 28 degrees C in Leibovitz-15 medium containing 10% foetal bovine serum. The cells have been sub-cultured at a ratio of 1:2 for more than 160 passages over a period of 3 years. Origin of the cultured cells was verified by comparison of their sequences of mitochondrial cytochrome oxidase subunit I genes (cox I) with the cox 1 sequence from cobia muscle tissue. The cell lines showed polyploidy. No mycoplasma contamination was detected. Susceptibility to grouper iridovirus was observed for the CB cell line but not the CF cell line. Both cell lines expressed green fluorescent protein after being transfected with green fluorescent reporter gene driven by the cytomegalovirus promoter.

In-line digital holographic sensor for monitoring and characterizing marine particulates

International Nuclear Information System (INIS)

Owen, Robert B.; Zozulya, Alex A.

2000-01-01

We report an in-line digital holographic sensor (DHS) for monitoring and characterizing marine particulates. This system images individual particles over a deep depth of field (>25 cm) with a resolution of 5 μm. The DHS projects a collimated beam through the water column and onto a lensless CCD array. Some light is diffracted by particulates and forms an object beam; the undeflected remainder constitutes the reference beam. The two beams combine at the CCD array and create an in-line hologram, which is then numerically reconstructed. The DHS eliminates many problems traditionally associated with holography. The CCD recording material considerably lowers the exposure time and eliminates most vibration problems. The laser power needs are low; the DHS uses a small 10-mW diode laser. Rapid numerical reconstruction eliminates photographic processing and optical reconstruction. We successfully operated the DHS underwater on a remotely operated vehicle; our test results include tracing a single particle from one hologram to the next, thus deriving a velocity vector for marine mass transport. We outline our digital holographic reconstruction procedure, and present our graphical user interface and user software tools. The DHS is particularly useful for providing in situ ground-truth measurements for environmental remote sensing. (c) 2000 Society of Photo-Optical Instrumentation Engineers

Characterization of bond line discontinuities in a high-Mn TWIP steel pipe welded by HF-ERW

Energy Technology Data Exchange (ETDEWEB)

Park, Gitae; Kim, Bongyoon; Kang, Yongjoon [Division of Materials Science and Engineering, Hanyang University, 222, Wangsimni-ro, Seongdong-gu, Seoul 04763 (Korea, Republic of); Kang, Heewoong [RD Team, Husteel, 131 Bugokgongdan-ro, Songak-eup, Dangjin-si, Chungnam 31721 (Korea, Republic of); Lee, Changhee, E-mail: chlee@hanyang.ac.kr [Division of Materials Science and Engineering, Hanyang University, 222, Wangsimni-ro, Seongdong-gu, Seoul 04763 (Korea, Republic of)

2016-08-15

In this work, the microstructure and defects in a high-frequency electrical resistance welded (HF-ERW) pipe of high-Mn twinning-induced plasticity (TWIP) steel were characterized. The microstructure of the base metal and the bond line were examined using both optical microscopy and scanning electron microscopy. The features of the bond line were similar to those of conventional steel. Simultaneously, the circumferential ductility was evaluated via a flaring test. It was concluded that the deterioration of the circumferential ductility in a high-Mn TWIP steel pipe was caused by irregular shaped oxide defects and a penetrator that had been formed during welding. Specifically, the penetrator, which is composed of MnO and Mn{sub 2}SiO{sub 4}, was found to be the most influential on the circumferential ductility of the welded pipe. The penetrator was analyzed using both an electron probe micro analyzer and transmission electron microscopy, and the formation sequence of the penetrator was evaluated. - Highlights: •This study focused on applying the HF-ERW process to the seam welding of expandable pipe using TWIP steels. •For improvement of the circumferential ductility, deterioration factors were characterized. •Penetrator which would mainly deteriorate the circumferential ductility consisted of round MnO and Mn{sub 2}SiO{sub 4}. •Metallurgical evidence of existing theory regarding the mechanism of defect formation during the HF-ERW was characterized.

Biochemical fingerprinting of Vibrio parahaemolyticus by the PhenePlate system: comparison between pandemic and non-pandemic serotypes.

Science.gov (United States)

Rahman, Mokhlasur; Bhuiyan, N A; Kuhn, I; Ramamurthy, T; Rahman, M; Mollby, R; Nair, G Balakrish

2006-10-01

During recent years a pandemic clone of Vibrio parahaemolyticus has emerged. Isolates of this clone are distributed among several serotypes, but are genotypically related. In the present study, a phenotyping method (biochemical fingerprinting) was used to characterize pandemic and non-pandemic isolates belonging to V. parahaemolyticus. It was found that the pandemic isolates showed a high level of phenotypic homogeneity and a majority of the pandemic isolates belonged to the same biochemical phenotype, whereas non-pandemic V. parahemolyticus isolates were more heterogeneous. In conclusion, biochemical fingerprinting of V. parahaemolyticus can be used as a first screening method to differentiate between pandemic and non-pandemic isolates of V. parahaemolyticus.

Characterizing subsurface water flow to artificial drain lines using fiber-optic distributed temperature sensing

Science.gov (United States)

Shults, D.; Brooks, E. S.; Heinse, R.; Keller, C. K.

2017-12-01

Over the last several years growers have experienced increasingly wet spring conditions in the Palouse Region located in North Idaho, Eastern Washington and Eastern Oregon. As a result more artificial drain lines are being installed so growers can access their fields earlier in the growing season. Additionally there has been increasing adoption of no-tillage practices among growers in order minimize erosion and runoff in the region. There is a growing body of evidence that suggests long-term no-tillage may lead to the establishment of large macropore networks through increased earthworm activity and the preservation of root channels. These macropore networks, in conjunctions with the presence of artificial drains lines, may create connected preferential flow paths from agricultural fields to receiving streams. This connectivity of flow paths from agricultural fields to receiving water bodies may increase the loading of nutrients and agricultural chemicals as some flow paths may largely bypass soil matrix interaction where materials can be sequestered. Our primary objective for this study was to characterize subsurface flow to two artificial drain lines, one under conventional tillage and the other under no-tillage, using distributed temperature sensing (DTS) technology. During the study (November 2016-April 2017) the near surface soil-water temperature was consistently colder than that of deeper depths. Temperature was thus used as a tracer as snow melt and soil-water moved from the near surface to the drain lines during snowmelt and precipitation events. The spatial and temporal variability of the temperature along the artificial drain line under no-tillage practices was found to be greater than that of the conventional tilled field. It is hypothesized that preferential flow paths are responsible for the increased variability of temperature seen in the drain line under long term no-till management. The temperature along the conventional till drain line showed a

BISEN: Biochemical simulation environment

NARCIS (Netherlands)

Vanlier, J.; Wu, F.; Qi, F.; Vinnakota, K.C.; Han, Y.; Dash, R.K.; Yang, F.; Beard, D.A.

2009-01-01

The Biochemical Simulation Environment (BISEN) is a suite of tools for generating equations and associated computer programs for simulating biochemical systems in the MATLAB® computing environment. This is the first package that can generate appropriate systems of differential equations for

Molecular and biochemical characterization of a novel intracellular invertase from Aspergillus niger with transfructosylating activity

NARCIS (Netherlands)

Goosen, C.; Yuan, X.L.; Munster, J.M. van; Ram, A.F.J.; Maarel, M.J.E.C. van der; Dijkhuizen, L.

2007-01-01

A novel subfamily of putative intracellular invertase enzymes (glycoside hydrolase family 32) has previously been identified in fungal genomes. Here, we report phylogenetic, molecular, and biochemical characteristics of SucB, one of two novel intracellular invertases identified in Aspergillus niger.

Comparative analysis and validation of the malachite green assay for the high throughput biochemical characterization of terpene synthases.

Science.gov (United States)

Vardakou, Maria; Salmon, Melissa; Faraldos, Juan A; O'Maille, Paul E

2014-01-01

Terpenes are the largest group of natural products with important and diverse biological roles, while of tremendous economic value as fragrances, flavours and pharmaceutical agents. Class-I terpene synthases (TPSs), the dominant type of TPS enzymes, catalyze the conversion of prenyl diphosphates to often structurally diverse bioactive terpene hydrocarbons, and inorganic pyrophosphate (PPi). To measure their kinetic properties, current bio-analytical methods typically rely on the direct detection of hydrocarbon products by radioactivity measurements or gas chromatography-mass spectrometry (GC-MS). In this study we employed an established, rapid colorimetric assay, the pyrophosphate/malachite green assay (MG), as an alternative means for the biochemical characterization of class I TPSs activity.•We describe the adaptation of the MG assay for turnover and catalytic efficiency measurements of TPSs.•We validate the method by direct comparison with established assays. The agreement of k cat/K M among methods makes this adaptation optimal for rapid evaluation of TPSs.•We demonstrate the application of the MG assay for the high-throughput screening of TPS gene libraries.

Body Composition and Ectopic Lipid Changes With Biochemical Control of Acromegaly.

Science.gov (United States)

Bredella, Miriam A; Schorr, Melanie; Dichtel, Laura E; Gerweck, Anu V; Young, Brian J; Woodmansee, Whitney W; Swearingen, Brooke; Miller, Karen K

2017-11-01

Acromegaly is characterized by growth hormone (GH) and insulinlike growth factor-1 (IGF-1) hypersecretion, and GH and IGF-1 play important roles in regulating body composition and glucose homeostasis. The purpose of our study was to investigate body composition including ectopic lipids, measures of glucose homeostasis, and gonadal steroids in patients with active acromegaly compared with age-, body mass index (BMI)-, and sex-matched controls and to determine changes in these parameters after biochemical control of acromegaly. Cross-sectional study of 20 patients with active acromegaly and 20 healthy matched controls. Prospective study of 16 patients before and after biochemical control of acromegaly. Body composition including ectopic lipids by magnetic resonance imaging/proton magnetic resonance spectroscopy; measures of glucose homeostasis by an oral glucose tolerance test; gonadal steroids. Patients with active acromegaly had lower mean intrahepatic lipid (IHL) and higher mean fasting insulin and insulin area under the curve (AUC) values than controls. Men with acromegaly had lower mean total testosterone, sex hormone-binding globulin, and estradiol values than male controls. After therapy, homeostasis model assessment of insulin resistance, fasting insulin level, and insulin AUC decreased despite an increase in IHL and abdominal and thigh adipose tissues and a decrease in muscle mass. Patients with acromegaly were characterized by insulin resistance and hyperinsulinemia but lower IHL compared with age-, BMI-, and sex-matched healthy controls. Biochemical control of acromegaly improved insulin resistance but led to a less favorable anthropometric phenotype with increased IHL and abdominal adiposity and decreased muscle mass. Copyright © 2017 Endocrine Society

Structural and Biochemical Characterization of Xylella fastidiosa DsbA Family Members: New insightsinto the Enzyme-Substrate Interaction

Energy Technology Data Exchange (ETDEWEB)

Rinaldi, F.; Meza, A; Gulmarges, B

2009-01-01

Disulfide oxidoreductase DsbA catalyzes disulfide bond formation in proteins secreted to the periplasm and has been related to the folding process of virulence factors in many organisms. It is among the most oxidizing of the thioredoxin-like proteins, and DsbA redox power is understood in terms of the electrostatic interactions involving the active site motif CPHC. The plant pathogen Xylella fastidiosa has two chromosomal genes encoding two oxidoreductases belonging to the DsbA family, and in one of them, the canonical motif CPHC is replaced by CPAC. Biochemical assays showed that both X. fastidiosa homologues have similar redox properties and the determination of the crystal structure of XfDsbA revealed substitutions in the active site of X. fastidiosa enzymes, which are proposed to compensate for the lack of the conserved histidine in XfDsbA2. In addition, electron density maps showed a ligand bound to the XfDsbA active site, allowing the characterization of the enzyme interaction with an 8-mer peptide. Finally, surface analysis of XfDsbA and XfDsbA2 suggests that X. fastidiosa enzymes may have different substrate specificities.

[Biochemical characterization of fractionated rat liver chromatin in experimental D-hypovitaminosis and after administration of steroidal drugs].

Science.gov (United States)

Levitskiĭ, E L; Kholodova, Iu D; Gubskiĭ, Iu I; Primak, R G; Chabannyĭ, V N; Kindruk, N L; Mozzhukhina, T G; Lenchevskaia, L K; Mironova, V N; Saad, L M

1993-01-01

Marked changes in the structural and functional characteristics of liver nuclear chromatin fractions are observed under experimental D-hypovitaminosis, which differ in the degree of transcriptional activity. DNA-polymerase activity and activity of the fraction, enriched with RNA-polymerase I, increases in the active fraction. Free radical LPO reactions are modified in the chromatin fraction with low activity and to the less degree in the active one. Disturbances of chromatine structural properties are caused with the change in the protein and lipid components of chromatin. Administration of ecdysterone preparations (separately and together with vitamin D3) has a partial corrective effect on structural and functional organization of nuclear chromatine. At the action of ecdysterone normalization of LPO reactions modified by pathological changes is observed in the chromatin fraction with low activity and to the less degree in the active one. This kind of influence corrects to the less degree chromatin functional activity and quantitative and qualitative modifications of its protein component. Simultaneous influence of ecdysterone and vitamin D3 leads to the partial normalization of the biochemical indices studied (except for those which characterize LPO reactions) mainly in the active chromatin fraction.

Development of a system for characterizing biomass quality of lignocellulosic feedstocks for biochemical conversion

Science.gov (United States)

Murphy, Patrick Thomas

The purpose of this research was twofold: (i) to develop a system for screening lignocellulosic biomass feedstocks for biochemical conversion to biofuels and (ii) to evaluate brown midrib corn stover as feedstock for ethanol production. In the first study (Chapter 2), we investigated the potential of corn stover from bm1-4 hybrids for increased ethanol production and reduced pretreatment intensity compared to corn stover from the isogenic normal hybrid. Corn stover from hybrid W64A X A619 and respective isogenic bm hybrids was pretreated by aqueous ammonia steeping using ammonium hydroxide concentrations from 0 to 30%, by weight, and the resulting residues underwent simultaneous saccharification and cofermentation (SSCF) to ethanol. Dry matter (DM) digested by SSCF increased with increasing ammonium hydroxide concentration across all genotypes (P>0.0001) from 277 g kg-1 DM in the control to 439 g kg-1 DM in the 30% ammonium hydroxide pretreatment. The bm corn stover materials averaged 373 g kg-1 DM of DM digested by SSCF compared with 335 g kg-1 DM for the normal corn stover (Pdetergent fiber (NDF) as a cell-wall isolation procedure, and (iii) elimination of the fermentation organism in the SSCF procedures used to determine biochemically available carbohydrates. The original and the HTP assay methods were compared using corn cobs, hybrid poplar, kenaf, and switchgrass. Biochemically available carbohydrates increased with the HTP methods in the corn cobs, hybrid poplar, and switchgrass, but remained the same in the kenaf. Total available carbohydrates increased and unavailable carbohydrates decreased with the HTP methods in the corn cobs and switchgrass and remained the same in the hybrid poplar and kenaf. There were no differences in total carbohydrates (CT) between the two methods. The final study evaluated the variability of biomass quality parameters in a set of corn stover samples, and developed calibration equations for determining parameter values using near

Characterization of a cultured human T-cell line with genetically altered ribonucleotide reductase activity. Model for immunodeficiency.

Science.gov (United States)

Waddell, D; Ullman, B

1983-04-10

From human CCRF-CEM T-cells growing in continuous culture, we have selected, isolated, and characterized a clonal cell line, APHID-D2, with altered ribonucleotide reductase activity. In comparative growth rate experiments, the APHID-D2 cell line is less sensitive than the parental cell line to growth inhibition by deoxyadenosine in the presence of 10 microM erythro-9-(2-hydroxy-3-nonyl)adenine, an inhibitor of adenosine deaminase. The APHID-D2 cell line has elevated levels of all four dNTPs. The resistance of the APHID-D2 cell line to growth inhibition by deoxyadenosine and the abnormal dNTP levels can be explained by the fact that the APHID-D2 ribonucleotide reductase, unlike the parental ribonucleotide reductase, is not normally sensitive to inhibition by dATP. These results suggest that the allosteric site of ribonucleotide reductase which binds both dATP and ATP is altered in the APHID-D2 line. The isolation of a mutant clone of human T-cells which contains a ribonucleotide reductase that has lost its normal sensitivity to dATP and which is resistant to deoxyadenosine-mediated growth inhibition suggests that a primary pathogenic target of accumulated dATP in lymphocytes from patients with adenosine deaminase deficiency may be the cellular ribonucleotide reductase.

Conventional cytogenetic characterization of a new cell line, ACP01, established from a primary human gastric tumor

Directory of Open Access Journals (Sweden)

E.M. Lima

2004-12-01

Full Text Available Gastric cancer is the second most frequent type of neoplasia and also the second most important cause of death in the world. Virtually all the established cell lines of gastric neoplasia were developed in Asian countries, and western countries have contributed very little to this area. In the present study we describe the establishment of the cell line ACP01 and characterize it cytogenetically by means of in vitro immortalization. Cells were transformed from an intestinal-type gastric adenocarcinoma (T4N2M0 originating from a 48-year-old male patient. This is the first gastric adenocarcinoma cell line established in Brazil. The most powerful application of the cell line ACP01 is in the assessment of cytotoxicity. Solid tumor cell lines from different origins have been treated with several conventional and investigational anticancer drugs. The ACP01 cell line is triploid, grows as a single, non-organized layer, similar to fibroblasts, with focus formation, heterogeneous division, and a cell cycle of approximately 40 h. Chromosome 8 trisomy, present in 60% of the cells, was the most frequent cytogenetic alteration. These data lead us to propose a multifactorial triggering of gastric cancer which evolves over multiple stages involving progressive genetic changes and clonal expansion.

Analysis of cell line variation in biochemical production of protoporphyrin IX

Science.gov (United States)

Gibbs, Summer L.; Chen, Bin; O'Hara, Julia A.; Hoopes, P. Jack; Hasan, Tayyaba; Pogue, Brian W.

2006-02-01

Protoporphyrin IX (PpIX) is produced via the heme synthesis pathway by the cell following administration of aminolevulinic acid (ALA). ALA synthase, the enzyme that produces ALA in the cell from glycine and succinyl-coenzyme A, is inhibited in a feedback mechanism by heme and thus is the rate limiting enzyme in the heme synthesis pathway. Since ALA is administered systemically, the rate limiting step that naturally exists in the cells is bypassed, however it is currently unclear why cells have different rate limiting steps in the ALA-PpIX synthesis pathway, and more specifically which types of cancer cells are most productive. It has been determined that when the same amount of ALA is administered to a wide panel of cancer cells in vitro that vastly differing amounts of PpIX are produced. The steps for the ALA-PpIX pathway occur in and around the mitochondria of the cell, but interestingly no correlation is seen between PpIX production and mitochondrial content of the cell, following ALA administration. However, total cell area shows positive correlation with PpIX production. Administration of the iron chelator, 1,2-dimethyl-3-hydroxy-4-pyridone (L1) in combination with ALA allows the final step in the heme synthesis pathway, conversion of PpIX to heme, to be delayed and thus increases the detectable amount of PpIX in each cell line. The cell lines that have the lowest PpIX production following administration of ALA alone show the largest increase in production following the combined administration of ALA and L1. PpIX fluorescence is thought to be a measure of cellular activity and the goal of the current study was to determine which cell lines would be the most promising targets for fluorescence detection or monitoring response to therapy. The results indicate that the cells with larger size and larger numbers of mitochondria may be good potential targets for this therapy. While this conclusion may appear obvious, it is not universally true, and cellular specific

Biochemical research elucidating metabolic pathways in Pneumocystis*

Directory of Open Access Journals (Sweden)

Kaneshiro E.S.

2010-12-01

Full Text Available Advances in sequencing the Pneumocystis carinii genome have helped identify potential metabolic pathways operative in the organism. Also, data from characterizing the biochemical and physiological nature of these organisms now allow elucidation of metabolic pathways as well as pose new challenges and questions that require additional experiments. These experiments are being performed despite the difficulty in doing experiments directly on this pathogen that has yet to be subcultured indefinitely and produce mass numbers of cells in vitro. This article reviews biochemical approaches that have provided insights into several Pneumocystis metabolic pathways. It focuses on 1 S-adenosyl-L-methionine (AdoMet; SAM, which is a ubiquitous participant in numerous cellular reactions; 2 sterols: focusing on oxidosqualene cyclase that forms lanosterol in P. carinii; SAM:sterol C-24 methyltransferase that adds methyl groups at the C-24 position of the sterol side chain; and sterol 14α-demethylase that removes a methyl group at the C-14 position of the sterol nucleus; and 3 synthesis of ubiquinone homologs, which play a pivotal role in mitochondrial inner membrane and other cellular membrane electron transport.

Aspects on the Physiological and Biochemical Foundations of Neurocritical Care

Directory of Open Access Journals (Sweden)

Carl-Henrik Nordström

2017-06-01

Full Text Available Neurocritical care (NCC is a branch of intensive care medicine characterized by specific physiological and biochemical monitoring techniques necessary for identifying cerebral adverse events and for evaluating specific therapies. Information is primarily obtained from physiological variables related to intracranial pressure (ICP and cerebral blood flow (CBF and from physiological and biochemical variables related to cerebral energy metabolism. Non-surgical therapies developed for treating increased ICP are based on knowledge regarding transport of water across the intact and injured blood–brain barrier (BBB and the regulation of CBF. Brain volume is strictly controlled as the BBB permeability to crystalloids is very low restricting net transport of water across the capillary wall. Cerebral pressure autoregulation prevents changes in intracranial blood volume and intracapillary hydrostatic pressure at variations in arterial blood pressure. Information regarding cerebral oxidative metabolism is obtained from measurements of brain tissue oxygen tension (PbtO2 and biochemical data obtained from intracerebral microdialysis. As interstitial lactate/pyruvate (LP ratio instantaneously reflects shifts in intracellular cytoplasmatic redox state, it is an important indicator of compromised cerebral oxidative metabolism. The combined information obtained from PbtO2, LP ratio, and the pattern of biochemical variables reveals whether impaired oxidative metabolism is due to insufficient perfusion (ischemia or mitochondrial dysfunction. Intracerebral microdialysis and PbtO2 give information from a very small volume of tissue. Accordingly, clinical interpretation of the data must be based on information of the probe location in relation to focal brain damage. Attempts to evaluate global cerebral energy state from microdialysis of intraventricular fluid and from the LP ratio of the draining venous blood have recently been presented. To be of clinical relevance

Biochemical Basis of Sestrin Physiological Activities

Energy Technology Data Exchange (ETDEWEB)

Ho, Allison; Cho, Chun-Seok; Namkoong, Sim; Cho, Uhn-Soo; Lee, Jun Hee (Michigan)

2016-05-10

Excessive accumulation of reactive oxygen species (ROS) and chronic activation of mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) are well-characterized promoters of aging and age-associated degenerative pathologies. Sestrins, a family of highly conserved stress-inducible proteins, are important negative regulators of both ROS and mTORC1 signaling pathways; however, the mechanistic basis of how Sestrins suppress these pathways remains elusive. In the past couple of years, breakthrough discoveries about Sestrin signaling and its molecular nature have markedly increased our biochemical understanding of Sestrin function. These discoveries have also uncovered new potential therapeutic strategies that may eventually enable us to attenuate aging and age-associated diseases.

Biochemical and molecular tools reveal two diverse Xanthomonas groups in bananas

DEFF Research Database (Denmark)

Adriko, John; Aritua, V.; Mortensen, Carmen Nieves

2016-01-01

Xanthomonas campestris pv. musacearum (Xcm) causing the banana Xanthomonas wilt (BXW) disease has been the main xanthomonad associated with bananas in East and Central Africa based on phenotypic and biochemical characteristics. However, biochemical methods cannot effectively distinguish between...... pathogenic and non-pathogenic xanthomonads. In this study, gram-negative and yellow-pigmented mucoid bacteria were isolated from BXW symptomatic and symptomless bananas collected from different parts of Uganda. Biolog, Xcm-specific (GspDm), Xanthomonas vasicola species-specific (NZ085) and Xanthomonas genus......-specific (X1623) primers in PCR, and sequencing of ITS region were used to identify and characterize the isolates. Biolog tests revealed several isolates as xanthomonads. The GspDm and NZ085 primers accurately identified three isolates from diseased bananas as Xcm and these were pathogenic when re...

Characterization of physical and biochemical changes in plasma treated spinach seed during germination

Science.gov (United States)

Hye Ji, Sang; Ki, Se Hoon; Kang, Min Ho; Choi, Jin Sung; Park, Yeunsoo; Oh, Jaesung; Kim, Seong Bong; Yoo, Suk Jae; Choi, Eun Ha; Park, Gyungsoon

2018-04-01

Despite the accumulating data on the effect of plasma on seed germination, mechanisms of plasma action need more extensive research. In a previous study, we observed that high voltage nanosecond pulsed plasma enhanced the germination of spinach seeds and subsequent seedling growth. As a follow-up study, we investigated the physico-chemical, biochemical, and molecular changes in seed after plasma treatment, focusing on the early germination stage, to elucidate mechanism(s) for the stimulating effects of plasma on seed germination. The primary radicle protruded from seeds exposed to high voltage nanosecond pulsed plasma (one shot) slightly faster than the control seeds. The hydrophilicity of the seed surface significantly increased after treatment with high voltage nanosecond pulsed plasma (one shot). However, a very subtle increase in water uptake by plasma treated seeds was observed. Raman and FTIR spectroscopy analyses on chloroform extract of seed coats demonstrated no significant chemical etching on the surface of plasma treated seeds. This may be related to no dramatic increase in water absorption by seeds. The level of GA hormone and starch hydrolysis inside the plasma treated seeds was significantly elevated within 24 h. Taken together, our results suggest that high voltage nanosecond pulsed plasma may not only enhance hydrophilicity of the seed surface but also stimulate biochemical and molecular processes inside seed, leading to enhanced embryonic development.

Structural And Biochemical Characterization of the Therapeutic A. Variabilis Phenylalanine Ammonia Lyase

Energy Technology Data Exchange (ETDEWEB)

Wang, L.; Gamez, A.; Archer, H.; Abola, E.E.; Sarkissian, C.N.; Fitzpatrick, P.; Wendt, D.; Zhang, Y.; Vellard, M.; Bliesath, J.; Bell, S.; Lemont, J.; Scriver, C.R.; Stevens, R.C.

2009-05-26

We have recently observed promising success in a mouse model for treating the metabolic disorder phenylketonuria with phenylalanine ammonia lyase (PAL) from Rhodosporidium toruloides and Anabaena variabilis. Both molecules, however, required further optimization in order to overcome problems with protease susceptibility, thermal stability, and aggregation. Previously, we optimized PAL from R. toruloides, and in this case we reduced aggregation of the A. variabilis PAL by mutating two surface cysteine residues (C503 and C565) to serines. Additionally, we report the structural and biochemical characterization of the A. variabilis PAL C503S/C565S double mutant and carefully compare this molecule with the R. toruloides engineered PAL molecule. Unlike previously published PAL structures, significant electron density is observed for the two active-site loops in the A. variabilis C503S/C565S double mutant, yielding a complete view of the active site. Docking studies and N-hydroxysuccinimide-biotin binding studies support a proposed mechanism in which the amino group of the phenylalanine substrate is attacked directly by the 4-methylidene-imidazole-5-one prosthetic group. We propose a helix-to-loop conformational switch in the helices flanking the inner active-site loop that regulates accessibility of the active site. Differences in loop stability among PAL homologs may explain the observed variation in enzyme efficiency, despite the highly conserved structure of the active site. A. variabilis C503S/C565S PAL is shown to be both more thermally stable and more resistant to proteolytic cleavage than R. toruloides PAL. Additional increases in thermal stability and protease resistance upon ligand binding may be due to enhanced interactions among the residues of the active site, possibly locking the active-site structure in place and stabilizing the tetramer. Examination of the A. variabilis C503S/C565S PAL structure, combined with analysis of its physical properties, provides

Biochemical characterization of enzyme fidelity of influenza A virus RNA polymerase complex.

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Shilpa Aggarwal

2010-04-01

Full Text Available It is widely accepted that the highly error prone replication process of influenza A virus (IAV, together with viral genome assortment, facilitates the efficient evolutionary capacity of IAV. Therefore, it has been logically assumed that the enzyme responsible for viral RNA replication process, influenza virus type A RNA polymerase (IAV Pol, is a highly error-prone polymerase which provides the genomic mutations necessary for viral evolution and host adaptation. Importantly, however, the actual enzyme fidelity of IAV RNA polymerase has never been characterized.Here we established new biochemical assay conditions that enabled us to assess both polymerase activity with physiological NTP pools and enzyme fidelity of IAV Pol. We report that IAV Pol displays highly active RNA-dependent RNA polymerase activity at unbiased physiological NTP substrate concentrations. With this robust enzyme activity, for the first time, we were able to compare the enzyme fidelity of IAV Pol complex with that of bacterial phage T7 RNA polymerase and the reverse transcriptases (RT of human immunodeficiency virus (HIV-1 and murine leukemia virus (MuLV, which are known to be low and high fidelity enzymes, respectively. We observed that IAV Pol displayed significantly higher fidelity than HIV-1 RT and T7 RNA polymerase and equivalent or higher fidelity than MuLV RT. In addition, the IAV Pol complex showed increased fidelity at lower temperatures. Moreover, upon replacement of Mg(++ with Mn(++, IAV Pol displayed increased polymerase activity, but with significantly reduced processivity, and misincorporation was slightly elevated in the presence of Mn(++. Finally, when the IAV nucleoprotein (NP was included in the reactions, the IAV Pol complex exhibited enhanced polymerase activity with increased fidelity.Our study indicates that IAV Pol is a high fidelity enzyme. We envision that the high fidelity nature of IAV Pol may be important to counter-balance the multiple rounds of

Molecular characterization and functional analysis of pteridine reductase in wild-type and antimony-resistant Leishmania lines.

Science.gov (United States)

de Souza Moreira, Douglas; Ferreira, Rafael Fernandes; Murta, Silvane M F

2016-01-01

Pteridine reductase (PTR1) is an NADPH-dependent reductase that participates in the salvage of pteridines, which are essential to maintain growth of Leishmania. In this study, we performed the molecular characterization of ptr1 gene in wild-type (WTS) and SbIII-resistant (SbR) lines from Leishmania guyanensis (Lg), Leishmania amazonensis (La), Leishmania braziliensis (Lb) and Leishmania infantum (Li), evaluating the chromosomal location, mRNA levels of the ptr1 gene and PTR1 protein expression. PFGE results showed that the ptr1 gene is located in a 797 kb chromosomal band in all Leishmania lines analyzed. Interestingly, an additional chromosomal band of 1070 kb was observed only in LbSbR line. Northern blot results showed that the levels of ptr1 mRNA are increased in the LgSbR, LaSbR and LbSbR lines. Western blot assays using the polyclonal anti-LmPTR1 antibody demonstrated that PTR1 protein is more expressed in the LgSbR, LaSbR and LbSbR lines compared to their respective WTS counterparts. Nevertheless, no difference in the level of mRNA and protein was observed between the LiWTS and LiSbR lines. Functional analysis of PTR1 enzyme was performed to determine whether the overexpression of ptr1 gene in the WTS L. braziliensis and L. infantum lines would change the SbIII-resistance phenotype of transfected parasites. Western blot results showed that the expression level of PTR1 protein was increased in the transfected parasites compared to the non-transfected ones. IC50 analysis revealed that the overexpression of ptr1 gene in the WTS L. braziliensis line increased 2-fold the SbIII-resistance phenotype compared to the non-transfected counterpart. Furthermore, the overexpression of ptr1 gene in the WTS L. infantum line did not change the SbIII-resistance phenotype. These results suggest that the PTR1 enzyme may be implicated in the SbIII-resistance phenotype in L. braziliensis line. Copyright © 2015 Elsevier Inc. All rights reserved.

Recent Advances on the Use of Biochemical Extracts as Filaricidal Agents

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Nazeh M. Al-Abd

2013-01-01

Full Text Available Lymphatic filariasis is a parasitic infection that causes a devastating public health and socioeconomic burden with an estimated infection of over 120 million individuals worldwide. The infection is caused by three closely related nematode parasites, namely, Wuchereria bancrofti, Brugia malayi, and B. timori, which are transmitted to human through mosquitoes of Anopheles, Culex, and Aedes genera. The species have many ecological variants and are diversified in terms of their genetic fingerprint. The rapid spread of the disease and the genetic diversification cause the lymphatic filarial parasites to respond differently to diagnostic and therapeutic interventions. This in turn prompts the current challenge encountered in its management. Furthermore, most of the chemical medications used are characterized by adverse side effects. These complications urgently warrant intense prospecting on bio-chemicals that have potent efficacy against either the filarial worms or thier vector. In lieu of this, we presented a review on recent literature that reported the efficacy of filaricidal biochemicals and those employed as vector control agents. In addition, methods used for biochemical extraction, screening procedures, and structure of the bioactive compounds were also presented.

Biochemical and chemical characterization of pink-pigmented oxidative bacteria.

Science.gov (United States)

Wallace, P L; Hollis, D G; Weaver, R E; Moss, C W

1990-04-01

The biochemical and chemical characteristics were determined for 156 clinical isolates of pink-pigmented bacteria that are similar to but distinct from Methylobacterium extorquens (synonymous with Pseudomonas mesophilica). These isolates were gram-negative, nonfermentative, usually nonvacuolated, coccoid rods; all grew at 35 degrees C and were catalase and urease positive; the majority grew on MacConkey agar and were variable for oxidase production and motility. On the basis of oxidation of xylose and mannitol and hydrolysis of esculin, these 156 strains were subdivided into four groups that were designated "pink coccoid" groups I, II, III, and IV. Groups I, II, and III are similar to an unnamed taxon described by Gilardi and Faur in 1984; only strains of group IV hydrolyze esculin. The cellular fatty acid compositions of strains of groups I, II, and III were essentially identical and differed from strains of group IV by the absence of 3-OH-C14:0 and the presence of C19:0 delta and 2-OH-C19:0 delta. The fatty acid composition of group IV strains was most similar to that of M. extorquens but differed by the presence of small amounts of two C17:1 acids, 3-OH-C16:0, and 2-OH-C18:1.

The alterations in biochemical signaling of hippocampal network activity in the autism brain The alterations in biochemical signaling of hippocampal network activity in the autism brain The alterations in biochemical signaling of hippocampal network activity in the autism brain

Institute of Scientific and Technical Information of China (English)

田允; 黄继云; 王锐; 陶蓉蓉; 卢应梅; 廖美华; 陆楠楠; 李静; 芦博; 韩峰

2012-01-01

Autism is a highly heritable neurodevelopmental condition characterized by impaired social interaction and communication. However, the role of synaptic dysfunction during development of autism remains unclear. In the present study, we address the alterations of biochemical signaling in hippocampal network following induction of the autism in experimental animals. Here, the an- imal disease model and DNA array being used to investigate the differences in transcriptome or- ganization between autistic and normal brain by gene co--expression network analysis.

Insight into Biochemical Characterization of Plant Sesquiterpene Synthases

DEFF Research Database (Denmark)

Manczak, Tom; Simonsen, Henrik Toft

2016-01-01

A fast and reproducible protocol was established for enzymatic characterization of plant sesquiterpene synthases that can incorporate radioactivity in their products. The method utilizes the 96-well format in conjunction with cluster tubes and enables processing of >200 samples a day. Along...... with reduced reagent usage, it allows further reduction in the use of radioactive isotopes and flammable organic solvents. The sesquiterpene synthases previously characterized were expressed in yeast, and the plant-derived Thapsia garganica kunzeaol synthase TgTPS2 was tested in this method. KM for TgTPS2...... was found to be 0.55 μM; the turnover number, kcat, was found to be 0.29 s-1, kcat for TgTPS2 is in agreement with that of terpene synthases of other plants, and kcat/KM was found to be 0.53 s-1 μM-1 for TgTPS2. The kinetic parameters were in agreement with previously published data....

Characterization of Camptothecin-induced Genomic Changes in the Camptothecin-resistant T-ALL-derived Cell Line CPT-K5

DEFF Research Database (Denmark)

Kjeldsen, Eigil; Nielsen, Christine J F; Roy, Amit

2018-01-01

-K5 and its parental cell line. We identified copy number alterations affecting genes important for maintaining genome integrity and reducing CPT-induced DNA damage. We show for the first time that short tandem repeats are targets for TOP1 cleavage, that can be differentially stimulated by CPT.......Acquisition of resistance to topoisomerase I (TOP1)-targeting camptothecin (CPT) derivatives is a major clinical problem. Little is known about the underlying chromosomal and genomic mechanisms. We characterized the CPT-K5 cell line expressing mutant CPT-resistant TOP1 and its parental T......-cell derived acute lymphoblastic leukemia CPT-sensitive RPMI-8402 cell line by karyotyping and molecular genetic methods, including subtractive oligo-based array comparative genomic hybridization (soaCGH) analysis. Karyotyping revealed that CPT-K5 cells had acquired additional structural aberrations...

Characterization of selection effects on broiler lines using DNA fingerprinting

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GS Schmidt

2003-05-01

Full Text Available The objective of this study was to evaluate the effect of selection for body weight on the genetic variability and diversity in broiler lines. Two paternal broiler lines (LL and LLc were used. LL line was selected for 12 generations for growth and carcass and reproduction characteristics. The LLc line was established from LL line in 1985 and mated at random. Blood samples from six chickens per line were collected and used for molecular analysis. Also, a DNA pool was made for each line to compare effects between lines. Data were analyzed considering the collected information on the presence or absence of DNA bands. Band sharing scores were calculated using the DICE coefficient. The pattern of the 21 most representative bands was used. DNA fingerprinting (DFP showed 90.48 % of polymorphism bands for both lines. Difference between lines was not due to the presence or absence of bands, but to the frequency of such bands in each genotype. Considering that both lines had the same genetic background, changes on band frequency were probably due to selection. Selection for body weight had an effect on the band frequency as evaluated by DFP, and for this reason this technique could be used as a tool in the selection process. Results also suggest that bands 4, 5 and 19 were linked to body weight traits, and bands 9, 10, 12, 13 and 21 were linked to reproductive traits such as egg production.

Experimental characterization of the flux-line lattice in superconducting V3Si

International Nuclear Information System (INIS)

Christen, D.; Chang, Y.; Kerchner, H.; Larson, B.; Narayan, J.; SeKula, S.

1983-01-01

Several microscopic properties of the flux-line lattice (FLL) in three separate single crystals of V 3 Si have been investigated by means of small-angle neutron diffraction. These low-field FLL characterizations have been correlated with the following material and superconducting properties: the real crystal symmetry parallel to the applied magnetic field; the micro-structure as determined by TEM; magnetic irreversibilities in the mixed state; reversible flux-line motion in ac response; martensitic structural transformation observed by X-ray diffraction. The three samples, V 3 Si-MP3, -MP4, and -MP5 possessed different defect structures, and this was manifested foremost in the FLL perfection. At low field (B 0-.5 T, but a highly mosaic, nearly polycrystalline FLL at lower fields. Sample MP4 contained large (500-1000 A) incoherent precipitates, and showed only a polycrystalline FLL at low field. In both MP3 and MP5, distinct anisotropic correlations were observed between the FLL morphology and the real-crystal direction along the applied field. The FLL perfection was strongly dependent on the growth history. The peak width history dependence for two different scattering geometries can be qualitatively modeled by proposed flux-pinning mechanisms. Quantitative comparisons with critical current measurements, however, are not totally reconcilable

Characterization of environmental chemicals with potential for DNA damage using isogenic DNA repair-deficient chicken DT40 cell lines.

Science.gov (United States)

Yamamoto, Kimiyo N; Hirota, Kouji; Kono, Koichi; Takeda, Shunichi; Sakamuru, Srilatha; Xia, Menghang; Huang, Ruili; Austin, Christopher P; Witt, Kristine L; Tice, Raymond R

2011-08-01

Included among the quantitative high throughput screens (qHTS) conducted in support of the US Tox21 program are those being evaluated for the detection of genotoxic compounds. One such screen is based on the induction of increased cytotoxicity in seven isogenic chicken DT40 cell lines deficient in DNA repair pathways compared to the parental DNA repair-proficient cell line. To characterize the utility of this approach for detecting genotoxic compounds and identifying the type(s) of DNA damage induced, we evaluated nine of 42 compounds identified as positive for differential cytotoxicity in qHTS (actinomycin D, adriamycin, alachlor, benzotrichloride, diglycidyl resorcinol ether, lovastatin, melphalan, trans-1,4-dichloro-2-butene, tris(2,3-epoxypropyl)isocyanurate) and one non-cytotoxic genotoxic compound (2-aminothiamine) for (1) clastogenicity in mutant and wild-type cells; (2) the comparative induction of γH2AX positive foci by melphalan; (3) the extent to which a 72-hr exposure duration increased assay sensitivity or specificity; (4) the use of 10 additional DT40 DNA repair-deficient cell lines to better analyze the type(s) of DNA damage induced; and (5) the involvement of reactive oxygen species in the induction of DNA damage. All compounds but lovastatin and 2-aminothiamine were more clastogenic in at least one DNA repair-deficient cell line than the wild-type cells. The differential responses across the various DNA repair-deficient cell lines provided information on the type(s) of DNA damage induced. The results demonstrate the utility of this DT40 screen for detecting genotoxic compounds, for characterizing the nature of the DNA damage, and potentially for analyzing mechanisms of mutagenesis. Copyright © 2011 Wiley-Liss, Inc.

Production and biochemical characterization of α-glucosidase from Aspergillus niger ITV-01 isolated from sugar cane bagasse.

Science.gov (United States)

Del Moral, S; Barradas-Dermitz, D M; Aguilar-Uscanga, M G

2018-01-01

Aspergillus niger ITV-01 presents amylolytic activity, identified as α-glucosidase, an enzyme that only produces α-d-glucose from soluble starch and that presents transglucosylase activity on α-d-glucopyranosyl-(1-4)-α-d-glucopyranose (maltose) (200 gL -1 ). Biochemical characterization was performed on A. niger ITV-01 α-glucosidase; its optimum parameters were pH 4.3, temperature 80 °C but stable at 40 °C, with an energy of activation (Ea) 176.25 kJ mol -1 . Using soluble starch as the substrate, K m and V max were 5 mg mL -1 and 1000 U mg -1 , respectively. As α-glucosidase is not a metalloenzyme, calcium and EDTA did not have any effect on its activity. The molecular weight was estimated by SDS-PAGE to be about 75 kDa. It was also active in methanol and ethanol. When ammonium sulfate (AS) and yeast extract (YE) nitrogen sources and calcium effect were evaluated, the greatest activity occurred using YE and calcium, as opposed to AS media where no activity was detected. The results obtained showed that this enzyme has industrial application potential in the processes to produce either ethanol or malto-oligosaccharides from α-d-glucopyranosyl-(1-4)-α-d-glucopyranose (maltose).

Genetic and Biochemical Characterization of the MinC-FtsZ Interaction in Bacillus subtilis

Science.gov (United States)

Castellen, Patricia; Nogueira, Maria Luiza C.; Bettini, Jefferson; Portugal, Rodrigo V.; Zeri, Ana Carolina M.; Gueiros-Filho, Frederico J.

2013-01-01

Cell division in bacteria is regulated by proteins that interact with FtsZ and modulate its ability to polymerize into the Z ring structure. The best studied of these regulators is MinC, an inhibitor of FtsZ polymerization that plays a crucial role in the spatial control of Z ring formation. Recent work established that E. coli MinC interacts with two regions of FtsZ, the bottom face of the H10 helix and the extreme C-terminal peptide (CTP). Here we determined the binding site for MinC on Bacillus subtilis FtsZ. Selection of a library of FtsZ mutants for survival in the presence of Min overexpression resulted in the isolation of 13 Min-resistant mutants. Most of the substitutions that gave rise to Min resistance clustered around the H9 and H10 helices in the C-terminal domain of FtsZ. In addition, a mutation in the CTP of B. subtilis FtsZ also produced MinC resistance. Biochemical characterization of some of the mutant proteins showed that they exhibited normal polymerization properties but reduced interaction with MinC, as expected for binding site mutations. Thus, our study shows that the overall architecture of the MinC-FtsZ interaction is conserved in E. coli and B. subtilis. Nevertheless, there was a clear difference in the mutations that conferred Min resistance, with those in B. subtilis FtsZ pointing to the side of the molecule rather than to its polymerization interface. This observation suggests that the mechanism of Z ring inhibition by MinC differs in both species. PMID:23577149

Cluster Analysis of Maize Inbred Lines

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Jiban Shrestha

2016-12-01

Full Text Available The determination of diversity among inbred lines is important for heterosis breeding. Sixty maize inbred lines were evaluated for their eight agro morphological traits during winter season of 2011 to analyze their genetic diversity. Clustering was done by average linkage method. The inbred lines were grouped into six clusters. Inbred lines grouped into Clusters II had taller plants with maximum number of leaves. The cluster III was characterized with shorter plants with minimum number of leaves. The inbred lines categorized into cluster V had early flowering whereas the group into cluster VI had late flowering time. The inbred lines grouped into the cluster III were characterized by higher value of anthesis silking interval (ASI and those of cluster VI had lower value of ASI. These results showed that the inbred lines having widely divergent clusters can be utilized in hybrid breeding programme.

Characterizing the Biochemical Response to Schistosoma mansoni Infection and Treatment with Praziquantel in Preschool and School Aged Children.

Science.gov (United States)

Panic, Gordana; Coulibaly, Jean T; Harvey, Nikita; Keiser, Jennifer; Swann, Jonathan

2018-05-21

Schistosomiasis is a widespread chronic neglected tropical disease prevalent mostly in children in under-resourced rural areas. Its pathological effects have been clinically characterized, yet the molecular-level effects are understudied. In this study, the biochemical effects of Schistosoma mansoni infection and praziquantel treatment were studied in 130 preschool aged and 159 school aged infected children and 11 noninfected children in Azaguié, Côte d'Ivoire. Urine samples were collected prior to receiving 20, 40, or 60 mg/kg of praziquantel or a placebo, as well as 24 h post-treatment, and at the 3-week follow up. Urinary metabolic phenotypes were measured using 1 H NMR spectroscopy, and metabolic variation associated with S. mansoni infection and praziquantel administration was identified using multivariate statistical techniques. Discriminatory metabolic signatures were detected between heavily infected and noninfected children at baseline as well as according to the dose of praziquantel administered 24 h post treatment. These signatures were primarily associated with the metabolic activity of the gut microbiota, gut health and growth biomarkers and energy and liver metabolism. These analyses provide insights into the metabolic phenotype of schistosomiasis and treatment with praziquantel in two important demographics.

Morphological and physico-biochemical characterization of various tomato cultivars in a simplified soilless media

Directory of Open Access Journals (Sweden)

Mahboob Ahmad Khan

2017-12-01

Full Text Available This study aimed to investigate nine commercial cultivars of tomato, in order to identify the most suitable cultivar in terms of morphological (plant height, fruit size, fruit weight and total yield and physico-biochemical (color, firmness, total soluble solid, titratable acidity, ascorbic acid, total sugar, reducing and non-reducing sugar, β-carotene and lycopene attributes. Plants were cultured hydroponically in the greenhouse. Results revealed that the morphological attributes of Beefsteak Group (BG of tomatoes was significantly better than that of Cherry Group (CG. In addition, CG has higher concentration of biochemical attributes, mainly β-carotene, sugars, total soluble solids (TSS and ascorbic acid contents. Within CG, cv. Aria was found to be the best for higher sugar contents, β-carotene and ascorbic acid contents; while, TSS was higher in the cv. Claree. Similarly in BG, cv. Sahel had the highest value of lycopene, β-carotene, TSS; whereas, lowest sugar contents were found in cv. Dirk. As far as firmness is concerned, cv. Naram (BG was found to be more firm, than cv. Aria (CG. The highest total yield was recorded for cv. Vernal; in BG and in cv. Claree for CG, depicting that BG had significantly higher total yield, compared with CG.

Development and characterization of a Psathyrostachys huashanica Keng 7Ns chromosome addition line with leaf rust resistance.

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Wanli Du

Full Text Available The aim of this study was to characterize a Triticum aestivum-Psathyrostachys huashanica Keng (2n = 2x = 14, NsNs disomic addition line 2-1-6-3. Individual line 2-1-6-3 plants were analyzed using cytological, genomic in situ hybridization (GISH, EST-SSR, and EST-STS techniques. The alien addition line 2-1-6-3 was shown to have two P. huashanica chromosomes, with a meiotic configuration of 2n = 44 = 22 II. We tested 55 EST-SSR and 336 EST-STS primer pairs that mapped onto seven different wheat chromosomes using DNA from parents and the P. huashanica addition line. One EST-SSR and nine EST-STS primer pairs indicated that the additional chromosome of P. huashanica belonged to homoeologous group 7, the diagnostic fragments of five EST-STS markers (BE404955, BE591127, BE637663, BF482781 and CD452422 were cloned, sequenced and compared. The results showed that the amplified polymorphic bands of P. huashanica and disomic addition line 2-1-6-3 shared 100% sequence identity, which was designated as the 7Ns disomic addition line. Disomic addition line 2-1-6-3 was evaluated to test the leaf rust resistance of adult stages in the field. We found that one pair of the 7Ns genome chromosomes carried new leaf rust resistance gene(s. Moreover, wheat line 2-1-6-3 had a superior numbers of florets and grains per spike, which were associated with the introgression of the paired P. huashanica chromosomes. These high levels of disease resistance and stable, excellent agronomic traits suggest that this line could be utilized as a novel donor in wheat breeding programs.

Comparison of biochemical and immunological profile of pediatric patients with acute myeloid leukemia in relation to healthy individuals

Directory of Open Access Journals (Sweden)

Fabiane L.F.Z. Sanches

2015-09-01

Conclusions: It was possible to characterize the biochemical and immunological profile of pediatric patients with AML, as well as highlight some significant differences in these parameters when comparing with healthy children and adolescents.

Characterization of a high-energy in-line phase contrast tomosynthesis prototype.

Science.gov (United States)

Wu, Di; Yan, Aimin; Li, Yuhua; Wong, Molly D; Zheng, Bin; Wu, Xizeng; Liu, Hong

2015-05-01

In this research, a high-energy in-line phase contrast tomosynthesis prototype was developed and characterized through quantitative investigations and phantom studies. The prototype system consists of an x-ray source, a motorized rotation stage, and a CMOS detector with a pixel pitch of 0.05 mm. The x-ray source was operated at 120 kVp for this study, and the objects were mounted on the rotation stage 76.2 cm (R1) from the source and 114.3 cm (R2) from the detector. The large air gap between the object and detector guarantees sufficient phase-shift effects. The quantitative evaluation of this prototype included modulation transfer function and noise power spectrum measurements conducted under both projection mode and tomosynthesis mode. Phantom studies were performed including three custom designed phantoms with complex structures: a five-layer bubble wrap phantom, a fishbone phantom, and a chicken breast phantom with embedded fibrils and mass structures extracted from an ACR phantom. In-plane images of the phantoms were acquired to investigate their image qualities through observation, intensity profile plots, edge enhancement evaluations, and/or contrast-to-noise ratio calculations. In addition, the robust phase-attenuation duality (PAD)-based phase retrieval method was applied to tomosynthesis for the first time in this research. It was utilized as a preprocessing method to fully exhibit phase contrast on the angular projection before reconstruction. The resolution and noise characteristics of this high-energy in-line phase contrast tomosynthesis prototype were successfully investigated and demonstrated. The phantom studies demonstrated that this imaging prototype can successfully remove the structure overlapping in phantom projections, obtain delineate interfaces, and achieve better contrast-to-noise ratio after applying phase retrieval to the angular projections. This research successfully demonstrated a high-energy in-line phase contrast tomosynthesis

Transmission line pulse system for avalanche characterization of high power semiconductor devices

Science.gov (United States)

Riccio, Michele; Ascione, Giovanni; De Falco, Giuseppe; Maresca, Luca; De Laurentis, Martina; Irace, Andrea; Breglio, Giovanni

2013-05-01

Because of the increasing in power density of electronic devices for medium and high power application, reliabilty of these devices is of great interest. Understanding the avalanche behaviour of a power device has become very important in these last years because it gives an indication of the maximum energy ratings which can be seen as an index of the device ruggedness. A good description of this behaviour is given by the static IV blocking characteristc. In order to avoid self heating, very relevant in high power devices, very short pulses of current have to be used, whose value can change from few milliamps up to tens of amps. The most used method to generate short pulses is the TLP (Transmission Line Pulse) test, which is based on charging the equivalent capacitance of a transmission line to high value of voltage and subsequently discharging it onto a load. This circuit let to obtain very short square pulses but it is mostly used for evaluate the ESD capability of semiconductor and, in this environment, it generates pulses of low amplitude which are not high enough to characterize the avalanche behaviour of high power devices . Advanced TLP circuit able to generate high current are usually very expensive and often suffer of distorption of the output pulse. In this article is proposed a simple, low cost circuit, based on a boosted-TLP configuration, which is capable to produce very square pulses of about one hundreds of nanosecond with amplitude up to some tens of amps. A prototype is implemented which can produce pulses up to 20A of amplitude with 200 ns of duration which can characterize power devices up to 1600V of breakdown voltage. Usage of microcontroller based logic make the circuit very flexible. Results of SPICE simulation are provided, together with experimental results. To prove the effectiveness of the circuit, the I-V blocking characteristics of two commercial devices, namely a 600V PowerMOS and a 1200V Trench-IGBT, are measured at different

[INVITED] Tilted fiber grating mechanical and biochemical sensors

Science.gov (United States)

Guo, Tuan; Liu, Fu; Guan, Bai-Ou; Albert, Jacques

2016-04-01

The tilted fiber Bragg grating (TFBG) is a new kind of fiber-optic sensor that possesses all the advantages of well-established Bragg grating technology in addition to being able to excite cladding modes resonantly. This device opens up a multitude of opportunities for single-point sensing in hard-to-reach spaces with very controllable cross-sensitivities, absolute and relative measurements of various parameters, and an extreme sensitivity to materials external to the fiber without requiring the fiber to be etched or tapered. Over the past five years, our research group has been developing multimodal fiber-optic sensors based on TFBG in various shapes and forms, always keeping the device itself simple to fabricate and compatible with low-cost manufacturing. This paper presents a brief review of the principle, fabrication, characterization, and implementation of TFBGs, followed by our progress in TFBG sensors for mechanical and biochemical applications, including one-dimensional TFBG vibroscopes, accelerometers and micro-displacement sensors; two-dimensional TFBG vector vibroscopes and vector rotation sensors; reflective TFBG refractometers with in-fiber and fiber-to-fiber configurations; polarimetric and plasmonic TFBG biochemical sensors for in-situ detection of cell, protein and glucose.

Elemental, stable isotopic and biochemical characterization of soil organic matter alteration across a natural peatland gradient

Science.gov (United States)

Cowie, G.; Mowbray, S.; Belyea, L.; Laing, C.; Allton, K.; Abbott, G.; Muhammad, A.

2010-12-01

Northern peatlands store around one third of global soil C and thus represent a key reservoir. To elucidate how these systems might respond to climate change, field- and laboratory-based experimental incubation studies are being conducted at sites across a natural peatland gradient in the boreonemoral zone of central Sweden (Ryggmossen). The site comprises four successional stages, from edge to centre; Swamp Forest (SF), Lagg Fen (LF), Bog Margin (BM) and Bog Plateau (BP). The well-preserved succession shows strong decreases in mineral cations and pH, and distinct changes in vegetation and water-table depth. As an underpinning to these experiments, comprehensive characterization of natural soil organic matter (SOM) alteration has been carried out through detailed analyses of vegetation and downcore profiles at contrasting topographic sites (hummock vs hollow) in each of the four locations. As illustrated in Figure 1, while some similarities occur in downcore trends, contrasts are observed in C and N elemental and stable isotopic compositions, between stages and, in some cases, between microtopographic settings. Downcore trends and intersite differences are also observed in biochemical yields and molecular composition (carbohydrates, amino acids, phenols, lipids and D/L amino acid ratios). These reflect SOM decay and alteration combined with the effects of contrasting hydrologic, redox and nutrient regimes and differing vegetation and microbial inputs at each of the study sites. Multivariate analysis is used to to elucidate compositional patterns that characterize and delineate progressive SOM decay, specific vegetation types, and the effects of contrasting environmental conditions at the different sites. Figure 1. A. Organic carbon content (wt %), B. Atomic ratio of organic C to total N, C. Stable C isotopic composition of organic C (d13Corg), and D. Stable N isotopic composition of total nitrogen (d15N), all for core profiles from contrasting settings (hummock and

A characterization of the ZFL cell line and primary hepatocytes as in vitro liver cell models for the zebrafish (Danio rerio)

International Nuclear Information System (INIS)

Eide, Marta; Rusten, Marte; Male, Rune; Jensen, Knut Helge Midtbø; Goksøyr, Anders

2014-01-01

Highlights: •The ZFL cell line and primary hepatocytes were characterized. •Basic and induced expression of nuclear receptors and target genes were found. •The ZFL cell line expresses very low basic levels of most genes. •The ZFL cells have low induction of gene expression following exposures. •Primary hepatocytes show large sex-dependent differences in gene expression. -- Abstract: The zebrafish (Danio rerio) is a widely used model species in biomedical research. The ZFL cell line, established from zebrafish liver, and freshly isolated primary hepatocytes from zebrafish have been used in several toxicological studies. However, no previous report has compared and characterized these two systems at the level of gene expression. The aim of this study was to evaluate the ZFL cell line in comparison to primary hepatocytes as in vitro models for studying effects of environmental contaminants in zebrafish liver. Using quantitative real-time PCR, the basal level and transcriptional induction potential of key genes involved in toxic responses in the ZFL cell line, primary hepatocytes and whole liver from zebrafish were compared. The study showed that the ZFL cells have lower levels of mRNA of most selected genes compared to zebrafish liver. The induced gene transcription following exposure to ligand was much lower in ZFL cells compared to zebrafish primary hepatocytes at the doses tested. Importantly, oestrogen receptor and vitellogenin genes showed low basal transcription and no induction response in the ZFL cell line. In conclusion, it appears that primary hepatocytes are well suited for studying environmental contaminants including xenoestrogens, but may show large sex-dependent differences in gene transcription. The ZFL cell line shows potential in toxicological studies involving the aryl hydrocarbon receptor pathway. However, low potential for transcriptional induction of genes in general should be expected, especially notable when studying estrogenic

A characterization of the ZFL cell line and primary hepatocytes as in vitro liver cell models for the zebrafish (Danio rerio)

Energy Technology Data Exchange (ETDEWEB)

Eide, Marta, E-mail: marta.eide@bio.uib.no [Department of Biology, University of Bergen, Bergen (Norway); Rusten, Marte; Male, Rune [Department of Molecular Biology, University of Bergen, Bergen (Norway); Jensen, Knut Helge Midtbø; Goksøyr, Anders [Department of Biology, University of Bergen, Bergen (Norway)

2014-02-15

Highlights: •The ZFL cell line and primary hepatocytes were characterized. •Basic and induced expression of nuclear receptors and target genes were found. •The ZFL cell line expresses very low basic levels of most genes. •The ZFL cells have low induction of gene expression following exposures. •Primary hepatocytes show large sex-dependent differences in gene expression. -- Abstract: The zebrafish (Danio rerio) is a widely used model species in biomedical research. The ZFL cell line, established from zebrafish liver, and freshly isolated primary hepatocytes from zebrafish have been used in several toxicological studies. However, no previous report has compared and characterized these two systems at the level of gene expression. The aim of this study was to evaluate the ZFL cell line in comparison to primary hepatocytes as in vitro models for studying effects of environmental contaminants in zebrafish liver. Using quantitative real-time PCR, the basal level and transcriptional induction potential of key genes involved in toxic responses in the ZFL cell line, primary hepatocytes and whole liver from zebrafish were compared. The study showed that the ZFL cells have lower levels of mRNA of most selected genes compared to zebrafish liver. The induced gene transcription following exposure to ligand was much lower in ZFL cells compared to zebrafish primary hepatocytes at the doses tested. Importantly, oestrogen receptor and vitellogenin genes showed low basal transcription and no induction response in the ZFL cell line. In conclusion, it appears that primary hepatocytes are well suited for studying environmental contaminants including xenoestrogens, but may show large sex-dependent differences in gene transcription. The ZFL cell line shows potential in toxicological studies involving the aryl hydrocarbon receptor pathway. However, low potential for transcriptional induction of genes in general should be expected, especially notable when studying estrogenic

Physiological and Biochemical characterization of Chlamydomonas sp. the Hydrogen Production's Strain

International Nuclear Information System (INIS)

Chader, S.; Belhamel, M.; H Hacene

2006-01-01

The hydrogen produced by biological way became, one of the most interesting subjects of research relating to development the energy system starting from renewable sources. This study describes the closed relation between the physiological behaviour, biochemical and rate of gases produced by Chlamydomonas sp. strain AT14, isolated in the area of Touat (the Sahara Algerian) and cultivated in a toric photo-bioreactor. A considerable growth was noted, where the concentration of the biomass double in only two days after incubation. The micro-algal cells present a 100% of viability, which relocate has satisfactory behaviour in the toric engine. In addition, the displacement water level in the system of measurement implies has gas production (0.1 ml) in coordination with the anaerobic period of the reactional enclosure. The yield of this way of hydrogen production is depending on the species used, the light intensity, and the conditions of culture. (authors)

Characterization and diversity assessment in coconut collections of ...

African Journals Online (AJOL)

Characterization and genetic diversity assessment among 30 conserved coconut accessions collected from Pacific Ocean Islands and Nicobar Islands for morphological traits, physico-chemical traits of tender nut water, leaf biochemical parameters and molecular characterization by using random amplified polymorphic ...

Characterization of pipes, drain lines, and ducts using the pipe explorer system

International Nuclear Information System (INIS)

Cremer, C.D.; Kendrick, D.T.; Cramer, E.

1997-01-01

As DOE dismantles its nuclear processing facilities, site managers must employ the best means of disposing or remediating hundreds of miles of potentially contaminated piping and duct work. Their interiors are difficult to access, and in many cases even the exteriors are inaccessible. Without adequate characterization, it must be assumed that the piping is contaminated, and the disposal cost of buried drain lines can be on the order of $1,200/ft and is often unnecessary as residual contamination levels often are below free release criteria. This paper describes the program to develop a solution to the problem of characterizing radioactive contamination in pipes. The technical approach and results of using the Pipe Explorer trademark system are presented. The heart of the system is SEA's pressurized inverting membrane adapted to transport radiation detectors and other tools into pipes. It offers many benefits over other pipe inspection approaches. It has video and beta/gamma detection capabilities, and the need for alpha detection has been addressed through the development of the Alpha Explorer trademark. These systems have been used during various stages of decontamination and decommissioning of DOE sites, including the ANL CP-5 reactor D ampersand D. Future improvements and extensions of their capabilities are discussed

Different energy metabolism in two human small cell lung cancer subpopulations examined by 31P magnetic resonance spectroscopy and biochemical analysis in vivo and in vitro

DEFF Research Database (Denmark)

Kristjansen, P E; Spang-Thomsen, M; Quistorff, B

1991-01-01

Two human small cell lung cancer tumor lines, maintained as solid tumor xenografts on nude mice and as in vitro cell cultures, were studied by in vivo 31P magnetic resonance spectroscopy and by biochemical analysis of extracts of solid tumors and cell cultures. The tumor lines CPH SCCL 54A and CPH...

Biochemical Network Stochastic Simulator (BioNetS: software for stochastic modeling of biochemical networks

Directory of Open Access Journals (Sweden)

Elston Timothy C

2004-03-01

Full Text Available Abstract Background Intrinsic fluctuations due to the stochastic nature of biochemical reactions can have large effects on the response of biochemical networks. This is particularly true for pathways that involve transcriptional regulation, where generally there are two copies of each gene and the number of messenger RNA (mRNA molecules can be small. Therefore, there is a need for computational tools for developing and investigating stochastic models of biochemical networks. Results We have developed the software package Biochemical Network Stochastic Simulator (BioNetS for efficientlyand accurately simulating stochastic models of biochemical networks. BioNetS has a graphical user interface that allows models to be entered in a straightforward manner, and allows the user to specify the type of random variable (discrete or continuous for each chemical species in the network. The discrete variables are simulated using an efficient implementation of the Gillespie algorithm. For the continuous random variables, BioNetS constructs and numerically solvesthe appropriate chemical Langevin equations. The software package has been developed to scale efficiently with network size, thereby allowing large systems to be studied. BioNetS runs as a BioSpice agent and can be downloaded from http://www.biospice.org. BioNetS also can be run as a stand alone package. All the required files are accessible from http://x.amath.unc.edu/BioNetS. Conclusions We have developed BioNetS to be a reliable tool for studying the stochastic dynamics of large biochemical networks. Important features of BioNetS are its ability to handle hybrid models that consist of both continuous and discrete random variables and its ability to model cell growth and division. We have verified the accuracy and efficiency of the numerical methods by considering several test systems.

Efficient simulation of intrinsic, extrinsic and external noise in biochemical systems

Science.gov (United States)

Pischel, Dennis; Sundmacher, Kai; Flassig, Robert J.

2017-01-01

Abstract Motivation: Biological cells operate in a noisy regime influenced by intrinsic, extrinsic and external noise, which leads to large differences of individual cell states. Stochastic effects must be taken into account to characterize biochemical kinetics accurately. Since the exact solution of the chemical master equation, which governs the underlying stochastic process, cannot be derived for most biochemical systems, approximate methods are used to obtain a solution. Results: In this study, a method to efficiently simulate the various sources of noise simultaneously is proposed and benchmarked on several examples. The method relies on the combination of the sigma point approach to describe extrinsic and external variability and the τ-leaping algorithm to account for the stochasticity due to probabilistic reactions. The comparison of our method to extensive Monte Carlo calculations demonstrates an immense computational advantage while losing an acceptable amount of accuracy. Additionally, the application to parameter optimization problems in stochastic biochemical reaction networks is shown, which is rarely applied due to its huge computational burden. To give further insight, a MATLAB script is provided including the proposed method applied to a simple toy example of gene expression. Availability and implementation: MATLAB code is available at Bioinformatics online. Contact: flassig@mpi-magdeburg.mpg.de Supplementary information: Supplementary data are available at Bioinformatics online. PMID:28881987

Simplifying biochemical models with intermediate species

DEFF Research Database (Denmark)

Feliu, Elisenda; Wiuf, Carsten

2013-01-01

techniques, we study systematically the effects of intermediate, or transient, species in biochemical systems and provide a simple, yet rigorous mathematical classification of all models obtained from a core model by including intermediates. Main examples include enzymatic and post-translational modification...... systems, where intermediates often are considered insignificant and neglected in a model, or they are not included because we are unaware of their existence. All possible models obtained from the core model are classified into a finite number of classes. Each class is defined by a mathematically simple...... canonical model that characterizes crucial dynamical properties, such as mono- and multistationarity and stability of steady states, of all models in the class. We show that if the core model does not have conservation laws, then the introduction of intermediates does not change the steady...

Molecular characterization of neoplastic and normal "sister" lymphoblastoid B-cell lines from chronic lymphocytic leukemia

DEFF Research Database (Denmark)

Lanemo Myhrinder, Anna; Hellqvist, Eva; Bergh, Ann-Charlotte

2013-01-01

Chronic lymphocytic leukemia (CLL) B-cells resemble self-renewing CD5 + B-cells carrying auto/xeno-antigen-reactive B-cell receptors (BCRs) and multiple innate pattern-recognition receptors, such as Toll-like receptors and scavenger receptors. Integration of signals from BCRs with multiple surface...... a comprehensive genotypic and phenotypic characterization of available CLL and normal B-cell-derived lymphoblastoid cell lines (LCLs) from the same individuals (n = 17). Authenticity and verification studies of CLL-patient origin were done by IGHV sequencing, fluorescence in situ hybridization (FISH) and DNA...

Biochemical and biological characterization of Bothriechis schlegelii snake venoms from Colombia and Costa Rica.

Science.gov (United States)

Prezotto-Neto, José P; Kimura, Louise F; Alves, André F; Gutiérrez, José María; Otero, Rafael; Suárez, Ana M; Santoro, Marcelo L; Barbaro, Katia C

2016-12-01

Snakebites inflicted by the arboreal viperid snake Bothriechis schlegelii in humans are characterized by pain, edema, and ecchymosis at the site of the bite, rarely with blisters, local necrosis, or defibrination. Herein, a comparative study of Bothriechis schlegelii snake venoms from Colombia (BsCo) and Costa Rica (BsCR) was carried out in order to compare their main activities and to verify the efficacy of Bothrops antivenom produced in Brazil to neutralize them. Biochemical (SDS-PAGE and zymography) and biological parameters (edematogenic, lethal, hemorrhagic, nociceptive, and phospholipase A 2 activities) induced by BsCo and BsCR snake venoms were evaluated. The presence of antibodies in Bothrops antivenom that recognize BsCo and BsCR snake venoms by enzyme-linked immunosorbent assay and Western blotting, as well as the ability of this antivenom to neutralize the toxic activities were also verified. SDS-PAGE showed differences between venoms. Distinctive caseinolytic and hyaluronidase patterns were detected by zymography. BsCo and BsCR showed similar phospholipase A 2 activity. Strong cross-reactivity between BsCo and BsCR was detected using Bothrops antivenom with many components located between 150 and 35 kDa. BsCR was more edematogenic and almost fourfold more hemorrhagic than BsCo, and both venoms induced nociception. BsCR (LD 50 5.60 mg/kg) was more lethal to mice than BsCo (LD 50 9.24 mg/kg). Bothrops antivenom was effective in the neutralization of lethal and hemorrhagic activities of BsCo and BsCR and was partially effective in the neutralization of edematogenic and nociceptive activities. In conclusion, geographic distribution influences the composition and activities of Bothriechis schlegelii venoms. Bothrops antivenom cross-reacted with these venoms and was partially effective in neutralizing some toxic activities of BsCo and BsCR.

Biochemical Characterization of a Family 15 Carbohydrate Esterase from a Bacterial Marine Arctic Metagenome.

Directory of Open Access Journals (Sweden)

Concetta De Santi

Full Text Available The glucuronoyl esterase enzymes of wood-degrading fungi (Carbohydrate Esterase family 15; CE15 form part of the hemicellulolytic and cellulolytic enzyme systems that break down plant biomass, and have possible applications in biotechnology. Homologous enzymes are predicted in the genomes of several bacteria, however these have been much less studied than their fungal counterparts. Here we describe the recombinant production and biochemical characterization of a bacterial CE15 enzyme denoted MZ0003, which was identified by in silico screening of a prokaryotic metagenome library derived from marine Arctic sediment. MZ0003 has high similarity to several uncharacterized gene products of polysaccharide-degrading bacterial species, and phylogenetic analysis indicates a deep evolutionary split between these CE15s and fungal homologs.MZ0003 appears to differ from previously-studied CE15s in some aspects. Some glucuronoyl esterase activity could be measured by qualitative thin-layer chromatography which confirms its assignment as a CE15, however MZ0003 can also hydrolyze a range of other esters, including p-nitrophenyl acetate, which is not acted upon by some fungal homologs. The structure of MZ0003 also appears to differ as it is predicted to have several large loop regions that are absent in previously studied CE15s, and a combination of homology-based modelling and site-directed mutagenesis indicate its catalytic residues deviate from the conserved Ser-His-Glu triad of many fungal CE15s. Taken together, these results indicate that potentially unexplored diversity exists among bacterial CE15s, and this may be accessed by investigation of the microbial metagenome. The combination of low activity on typical glucuronoyl esterase substrates, and the lack of glucuronic acid esters in the marine environment suggest that the physiological substrate of MZ0003 and its homologs is likely to be different from that of related fungal enzymes.

Proximate and biochemical characterization of burrito (Bachydeuterus auritus) and flying gurnard (Dactylopterus volitans).

Science.gov (United States)

Abbey, Lawrence D; Glover-Amengor, Mary; Atikpo, Margaret Ottah; Howell, Nazlin K

2017-05-01

With limited protein resources and depleting commercial fish species there is the need to improve utilization of some of the lesser known species which are underutilized, for example, big eye grunt (burrito), Bachydeuterus auritus, and the flying gurnard ( Dactylopterus volitans ), (other names Cephalocanthus volitans (local) Pampansre ). This study was to characterize some of the proximate and biochemical properties of burrito and the flying gurnard so as to evaluate their potential for use in human nutrition and other value-added products. Proximate and chemical analysis were determined by the methods of AOAC. Fatty acid profiles were determined following the method of Saaed and Howell (1999). Amino acid profiles for the species were determined according to Bidlingmeyer et al. (1987). The protein content of both the water soluble and salt soluble protein extracts of the fish species were determined by the Bradford Protein Assay method (Bradford 1976). Rancidity of the fish species was assessed by thiobarbituric acid reactive substances (TBARS) and Peroxide value (PV) as described by Saeed and Howell (1999). Burrito contained 18% protein, whereas the flying gurnard contained 22.3%. Calcium content was 296 mg/100 g for burrito and 185 mg/100 g for flying gurnard, whereas iron content was 4.1 mg/100 g and 1.0 mg/100 g for burrito and the flying gurnard, respectively. Palmitic acid (C16) was 27% and 14.3% for the flying gurnard and burrito, respectively. C17: 1ω8 was 3% in the flying gurnard and 0.2% in burrito. Oleic (C18:1ω9) was 17% in the flying gurnard and 6% in burrito. C20:4ω6 was 1.6% in the flying gurnard and 3% in burrito. Docosahexaenoic acid (C22:6ω3) was 4.9% in the flying gurnard and 4.0% in burrito. Both burrito and the flying gurnard are of high nutritional quality as they had a high protein content, good general amino acid profile and abundance of polyunsaturated fatty acids.

Selection and characterizations of radiation-induced salinity-tolerant lines in rice

International Nuclear Information System (INIS)

Lee, I.S.; Kim, D.S.; Lee, S.J.; Song, H.S.; Lim, Y.P.; Lee, Y.I.

2003-01-01

NaCl tolerant cell lines were selected from irradiated callus, which were generated from seed cultures. M 1 -regenerates were obtained from the salt-tolerant callus cultured on the auxin-free medium for 30 days. Some regenerants were more tolerant than the parent variety (Dongjinbyeo) on a medium containing 0.75 % NaCl. Seeds (M 3 5,000 lines) derived from M 2 lines were grown to the 3 leaf stage. M 3 lines were soaked with a 0.75 % salt solution for 3 weeks and 350 salt-tolerant genotypes were selected. Among the M 3 350 lines, forty tolerant lines were selected from a saline field (10~14 mS) near the sea coast. Of the forty lines, two lines (18-1 and 50-1) showed more improved plant height, panicle length, tillering number, spikelet number and yield than those of the original variety. Thirty primers were screened and two RAPD markers were identified, which appeared in both the salt-tolerant lines (18-1 and 50-1). From DNA-hybridization experiments, it appeared that the fragment arose from the middle-repetitive copy sequences. The transcript involved in the marker showed a higher expression in the salt-tolerant lines than the sensitive lines. The salt-tolerant lines would be useful as a resource for salt-tolerant breeding. (author)

Production and molecular characterization of bread wheat lines with reduced amount of α-type gliadins.

Science.gov (United States)

Camerlengo, Francesco; Sestili, Francesco; Silvestri, Marco; Colaprico, Giuseppe; Margiotta, Benedetta; Ruggeri, Roberto; Lupi, Roberta; Masci, Stefania; Lafiandra, Domenico

2017-12-19

Among wheat gluten proteins, the α-type gliadins are the major responsible for celiac disease, an autoimmune disorder that affects about 1% of the world population. In fact, these proteins contain several toxic and immunogenic epitopes that trigger the onset of the disease. The α-type gliadins are a multigene family, encoded by genes located at the complex Gli-2 loci. Here, three bread wheat deletion lines (Gli-A2, Gli-D2 and Gli-A2/Gli-D2) at the Gli-2 loci were generated by the introgression in the bread wheat cultivar Pegaso of natural mutations, detected in different bread wheat cultivars. The molecular characterization of these lines allowed the isolation of 49 unique expressed genes coding α-type gliadins, that were assigned to each of the three Gli-2 loci. The number and the amount of α-type gliadin transcripts were drastically reduced in the deletion lines. In particular, the line Gli-A2/Gli-D2 contained only 12 active α-type gliadin genes (-75.6% respect to the cv. Pegaso) and a minor level of transcripts (-80% compared to cv. Pegaso). Compensatory pleiotropic effects were observed in the two other classes of gliadins (ω- and γ-gliadins) either at gene expression or protein levels. Although the comparative analysis of the deduced amino acid sequences highlighted the typical structural features of α-type gliadin proteins, substantial differences were displayed among the 49 proteins for the presence of toxic and immunogenic epitopes. The deletion line Gli-A2/Gli-D2 did not contain the 33-mer peptide, one of the major epitopes triggering the celiac disease, representing an interesting material to develop less "toxic" wheat varieties.

Biochemical characterization of MLH3 missense mutations does not reveal an apparent role of MLH3 in Lynch syndrome

DEFF Research Database (Denmark)

Ou, Jianghua; Rasmussen, Merete; Westers, Helga

2009-01-01

for eight of these MLH3 UVs identified in suspected Lynch syndrome patients, we performed several biochemical tests. We determined the protein expression and stability, protein localization and interaction of the mutant MLH3 proteins with wildtype MLH1. All eight MLH3 UVs gave protein expression levels...... comparable with wildtype MLH3. Furthermore, the UV-containing proteins, in contrast to previous studies, were all localized normally in the nucleus and they interacted normally with wildtype MLH1. Our different biochemical assays yielded no evidence that the eight MLH3 UVs tested are the cause of hereditary...

Power line communications theory and applications for narrowband and broadband communications over power lines

CERN Document Server

Ferreira, Hendrik C; Newbury, John; Swart, Theo G

2010-01-01

Power Line Communications (PLC) is a promising emerging technology, which has attracted much attention due to the wide availability of power distribution lines. This book provides a thorough introduction to the use of power lines for communication purposes, ranging from channel characterization, communications on the physical layer and electromagnetic interference, through to protocols, networks, standards and up to systems and implementations. With contributions from many of the most prominent international PLC experts from academia and industry, Power Line Communications brings togeth

Establishment and characterization of a fin cell line from blunt snout bream, Megalobrama amblycephala.

Science.gov (United States)

Zhu, Dong-Mei; Yang, Kun; Wang, Wei-Min; Song, Wen

2013-12-01

This study established and characterized a new cell line (MAF) from the fin of blunt snout bream (Megalobrama amblycephala), a freshwater fish cultivated in China. MAF cells proliferated well in medium 199 supplemented with 10 % fetal bovine serum at 28 °C and have been subcultured more than 95 times in almost a year. MAF cells were revived at 90-95 % viability after 3-6 months of storage in liquid nitrogen. Karyotyping indicated that the modal chromosome number of MAF cells was 48. The MAF cell line consisted predominantly of fibroblastic and epithelial-like cells from M. amblycephala, which was confirmed by immunofluorescence and mitochondrial 12s rRNA sequencing. Viral susceptibility tests showed that MAF cells were susceptible to infection by snakehead rhabdovirus, spring viremia carp virus, and channel catfish virus, which was demonstrated by the presence of cytopathic effect, high viral titers, and PCR products. Bacterial cytotoxicity studies showed that extracellular products from Aeromonas hydrophila were toxic to MAF cells. Cu²⁺ was also cytotoxic to MAF cells, and the 24-h IC₅₀ value was 144.48 μmol/l. When MAF cells were transfected with pEGFP-N1 plasmid, bright fluorescent signals were observed, and the transfection efficiency reached up to 5 %. These results suggest that the MAF cell line may provide a valuable tool for studying virus pathogenesis, as well as cytotoxicity testing and genetic manipulation studies.

Biochemical transformation of deoxythymidine kinase-deficient mouse cells with uv-irradiated equine herpesvirus type 1

International Nuclear Information System (INIS)

Allen, G.P.; McGowan, J.J.; Gentry, G.A.; Randall, C.C.

1978-01-01

A line of 3T3 mouse cells lacking deoxythymidine kinase (dTK - ) was stably transformed to the dTK + phenotype after exposure to uv-irradiated equine herpesvirus type 1 (EHV-1). Biochemical transformants were isolated in a system selective for the dTK + phenotype (Eagle minimal essential medium containing 10 -4 M hypoxanthine, 6 x 10 -7 M aminopterin, and 2 x 10 -5 M deoxythymidine). Transformation was accompanied by the acquisition of a dTK activity with immunological, electrophoretic, and biochemical characteristics identical to those of the dTK induced by EHV-1 during productive infection. The transformed cells have been maintained in selective culture medium for more than 50 passages and have retained the capacity to express EHV-1-specific antigens. Spontaneous release of infectious virus has not been detected in the transformed lines, and the cells were not oncogenic for athymic nude mice. In contrast to normal dTK + 3T3 cells, EHV-1 transformants were unable to grow in the presence of arabinosylthymine, a drug selectively phosphorylated by herpesvirus-coded dTK's. These results indicate that a portion of the EHV-1 genome is able to persist in the transformed cells for many generations and be expressed as an enzymatically active viral gene product

Genetic and biochemical characterization of rhizobacterial strains and their potential use in combination with chelants for assisted phytoremediation.

Science.gov (United States)

Cicatelli, Angela; Guarino, Francesco; Baldan, Enrico; Castiglione, Stefano

2017-03-01

Copper and zinc are essential micronutrients in plants but, at high concentrations, they are toxic. Assisted phytoremediation is an emerging "green" technology that aims to improve the efficiency of tolerant species to remove metals from soils through the use of chelants or microorganisms. Rhizobacteria can promote plant growth and tolerance and also affect the mobility, bioavailability, and complexation of metals. A pot experiment was conducted to evaluate the phytoremediation effectiveness of sunflowers cultivated in a Cu- and Zn-spiked soil, in the presence or absence of bacterial consortium and/or chelants. The consortium was constituted of two Stenotrophomonas maltophilia strains and one of Agrobacterium sp. These strains were previously isolated from the rhizosphere of maize plants cultivated on a metal-polluted soil and here molecularly and biochemically characterized. Results showed that the consortium improved sunflower growth and biomass production on the spiked soils. Sunflowers accumulated large amounts of metals in their roots and leaves; however, neither the bacterial consortium nor the chelants, singularly added to pots, influenced significantly Cu and Zn plant uptake. Furthermore, the consecutive soil amendment with the EDTA and bacterial consortium determined a consistent accumulation of metals in sunflowers, and it might be an alternative strategy to limit the use of EDTA and its associated environmental risks in phytoremediation.

Biochemical Responses of Peach Leaves Infected with Taphrina Deformans Berk/Tul.

Directory of Open Access Journals (Sweden)

Lyubka Koleva-Valkova

2017-01-01

Full Text Available The phytopathogenic fungus Taphrina deformans causing the so called “leaf curl disease” in peach trees leads to severe yield losses due to the development of leaf hypertrophy and subsequent necrosis and scission. Because of its economic importance, the molecular mechanisms underlying the onset and progression of the disease are of considerable interest to the agricultural science. In this study various biochemical parameters, including the activities of the antioxidant enzymes guaiacol peroxidase, syringaldazine peroxidase and catalase, total polyphenols and anthocyanin content, concentration of free proline, antiradical activity and quantity of plastid pigments, were characterized. All these were measured in both leaves with clear symptoms and distally situated leaves from the same plant that show no signs of the infection. The results demonstrate that the pathogen induces considerable biochemical changes concerning enzymatic and non‑enzymatic elements of the plant defense and antioxidant systems. Moreover, it seems that the fungus provokes a systemic response detectable even in the tissues without observable symptoms.

Establishment and characterization of novel epithelial-like cell lines derived from human periodontal ligament tissue in vitro.

Science.gov (United States)

Tansriratanawong, Kallapat; Ishikawa, Hiroshi; Toyomura, Junko; Sato, Soh

2017-10-01

In this study, novel human-derived epithelial-like cells (hEPLCs) lines were established from periodontal ligament (PDL) tissues, which were composed of a variety of cell types and exhibited complex cellular activities. To elucidate the putative features distinguishing these from epithelial rest of Malassez (ERM), we characterized hEPLCs based on cell lineage markers and tight junction protein expression. The aim of this study was, therefore, to establish and characterize hEPLCs lines from PDL tissues. The hEPLCs were isolated from PDL of third molar teeth. Cellular morphology and cell organelles were observed thoroughly. The characteristics of epithelial-endothelial-mesenchymal-like cells were compared in several markers by gene expression and immunofluorescence, to ERM and human umbilical-vein endothelial cells (HUVECs). The resistance between cellular junctions was assessed by transepithelial electron resistance, and inflammatory cytokines were detected by ELISA after infecting hEPLCs with periodontopathic bacteria. The hEPLCs developed into small epithelial-like cells in pavement appearance similar to ERM. However, gene expression patterns and immunofluorescence results were different from ERM and HUVECs, especially in tight junction markers (Claudin, ZO-1, and Occludins), and endothelial markers (vWF, CD34). The transepithelial electron resistance indicated higher resistance in hEPLCs, as compared to ERM. Periodontopathic bacteria were phagocytosed with upregulation of inflammatory cytokine secretion within 24 h. In conclusion, hEPLCs that were derived using the single cell isolation method formed tight multilayers colonies, as well as strongly expressed tight junction markers in gene expression and immunofluorescence. Novel hEPLCs lines exhibited differently from ERM, which might provide some specific functions such as metabolic exchange and defense mechanism against bacterial invasion in periodontal tissue.

Characterization of a mutant rat kangaroo cell line with alterations in the cell cycle and DNA repair

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Miyaji E.N.

2000-01-01

Full Text Available Using a positive selection system for isolating DNA replication and repair related mutants, we isolated a clone from a rat kangaroo cell line (PtK2 that has increased sensitivity to UV light. Characterization of this clone indicated normal post-replication repair after UV irradiation, and normal removal rates of cyclobutane pyrimidine dimers and pyrimidine(6-4pyrimidone photoproducts by excision repair. However, this cell line has decreased ability to make early incisions on damaged DNA, possibly indicating a defect in preferential repair of actively transcribed genes, and a slower cell proliferation rate, including a longer S-phase. This phenotype reinforces the present notion that control of key mechanisms in cell metabolism, such as cell cycle control, repair, transcription and cell death, can be linked.

Establishment and characterization of a spontaneously immortalized trophoblast cell line (HPT-8) and its hepatitis B virus-expressing clone.

Science.gov (United States)

Zhang, Lei; Zhang, Weilu; Shao, Chen; Zhang, Jingxia; Men, Ke; Shao, Zhongjun; Yan, Yongping; Xu, Dezhong

2011-08-01

Most trophoblast cell lines currently available to study vertical transmission of hepatitis B virus (HBV) are immortalized by viral transformation. Our goal was to establish and characterize a spontaneously immortalized human first-trimester trophoblast cell line and its HBV-expressing clone. Chorionic villi of Asian human first-trimester placentae were digested with trypsin and collagenase I to obtain the primary trophoblast cell culture. A spontaneously immortalized trophoblast cell line (HPT-8) was analyzed by scanning and transmission electron microscopy, cell cycle analysis, immunohistochemistry and immunofluorescence. HPT-8 cells were stably transfected with the adr subtype of HBV (HPT-8-HBV) and characterized by PCR and enzyme-linked immunosorbent assay. We obtained a clonal derivative of a spontaneously immortalized primary cell clone (HPT-8). HPT-8 cells were epithelioid and polygonal, and formed multinucleate, giant cells. They exhibited microvilli, distinct desmosomes between adjacent cells, abundant endoplasm, lipid inclusions and glycogen granules, which are all characteristic of cytotrophoblasts. HPT-8 cells expressed cytokeratin 7, cytokeratin 18, vimentin, cluster of differentiation antigen 9, epidermal growth factor receptor, stromal cell-derived factor 1 and placental alkaline phosphatase. They secreted prolactin, estradiol, progesterone and hCG, and were positive for HLA-G, a marker of extravillous trophoblasts. HPT-8-HBV cells were positive for HBV relaxed-circular, covalently closed circular DNA and pre-S sequence. HPT-8-HBV cells also produced and secreted HBV surface antigen and HBV e antigen. We established a trophoblast cell line, HPT-8 and its HBV-expressing clone which could be valuable in exploring the mechanism of HBV viral integration in human trophoblasts during intrauterine infection.

Biochemical Changes in the Fermentation of the Soy Sauce Prepared with Bittern

OpenAIRE

Chunqi Mao; Guoqing He; Xinyong Du; Meilin Cui; Shiyang Gao

2013-01-01

A new bittern soy sauce was developed; the overall aim of this study was to characterize the quality of the soy sauce. Biochemical changes occurring during experimental soy sauce fermentations were described. Results revealed that, the contents of amino nitrogen, reducing sugars and enzyme activity were slightly lower in soy sauce prepared with bittern than with traditional one. No marked difference in pH was noted between the two types of soy sauce.

Biochemical characterization of the plasma membrane H+ - ATPase from red beet (Beta vulgaris) hypocotyl tissue

International Nuclear Information System (INIS)

Oleski, N.A.

1986-01-01

Several biochemical techniques including selective solubilization followed by gel filtration or various types of affinity chromatography, and antibody production were employed in an attempt to purify the plasma membrane H + - ATPase from red beet hypocotyl tissue. While the enzyme could not be purified using any of these methods, it was possible to successfully conduct a more detailed biochemical analysis of the H + - ATPase. The molecular weight and isoelectric point of the enzyme were determined using N,N'dicyclohexylcarbodiimide (DCCD) and a H + - ATPase antibody. When plasma membrane vesicles were incubated with 20 μM [ 14 C]-DCCD at 0 C, a single 97,000 dalton protein was apparent on a fluorograph. A close correlation between [ 14 C]-DCCD labelling of the 97,000 dalton protein and the extent of ATPase inhibition over a range of DCCD concentrations suggests that this 97,000 dalton protein is a component of the plasma membrane H + - ATPase. An antibody raised against the plasma membrane H + - ATPase of Neurospora crassa cross-reacted with the 97,000 dalton DCCD-binding protein, further supporting the identity of this protein. Immunoblots of two dimensional gels of red beet plasma membrane vesicles indicated the isoelectric point of the enzyme to be pH 6.5

Characterization of the Goubau line for testing beam diagnostic instruments

Science.gov (United States)

Kim, S. Y.; Stulle, F.; Sung, C. K.; Yoo, K. H.; Seok, J.; Moon, K. J.; Choi, C. U.; Chung, Y.; Kim, G.; Woo, H. J.; Kwon, J.; Lee, I. G.; Choi, E. M.; Chung, M.

2017-12-01

One of the main characteristics of the Goubau line is that it supports a low-loss, non-radiated surface wave guided by a dielectric-coated metal wire. The dominant mode of the surface wave along the Goubau line is a TM01 mode, which resembles the pattern of the electromagnetic fields induced in the metallic beam pipe when the charged particle beam passes through it. Therefore, the Goubau line can be used for the preliminary bench test and performance optimization of the beam diagnostic instruments without requiring charged particle beams from the accelerators. In this paper, we discuss the basic properties of the Goubau line for testing beam diagnostic instruments and present the initial test results for button-type beam position monitors (BPMs). The experimental results are consistent with the theoretical estimations, which indicates that Goubau line allows effective testing of beam diagnostic equipment.

Lactation Defect in a Widely Used MMTV-Cre Transgenic Line of Mice

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Yuan, Taichang; Wang, Yongping; Pao, Lily; Anderson, Steve M.; Gu, Haihua

2011-01-01

Background MMTV-Cre mouse lines have played important roles in our understanding about the functions of numerous genes in mouse mammary epithelial cells during mammary gland development and tumorigenesis. However, numerous studies have not included MMTV-Cre mice as controls, and many investigators have not indicated which of the different MMTV-Cre founder lines were used in their studies. Here, we describe a lactation defect that severely limits the use of one of the most commonly used MMTV-Cre founder lines. Methodology/Principal Findings To explore the role of protein tyrosine phosphatase Shp1 in mammary gland development, mice bearing the floxed Shp1 gene were crossed with MMTV-Cre mice and mammary gland development was examined by histological and biochemical techniques, while lactation competency was assessed by monitoring pup growth. Surprisingly, both the Shp1fl/+;MMTV-Cre and MMTV-Cre female mice displayed a severe lactation defect when compared to the Shp1 fl/+ control mice. Histological and biochemical analyses reveal that female mice expressing the MMTV-Cre transgene, either alone or in combination with floxed genes, exhibit defects in lobuloalveolar expansion, presence of large cytoplasmic lipid droplets in luminal alveolar epithelial cells postpartum, and precocious induction of involution. Using a PCR-based genotyping method, the three different founder lines can be distinguished, and we determined that the MMTV-Cre line A, the most widely used MMTV-Cre founder line, exhibits a profound lactation defect that limits its use in studies on mammary gland development. Conclusions/Significance The identification of a lactation defect in the MMTV-Cre line A mice indicates that investigators must use MMTV-Cre alone mice as control in studies that utilize Cre recombinase to excise genes of interest from mammary epithelial cells. Our results also suggest that previous results obtained in studies using the MMTV-Cre line A line should be re-evaluated if the

Bioanalytical Applications of Fluorescence Line-Narrowing and Non-Line-Narrowing Spectroscopy Interfaced with Capillary Electrophoresis and High-Performance Liquid Chromatography

Energy Technology Data Exchange (ETDEWEB)

Roberts, Kenneth Paul [Iowa State Univ., Ames, IA (United States)

2001-01-01

Capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC) are widely used analytical separation techniques with many applications in chemical, biochemical, and biomedical sciences. Conventional analyte identification in these techniques is based on retention/migration times of standards; requiring a high degree of reproducibility, availability of reliable standards, and absence of coelution. From this, several new information-rich detection methods (also known as hyphenated techniques) are being explored that would be capable of providing unambiguous on-line identification of separating analytes in CE and HPLC. As further discussed, a number of such on-line detection methods have shown considerable success, including Raman, nuclear magnetic resonance (NMR), mass spectrometry (MS), and fluorescence line-narrowing spectroscopy (FLNS). In this thesis, the feasibility and potential of combining the highly sensitive and selective laser-based detection method of FLNS with analytical separation techniques are discussed and presented. A summary of previously demonstrated FLNS detection interfaced with chromatography and electrophoresis is given, and recent results from on-line FLNS detection in CE (CE-FLNS), and the new combination of HPLC-FLNS, are shown.

RMBNToolbox: random models for biochemical networks

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Niemi Jari

2007-05-01

Full Text Available Abstract Background There is an increasing interest to model biochemical and cell biological networks, as well as to the computational analysis of these models. The development of analysis methodologies and related software is rapid in the field. However, the number of available models is still relatively small and the model sizes remain limited. The lack of kinetic information is usually the limiting factor for the construction of detailed simulation models. Results We present a computational toolbox for generating random biochemical network models which mimic real biochemical networks. The toolbox is called Random Models for Biochemical Networks. The toolbox works in the Matlab environment, and it makes it possible to generate various network structures, stoichiometries, kinetic laws for reactions, and parameters therein. The generation can be based on statistical rules and distributions, and more detailed information of real biochemical networks can be used in situations where it is known. The toolbox can be easily extended. The resulting network models can be exported in the format of Systems Biology Markup Language. Conclusion While more information is accumulating on biochemical networks, random networks can be used as an intermediate step towards their better understanding. Random networks make it possible to study the effects of various network characteristics to the overall behavior of the network. Moreover, the construction of artificial network models provides the ground truth data needed in the validation of various computational methods in the fields of parameter estimation and data analysis.

Temperature dependence of dynamical permeability characterization of magnetic thin films using shorted microstrip line probe

International Nuclear Information System (INIS)

Li, Xiling; Li, Chengyi; Chai, Guozhi

2017-01-01

A temperature dependence microwave permeability characterization system of magnetic thin film up to 10 GHz is designed and fabricated. This system can be used at temperatures ranging from room temperature to 200 °C, and is based on a shorted microstrip probe, which is made by microwave printed circuit board. Without contacting the magnetic thin films to the probe, the microwave permeability of the film can be detected without any limitations of sample size and with almost the same accuracy, as shown by comparison with the results obtained from a shorted microstrip transmission-line fixture. The complex permeability can be deduced by an analytical approach from the measured reflection coefficient of a strip line ( S 11 ) with and without a ferromagnetic film material on it. The procedures are the same with the shorted microstrip transmission-line method. The microwave permeability of an oblique deposited CoZr thin film was investigated with this probe. The results show that the room temperature dynamic permeability of the CoZr film is in good agreement with the results obtained from the established short-circuited microstrip perturbation method. The temperature dependence permeability results fit well with the Landau–Lifshitz–Gilbert equation. Development of the temperature-dependent measurement of the magnetic properties of magnetic thin film may be useful for the high-frequency application of magnetic devices at high temperatures. (paper)

Enzymatic and biochemical characterization of Bungarus sindanus snake venom acetylcholinesterase

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M Ahmed

2012-01-01

Full Text Available This study analyses venom from the elapid krait snake Bungarus sindanus, which contains a high level of acetylcholinesterase (AChE activity. The enzyme showed optimum activity at alkaline pH (8.5 and 45ºC. Krait venom AChE was inhibited by substrate. Inhibition was significantly reduced by using a high ionic strength buffer; low ionic strength buffer (10 mM PO4 pH 7.5 inhibited the enzyme by 1. 5mM AcSCh, while high ionic strength buffer (62 mM PO4 pH 7.5 inhibited it by 1 mM AcSCh. Venom acetylcholinesterase was also found to be thermally stable at 45ºC; it only lost 5% of its activity after incubation at 45ºC for 40 minutes. The Michaelis-Menten constant (Km for acetylthiocholine iodide hydrolysis was found to be 0.068 mM. Krait venom acetylcholinesterase was also inhibited by ZnCl2, CdCl2, and HgCl2 in a concentrationdependent manner. Due to the elevated levels of AChE with high catalytic activity and because it is more stable than any other sources, Bungarus sindanus venom is highly valuable for biochemical studies of this enzyme.

Isolation and characterization of a radiosensitive Chinese hamster ovary cell line

International Nuclear Information System (INIS)

Fuller, L.F.

1987-01-01

A x-ray sensitive Chinese hamster ovary cell line was isolated using a semi-automated procedure in which mutagenized CHO cells were allowed to form colonies on top of agar, x-irradiated, then photographed at two later times. Comparison of the photographs allowed the identification of colonies which displayed significant growth arrest. One of the colonies identified in this manner produced a stable, radiosensitive line. This cell line is normal in x-ray induced inhibition of DNA synthesis, and single- and double-strand break repair, and is moderately sensitive to ethyl methane sulfonate and UV light. The sensitive line performs only half as much x-ray-induced repair replication as the parental line and this deficiency is believed to be the primary cause of its radiosensitivity. The sensitive line produces significantly higher numbers of x-ray-induced chromosome and chromatid aberrations including chromatid aberrations following exposure during the G 1 phase of the cell cycle. The line is hypomutable compared to the parental line with x-ray exposure inducing only one-third as many 6-thioguanine resistant colonies

Involvement of immunologic and biochemical mechanisms in the pathogenesis of Tourette's syndrome

Science.gov (United States)

Landau, Yuval Eliahu; Steinberg, Tamar; Richmand, Brian; Leckman, James Frederick; Apter, Alan

2014-01-01

Tourette's syndrome is a neurodevelopmental disorder clinically characterized by multiple motor and phonic tics. It is likely that a neurobiological susceptibility to the disorder is established during development by the interaction of genetic, biochemical, immunological, and environmental factors. This study sought to investigate the possible correlation of several immunological and biochemical markers with Tourette's syndrome. Children with Tourette's syndrome attending a tertiary pediatric medical center from May 2008 to April 2010, and healthy age-matched control subjects underwent a comprehensive biochemical and immunological work-up. Demographic data were abstracted from the medical records. Findings were compared between the groups and analyzed statistically. Sixty-eight children with Tourette's syndrome (58 males, 85.3%) and 36 healthy children (25 males, 69.4%) were recruited. Compared with the control group, the Tourette's syndrome group had significantly higher levels of ferritin (p = 0.01) and hemoglobin (p = 0.02), a lower level of zinc (p = 0.05), and a lower percentage of non-ceruloplasmin copper (p = 0.01). Analysis of the immunological markers revealed no significant between-group differences in IgA, IgM or IgG; however, IgE and IgG-4 levels were significantly higher in the Tourette's syndrome group (p = 0.04 and p = 0.02, respectively). Children with Tourette's syndrome have high levels of biochemical indices of oxidative stress and the quantitative immunoglobulins. These findings add to the still-limited knowledge on the pathogenesis of Tourette's syndrome and may have implications for the development of novel therapeutic modalities. PMID:22139323

Establishment and characterization of two human breast carcinoma cell lines by spontaneous immortalization: Discordance between Estrogen, Progesterone and HER2/neu receptors of breast carcinoma tissues with derived cell lines

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Kamalidehghan Behnam

2012-10-01

Full Text Available Abstract Background Breast cancer is one of the most common cancers among women throughout the world. Therefore, established cell lines are widely used as in vitro experimental models in cancer research. Methods Two continuous human breast cell lines, designated MBC1 and MBC2, were successfully established and characterized from invasive ductal breast carcinoma tissues of Malaysian patients. MBC1 and MBC2 have been characterized in terms of morphology analysis, population doubling time, clonogenic formation, wound healing assay, invasion assay, cell cycle, DNA profiling, fluorescence immunocytochemistry, Western blotting and karyotyping. Results MBC1 and MBC2 exhibited adherent monolayer epithelial morphology at a passage number of 150. Receptor status of MBC1 and MBC2 show (ER+, PR+, HER2+ and (ER+, PR-, HER2+, respectively. These results are in discordance with histopathological studies of the tumoral tissues, which were triple negative and (ER-, PR-, HER2+ for MBC1 and MBC2, respectively. Both cell lines were capable of growing in soft agar culture, which suggests their metastatic potential. The MBC1 and MBC2 metaphase spreads showed an abnormal karyotype, including hyperdiploidy and complex rearrangements with modes of 52–58 chromosomes per cell. Conclusions Loss or gain in secondary properties, deregulation and specific genetic changes possibly conferred receptor changes during the culturing of tumoral cells. Thus, we hypothesize that, among heterogenous tumoral cells, only a small minority of ER+/PR+/HER2+ and ER+/PR-/HER2+ cells with lower energy metabolism might survive and adjust easily to in vitro conditions. These cell lines will pave the way for new perspectives in genetic and biological investigations, drug resistance and chemotherapy studies, and would serve as prototype models in Malaysian breast carcinogenesis investigations.

Isolation and characterization of a spontaneously immortalized bovine retinal pigmented epithelial cell line

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Griffiths T Daniel

2009-05-01

Full Text Available Abstract Background The Retinal Pigmented Epithelium (RPE is juxtaposed with the photoreceptor outer segments of the eye. The proximity of the photoreceptor cells is a prerequisite for their survival, as they depend on the RPE to remove the outer segments and are also influenced by RPE cell paracrine factors. RPE cell death can cause a progressive loss of photoreceptor function, which can diminish vision and, over time, blindness ensues. Degeneration of the retina has been shown to induce a variety of retinopathies, such as Stargardt's disease, Cone-Rod Dystrophy (CRD, Retinitis Pigmentosa (RP, Fundus Flavimaculatus (FFM, Best's disease and Age-related Macular Degeneration (AMD. We have cultured primary bovine RPE cells to gain a further understanding of the mechanisms of RPE cell death. One of the cultures, named tRPE, surpassed senescence and was further characterized to determine its viability as a model for retinal diseases. Results The tRPE cell line has been passaged up to 150 population doublings and was shown to be morphologically similar to primary cells. They have been characterized to be of RPE origin by reverse transcriptase PCR and immunocytochemistry using the RPE-specific genes RPE65 and CRALBP and RPE-specific proteins RPE65 and Bestrophin. The tRPE cells are also immunoreactive to vimentin, cytokeratin and zonula occludens-1 antibodies. Chromosome analysis indicates a normal diploid number. The tRPE cells do not grow in suspension or in soft agar. After 3H thymidine incorporation, the cells do not appear to divide appreciably after confluency. Conclusion The tRPE cells are immortal, but still exhibit contact inhibition, serum dependence, monolayer growth and secrete an extra-cellular matrix. They retain the in-vivo morphology, gene expression and cell polarity. Additionally, the cells endocytose exogenous melanin, A2E and purified lipofuscin granules. This cell line may be a useful in-vitro research model for retinal

Differences in sialic acid residues among bone alkaline phosphatase isoforms: a physical, biochemical, and immunological characterization.

Science.gov (United States)

Magnusson, P; Farley, J R

2002-12-01

High-performance liquid chromatography (HPLC) separates three human bone alkaline phosphatase (BALP) isoforms in serum; two major BALP isoforms, B1 and B2, and a minor fraction, B/I, which is composed on average of 70% bone and 30% intestinal ALP. The current studies were intended to identify an in vitro source of the BALP isoforms for physical, biochemical, and immunological characterizations. The three BALP isoforms were identified in extracts of human osteosarcoma (SaOS-2) cells, by HPLC, after separation by anion-exchange chromatography. All three BALP isoforms were similar with respect to freeze-thaw stability, solubility, heat inactivation, and inhibition by L-phenylalanine, L-homoarginine, and levamisole. The isoforms were also kinetically similar (i.e., maximal velocity and KM at pH 8.8 and pH 10.0). The isoforms differed, however, with respect to sensitivity to precipitation with wheat germ agglutinin (WGA), P acid residues was estimated to be 29 and 45, for each B1 and B2 homodimer, respectively. Apparent discrepancies between these estimates of molecular weight and estimates based on gel filtration chromatography were attributed to nonspecific interactions between carbohydrate residues and the gel filtration beads. All three BALP isoforms showed similar dose-dependent linearity in the commercial Alkphase-B and Tandem-MP Ostase immunoassays, r = 0.944 and r = 0.985, respectively (P acid residues compared with B/I, which mainly explains the apparent differences in molecular weight. Future investigations will focus on the clinical and functional significance of the revealed differences in sialic acid residues.

Derivation and characterization of novel nonhuman primate embryonic stem cell lines from in vitro-fertilized baboon preimplantation embryos.

Science.gov (United States)

Chang, Tien-Cheng; Liu, Ya-Guang; Eddy, Carlton A; Jacoby, Ethan S; Binkley, Peter A; Brzyski, Robert G; Schenken, Robert S

2011-06-01

The development of nonhuman primate (NHP) embryonic stem cell (ESC) models holds great promise for cell-mediated treatment of debilitating diseases and to address numerous unanswered questions regarding the therapeutic efficacy of ESCs while supplanting ethical considerations involved with human studies. Here we report successful establishment and characterization of 3 novel baboon (Papio cynocephalus) ESC lines from the inner cell mass of intracytoplasmic sperm injection-derived blastocysts. Embryos were cultured in an improved baboon embryo in vitro culture protocol. The inner cell mass of blastocyst was laser-dissected and plated on mouse embryonic fibroblast feeder cell monolayer in the NHP ESC culture medium. Three cell lines with characteristic ESC morphology have been cultured through an extended period (>14 months), with 2 male cell lines (UT-1 and -2) and 1 female cell line (UT-3) displaying normal baboon karyotypes. Reverse transcription-polymerase chain reaction analysis confirmed that all 3 lines express primate ESC pluripotency markers, including OCT-4, NANOG, SOX-2, TERT, TDGF, LEFTYA, and REX-1. All 3 lines demonstrated positive immunocytochemical staining for OCT-4, stage-specific embryonic antigen-3, stage-specific embryonic antigen-4, TRA-1-60, and TRA-1-81. Baboon ESCs injected into NOD/SCID mice formed teratomas with all 3 germ layers. In addition, embryoid body-like spherical structures were derived and initial outgrowth was observed when embedded into extracellular matrix Matrigel. The ESC lines established in this NHP model have the potential to extend our knowledge in the fields of developmental biology, regenerative medicine, and future applications, including preclinical safety assessment of in vivo stem cell therapy.

Crystal Structure and Biochemical Characterization of a Mycobacterium smegmatis AAA-Type Nucleoside Triphosphatase Phosphohydrolase (Msm0858).

Science.gov (United States)

Unciuleac, Mihaela-Carmen; Smith, Paul C; Shuman, Stewart

2016-05-15

AAA proteins (ATPases associated with various cellular activities) use the energy of ATP hydrolysis to drive conformational changes in diverse macromolecular targets. Here, we report the biochemical characterization and 2.5-Å crystal structure of a Mycobacterium smegmatis AAA protein Msm0858, the ortholog of Mycobacterium tuberculosis Rv0435c. Msm0858 is a magnesium-dependent ATPase and is active with all nucleoside triphosphates (NTPs) and deoxynucleoside triphosphates (dNTPs) as substrates. The Msm0858 structure comprises (i) an N-terminal domain (amino acids [aa] 17 to 201) composed of two β-barrel modules and (ii) two AAA domains, D1 (aa 212 to 473) and D2 (aa 476 to 744), each of which has ADP in the active site. Msm0858-ADP is a monomer in solution and in crystallized form. Msm0858 domains are structurally homologous to the corresponding modules of mammalian p97. However, the position of the N-domain modules relative to the AAA domains in the Msm0858-ADP tertiary structure is different and would impede the formation of a p97-like hexameric quaternary structure. Mutational analysis of the A-box and B-box motifs indicated that the D1 and D2 AAA domains are both capable of ATP hydrolysis. Simultaneous mutations of the D1 and D2 active-site motifs were required to abolish ATPase activity. ATPase activity was effaced by mutation of the putative D2 arginine finger, suggesting that Msm0858 might oligomerize during the ATPase reaction cycle. A truncated variant Msm0858 (aa 212 to 745) that lacks the N domain was characterized as a catalytically active homodimer. Recent studies have underscored the importance of AAA proteins (ATPases associated with various cellular activities) in the physiology of mycobacteria. This study reports the ATPase activity and crystal structure of a previously uncharacterized mycobacterial AAA protein, Msm0858. Msm0858 consists of an N-terminal β-barrel domain and two AAA domains, each with ADP bound in the active site. Msm0858 is a

Genetic and biochemical characterization of HMB-1, a novel subclass B1 metallo-β-lactamase found in a Pseudomonas aeruginosa clinical isolate.

Science.gov (United States)

Pfennigwerth, Niels; Lange, Felix; Belmar Campos, Cristina; Hentschke, Moritz; Gatermann, Sören G; Kaase, Martin

2017-04-01

To characterize a novel subclass B1 metallo-β-lactamase (MBL) found in an MDR Pseudomonas aeruginosa clinical isolate. The isolate P. aeruginosa NRZ-03096 was recovered in 2012 from an anal swab from a patient hospitalized in Northern Germany and showed high MICs of carbapenems. MBL production was analysed by several phenotypic tests. Genetic characterization of the novel bla gene and MLST was performed by WGS. The novel bla gene was expressed in Escherichia coli TOP10 and the enzyme was subjected to biochemical characterization to determine the kinetic parameters K m and k cat . P. aeruginosa NRZ-03096 was resistant to all tested β-lactams and showed an MBL phenotype. Shotgun cloning experiments yielded a clone producing a novel subclass B1 enzyme with only 74.3% identity to the next nearest relative, KHM-1. The novel MBL was named HMB-1 (for Hamburg MBL). Analysis of WGS data showed that the bla HMB-1 gene was chromosomally located as part of a Tn 3 family transposon that was named Tn 6345 . Expression of bla HMB-1 in E. coli TOP10 led to increased resistance to β-lactams. Determination of K m and k cat revealed that HMB-1 had different hydrolytic characteristics compared with KHM-1, with lower hydrolytic rates for cephalosporins and a higher rate for imipenem. The identification of HMB-1 further underlines the ongoing spread and diversification of carbapenemases in Gram-negative human pathogens and especially in P. aeruginosa . © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Morphological and Biochemical Characterization of Bovine Congenital Psudomyotonia

OpenAIRE

Dorotea, Tiziano

2015-01-01

The Ca2+-ATPase of sarco(endo)plasmic reticulum (SERCA) is a protein of about 110 kDa member of the P-type ATPases family. SERCA pumps utilize the energy derived from the hydrolysis of a molecule of ATP to transport two Ca2+ ions across the Sarcoplasmic Reticulum (SR) membrane to decrease the Ca2+ concentration in the cytosol. SERCA isoform 1a (SERCA1a) is the mainly expressed isoform in adult fast-twitch muscle fibre and it is both structurally and functionally the best characterized member ...

In-line quantification and characterization of membrane fouling

KAUST Repository

Bucs, Szilard

2016-06-16

Methods of detecting, quantifying and/or characterizing the fouling of a device from a combination of pressure and spectroscopic data are provided. The device can be any device containing components susceptible to fouling. Components can include membranes, pipes, or reactors. Suitable devices include membrane devices, heat exchangers, and chemical or bio-reactors. Membrane devices can include, for example, microfiltration devices, ultrafiltration devices, nanofiltration devices, reverse osmosis, forward osmosis, osmosis, reverse electrodialysis, electro- deionisation or membrane distillation devices. The methods can be applied to any type of membrane, including tubular, spiral, hollow fiber, flat sheet, and capillary membranes. The spectroscopic characterization can include measuring one or more of the absorption, fluorescence, or raman spectroscopic data of one or more foulants. The methods can allow for the early detection and/or characterization of fouling. The characterization can include determining the specific foulant(s) or type of foulant(s) present. The characterization of fouling can allow for the selection of an appropriate de-fouling method and timing.

Generation and characterization of rat liver stem cell lines and their engraftment in a rat model of liver failure

Science.gov (United States)

Kuijk, Ewart W.; Rasmussen, Shauna; Blokzijl, Francis; Huch, Meritxell; Gehart, Helmuth; Toonen, Pim; Begthel, Harry; Clevers, Hans; Geurts, Aron M.; Cuppen, Edwin

2016-01-01

The rat is an important model for liver regeneration. However, there is no in vitro culture system that can capture the massive proliferation that can be observed after partial hepatectomy in rats. We here describe the generation of rat liver stem cell lines. Rat liver stem cells, which grow as cystic organoids, were characterized by high expression of the stem cell marker Lgr5, by the expression of liver progenitor and duct markers, and by low expression of hepatocyte markers, oval cell markers, and stellate cell markers. Prolonged cultures of rat liver organoids depended on high levels of WNT-signalling and the inhibition of BMP-signaling. Upon transplantation of clonal lines to a Fah−/− Il2rg−/− rat model of liver failure, the rat liver stem cells engrafted into the host liver where they differentiated into areas with FAH and Albumin positive hepatocytes. Rat liver stem cell lines hold potential as consistent reliable cell sources for pharmacological, toxicological or metabolic studies. In addition, rat liver stem cell lines may contribute to the development of regenerative medicine in liver disease. To our knowledge, the here described liver stem cell lines represent the first organoid culture system in the rat. PMID:26915950

Characterization of the early events in dengue virus cell entry by biochemical assays and single-virus tracking

NARCIS (Netherlands)

van der Schaar, Hilde M.; Rust, Michael J.; Waarts, Barry-Lee; van der Ende-Metselaarl, Heidi; Kuhn, Richard J.; Wilschut, Jan; Zhuang, Xiaowei; Smit, Jolanda M.

In this study, we investigated the cell entry characteristics of dengue virus (DENV) type 2 strain SI on mosquito, BHK-15, and BS-C-1 cells. The concentration of virus particles measured by biochemical assays was found to be substantially higher than the number of infectious particles determined by

Characterization of the early events in dengue virus cell entry by biochemical assays and single-virus tracking

NARCIS (Netherlands)

van der Schaar, Hilde M.; Rust, Michael J.; Waarts, Barry-Lee; van der Ende-Metselaarl, Heidi; Kuhn, Richard J.; Wilschut, Jan; Zhuang, Xiaowei; Smit, Jolanda M.

2007-01-01

In this study, we investigated the cell entry characteristics of dengue virus (DENV) type 2 strain SI on mosquito, BHK-15, and BS-C-1 cells. The concentration of virus particles measured by biochemical assays was found to be substantially higher than the number of infectious particles determined by

PIXE analysis of U and Pu from hot particles: K-lines vs L-lines

DEFF Research Database (Denmark)

Jiménez-Ramos, M.C.; García López, J.; Eriksson, M.

2012-01-01

. (2007) [1] and Jiménez-Ramos et al. (2010) [2], radioactive particles from aircraft accidents in Palomares and Thule have been characterized. Using micro-Proton Induced X-ray Emission (μ-PIXE) and confocal X-ray fluorescence microprobe (μ-XRF), the L-lines of the main components in these particles, U......-lines present not overlapping between the main peaks in the spectra and the X-ray absorption in the sample is much lower than for L-lines....

Structural and biochemical characterization of 3-hydroxybenzoate 6-hydroxylase

NARCIS (Netherlands)

Montersino, S.

2012-01-01

The thesis deals with the characterization of a new flavoprotein hydroxylase 3 hydroxybenzoate 6-hydroxylase (3HB6H) from Rhodococcus jostii RHA1. 3HB6H is able to insert exclusively oxygen in para-position and the enzyme has been chosen to study the structural basis of such regioselectivity. As

On the Adaptive Design Rules of Biochemical Networks in Evolution

Directory of Open Access Journals (Sweden)

Bor-Sen Chen

2007-01-01

Full Text Available Biochemical networks are the backbones of physiological systems of organisms. Therefore, a biochemical network should be sufficiently robust (not sensitive to tolerate genetic mutations and environmental changes in the evolutionary process. In this study, based on the robustness and sensitivity criteria of biochemical networks, the adaptive design rules are developed for natural selection in the evolutionary process. This will provide insights into the robust adaptive mechanism of biochemical networks in the evolutionary process. We find that if a mutated biochemical network satisfies the robustness and sensitivity criteria of natural selection, there is a high probability for the biochemical network to prevail during natural selection in the evolutionary process. Since there are various mutated biochemical networks that can satisfy these criteria but have some differences in phenotype, the biochemical networks increase their diversities in the evolutionary process. The robustness of a biochemical network enables co-option so that new phenotypes can be generated in evolution. The proposed robust adaptive design rules of natural selection gain much insight into the evolutionary mechanism and provide a systematic robust biochemical circuit design method of biochemical networks for biotechnological and therapeutic purposes in the future.

In-line digital holography with phase-shifting Greek-ladder sieves

Science.gov (United States)

Xie, Jing; Zhang, Junyong; Zhang, Yanli; Zhou, Shenlei; Zhu, Jianqiang

2018-04-01

Phase shifting is the key technique in in-line digital holography, but traditional phase shifters have their own limitations in short wavelength regions. Here, phase-shifting Greek-ladder sieves with amplitude-only modulation are introduced into in-line digital holography, which are essentially a kind of diffraction lens with three-dimensional array diffraction-limited foci. In the in-line digital holographic experiment, we design two kinds of sieves by lithography and verify the validity of their phase-shifting function by measuring a 1951 U.S. Air Force resolution test target and three-dimensional array foci. With advantages of high resolving power, low cost, and no limitations at shorter wavelengths, phase-shifting Greek-ladder sieves have great potential in X-ray holography or biochemical microscopy for the next generation of synchrotron light sources.

Genetic variability among advanced lines of brassica

International Nuclear Information System (INIS)

Ullah, N.; Farhatullah, A.; Rahman, H.U.; Fayyaz, L.

2015-01-01

Genetic variability for morphological and biochemical traits among six advanced lines (F10:11) of brassica was studied at The University of Agriculture Peshawar during crop season of 2012-13. These lines were developed through interspecific hybridization. Significant differences at (p=0.01) for plant height, main, pods main raceme-1, pod length, seed yield plant-1 and protein content at (p=0.05) for 100-seed weight, oil content were recorded. The advanced line, AUP-05 produced the maximum seed yield plant-1 (19.73 g), protein content (24.56%), 100-seed weight (0.64 g). Advanced line AUP-04 had the highest erucic acid (50.31%), linolenic acid (10.60%) and was late maturing (179.33). Advanced line AUP-06 produced the high oil content (48.82%). Advanced line AUP-03 produced comparatively longer main raceme (69.32 cm). Environmental variance was smaller than genotypic variance for majority of the traits. Genotypic and phenotypic coefficients of variation ranged from 2.45 to 25.67% and 2.50 to 27.68%, respectively. Heritability was high for majority of the traits. The maximum heritability was recorded for plant height (0.61), main raceme length (0.81), pods main raceme-1 (0.74), seed yield plant-1 (0.86) and protein content (0.77). Moderate heritability was observed for oil (0.58) contents. Heritability for 100-seed weight (0.30) was the lowest. These lines may be released as new improved varieties for specific parameters. (author)

Isolation, Characterization, and Establishment of Spontaneously Immortalized Cell Line HRPE-2S With Stem Cell Properties.

Science.gov (United States)

Shams Najafabadi, Hoda; Soheili, Zahra-Soheila; Samiei, Shahram; Ahmadieh, Hamid; Ranaei Pirmardan, Ehsan; Masoumi, Maryam

2017-10-01

The retinal pigment epithelium is a monolayer of highly specialized pigmented cells located between the neural retina and the Bruch's membrane of the choroid. RPE cells play a crucial role in the maintenance and function of the underlying photoreceptors. This study introduces a spontaneously arising human retinal pigment epithelial cell line, HRPE-2S, which was isolated from primary RPE cell culture of 2 days old male donor. We characterized morphology and functional properties of the new cell line. The immortalized cell line was maintained in culture for more than 70 passages and 240 divisions. The average doubling time of the cells was approximately 22 h and got freezed at 26th passage. The cell line expressed RPE-specific markers RPE65 and cell junction protein ZO1 as an epithelial cell marker. It also expressed CHX10, PAX6, Nestin, SOX2 as stem and retinal progenitor cell markers. Ki67 as a marker of cell proliferation was expressed in all HRPE-2S cells. It represented typical epithelial cobblestone morphology and did not phenotypically change through several passages. Stem cell-like aggregations (neurospheres) were observed in SEM microscopy. The cells represented high mitotic index. They could be viable under hypoxic conditions and serum deprivation. According to functional studies, the cell line exhibited stem cell-like behaviors with particular emphasis on its self-renewal capacity. LDH isoenzymes expression pattern confirmed the same cellular source for both of the HRPE-2S cells and primary RPE cells. Characteristics of HRPE-2S cells promise it as an in vitro model for RPE stem cell-based researches. J. Cell. Physiol. 232: 2626-2640, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

PREDICTORS OF BIOCHEMICAL REMISSION AND RECURRENCE AFTER SURGICAL AND RADIATION TREATMENTS OF CUSHING DISEASE: A SYSTEMATIC REVIEW AND META-ANALYSIS.

Science.gov (United States)

Abu Dabrh, Abd Moain Abu; Singh Ospina, Naykky M; Al Nofal, Alaa; Farah, Wigdan H; Barrionuevo, Patricia; Sarigianni, Maria; Mohabbat, Arya B; Benkhadra, Khalid; Carranza Leon, Barbara G; Gionfriddo, Michael R; Wang, Zhen; Mohammed, Khaled; Ahmed, Ahmed T; Elraiyah, Tarig A; Haydour, Qusay; Alahdab, Fares; Prokop, Larry J; Murad, Mohammad Hassan

2016-04-01

We conducted a systematic review and meta-analysis to synthesize the evidence about predictors that may affect biochemical remission and recurrence after transsphenoidal surgery (TSS), radiosurgery (RS), and radiotherapy (RT) in Cushing disease. We searched multiple databases through December 2014 including original controlled and uncontrolled studies that enrolled patients with Cushing disease who received TSS (first-line), RS, or RT. We extracted data independently, in duplicates. Outcomes of interest were biochemical remission and recurrence. A meta-analysis was conducted using the random-effects model to estimate event rates with 95% confidence intervals (CIs). First-line TSS was associated with high remission (76% [95% CI, 72 to 79%]) and low recurrence rates (10% [95% CI, 6 to 16%]). Remission after TSS was higher in patients with microadenomas or positive-adrenocorticotropic hormone tumor histology. RT was associated with a high remission rate (RS, 68% [95% CI, 61 to 77%]; RT, 66% [95% CI, 58 to 75%]) but also with a high recurrence rate (RS, 32% [95% CI, 16 to 60%]; RT, 26% [95% CI, 14 to 48%]). Remission after RS was higher at short-term follow-up (≤2 years) and with high-dose radiation, while recurrence was higher in women and with lower-dose radiation. Remission was after RT in adults who received TSS prior to RT, and with lower radiation doses. There was heterogeneity (nonstandardization) in the criteria and cutoff points used to define biochemical remission and recurrence. First-line TSS is associated with high remission and low recurrence, while RS and RT are associated with reasonable remission rates but important recurrence rates. The current evidence warrants low confidence due to the noncomparative nature of the studies, high heterogeneity, and imprecision.

Raman spectroscopic biochemical mapping of tissues

Science.gov (United States)

Stone, Nicholas; Hart Prieto, Maria C.; Kendall, Catherine A.; Shetty, Geeta; Barr, Hugh

2006-02-01

Advances in technologies have brought us closer to routine spectroscopic diagnosis of early malignant disease. However, there is still a poor understanding of the carcinogenesis process. For example it is not known whether many cancers follow a logical sequence from dysplasia, to carcinoma in situ, to invasion. Biochemical tissue changes, triggered by genetic mutations, precede morphological and structural changes. These can be probed using Raman or FTIR microspectroscopy and the spectra analysed for biochemical constituents. Local microscopic distribution of various constituents can then be visualised. Raman mapping has been performed on a number of tissues including oesophagus, breast, bladder and prostate. The biochemical constituents have been calculated at each point using basis spectra and least squares analysis. The residual of the least squares fit indicates any unfit spectral components. The biochemical distribution will be compared with the defined histopathological boundaries. The distribution of nucleic acids, glycogen, actin, collagen I, III, IV, lipids and others appear to follow expected patterns.

Biochemical characterization and anti-inflammatory properties of an isothiocyanate-enriched moringa (Moringa oleifera) seed extract.

Science.gov (United States)

Jaja-Chimedza, Asha; Graf, Brittany L; Simmler, Charlotte; Kim, Youjin; Kuhn, Peter; Pauli, Guido F; Raskin, Ilya

2017-01-01

Moringa oleifera Lam. is a tropical plant, used for centuries as food and traditional medicine. The aim of this study was to develop, validate and biochemically characterize an isothiocyanate-enriched moringa seed extract (MSE), and to compare the anti-inflammatory effects of MSE-containing moringa isothiocyanate-1 (MIC-1) with a curcuminoid-enriched turmeric extract (CTE), and a material further enriched in its primary phytochemical, curcumin (curcumin-enriched material; CEM). MSE was prepared by incubating ground moringa seeds with water to allow myrosinase-catalyzed enzymatic formation of bioactive MIC-1, the predominant isothiocyanate in moringa seeds. Optimization of the extraction process yielded an extract of 38.9% MIC-1. Phytochemical analysis of MSE revealed the presence of acetylated isothiocyanates, phenolic glycosides unique to moringa, flavonoids, fats and fatty acids, proteins and carbohydrates. MSE showed a reduction in the carrageenan-induced rat paw edema (33% at 500 mg/kg MIC-1) comparable to aspirin (27% at 300 mg/kg), whereas CTE did not have any significant effect. In vitro, MIC-1 at 1 μM significantly reduced the production of nitric oxide (NO) and at 5 μM, the gene expression of LPS-inducible nitric oxide synthase (iNOS) and interleukins 1β and 6 (IL-1β and IL-6), whereas CEM did not show any significant activity at all concentrations tested. MIC-1 (10μM) was also more effective at upregulating the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) target genes NAD(P)H:quinone oxidoreductase 1 (NQO1), glutathione S-transferase pi 1 (GSTP1), and heme oxygenase 1 (HO1) than the CEM. Thus, in contrast to CTE and CEM, MSE and its major isothiocyanate MIC-1 displayed strong anti-inflammatory and antioxidant properties in vivo and in vitro, making them promising botanical leads for the mitigation of inflammatory-mediated chronic disorders.

Biochemical characterization and anti-inflammatory properties of an isothiocyanate-enriched moringa (Moringa oleifera seed extract.

Directory of Open Access Journals (Sweden)

Asha Jaja-Chimedza

Full Text Available Moringa oleifera Lam. is a tropical plant, used for centuries as food and traditional medicine. The aim of this study was to develop, validate and biochemically characterize an isothiocyanate-enriched moringa seed extract (MSE, and to compare the anti-inflammatory effects of MSE-containing moringa isothiocyanate-1 (MIC-1 with a curcuminoid-enriched turmeric extract (CTE, and a material further enriched in its primary phytochemical, curcumin (curcumin-enriched material; CEM. MSE was prepared by incubating ground moringa seeds with water to allow myrosinase-catalyzed enzymatic formation of bioactive MIC-1, the predominant isothiocyanate in moringa seeds. Optimization of the extraction process yielded an extract of 38.9% MIC-1. Phytochemical analysis of MSE revealed the presence of acetylated isothiocyanates, phenolic glycosides unique to moringa, flavonoids, fats and fatty acids, proteins and carbohydrates. MSE showed a reduction in the carrageenan-induced rat paw edema (33% at 500 mg/kg MIC-1 comparable to aspirin (27% at 300 mg/kg, whereas CTE did not have any significant effect. In vitro, MIC-1 at 1 μM significantly reduced the production of nitric oxide (NO and at 5 μM, the gene expression of LPS-inducible nitric oxide synthase (iNOS and interleukins 1β and 6 (IL-1β and IL-6, whereas CEM did not show any significant activity at all concentrations tested. MIC-1 (10μM was also more effective at upregulating the nuclear factor (erythroid-derived 2-like 2 (Nrf2 target genes NAD(PH:quinone oxidoreductase 1 (NQO1, glutathione S-transferase pi 1 (GSTP1, and heme oxygenase 1 (HO1 than the CEM. Thus, in contrast to CTE and CEM, MSE and its major isothiocyanate MIC-1 displayed strong anti-inflammatory and antioxidant properties in vivo and in vitro, making them promising botanical leads for the mitigation of inflammatory-mediated chronic disorders.

Biochemical characterization of prostate-specific membrane antigen from canine prostate carcinoma cells.

Science.gov (United States)

Wu, Lisa Y; Johnson, Jacqueline M; Simmons, Jessica K; Mendes, Desiree E; Geruntho, Jonathan J; Liu, Tiancheng; Dirksen, Wessel P; Rosol, Thomas J; Davis, William C; Berkman, Clifford E

2014-05-01

Prostate-specific membrane antigen (PSMA) remains an important target for diagnostic and therapeutic application for human prostate cancer. Model cell lines have been recently developed to study canine prostate cancer but their PSMA expression and enzymatic activity have not been elucidated. The present study was focused on determining PSMA expression in these model canine cell lines and the use of fluorescent small-molecule enzyme inhibitors to detect canine PSMA expression by flow cytometry. Western blot and RT-PCR were used to determine the transcriptional and translational expression of PSMA on the canine cell lines Leo and Ace-1. An endpoint HPLC-based assay was used to monitor the enzymatic activity of canine PSMA and the potency of enzyme inhibitors. Flow cytometry was used to detect the PSMA expressed on Leo and Ace-1 cells using a fluorescently tagged PSMA enzyme inhibitor. Canine PSMA expression on the Leo cell line was confirmed by Western blot and RT-PCR, the enzyme activity, and flow cytometry. Kinetic parameters Km and Vmax of PSMA enzymatic activity for the synthetic substrate (PABGγG) were determined to be 393 nM and 220 pmol min(-1)  mg protein(-1) , respectively. The inhibitor core 1 and fluorescent inhibitor 2 were found to be potent reversible inhibitors (IC50  = 13.2 and 1.6 nM, respectively) of PSMA expressed on the Leo cell line. Fluorescent labeling of Leo cells demonstrated that the fluorescent PSMA inhibitor 2 can be used for the detection of PSMA-positive canine prostate tumor cells. Expression of PSMA on Ace-1 was low and not detectable by flow cytometry. The results described herein have demonstrated that PSMA is expressed on canine prostate tumor cells and exhibits similar enzymatic characteristics as human PSMA. The findings show that the small molecule enzyme inhibitors currently being studied for use in diagnosis and therapy of human prostate cancer can also be extended to include canine prostate cancer. Importantly

Experimental characterization of the flux-line lattice in superconducting V3Si

International Nuclear Information System (INIS)

Christen, D.K.; Kerchner, H.R.; Narayan, J.; Chang, Y.K.; Larson, B.C.; Sekula, S.T.

1982-01-01

Several microscopic properties of the flux-line lattice (FLL) in three separate single crystals of V 3 Si have been investigated by means of small-angle neutron diffraction. These low-field FLL characterizations have been correlated with the following material and superconducting properties: (1) the real crystal symmetry parallel to the applied magnetic field; (2) the micro-structure as determined by TEM; (3) magnetic irreversibilities in the mixed state; (4) reversible flux-line motion in ac response; and (5) martensitic structural transformation observed by x-ray diffraction. The three samples, V 3 Si-MP3, -MP4, and -MP5 possessed different defect structures, and this was manifested foremost in the FLL perfection. At low field (B 0 K, only MP3, which is free of second-phase precipitates, showed a highly resolved FLL. Sample MP5 contains a low density of small (200 A) coherent precipitates, and revealed well-defined FLL Bragg peaks for B greater than or equal to 0.5 T, but a highly mosaic, nearly polycrystalline FLL at lower fields. Sample MP4 contained large (500 to 1000 A) incoherent precipitates, and showed only a polycrystalline FLL at low field. In both MP3 and MP5, distinct anisotropic correlations were observed between the FLL morphology and the real-crystal direction along the applied field. The FLL perfection was strongly dependent on the growth history. The peak width history dependence for two different scattering geometries can be qualitatively modeled by proposed flux-pinning mechanisms. Quantitative comparisons with critical current measurements, however, are not totally reconcilable

Biochemical characterization of thermophilic lignocellulose degrading enzymes and their potential for biomass bioprocessing

Energy Technology Data Exchange (ETDEWEB)

Zambare, Vasudeo; Zambare, Archana; Christopher, Lew P. [Center for Bioprocessing Research & Development, South Dakota School of Mines and Technology, Rapid City 57701, SD (United States); Muthukumarappan, Kasiviswanath [Center for Bioprocessing Research & Development, South Dakota State University, Brookings 57007, SD (United States)

2011-07-01

. This could have important implications in the enzymatic breakdown of lignocellulosic biomass for the establishment of a robust and cost-efficient process for production of cellulosic ethanol. To the best of our knowledge, this work represents the first report in literature on biochemical characterization of lignocellulose-degrading enzymes from a thermophilic microbial consortium.

(Hygienic quality of lakes which are used as open-air bath. 2. Communication: Comparison of biochemical typed Escherichia coli strains isolated from lake water and bird excrements (author's transl))

Energy Technology Data Exchange (ETDEWEB)

Dott, W.; Wolff, H.J.; Botzenhart, K.

1981-01-01

368 strains of Escherichia coli were isolated from lakewaters and bird excrements (mainly black-headed gulls) using mandatory methods for drinking water examination. All strains were further characterized by 43 biochemical physiological and 30 morphological features. Besides the API 30 E Enterobacteriaceae system and the Roche Enterotube system of biochemical testing were used comparatively. The analysis using numerical methods for taxonomy pointed out that a single biotop could be characterized by the small physiological variation of the strains.

On-line characterization of emulsions aiming equipment sizing and definition of primary processing conditions; Caracterizacao em tempo real de emulsoes visando o dimensionamento de equipamentos e a definicao das condicoes de processamento primario

Energy Technology Data Exchange (ETDEWEB)

Lucas, Gloria M.S. [Fundacao Gorceix, Ouro Preto, MG (Brazil); Oliveira, Roberto C.G. de [PETROBRAS, Rio de Janeiro, RJ (Brazil)

2008-07-01

In this work are presented the results of the test of emulsion generation, carried out in the Experimental Nucleus of PETROBRAS in Atalaia city Brazil, in which equipment for on-line measurement of droplet size distribution was tested, whose operation principle is of acoustic attenuation. One of the objectives of this study was the validation of this technology for on-line characterization of petroleum emulsions, aiming at the optimization of the sub sea or top side processing equipment. The emulsions had been on-line monitored by the acoustic spectrometer, installed downstream of a choke valve. Samples of generated emulsions had been collected in different points of the circuit and characterized off-line how much its stability, droplet size distribution (DSD), water cut and viscosity. Moreover, in some stages of the test a demulsifier was injected so as to verify the behavior of emulsions. The acoustic spectrometry technique demonstrated to be efficient in the on-line characterization of the droplet size distributions of emulsions generated in different operational conditions. The acoustic technique presented satisfactory performance, in view of the comparison of the values of median gotten on-line and off-line. (author)

Effect of Substrate-Inoculum Ratio on the Biochemical Methane Potential of Municipal Biowastes

OpenAIRE

Parra-Orobio Brayan Alexis; Torres-Lozada Patricia; Marmolejo-Rebellón Luis Fernando; Cárdenas-Cleves Lina Marcela; Vásquez-Franco Carlos; Torres-López Wilmar Alexander; Ordoñez-Andrade José Abdón

2015-01-01

Biowastes are the predominant fraction of municipal solid waste (MSW) of developing countries. Biowastes are characterized by a high content of organic matter; characteristics which facilitates its transformation through biological processes such as anaerobic digestion (AD). Using biochemical methane potential (PBM) assays, in this study we evaluated the influence of substrate-inoculum (S/I) ratio on the AD of biowaste from MSW. The S/I ranged between 0.25 to 9 gSVsubstrate * ...

Establishment and characterization of a new human pancreatic adenocarcinoma cell line with high metastatic potential to the lung

International Nuclear Information System (INIS)

Kalinina, Tatyana; Simon, Ronald; Otto, Benjamin; Dierlamm, Judith; Schwarzenbach, Heidi; Effenberger, Katharina E; Bockhorn, Maximilian; Izbicki, Jakob R; Yekebas, Emre F; Güngör, Cenap; Thieltges, Sabrina; Möller-Krull, Maren; Murga Penas, Eva Maria; Wicklein, Daniel; Streichert, Thomas; Schumacher, Udo; Kalinin, Viacheslav

2010-01-01

Pancreatic cancer is still associated with devastating prognosis. Real progress in treatment options has still not been achieved. Therefore new models are urgently needed to investigate this deadly disease. As a part of this process we have established and characterized a new human pancreatic cancer cell line. The newly established pancreatic cancer cell line PaCa 5061 was characterized for its morphology, growth rate, chromosomal analysis and mutational analysis of the K-ras, EGFR and p53 genes. Gene-amplification and RNA expression profiles were obtained using an Affymetrix microarray, and overexpression was validated by IHC analysis. Tumorigenicity and spontaneous metastasis formation of PaCa 5061 cells were analyzed in pfp -/- /rag2 -/- mice. Sensitivity towards chemotherapy was analysed by MTT assay. PaCa 5061 cells grew as an adhering monolayer with a doubling time ranging from 30 to 48 hours. M-FISH analyses showed a hypertriploid complex karyotype with multiple numerical and unbalanced structural aberrations. Numerous genes were overexpressed, some of which have previously been implicated in pancreatic adenocarcinoma (GATA6, IGFBP3, IGFBP6), while others were detected for the first time (MEMO1, RIOK3). Specifically highly overexpressed genes (fold change > 10) were identified as EGFR, MUC4, CEACAM1, CEACAM5 and CEACAM6. Subcutaneous transplantation of PaCa 5061 into pfp -/- /rag2 -/- mice resulted in formation of primary tumors and spontaneous lung metastasis. The established PaCa 5061 cell line and its injection into pfp -/- /rag2 -/- mice can be used as a new model for studying various aspects of the biology of human pancreatic cancer and potential treatment approaches for the disease

In vitro phenotypic, genomic and proteomic characterization of a cytokine-resistant murine β-TC3 cell line.

Directory of Open Access Journals (Sweden)

Antonina Coppola

Full Text Available Type 1 diabetes mellitus (T1DM is caused by the selective destruction of insulin-producing β-cells. This process is mediated by cells of the immune system through release of nitric oxide, free radicals and pro-inflammatory cytokines, which induce a complex network of intracellular signalling cascades, eventually affecting the expression of genes involved in β-cell survival.The aim of our study was to investigate possible mechanisms of resistance to cytokine-induced β-cell death. To this purpose, we created a cytokine-resistant β-cell line (β-TC3R by chronically treating the β-TC3 murine insulinoma cell line with IL-1β + IFN-γ. β-TC3R cells exhibited higher proliferation rate and resistance to cytokine-mediated cell death in comparison to the parental line. Interestingly, they maintained expression of β-cell specific markers, such as PDX1, NKX6.1, GLUT2 and insulin. The analysis of the secretory function showed that β-TC3R cells have impaired glucose-induced c-peptide release, which however was only moderately reduced after incubation with KCl and tolbutamide. Gene expression analysis showed that β-TC3R cells were characterized by downregulation of IL-1β and IFN-γ receptors and upregulation of SOCS3, the classical negative regulator of cytokines signaling. Comparative proteomic analysis showed specific upregulation of 35 proteins, mainly involved in cell death, stress response and folding. Among them, SUMO4, a negative feedback regulator in NF-kB and JAK/STAT signaling pathways, resulted hyper-expressed. Silencing of SUMO4 was able to restore sensitivity to cytokine-induced cell death in β-TC3R cells, suggesting it may play a key role in acquired cytokine resistance by blocking JAK/STAT and NF-kB lethal signaling.In conclusion, our study represents the first extensive proteomic characterization of a murine cytokine-resistant β-cell line, which might represent a useful tool for studying the mechanisms involved in resistance to

Molecular and biochemical characterizations of three fructose-1,6-bisphosphate aldolases from Clonorchis sinensis.

Science.gov (United States)

Li, Shan; Bian, Meng; Wang, Xiaoyun; Chen, Xueqing; Xie, Zhizhi; Sun, Hengchang; Jia, Feifei; Liang, Pei; Zhou, Chenhui; He, Lei; Mao, Qiang; Huang, Bo; Liang, Chi; Wu, Zhongdao; Li, Xuerong; Xu, Jin; Huang, Yan; Yu, Xinbing

2014-01-01

Fructose-1,6-bisphosphate aldolase (FbA) is a ubiquitous enzyme in glycolysis. In the present study, we screened out three distinct genes encoding FbA isozymes (CsFbAs, CsFbA-1/2/3) from Clonorchis sinensis (C. sinensis) and characterized their sequences and structures profiles as well as biochemical properties. The amino acid sequences of CsFbAs shared homology with those of Class I FbAs from other species. The putative quaternary structures revealed that CsFbA-2 and CsFbA-3 were tetramers, while CsFbA-1 was dimer. Recombinant CsFbA-2 and CsFbA-3 (rCsFbA-2/3) were confirmed to be Class I FbAs for their stable enzymatic activities in the presence of EDTA or metal ions. However, recombinant CsFbA-1 (rCsFbA-1) did not show the catalytic activity, which might be due to the inappropriate fold and interaction between its subunits. Both rCsFbA-2 and rCsFbA-3 showed similar enzymatic properties such as optimal temperatures and broad pH ranges that similar to human FbA isozymes. They showed relatively higher affinities for fructose-1,6-bisphosphate (FBP) than fructose-1-phosphate (F-1-P). Their kcat ratios of FBP to F-1-P were in accordance with those of human FbA-A or C. In addition, CsFbAs were differentially transcribed in the developmental stages of C. sinensis, suggesting their essential roles throughout the life stages. Extensive distribution of CsFbAs in adult worms indicated that ubiquitous activities of CsFbAs took place in these organs. Collectively, these results suggested that long-term parasitic environment might adapt these isozymes similar to host FbAs for metabolic requirement. Our study will provide new insight into CsFbAs in the glycometabolism of C. sinensis and relationship between the host and the parasite. Copyright © 2014 Elsevier B.V. All rights reserved.

Text-based phenotypic profiles incorporating biochemical phenotypes of inborn errors of metabolism improve phenomics-based diagnosis.

Science.gov (United States)

Lee, Jessica J Y; Gottlieb, Michael M; Lever, Jake; Jones, Steven J M; Blau, Nenad; van Karnebeek, Clara D M; Wasserman, Wyeth W

2018-05-01

Phenomics is the comprehensive study of phenotypes at every level of biology: from metabolites to organisms. With high throughput technologies increasing the scope of biological discoveries, the field of phenomics has been developing rapid and precise methods to collect, catalog, and analyze phenotypes. Such methods have allowed phenotypic data to be widely used in medical applications, from assisting clinical diagnoses to prioritizing genomic diagnoses. To channel the benefits of phenomics into the field of inborn errors of metabolism (IEM), we have recently launched IEMbase, an expert-curated knowledgebase of IEM and their disease-characterizing phenotypes. While our efforts with IEMbase have realized benefits, taking full advantage of phenomics requires a comprehensive curation of IEM phenotypes in core phenomics projects, which is dependent upon contributions from the IEM clinical and research community. Here, we assess the inclusion of IEM biochemical phenotypes in a core phenomics project, the Human Phenotype Ontology. We then demonstrate the utility of biochemical phenotypes using a text-based phenomics method to predict gene-disease relationships, showing that the prediction of IEM genes is significantly better using biochemical rather than clinical profiles. The findings herein provide a motivating goal for the IEM community to expand the computationally accessible descriptions of biochemical phenotypes associated with IEM in phenomics resources.

Design of an expert system based on neuro-fuzzy inference analyzer for on-line microstructural characterization using magnetic NDT method

International Nuclear Information System (INIS)

Ghanei, S.; Vafaeenezhad, H.; Kashefi, M.; Eivani, A.R.; Mazinani, M.

2015-01-01

Tracing microstructural evolution has a significant importance and priority in manufacturing lines of dual-phase steels. In this paper, an artificial intelligence method is presented for on-line microstructural characterization of dual-phase steels. A new method for microstructure characterization based on the theory of magnetic Barkhausen noise nondestructive testing method is introduced using adaptive neuro-fuzzy inference system (ANFIS). In order to predict the accurate martensite volume fraction of dual-phase steels while eliminating the effect and interference of frequency on the magnetic Barkhausen noise outputs, the magnetic responses were fed into the ANFIS structure in terms of position, height and width of the Barkhausen profiles. The results showed that ANFIS approach has the potential to detect and characterize microstructural evolution while the considerable effect of the frequency on magnetic outputs is overlooked. In fact implementing multiple outputs simultaneously enables ANFIS to approach to the accurate results using only height, position and width of the magnetic Barkhausen noise peaks without knowing the value of the used frequency. - Highlights: • New NDT system for microstructural evaluation based on MBN using ANFIS modeling. • Sensitivity of magnetic Barkhausen noise to microstructure changes of the DP steels. • Accurate prediction of martensite by feeding multiple MBN outputs simultaneously. • Obtaining the modeled output without knowing the amount of the used frequency

Design of an expert system based on neuro-fuzzy inference analyzer for on-line microstructural characterization using magnetic NDT method

Energy Technology Data Exchange (ETDEWEB)

Ghanei, S., E-mail: Sadegh.Ghanei@yahoo.com [Department of Materials Engineering, Faculty of Engineering, Ferdowsi University of Mashhad, Azadi Square, Mashhad (Iran, Islamic Republic of); Vafaeenezhad, H. [Centre of Excellence for High Strength Alloys Technology (CEHSAT), School of Metallurgical and Materials Engineering, Iran University of Science and Technology (IUST), Narmak, Tehran (Iran, Islamic Republic of); Kashefi, M. [Department of Materials Engineering, Faculty of Engineering, Ferdowsi University of Mashhad, Azadi Square, Mashhad (Iran, Islamic Republic of); Eivani, A.R. [Centre of Excellence for High Strength Alloys Technology (CEHSAT), School of Metallurgical and Materials Engineering, Iran University of Science and Technology (IUST), Narmak, Tehran (Iran, Islamic Republic of); Mazinani, M. [Department of Materials Engineering, Faculty of Engineering, Ferdowsi University of Mashhad, Azadi Square, Mashhad (Iran, Islamic Republic of)

2015-04-01

Tracing microstructural evolution has a significant importance and priority in manufacturing lines of dual-phase steels. In this paper, an artificial intelligence method is presented for on-line microstructural characterization of dual-phase steels. A new method for microstructure characterization based on the theory of magnetic Barkhausen noise nondestructive testing method is introduced using adaptive neuro-fuzzy inference system (ANFIS). In order to predict the accurate martensite volume fraction of dual-phase steels while eliminating the effect and interference of frequency on the magnetic Barkhausen noise outputs, the magnetic responses were fed into the ANFIS structure in terms of position, height and width of the Barkhausen profiles. The results showed that ANFIS approach has the potential to detect and characterize microstructural evolution while the considerable effect of the frequency on magnetic outputs is overlooked. In fact implementing multiple outputs simultaneously enables ANFIS to approach to the accurate results using only height, position and width of the magnetic Barkhausen noise peaks without knowing the value of the used frequency. - Highlights: • New NDT system for microstructural evaluation based on MBN using ANFIS modeling. • Sensitivity of magnetic Barkhausen noise to microstructure changes of the DP steels. • Accurate prediction of martensite by feeding multiple MBN outputs simultaneously. • Obtaining the modeled output without knowing the amount of the used frequency.

Similarity law for Widom lines and coexistence lines.

Science.gov (United States)

Banuti, D T; Raju, M; Ihme, M

2017-05-01

The coexistence line of a fluid separates liquid and gaseous states at subcritical pressures, ending at the critical point. Only recently, it became clear that the supercritical state space can likewise be divided into regions with liquidlike and gaslike properties, separated by an extension to the coexistence line. This crossover line is commonly referred to as the Widom line, and is characterized by large changes in density or enthalpy, manifesting as maxima in the thermodynamic response functions. Thus, a reliable representation of the coexistence line and the Widom line is important for sub- and supercritical applications that depend on an accurate prediction of fluid properties. While it is known for subcritical pressures that nondimensionalization with the respective species critical pressures p_{cr} and temperatures T_{cr} only collapses coexistence line data for simple fluids, this approach is used for Widom lines of all fluids. However, we show here that the Widom line does not adhere to the corresponding states principle, but instead to the extended corresponding states principle. We resolve this problem in two steps. First, we propose a Widom line functional based on the Clapeyron equation and derive an analytical, species specific expression for the only parameter from the Soave-Redlich-Kwong equation of state. This parameter is a function of the acentric factor ω and compares well with experimental data. Second, we introduce the scaled reduced pressure p_{r}^{*} to replace the previously used reduced pressure p_{r}=p/p_{cr}. We show that p_{r}^{*} is a function of the acentric factor only and can thus be readily determined from fluid property tables. It collapses both subcritical coexistence line and supercritical Widom line data over a wide range of species with acentric factors ranging from -0.38 (helium) to 0.34 (water), including alkanes up to n-hexane. By using p_{r}^{*}, the extended corresponding states principle can be applied within

Characterization and optimization of the R A-6 s on-line neutron radiography facility

International Nuclear Information System (INIS)

Mezio Guanes, Federico Andres

2007-01-01

With the objective of characterizing and optimizing the radiation-field filters behavior in the beam of the R A-6 on-line Neutron Radiography facility, some improvements have been done to the facility devices.We studied the camera sensibility, the best camera acquisition software configuration, the best depth of field, we increased the system tuning efficiency.We also studied the linearity of the facility vs the reactor core neutron fluence and finally we constructed a device that ensure the repeatability of the measurements.The main parameters chosen to represent the best radiation-field set-up are the thermal neutron flux and dose in the position of the camera.Finally, a camera shield optimization haven been done in function of its position [es

Characterization of cetuximab F{sub c/2} dimers by off-line CZE-MS

Energy Technology Data Exchange (ETDEWEB)

François, Yannis-Nicolas, E-mail: yfrancois@unistra.fr [Laboratoire de Spectrométrie de Masse des Interactions et des Systèmes (LSMIS), UDS-CNRS UMR 7140, Université de Strasbourg, Strasbourg (France); Biacchi, Michael; Said, Nassur; Renard, Charly [Laboratoire de Spectrométrie de Masse des Interactions et des Systèmes (LSMIS), UDS-CNRS UMR 7140, Université de Strasbourg, Strasbourg (France); Beck, Alain [Centre d' immunologie Pierre Fabre, Saint-Julien-en-Genevois (France); Gahoual, Rabah [Laboratoire de Spectrométrie de Masse des Interactions et des Systèmes (LSMIS), UDS-CNRS UMR 7140, Université de Strasbourg, Strasbourg (France); Laboratory of Bioanalytical Chemistry and Spectroscopy, Vrije Universiteit, Amsterdam (Netherlands); Leize-Wagner, Emmanuelle [Laboratoire de Spectrométrie de Masse des Interactions et des Systèmes (LSMIS), UDS-CNRS UMR 7140, Université de Strasbourg, Strasbourg (France)

2016-02-18

Monoclonal antibody (mAb) therapeutics attract the largest concern due to their strong therapeutic potency and specificity. The Fc region of mAbs is common to many new biotherapeutics as biosimilar, antibody drug conjugate or fusion protein. Fc region has consequences for Fc-mediated effector functions that might be desirable for therapeutic applications. As a consequence, there is a continuous need for improvement of analytical methods to enable fast and accurate characterization of biotherapeutics. Capillary zone electrophoresis-Mass spectrometry couplings (CZE-MS) appear really attractive methods for the characterization of biological samples. In this report, we used CZE-MS systems developed in house and native MS infusion to allow precise middle-up characterization of F{sub c/2} variant of cetuximab. Molecular weights were measured for three F{sub c/2} charge variants detected in the CZE separation of cetuximab subunits. Two F{sub c/2} C-terminal lysine variants were identified and separated. As the aim is to understand the presence of three peaks in the CZE separation for two F{sub c/2} subunits, we developed a strategy using CZE-UV/MALDI-MS and CZE-UV/ESI-MS to evaluate the role of N-glycosylation and C-terminal lysine truncation on the CZE separation. The chemical structure of N-glycosylation expressed on the Fc region of cetuximab does not influence CZE separation while C-terminal lysine is significantly influencing separation. In addition, native MS infusion demonstrated the characterization of F{sub c/2} dimers at pH 5.7 and 6.8 and the first separation of these dimers using CZE-MS. - Highlights: • We confirm the power of off-line CE/MS coupling for the separation of mAb isoforms. • We demonstrate the role of C-terminal lysine truncation in the separation process. • We demonstrate the presence of non-covalent interactions between Fc/2 subunit. • We identify Fc/2 homo- and heterodimers.

Biophysical and biochemical constraints imposed by salt stress:Learning from halophyte

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Bernardo eDuarte

2014-12-01

Full Text Available Soil salinization is one of the most important factors impacting plant productivity. About 3.6 billion of the world’s 5.2 billion ha of agricultural dryland have already suffered erosion, degradation and salinization. Halophytes typically are considered as plants able to complete their life cycle in environments where the salt concentration is 200 mM NaCl or higher. Different strategies are known to overcome salt stress, as adaptation mechanisms from this type of plants. Salinity adjustment is a complex phenomenon characterized by both biochemical and biophysical adaptations. As photosynthesis is a prerequisite for biomass production, halophytes adapted their electronic transduction pathways and the entire energetic metabolism to overcome the salt excess. The maintenance of ionic homeostasis is in the basis of all cellular stress in particular in terms of redox potential and energy transduction. In the present work the biophysical mechanisms underlying energy capture and transduction in halophytes are discussed alongside with their relation to biochemical mechanisms, integrating data from photosystem light harvesting complexes, electronic transport chains to the quinone pools, carbon harvesting and energy dissipation metabolism.

Biochemical characterization of sap (latex) of a few Indian mango varieties.

Science.gov (United States)

John, K Saby; Bhat, S G; Prasada Rao, U J S

2003-01-01

Mango sap (latex) from four Indian varieties was studied for its composition. Sap was separated into non-aqueous and aqueous phases. Earlier, we reported that the non-aqueous phase contained mainly mono-terpenes having raw mango aroma (Phytochemistry 52 (1999) 891). In the present study biochemical composition of the aqueous phase was studied. Aqueous phase contained little amount of protein (2.0-3.5 mg/ml) but showed high polyphenol oxidase (147-214 U/mg protein) and peroxidase (401-561 U/mg protein) activities. It contained low amounts of polyphenols and protease activities. On native PAGE, all the major protein bands exhibited both polyphenol oxidase and peroxidase activities. Both polyphenol oxidase and peroxidase activities were found to be stable in the aqueous phase of sap at 4 degrees C. Sap contained large amount of non-dialyzable and non-starchy carbohydrate (260-343 mg/ml sap) which may be responsible for maintaining a considerable pressure of fluid in the ducts. Thus, the mango sap could be a valuable by-product in the mango industry as it contains some of the valuable enzymes and aroma components.

Biochemical and morphological characterization of light and heavy sarcoplasmic reticulum vesicles

Energy Technology Data Exchange (ETDEWEB)

Campbell, K.P.

1978-01-01

Light and heavy sarcoplasmic reticulum vesicles isolated from rabbit leg muscle have been used in a study of chloride-induced calcium release. The biochemical and morphological data indicate that light sarcoplasmic reticulum vesicles are derived from the longitudinal reticulum and heavy sarcoplasmic reticulum vesicles are derived from the terminal cisternae of the sarcoplasmic reticulum. The light and heavy sarcoplasmic reticulum vesicles were both able to accumulate calcium in the presence of ATP to amounts greater than 100 nmoles Ca/sup + +/ per mg of protein in less than one minute. Light and heavy sarcoplasmic reticulum vesicles each had a biphasic time course of calcium uptake. The initial uptake was followed by a rapid release after approximately one minute, of 30 to 40% of the accumulated calcium, which was then followed by a slower phase of calcium accumulation. Results indicate that the chloride induced release of calcium may be acting by two mechanisms, osmotic swelling and depolarization. The release of calcium from the light SR vesicles is probably due to osmotic swelling and the release of calcium from the heavy SR vesicles is probably due to depolarization.

Functional somatostatin receptors on a rat pancreatic acinar cell line

International Nuclear Information System (INIS)

Viguerie, N.; Tahiri-Jouti, N.; Esteve, J.P.; Clerc, P.; Logsdon, C.; Svoboda, M.; Susini, C.; Vaysse, N.; Ribet, A.

1988-01-01

Somatostatin receptors from a rat pancreatic acinar cell line, AR4-2J, were characterized biochemically, structurally, and functionally. Binding of 125 I-[Tyr 11 ]Somatostatin to AR4-2J cells was saturable, exhibiting a single class of high-affinity binding sites with a maximal binding capacity of 258 ± 20 fmol/10 6 cells. Somatostatin receptor structure was analyzed by covalently cross-linking 125 I-[Tyr 11 ]somatostatin to its plasma membrane receptors. Gel electrophoresis and autoradiography of cross-linked proteins revealed a peptide containing the somatostatin receptor. Somatostatin inhibited vasoactive intestinal peptide (VIP)-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) formation in a dose-dependent manner. The concentration of somatostatin that caused half-maximal inhibition of cAMP formation was close to the receptor affinity for somatostatin. Pertussis toxin pretreatment of AR4-2J cells prevented somatostatin inhibition of VIP-stimulated cAMP formation as well as somatostatin binding. The authors conclude that AR4-2J cells exhibit functional somatostatin receptors that retain both specificity and affinity of the pancreatic acinar cell somatostatin receptors and act via the pertussis toxin-sensitive guanine nucleotide-binding protein N i to inhibit adenylate cyclase

MEMS-based transmission lines for microwave applications

Science.gov (United States)

Wu, Qun; Fu, Jiahui; Gu, Xuemai; Shi, Huajuan; Lee, Jongchul

2003-04-01

This paper mainly presents a briefly review for recent progress in MEMS-based transmission lines for use in microwave and millimeterwave range. MEMS-based transmission lines including different transmission line structure such as membrane-supported microstrip line microstrip line, coplanar microshield transmission line, LIGA micromachined planar transmission line, micromachined waveguides and coplanar waveguide are discussed. MEMS-based transmission lines are characterized by low propagation loss, wide operation frequency band, low dispersion and high quality factor, in addition, the fabrication is compatible with traditional processing of integrated circuits (IC"s). The emergence of MEMS-based transmission lines provided a solution for miniaturizing microwave system and monolithic microwave integrated circuits.

Acetylcholine receptors and cholinergic ligands: biochemical and genetic aspects in Torpedo californica and Drosophila melanogaster

International Nuclear Information System (INIS)

Rosenthal, L.S.

1987-01-01

This study evaluates the biochemical and genetic aspects of the acetylcholine receptor proteins and cholinergic ligands in Drosophila melanogaster and Torpedo californica. Included are (1) a comparative study of nicotinic ligand-induced cation release from acetylcholine receptors isolated from Torpedo californica and from Drosophila melanogaster, (2) solution studies of the cholinergic ligands, nikethamide and ethamivan, aimed at measuring internal molecular rotational barriers in solvents of different polarity; and (3) the isolation and characterization of the gene(s) for the acetylcholine receptor in Drosophila melasogaster. Acetylcholine receptor proteins isolated from Drosphila melanogaster heads were found to behave kinetically similar (with regards to cholinergic ligand-induced 155 Eu: 3+ displacement from prelabeled proteins) to receptor proteins isolated from Torpedo californica electric tissue, providing additional biochemical evidence for the existence of a Drosophila acetylcholine receptor

Biochemical and genetic analyses of the oomycete Pythium insidiosum provide new insights into clinical identification and urease-based evolution of metabolism-related traits

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Theerapong Krajaejun

2018-06-01

Full Text Available The oomycete microorganism, Pythium insidiosum, causes the life-threatening infectious condition, pythiosis, in humans and animals worldwide. Affected individuals typically endure surgical removal of the infected organ(s. Detection of P. insidiosum by the established microbiological, immunological, or molecular methods is not feasible in non-reference laboratories, resulting in delayed diagnosis. Biochemical assays have been used to characterize P. insidiosum, some of which could aid in the clinical identification of this organism. Although hydrolysis of maltose and sucrose has been proposed as the key biochemical feature useful in discriminating P. insidiosum from other oomycetes and fungi, this technique requires a more rigorous evaluation involving a wider selection of P. insidiosum strains. Here, we evaluated 10 routinely available biochemical assays for characterization of 26 P. insidiosum strains, isolated from different hosts and geographic origins. Initial assessment revealed diverse biochemical characteristics across the P. insidiosum strains tested. Failure to hydrolyze sugars is observed, especially in slow-growing strains. Because hydrolysis of maltose and sucrose varied among different strains, use of the biochemical assays for identification of P. insidiosum should be cautioned. The ability of P. insidiosum to hydrolyze urea is our focus, because this metabolic process relies on the enzyme urease, an important virulence factor of other pathogens. The ability to hydrolyze urea varied among P. insidiosum strains and was not associated with growth rates. Genome analyses demonstrated that urease- and urease accessory protein-encoding genes are present in both urea-hydrolyzing and non-urea-hydrolyzing strains of P. insidiosum. Urease genes are phylogenetically conserved in P. insidiosum and related oomycetes, while the presence of urease accessory protein-encoding genes is markedly diverse in these organisms. In summary, we dissected

Biochemical and nutritional characterization of coconut (Cocos nucifera L.) haustorium.

Science.gov (United States)

Manivannan, Arivalagan; Bhardwaj, Rakesh; Padmanabhan, Sugatha; Suneja, Poonam; Hebbar, K B; Kanade, Santosh R

2018-01-01

Study was conducted to determine the biochemical constituents in coconut (Cocos nucifera L.) haustorium, a spongy tissue formed during coconut germination. Results indicated that 100g of dried coconut haustorium contained 1.05±0.2% ash, 44.2±4.6% soluble sugar, 24.5±3.2% starch, 5.50±0.3% protein, 1.99±0.9% fat, 5.72±0.4% soluble dietary fibre, 20.3±1.9% insoluble dietary fibre, and 146±14.3mg phenolics. Mineral profiling showed that it contained 145±8.6, 104±9.6, 33.9±8.2, 30.9±1.9, 9.45±2.1, 0.292±0.1, 2.53±0.2 and 1.20±0.1mg of K, Mg, Ca, P, Mn, Cu, Fe and Zn, respectively. Antioxidant activity assay indicated that 100g haustorium was equivalent to 1918±173, 170±20.4, 72.8±14.7 and 860±116mg of Trolox as measured by CUPRAC, FRAP, DPPH and ABTS, respectively. Amino acid score indicated that methionine+cysteine (57.6%), phenylalanine+tyrosine (32.6%), leucine (45.7%) and isoleucine (68%) are found less in haustorium. Further studies needed in developing nutritionally balanced formulations using coconut haustorium, which will be useful for lactose intolerant children. Copyright © 2016 Elsevier Ltd. All rights reserved.

Conventional and molecular cytogenetics of human non-medullary thyroid carcinoma: characterization of eight cell line models and review of the literature on clinical samples

International Nuclear Information System (INIS)

Ribeiro, Franclim Ricardo; Meireles, Ana Margarida; Rocha, Ana Sofia; Teixeira, Manuel Rodrigues

2008-01-01

Cell lines are often poorly characterized from a genetic point of view, reducing their usefulness as tumor models. Our purpose was to assess the genetic background of eight commonly used human thyroid carcinoma models and to compare the findings with those reported for primary tumors of the gland. We used chromosome banding analysis and comparative genomic hybridization to profile eight non-medullary thyroid carcinoma cell lines of papillary (TPC-1, FB2, K1 and B-CPAP), follicular (XTC-1) or anaplastic origin (8505C, C643 and HTH74). To assess the representativeness of the findings, we additionally performed a thorough review of cytogenetic (n = 125) and DNA copy number information (n = 270) available in the literature on clinical samples of thyroid carcinoma. The detailed characterization of chromosomal markers specific for each cell line revealed two cases of mistaken identities: FB2 was shown to derive from TPC-1 cells, whereas K1 cells have their origin in cell line GLAG-66. All cellular models displayed genomic aberrations of varying complexity, and recurrent gains at 5p, 5q, 8q, and 20q (6/7 cell lines) and losses at 8p, 13q, 18q, and Xp (4/7 cell lines) were seen. Importantly, the genomic profiles were compatible with those of the respective primary tumors, as seen in the meta-analysis of the existing literature data. We provide the genomic background of seven independent thyroid carcinoma models representative of the clinical tumors of the corresponding histotypes, and highlight regions of recurrent aberrations that may guide future studies aimed at identifying target genes. Our findings further support the importance of routinely performing cytogenetic studies on cell lines, to detect cross-contamination mishaps such as those identified here

Revisiting spatial distribution and biochemical composition of calcium-containing crystals in human osteoarthritic articular cartilage.

OpenAIRE

Nguyen, C.; Bazin, D.; Daudon, M.; Chatron-Colliet, A.; Hannouche, D.; Bianchi, A.; Côme, D.; So, A.; Busso, N.; Lioté, F.; Ea, H.K.

2013-01-01

International audience; INTRODUCTION: Calcium-containing (CaC) crystals, including basic calcium phosphate (BCP) and calcium pyrophosphate dihydrate (CPP), are associated with destructive forms of osteoarthritis (OA). We assessed their distribution and biochemical and morphologic features in human knee OA cartilage. METHODS: We prospectively included 20 patients who underwent total knee replacement (TKR) for primary OA. CaC crystal characterization and identification involved Fourier-transfor...

Characterizing Exoplanet Atmospheres : A Complete Line List for Phosphine

Science.gov (United States)

Sousa-Silva, C.; Yurchenko, S. N.; Tennyson, J.

2013-09-01

The ability to characterise the atmospheres of cool stars, brown dwarfs and exoplanets requires fundamental data for all species contributing significantly to their opacity. However, with notable exceptions such as water and ammonia, existing molecular line lists are not sufficiently accurate or complete to allow for a full spectroscopic analysis of these bodies. ExoMol (www.exomol.com [1]) is a project that aims to rectify this by generating comprehensive line lists for all molecules likely to be detected in the atmospheres of cool astrophysical objects in the foreseeable future. The spectral data is generated by employing ab initio quantum mechanical methods, performing empirical refinement based on experimental spectroscopic data and harnessing high performance computing. Here we present our work on phosphine, (PH3), an equilateral pyramidal molecule (the phosphorus analogue to ammonia). Phosphine is known to be important for the atmospheres of giant-planets, cool stars and many other astronomical bodies. Rotational transition features of phosphine have been found in the far- infrared spectra of Saturn and Jupiter [2, 3], where it is a marker for vertical convection zones. A computed room temperature line list of phosphine is presented here [4], illustrated in the accompanying figure 1. This line list is a precursor to a high temperature equivalent to be produced in the near future, necessary for the analysis of cool stars and brown dwarfs. All the transitions' energy levels and Einstein A-coefficients were computed using the program TROVE [5].

Establishment and partial characterization of a cell line from burbot Lota lota maculosa: susceptibility to IHNV, IPNV and VHSV.

Science.gov (United States)

Polinski, Mark P; Drennan, John D; Batts, William N; Ireland, Susan C; Cain, Kenneth D

2010-05-18

This study describes the development and partial characterization of a continuous fibroblastic-like cell line (BEF-1) developed from late stage embryos of North American burbot Lota lota maculosa. This cell line has been maintained for over 5 yr and 100 passages in vitro. Cells were cultured using Eagle's minimum essential medium with Earle's salts (MEM) supplemented with GlutaMAX, and 10% fetal bovine serum (FBS), pH 7.4. The addition of penicillin-streptomycin-neomycin (PSN) antibiotic mixture (0.05, 0.05, 0.1 mg m(-1), respectively) did not negatively influence cell replication; however, the antimycotic FungizoneTM (2.5 microg m(-1), amphotericin B) caused cell rounding and resulted in a severe decrease in cell proliferation. Optimal incubation temperature has been observed between 15 and 23 degrees C, and at these temperatures cultures are routinely passed using standard trypsinization methods every 5 to 7 d at a split ratio of 1:3 or 1:4. The cell line was susceptible to isolates of the M and U North American genotypes of infectious hematopoietic necrosis virus (IHNV), and to isolates of genotypes I, IVa, and IVb of viral hemorrhagic septicemia virus (VHSV). In contrast, the cell line was refractory to infection by 2 North American isolates of infectious pancreatic necrosis virus (IPNV) from serotypes A1 and A9. This cell line provides a new laboratory tool, will allow further investigation into viral diseases of burbot and possibly other species, and is the first immortalized cell line reported from a species in the Gadidae (cod) family.

Establishment and partial characterization of a cell line from burbot Lota lota maculosa: susceptibility to IHNV, IPNV and VHSV.

Science.gov (United States)

Batts, William N.; Polinski, Mark P.; Drennan, John D.; Ireland, Susan C.; Cain, Kenneth D.

2010-01-01

This study describes the development and partial characterization of a continuous fibroblastic-like cell line (BEF-1) developed from late stage embryos of North American burbot Lota lota maculosa. This cell line has been maintained for over 5 yr and 100 passages in vitro. Cells were cultured using Eagle’s minimum essential medium with Earle’s salts (MEM) supplemented with GlutaMAX™, and 10% fetal bovine serum (FBS), pH 7.4. The addition of penicillin-streptomycin-neomycin (PSN) antibiotic mixture (0.05, 0.05, 0.1 mg ml–1, respectively) did not negatively influence cell replication; however, the antimycotic Fungizone™ (2.5 µg ml–1, amphotericin B) caused cell rounding and resulted in a severe decrease in cell proliferation. Optimal incubation temperature has been observed between 15 and 23°C, and at these temperatures cultures are routinely passed using standard trypsinization methods every 5 to 7 d at a split ratio of 1:3 or 1:4. The cell line was susceptible to isolates of the M and U North American genotypes of infectious hematopoietic necrosis virus (IHNV), and to isolates of genotypes I, IVa, and IVb of viral hemorrhagic septicemia virus (VHSV). In contrast, the cell line was refractory to infection by 2 North American isolates of infectious pancreatic necrosis virus (IPNV) from serotypes A1 and A9. This cell line provides a new laboratory tool, will allow further investigation into viral diseases of burbot and possibly other species, and is the first immortalized cell line reported from a species in the Gadidae (cod) family.

Interactions of Cannabinoids With Biochemical Substrates

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Brian F Thomas

2017-05-01

Full Text Available Recent decades have seen much progress in the identification and characterization of cannabinoid receptors and the elucidation of the mechanisms by which derivatives of the Cannabis sativa plant bind to receptors and produce their physiological and psychological effects. The information generated in this process has enabled better understanding of the fundamental physiological and psychological processes controlled by the central and peripheral nervous systems and has fostered the development of natural and synthetic cannabinoids as therapeutic agents. A negative aspect of this decades-long effort is the proliferation of clandestinely synthesized analogs as recreational street drugs with dangerous effects. Currently, the interactions of cannabinoids with their biochemical substrates are extensively but inadequately understood, and the clinical application of derived and synthetic receptor ligands remains quite limited. The wide anatomical distribution and functional complexity of the cannabinoid system continue to indicate potential for both therapeutic and side effects, which offers challenges and opportunities for medicinal chemists involved in drug discovery and development.

Biochemical Process Development and Integration | Bioenergy | NREL

Science.gov (United States)

Biochemical Process Development and Integration Biochemical Process Development and Integration Our conversion and separation processes to pilot-scale integrated process development and scale up. We also Publications Accounting for all sugar produced during integrated production of ethanol from lignocellulosic

Combination of biochemical and mechanical cues for tendon tissue engineering.

Science.gov (United States)

Testa, Stefano; Costantini, Marco; Fornetti, Ersilia; Bernardini, Sergio; Trombetta, Marcella; Seliktar, Dror; Cannata, Stefano; Rainer, Alberto; Gargioli, Cesare

2017-11-01

Tendinopathies negatively affect the life quality of millions of people in occupational and athletic settings, as well as the general population. Tendon healing is a slow process, often with insufficient results to restore complete endurance and functionality of the tissue. Tissue engineering, using tendon progenitors, artificial matrices and bioreactors for mechanical stimulation, could be an important approach for treating rips, fraying and tissue rupture. In our work, C3H10T1/2 murine fibroblast cell line was exposed to a combination of stimuli: a biochemical stimulus provided by Transforming Growth Factor Beta (TGF-β) and Ascorbic Acid (AA); a three-dimensional environment represented by PEGylated-Fibrinogen (PEG-Fibrinogen) biomimetic matrix; and a mechanical induction exploiting a custom bioreactor applying uniaxial stretching. In vitro analyses by immunofluorescence and mechanical testing revealed that the proposed combined approach favours the organization of a three-dimensional tissue-like structure promoting a remarkable arrangement of the cells and the neo-extracellular matrix, reflecting into enhanced mechanical strength. The proposed method represents a novel approach for tendon tissue engineering, demonstrating how the combined effect of biochemical and mechanical stimuli ameliorates biological and mechanical properties of the artificial tissue compared to those obtained with single inducement. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Studies on maize inbred lines susceptibility to herbicides

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Stefanović Lidija

2010-01-01

Full Text Available This paper presents the analysis of results obtained during long- term studies on the response of maize inbred lines to herbicides. Under the agroecological conditions of Zemun Polje the response (reaction of maize inbred lines to herbicides of different classes was investigated. Biological tests were performed and some agronomic, morphological, physiological and biochemical parameters were determined when the response of maize inbred lines to herbicides was estimated. The use of active ingredients of herbicides from triazine, acetanilide, thiocarbamate to new chemical groups (sulfonylurea etc., have been resulted in changes in weed suppression and susceptibility of inbred lines. Obtained results show that effects of herbicides on susceptible maize genotypes can be different: they can slowdown the growth and development and affect the plant height; they can also affect the stages of the tassel and ear development and at the end they can reduced grain yield of the tested inbreds. Numerous studies confirmed the existence of differences in susceptibility level of maize genotypes in relation to herbicides. According to gained results the recommendations for growers are made on the possibility of the application of new herbicides in the hybrid seed production.

Characterization and classification of one new cytoplasmic male sterility (CMS) line based on morphological, cytological and molecular markers in non-heading Chinese cabbage (Brassica rapa L.).

Science.gov (United States)

Heng, Shuangping; Shi, Dianyi; Hu, Zhenhua; Huang, Tao; Li, Jinping; Liu, Liyan; Xia, Chunxiu; Yuan, Zhenzhen; Xu, Yuejin; Fu, Tingdong; Wan, Zhengjie

2015-09-01

A new non-heading Chinese cabbage CMS line M119A was characterized and specific molecular markers were developed to classify different CMS types. One new non-heading Chinese cabbage (Brassica rapa L.) cytoplasmic male sterile (CMS) line M119A was obtained by interspecific crosses between the recently discovered hau CMS line of Brassica juncea and B. rapa. Furthermore, the line was characterized and compared with other five isonuclear-alloplasmic CMS lines. The M119A line produced six stamens without pollen and only two stamen fused together in fewer flowers. Tissue section indicated that anther abortion in M119A may have occurred during differentiation of the archesporial cells without pollen sac. All the six CMS lines were grouped into three types based on the presence of three PCR fragments of 825, 465 and 772 bp amplified with different mitochondrial genes specific primers. The 825-bp fragment was amplified both in 09-10A and H201A using the specific primer pair P-orf224-atp6, and showed 100 % identity with the mitochondrial gene of pol CMS. The 465-bp fragment was amplified in 30A and 105A using the primer pair P-orf138 and shared 100 % identity with the mitochondrial gene of ogu CMS. The 772-bp fragment was amplified in M119A and H203A using the primer pair P-orf288 and showed 100 % identity with the mitochondrial gene of hau CMS. Therefore, these markers could efficiently distinguish different types of isonuclear-alloplasmic CMS lines of non-heading Chinese cabbage, which were useful for improving the efficiency of cross-breeding and heterosis utilization in cruciferous vegetables.

Generation and Characterization of Vascular Smooth Muscle Cell Lines Derived from a Patient with a Bicuspid Aortic Valve

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Pamela Lazar-Karsten

2016-04-01

Full Text Available Thoracic aortic dilation is the most common malformation of the proximal aorta and is responsible for 1%–2% of all deaths in industrialized countries. In approximately 50% of patients with a bicuspid aortic valve (BAV, dilation of any or all segments of the aorta occurs. BAV patients with aortic dilation show an increased incidence of cultured vascular smooth muscle cell (VSMC loss. In this study, VSMC, isolated from the ascending aorta of BAV, was treated with Simian virus 40 to generate a BAV-originated VSMC cell line. To exclude any genomic DNA or cross-contamination, highly polymorphic short tandem repeats of the cells were profiled. The cells were then characterized using flow cytometry and karyotyping. The WG-59 cell line created is the first reported VSMC cell line isolated from a BAV patient. Using an RT2 Profiler PCR Array, genes within the TGFβ/BMP family that are dependent on losartan treatment were identified. Endoglin was found to be among the regulated genes and was downregulated in WG-59 cells following treatment with different losartan concentrations, when compared to untreated WG-59 cells.

Biochemical characterization of a new type of intracellular PHB depolymerase from Rhodospirillum rubrum with high hydrolytic activity on native PHB granules.

Science.gov (United States)

Sznajder, Anna; Jendrossek, Dieter

2011-03-01

A Rhodospirillum rubrum gene that is predicted to code for an extracellular poly(3-hydroxybutyrate) (PHB) depolymerase by the recently published polyhydroxyalkanoates (PHA) depolymerase engineering database was cloned. The gene product (PhaZ3( Rru )) was expressed in recombinant E. coli, purified and biochemically characterized. PhaZ3( Rru ) turned out, however, to share characteristics of intracellular PHB depolymerases and revealed a combination of properties that have not yet been described for other PHB depolymerases. A fusion of PhaZ3( Rru )with the enhanced cyan fluorescent protein was able to bind to PHB granules in vivo and supported the function as an intracellular PHB depolymerase. Purified PhaZ3( Rru ) was specific for short-chain-length polyhydroxyalkanoates (PHA(SCL)) and hydrolysed both untreated native PHB granules as well as trypsin-activated native PHB granules to a mixture of mono- and dimeric 3-hydroxybutyrate. Crystalline (denatured) PHB granules were not hydrolysed by PhayZ3( Rru ). Low concentrations of calcium or magnesium ions (1-5 mM) reversibly (EDTA) inhibited the enzyme. Our data suggest that PhaZ3( Rru ) is the representative of a new type of the growing number of intracellular PHB depolymerases.

Improved cell line IPEC-J2, characterized as a model for porcine jejunal epithelium.

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Silke S Zakrzewski

Full Text Available Cell lines matching the source epithelium are indispensable for investigating porcine intestinal transport and barrier properties on a subcellular or molecular level and furthermore help to reduce animal usage. The porcine jejunal cell line IPEC-J2 is established as an in vitro model for porcine infection studies but exhibits atypically high transepithelial resistances (TER and only low active transport rates so that the effect of nutritional factors cannot be reliably investigated. This study aimed to properly remodel IPEC-J2 and then to re-characterize these cells regarding epithelial architecture, expression of barrier-relevant tight junction (TJ proteins, adequate TER and transport function, and reaction to secretagogues. For this, IPEC-J2 monolayers were cultured on permeable supports, either under conventional (fetal bovine serum, FBS or species-specific (porcine serum, PS conditions. Porcine jejunal mucosa was analyzed for comparison. Main results were that under PS conditions (IPEC-J2/PS, compared to conventional FBS culture (IPEC-J2/FBS, the cell height increased 6-fold while the cell diameter was reduced by 50%. The apical cell membrane of IPEC-J2/PS exhibited typical microvilli. Most importantly, PS caused a one order of magnitude reduction of TER and of trans- and paracellular resistance, and a 2-fold increase in secretory response to forskolin when compared to FBS condition. TJ ultrastructure and appearance of TJ proteins changed dramatically in IPEC-J2/PS. Most parameters measured under PS conditions were much closer to those of typical pig jejunocytes than ever reported since the cell line's initial establishment in 1989. In conclusion, IPEC-J2, if cultured under defined species-specific conditions, forms a suitable model for investigating porcine paracellular intestinal barrier function.

Immunological and biochemical characterization of extracellular polysaccharides of mucoralean moulds

NARCIS (Netherlands)

Ruiter, de G.A.

1993-01-01

In this thesis the characterization is described of the antigenic determinants (epitopes) of the extracellular polysaccharides (EPSs) from moulds belonging to the order of Mucorales. Detailed knowledge of the structure of these epitopes allows for further development of a new generation of

In vitro characterization and inhibition of the CXCR4/CXCL12 chemokine axis in human uveal melanoma cell lines

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Antecka Emilia

2007-11-01

Full Text Available Abstract Purpose The CXCR4/CXCL12 chemokine axis may play a critical role in guiding CXCR4+ circulating malignant cells to organ specific locations that actively secrete its ligand CXCL12 (SDF-1 such as bone, brain, liver, and lungs. We sought to characterize the presence of the CXCR4/CXCL12 axis in five uveal melanoma (UM cell lines in vitro. The ability of TN14003, a synthetic peptide inhibitor that targets the CXCR4 receptor complex, to inhibit this axis was also assessed. Methods Immunocytochemistry was performed against CXCR4 to confirm expression of this chemokine receptor in all five UM cell lines. Flow cytometry was preformed to evaluate CXCR4 cell surface expression on all five UM cell lines. A proliferation assay was also used to test effects TN14003 would have on cellular proliferation. Inhibition of cellular migration by specifically inhibiting the CXCR4/CXCL12 axis with TN14003 was also investigated. The binding efficacy of TN14003 to the CXCR4 receptor was assessed through flow cytometric methods. Results The CXCR4 receptor was present on all five UM cell lines. All five cell lines expressed different relative levels of surface CXCR4. TN14003 did not affect the proliferation of the five cell lines (p > 0.05. All cell lines migrated towards the chemokine CXCL12 at a level greater than the negative control (p Conclusion Interfering with the CXCR4/CXCL12 axis, using TN14003 was shown to effectively down regulate UM cell migration in vitro. Knowing that UM expresses the CXCR4 receptor, these CXCR4+ cells may be less likely to colonize distant organs that secrete the CXCL12 ligand, if treated with an inhibitor that binds CXCR4. Further studies should be pursued in order to test TN14003 efficacy in vivo.

Self-organizing ontology of biochemically relevant small molecules.

Science.gov (United States)

Chepelev, Leonid L; Hastings, Janna; Ennis, Marcus; Steinbeck, Christoph; Dumontier, Michel

2012-01-06

The advent of high-throughput experimentation in biochemistry has led to the generation of vast amounts of chemical data, necessitating the development of novel analysis, characterization, and cataloguing techniques and tools. Recently, a movement to publically release such data has advanced biochemical structure-activity relationship research, while providing new challenges, the biggest being the curation, annotation, and classification of this information to facilitate useful biochemical pattern analysis. Unfortunately, the human resources currently employed by the organizations supporting these efforts (e.g. ChEBI) are expanding linearly, while new useful scientific information is being released in a seemingly exponential fashion. Compounding this, currently existing chemical classification and annotation systems are not amenable to automated classification, formal and transparent chemical class definition axiomatization, facile class redefinition, or novel class integration, thus further limiting chemical ontology growth by necessitating human involvement in curation. Clearly, there is a need for the automation of this process, especially for novel chemical entities of biological interest. To address this, we present a formal framework based on Semantic Web technologies for the automatic design of chemical ontology which can be used for automated classification of novel entities. We demonstrate the automatic self-assembly of a structure-based chemical ontology based on 60 MeSH and 40 ChEBI chemical classes. This ontology is then used to classify 200 compounds with an accuracy of 92.7%. We extend these structure-based classes with molecular feature information and demonstrate the utility of our framework for classification of functionally relevant chemicals. Finally, we discuss an iterative approach that we envision for future biochemical ontology development. We conclude that the proposed methodology can ease the burden of chemical data annotators and

Self-organizing ontology of biochemically relevant small molecules

Science.gov (United States)

2012-01-01

Background The advent of high-throughput experimentation in biochemistry has led to the generation of vast amounts of chemical data, necessitating the development of novel analysis, characterization, and cataloguing techniques and tools. Recently, a movement to publically release such data has advanced biochemical structure-activity relationship research, while providing new challenges, the biggest being the curation, annotation, and classification of this information to facilitate useful biochemical pattern analysis. Unfortunately, the human resources currently employed by the organizations supporting these efforts (e.g. ChEBI) are expanding linearly, while new useful scientific information is being released in a seemingly exponential fashion. Compounding this, currently existing chemical classification and annotation systems are not amenable to automated classification, formal and transparent chemical class definition axiomatization, facile class redefinition, or novel class integration, thus further limiting chemical ontology growth by necessitating human involvement in curation. Clearly, there is a need for the automation of this process, especially for novel chemical entities of biological interest. Results To address this, we present a formal framework based on Semantic Web technologies for the automatic design of chemical ontology which can be used for automated classification of novel entities. We demonstrate the automatic self-assembly of a structure-based chemical ontology based on 60 MeSH and 40 ChEBI chemical classes. This ontology is then used to classify 200 compounds with an accuracy of 92.7%. We extend these structure-based classes with molecular feature information and demonstrate the utility of our framework for classification of functionally relevant chemicals. Finally, we discuss an iterative approach that we envision for future biochemical ontology development. Conclusions We conclude that the proposed methodology can ease the burden of

Self-organizing ontology of biochemically relevant small molecules

Directory of Open Access Journals (Sweden)

Chepelev Leonid L

2012-01-01

Full Text Available Abstract Background The advent of high-throughput experimentation in biochemistry has led to the generation of vast amounts of chemical data, necessitating the development of novel analysis, characterization, and cataloguing techniques and tools. Recently, a movement to publically release such data has advanced biochemical structure-activity relationship research, while providing new challenges, the biggest being the curation, annotation, and classification of this information to facilitate useful biochemical pattern analysis. Unfortunately, the human resources currently employed by the organizations supporting these efforts (e.g. ChEBI are expanding linearly, while new useful scientific information is being released in a seemingly exponential fashion. Compounding this, currently existing chemical classification and annotation systems are not amenable to automated classification, formal and transparent chemical class definition axiomatization, facile class redefinition, or novel class integration, thus further limiting chemical ontology growth by necessitating human involvement in curation. Clearly, there is a need for the automation of this process, especially for novel chemical entities of biological interest. Results To address this, we present a formal framework based on Semantic Web technologies for the automatic design of chemical ontology which can be used for automated classification of novel entities. We demonstrate the automatic self-assembly of a structure-based chemical ontology based on 60 MeSH and 40 ChEBI chemical classes. This ontology is then used to classify 200 compounds with an accuracy of 92.7%. We extend these structure-based classes with molecular feature information and demonstrate the utility of our framework for classification of functionally relevant chemicals. Finally, we discuss an iterative approach that we envision for future biochemical ontology development. Conclusions We conclude that the proposed methodology

Formation and characterization of FeLV iscoms.

NARCIS (Netherlands)

L. Akerblom; K. Strö mstedt; S. Hö glund; A.D.M.E. Osterhaus (Albert); B. Morein (Bror)

1989-01-01

textabstractImmunostimulating complexes (ISCOMs) have been prepared from feline leukaemia virus (FeLV) envelope proteins. The ISCOMs were characterized biochemically in SDS-polyacrylamide gel electrophoresis showing the presence of proteins of estimated molecular weights of 15,000, 27,000 and

Biochemical basis for the action of radioprotective drugs

International Nuclear Information System (INIS)

Romantsev, E.F.; Blokhina, V.D.; Zhulanova, Z.I.; Koshcheenko, N.N.; Filippovich, I.V.

1977-01-01

The hypothesis of complex biochemical mechanism of action of radioprotective drugs is described. Shortly after injection of radioprotective aminothiols into animals the inhibition of radiosensitive biochemical processes: DNA and RNA synthesis, protein synthesis and oxidative phosphorylation has been observed. The molecular mechanism of these phenomena consists of radioprotectors ability to form adsorption, thioester, amide, and disulphide bonds with appropriate enzymes. The curve reflecting the formation and breakdown of mixed disulphides between radioprotectors and proteins coincides well with that reflecting the radioprotective effect dependence on time. The radiobiological significance of molecular interactions observed may be interpreted as the diminution in ''spoiled'' molecules formation (inhibition of replication) and elevation in repartion rate. The inhibition of biochemical processes has the reversible nature and last for short time. The drugs acting according to so-called oxygen effect protect also by means of biochemical mechanisms. The molecular mechanism is mediated through their ability to bind to receptors, and biologically important molecules and macromolecules. As a result the inhibition of radiosensitive processes occurs, the ''spoiled'' molecules number is diminished and reparation takes place more easily. The idea on the complex biochemical mechanism of action of radioprotectors correlates with the proposal on complex biochemical mechanism responsible for interphase death occured after irradiation

Genome-wide characterization of genetic variants and putative regions under selection in meat and egg-type chicken lines.

Science.gov (United States)

Boschiero, Clarissa; Moreira, Gabriel Costa Monteiro; Gheyas, Almas Ara; Godoy, Thaís Fernanda; Gasparin, Gustavo; Mariani, Pilar Drummond Sampaio Corrêa; Paduan, Marcela; Cesar, Aline Silva Mello; Ledur, Mônica Corrêa; Coutinho, Luiz Lehmann

2018-01-25

Meat and egg-type chickens have been selected for several generations for different traits. Artificial and natural selection for different phenotypes can change frequency of genetic variants, leaving particular genomic footprints throghtout the genome. Thus, the aims of this study were to sequence 28 chickens from two Brazilian lines (meat and white egg-type) and use this information to characterize genome-wide genetic variations, identify putative regions under selection using Fst method, and find putative pathways under selection. A total of 13.93 million SNPs and 1.36 million INDELs were identified, with more variants detected from the broiler (meat-type) line. Although most were located in non-coding regions, we identified 7255 intolerant non-synonymous SNPs, 512 stopgain/loss SNPs, 1381 frameshift and 1094 non-frameshift INDELs that may alter protein functions. Genes harboring intolerant non-synonymous SNPs affected metabolic pathways related mainly to reproduction and endocrine systems in the white-egg layer line, and lipid metabolism and metabolic diseases in the broiler line. Fst analysis in sliding windows, using SNPs and INDELs separately, identified over 300 putative regions of selection overlapping with more than 250 genes. For the first time in chicken, INDEL variants were considered for selection signature analysis, showing high level of correlation in results between SNP and INDEL data. The putative regions of selection signatures revealed interesting candidate genes and pathways related to important phenotypic traits in chicken, such as lipid metabolism, growth, reproduction, and cardiac development. In this study, Fst method was applied to identify high confidence putative regions under selection, providing novel insights into selection footprints that can help elucidate the functional mechanisms underlying different phenotypic traits relevant to meat and egg-type chicken lines. In addition, we generated a large catalog of line-specific and common

CD147 expression predicts biochemical recurrence after prostatectomy independent of histologic and pathologic features.

Science.gov (United States)

Bauman, Tyler M; Ewald, Jonathan A; Huang, Wei; Ricke, William A

2015-07-25

CD147 is an MMP-inducing protein often implicated in cancer progression. The purpose of this study was to investigate the expression of CD147 in prostate cancer (PCa) progression and the prognostic ability of CD147 in predicting biochemical recurrence after prostatectomy. Plasma membrane-localized CD147 protein expression was quantified in patient samples using immunohistochemistry and multispectral imaging, and expression was compared to clinico-pathological features (pathologic stage, Gleason score, tumor volume, preoperative PSA, lymph node status, surgical margins, biochemical recurrence status). CD147 specificity and expression were confirmed with immunoblotting of prostate cell lines, and CD147 mRNA expression was evaluated in public expression microarray datasets of patient prostate tumors. Expression of CD147 protein was significantly decreased in localized tumors (pT2; p = 0.02) and aggressive PCa (≥pT3; p = 0.004), and metastases (p = 0.001) compared to benign prostatic tissue. Decreased CD147 was associated with advanced pathologic stage (p = 0.009) and high Gleason score (p = 0.02), and low CD147 expression predicted biochemical recurrence (HR 0.55; 95 % CI 0.31-0.97; p = 0.04) independent of clinico-pathologic features. Immunoblot bands were detected at 44 kDa and 66 kDa, representing non-glycosylated and glycosylated forms of CD147 protein, and CD147 expression was lower in tumorigenic T10 cells than non-tumorigenic BPH-1 cells (p = 0.02). Decreased CD147 mRNA expression was associated with increased Gleason score and pathologic stage in patient tumors but is not associated with recurrence status. Membrane-associated CD147 expression is significantly decreased in PCa compared to non-malignant prostate tissue and is associated with tumor progression, and low CD147 expression predicts biochemical recurrence after prostatectomy independent of pathologic stage, Gleason score, lymph node status, surgical margins, and tumor volume in multivariable

[Signal transduction in plant development: Chemical and biochemical approaches to receptor identification]. Progress report, [May 15, 1993--May 14, 1994

Energy Technology Data Exchange (ETDEWEB)

1994-06-01

Progress is reported on studies concerning NAD(P)H-2,6-DMBQ oxidoreductase of Striga asiatica aimed at elucidating basic biochemical parameters of Striga. Reported studies include characterization of the enzyme, development of Striga molecular genetics, and development of a redox model for germination control.

Characterization and performance of 16 new inbred lines of lettuce

Science.gov (United States)

The Agricultural Research Service, U.S. Department of Agriculture announces the release of sixteen breeding lines of lettuce (Lactuca sativa L.). Five (SM13-I1, SM13-I2, SM13-I3, SM13-I4, and SM13-I5) of the six iceberg breeding lines can be used for whole head or salad blend production; the sixth i...

In vitro radiation and chemotherapy sensitivity of established cell lines of human small cell lung cancer and its large cell morphological variants

International Nuclear Information System (INIS)

Carney, D.N.; Mitchell, J.B.; Kinsella, T.J.

1983-01-01

The in vitro response to radiation and chemotherapeutic drugs of cell lines established from 7 patients with small cell (SC) lung cancer were tested using a soft agarose clonogenic assay. Five cell lines retained the typical morphological and biochemical amine precursor uptake decarboxylation characteristics of SC, while two cell lines had undergone ''transformation'' to large cell (LC) morphological variants with loss of amine precursor uptake decarboxylation cell characteristics of SC. The radiation survival curves for the SC lines were characterized by D0 values ranging from 51 to 140 rads and extrapolation values (n) ranging from 1.0 to 3.3. While the D0 values of the radiation survival curves of the LC variants were similar (91 and 80 rads), the extrapolation values were 5.6 and 11.1 In vitro chemosensitivity testing of the cell lines revealed an excellent correlation between prior treatment status of the patient and in vitro sensitivity or resistance. No correlation was observed between in vitro chemosensitivity and radiation response. These data suggest that transformation of SC to LC with loss of amine precursor uptake and decarboxylation characteristics is associated with a marked increase in radiation resistance (n) in vitro. The observation of a 2- to 5-fold increase in survival of the LC compared to the SC lines following 200 rads suggests that the use of larger daily radiation fractions and/or radiation-sensitizing drugs might lead to a significantly greater clinical response in patients with LC morphology. This clinical approach may have a major impact on patient response and survival

Biochemical reactions of the organism

International Nuclear Information System (INIS)

Fedorova, A.V.

1984-01-01

Effects of mercury, strontium chloride, GMDA, trichlorfon as well as some radionuclides ( 89 Sr, 137 Cs, 203 Hg) were studied on rats. Changes in biochemical parameters (histamine content, activity of cholinesterase and histaminase) are noted. Most noticeable changes were observed in enzymatic activity. Distortion of enzymatic systems and accumulation of intermediate exchange and decay products of tissues in excess quantities affecting other systems can be the reason for changes in the organism. The observed changes in biochemical parameters should be necessarily taken into account at hygienic regulations of harmful effects of enviroment

In Vitro Biochemical Characterization of All Barley Endosperm Starch Synthases

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Jose Antonio Cuesta-Seijo

2016-01-01

Full Text Available Starch is the main storage polysaccharide in cereals and the major source of calories in the human diet. It is synthesized by a panel of enzymes including five classes of starch synthases (SSs. While the overall starch synthase (SS reaction is known, the functional differences between the five SS classes are poorly understood. Much of our knowledge comes from analyzing mutant plants with altered SS activities, but the resulting data are often difficult to interpret as a result of pleitropic effects, competition between enzymes, overlaps in enzyme activity and disruption of multi-enzyme complexes. Here we provide a detailed biochemical study of the activity of all five classes of SSs in barley endosperm. Each enzyme was produced recombinantly in E. coli and the properties and modes of action in vitro were studied in isolation from other SSs and other substrate modifying activities. Our results define the mode of action of each SS class in unprecedented detail; we analyze their substrate selection, temperature dependence and stability, substrate affinity and temporal abundance during barley development. Our results are at variance with some generally accepted ideas about starch biosynthesis and might lead to the reinterpretation of results obtained in planta. In particular, they indicate that granule bound SS is capable of processive action even in the absence of a starch matrix, that SSI has no elongation limit, and that SSIV, believed to be critical for the initiation of starch granules, has maltoligosaccharides and not polysaccharides as its preferred substrates.

Purification and biochemical characterization of Mur ligases from Staphylococcus aureus.

Science.gov (United States)

Patin, Delphine; Boniface, Audrey; Kovač, Andreja; Hervé, Mireille; Dementin, Sébastien; Barreteau, Hélène; Mengin-Lecreulx, Dominique; Blanot, Didier

2010-12-01

The Mur ligases (MurC, MurD, MurE and MurF) catalyze the stepwise synthesis of the UDP-N-acetylmuramoyl-pentapeptide precursor of peptidoglycan. The murC, murD, murE and murF genes from Staphylococcus aureus, a major pathogen, were cloned and the corresponding proteins were overproduced in Escherichia coli and purified as His(6)-tagged forms. Their biochemical properties were investigated and compared to those of the E. coli enzymes. Staphylococcal MurC accepted L-Ala, L-Ser and Gly as substrates, as the E. coli enzyme does, with a strong preference for L-Ala. S. aureus MurE was very specific for L-lysine and in particular did not accept meso-diaminopimelic acid as a substrate. This mirrors the E. coli MurE specificity, for which meso-diaminopimelic acid is the preferred substrate and L-lysine a very poor one. S. aureus MurF appeared less specific and accepted both forms (L-lysine and meso-diaminopimelic acid) of UDP-MurNAc-tripeptide, as the E. coli MurF does. The inverse and strict substrate specificities of the two MurE orthologues is thus responsible for the presence of exclusively meso-diaminopimelic acid and L-lysine at the third position of the peptide in the peptidoglycans of E. coli and S. aureus, respectively. The specific activities of the four Mur ligases were also determined in crude extracts of S. aureus and compared to cell requirements for peptidoglycan biosynthesis. Copyright © 2010 Elsevier Masson SAS. All rights reserved.

Generation and characterization of the first immortalized alpaca cell line suitable for diagnostic and immunization studies.

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Valentina Franceschi

Full Text Available Raising of alpacas as exotic livestock for wool and meat production and as companion animals is growing in importance in the United States, Europe and Australia. Furthermore the alpaca, as well as the rest of the camelids, possesses the peculiarity of producing single-chain antibodies from which nanobodies can be generated. Nanobodies, due to their structural simplicity and reduced size, are very versatile in terms of manipulation and bio-therapeutic exploitation. In fact the biotech companies involved in nanobody production and application continue to grow in number and size. Hence, the development of reagents and tools to assist in the further growth of this new scientific and entrepreneurial reality is becoming a necessity. These are needed mainly to address alpaca disease diagnosis and prophylaxis, and to develop alpaca immunization strategies for nanobody generation. For instance an immortalized alpaca cell line would be extremely valuable. In the present work the first stabilized alpaca cell line from alpaca skin stromal cells (ASSCs was generated and characterized. This cell line was shown to be suitable for replication of viruses bovine herpesvirus-1, bovine viral diarrhea virus and caprine herpesvirus-1 and the endocellular parasite Neospora caninum. Moreover ASSCs were easy to transfect and transduce by several methods. These two latter characteristics are extremely useful when recombinant antigens need to be produced in a host homologous system. This work could be considered as a starting point for the expansion of the biotechnologies linked to alpaca farming and industry.

Characterization of superconducting transmission line resonators

Energy Technology Data Exchange (ETDEWEB)

Goetz, Jan; Summer, Philipp; Meier, Sebastian; Haeberlein, Max; Wulschner, Karl Friedrich; Eder, Peter; Fischer, Michael; Schwarz, Manuel; Deppe, Frank; Fedorov, Kirill; Huebl, Hans; Menzel, Edwin [Walther-Meissner-Institut, Bayerische Akademie der Wissenschaften, Garching (Germany); Physik-Department, TU Muenchen, Garching (Germany); Krawczyk, Marta; Marx, Achim [Walther-Meissner-Institut, Bayerische Akademie der Wissenschaften, Garching (Germany); Baust, Alexander; Xie, Edwar; Zhong, Ling; Gross, Rudolf [Walther-Meissner-Institut, Bayerische Akademie der Wissenschaften, Garching (Germany); Physik-Department, TU Muenchen, Garching (Germany); Nanosystems Initiative Munich (NIM), Muenchen (Germany)

2015-07-01

Superconducting transmission line resonators are widely used in circuit quantum electrodynamics experiments as quantum bus or storage devices. For these applications, long coherence times, which can be linked to the internal quality factor of the resonators, are crucial. Here, we show a systematic study of the internal quality factor of niobium thin film resonators. We analyze different cleaning methods and substrate parameters for coplanar waveguide as well as microstrip geometries. In addition, we investigate the impact of a niobium-aluminum interface which is necessary for galvanically coupled flux qubits made from aluminum. This interface can be avoided by fabricating the complete resonator-qubit structure using Al/AlO{sub x}/Al technology during fabrication.

Biochemical, Kinetic, and Spectroscopic Characterization of Ruegeria pomeroyi DddW--A Mononuclear Iron-Dependent DMSP Lyase.

Directory of Open Access Journals (Sweden)

Adam E Brummett

Full Text Available The osmolyte dimethylsulfoniopropionate (DMSP is a key nutrient in marine environments and its catabolism by bacteria through enzymes known as DMSP lyases generates dimethylsulfide (DMS, a gas of importance in climate regulation, the sulfur cycle, and signaling to higher organisms. Despite the environmental significance of DMSP lyases, little is known about how they function at the mechanistic level. In this study we biochemically characterize DddW, a DMSP lyase from the model roseobacter Ruegeria pomeroyi DSS-3. DddW is a 16.9 kDa enzyme that contains a C-terminal cupin domain and liberates acrylate, a proton, and DMS from the DMSP substrate. Our studies show that as-purified DddW is a metalloenzyme, like the DddQ and DddP DMSP lyases, but contains an iron cofactor. The metal cofactor is essential for DddW DMSP lyase activity since addition of the metal chelator EDTA abolishes its enzymatic activity, as do substitution mutations of key metal-binding residues in the cupin motif (His81, His83, Glu87, and His121. Measurements of metal binding affinity and catalytic activity indicate that Fe(II is most likely the preferred catalytic metal ion with a nanomolar binding affinity. Stoichiometry studies suggest DddW requires one Fe(II per monomer. Electronic absorption and electron paramagnetic resonance (EPR studies show an interaction between NO and Fe(II-DddW, with NO binding to the EPR silent Fe(II site giving rise to an EPR active species (g = 4.29, 3.95, 2.00. The change in the rhombicity of the EPR signal is observed in the presence of DMSP, indicating that substrate binds to the iron site without displacing bound NO. This work provides insight into the mechanism of DMSP cleavage catalyzed by DddW.

Use of GEANT4 vs. MCNPX for the characterization of a boron-lined neutron detector

Energy Technology Data Exchange (ETDEWEB)

Ende, B.M. van der; Atanackovic, J.; Erlandson, A.; Bentoumi, G.

2016-06-01

This work compares GEANT4 with MCNPX in the characterization of a boron-lined neutron detector. The neutron energy ranges simulated in this work (0.025 eV to 20 MeV) are the traditional domain of MCNP simulations. This paper addresses the question, how well can GEANT4 and MCNPX be employed for detailed thermal neutron detector characterization? To answer this, GEANT4 and MCNPX have been employed to simulate detector response to a {sup 252}Cf energy spectrum point source, as well as to simulate mono-energetic parallel beam source geometries. The {sup 252}Cf energy spectrum simulation results demonstrate agreement in detector count rate within 3% between the two packages, with the MCNPX results being generally closer to experiment than are those from GEANT4. The mono-energetic source simulations demonstrate agreement in detector response within 5% between the two packages for all neutron energies, and within 1% for neutron energies between 100 eV and 5 MeV. Cross-checks between the two types of simulations using ISO-8529 {sup 252}Cf energy bins demonstrates that MCNPX results are more self-consistent than are GEANT4 results, by 3–4%.

Biochemical signatures of in vitro radiation response in human lung, breast and prostate tumour cells observed with Raman spectroscopy

International Nuclear Information System (INIS)

Matthews, Q; Jirasek, A; Lum, J J; Brolo, A G

2011-01-01

This work applies noninvasive single-cell Raman spectroscopy (RS) and principal component analysis (PCA) to analyze and correlate radiation-induced biochemical changes in a panel of human tumour cell lines that vary by tissue of origin, p53 status and intrinsic radiosensitivity. Six human tumour cell lines, derived from prostate (DU145, PC3 and LNCaP), breast (MDA-MB-231 and MCF7) and lung (H460), were irradiated in vitro with single fractions (15, 30 or 50 Gy) of 6 MV photons. Remaining live cells were harvested for RS analysis at 0, 24, 48 and 72 h post-irradiation, along with unirradiated controls. Single-cell Raman spectra were acquired from 20 cells per sample utilizing a 785 nm excitation laser. All spectra (200 per cell line) were individually post-processed using established methods and the total data set for each cell line was analyzed with PCA using standard algorithms. One radiation-induced PCA component was detected for each cell line by identification of statistically significant changes in the PCA score distributions for irradiated samples, as compared to unirradiated samples, in the first 24-72 h post-irradiation. These RS response signatures arise from radiation-induced changes in cellular concentrations of aromatic amino acids, conformational protein structures and certain nucleic acid and lipid functional groups. Correlation analysis between the radiation-induced PCA components separates the cell lines into three distinct RS response categories: R1 (H460 and MCF7), R2 (MDA-MB-231 and PC3) and R3 (DU145 and LNCaP). These RS categories partially segregate according to radiosensitivity, as the R1 and R2 cell lines are radioresistant (SF 2 > 0.6) and the R3 cell lines are radiosensitive (SF 2 < 0.5). The R1 and R2 cell lines further segregate according to p53 gene status, corroborated by cell cycle analysis post-irradiation. Potential radiation-induced biochemical response mechanisms underlying our RS observations are proposed, such as (1) the

Metabolic characterization of invaded cells of the pancreatic cancer cell line, PANC-1.

Science.gov (United States)

Fujita, Mayumi; Imadome, Kaori; Imai, Takashi

2017-05-01

We previously reported that about 0.4% of cells in the cultured human pancreatic cancer cell line, PANC-1, can invade matrigel during the transwell invasion assay, suggesting that these invaded PANC-1 cells may have specific characteristics to keep their invasive potential. To identify the metabolic characterization specific in the invaded PANC-1 cells, metabolome analysis of the invaded PANC-1 compared with the whole cultured PANC-1 was performed using CE-TOFMS, and concentrations of 110 metabolites were measured. In contrast to the whole cultured cells, the invaded PANC-1 was characterized as a population with reduced levels of amino acids and TCA cycle intermediates, and decreased and increased intermediates in glycolysis and nucleic acid metabolism. In particular, the ratio of both adenosine and guanosine energy charge was reduced in the invaded cells, revealing that the consumption of ATP and GTP was high in the invaded cells, and thus suggesting that ATP- or GTP-generating pathways are stimulated. In addition, the GSH/GSSG ratio was low in the invaded cells, but these cells had a higher surviving fraction after exposure to hydrogen peroxide. Thus, the invaded cells were the population resistant to oxidative stress. Furthermore, reduction in intracellular GSH content inhibited PANC-1 invasiveness, indicated that GSH has an important role in PANC-1 invasiveness. Overall, we propose the invaded cells have several unique metabolic profiles. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Prevalence of biochemical and immunological abnormalities in ...

African Journals Online (AJOL)

Tile prevalence of biochemical and immunological abnormalities was studied in a group of 256 patients with rheumatoid arthritis (104 coloureds, 100 whites and 52 blacks). The most common biochemical abnormalities detected were a reduction in the serum creatinine value (43,4%), raised globulins (39,7%), raised serum ...

[Biochemical diagnostics of fatal opium intoxication].

Science.gov (United States)

Papyshev, I P; Astashkina, O G; Tuchik, E S; Nikolaev, B S; Cherniaev, A L

2013-01-01

Biochemical diagnostics of fatal opium intoxication remains a topical problem in forensic medical science and practice. We investigated materials obtained in the course of forensic medical expertise of the cases of fatal opium intoxication. The study revealed significant differences between myoglobin levels in blood, urine, myocardium, and skeletal muscles. The proposed approach to biochemical diagnostics of fatal opium intoxication enhances the accuracy and the level of evidence of expert conclusions.

Yellow Rust Resistance in Advanced Lines and Commercial ...

African Journals Online (AJOL)

The objective of this study was to characterize seedling yellow rust resistance in 21 advanced bread wheat lines and 20 cultivars from Ethiopia. Yellow rust infection types (ITs) produced on test wheat lines and cultivars from nine yellow rust races were compared with ITs produced on standard differential lines that differed ...

Characterization of cells recovered from the xenotransplanted NG97 human-derived glioma cell line subcultured in a long-term in vitro

Directory of Open Access Journals (Sweden)

Heinrich Juliana K

2008-10-01

Full Text Available Abstract Background In order to elucidate tumoral progression and drug resistance, cultured cell lines are valuable tools applied on tumor related assays provided they are well established and characterized. Our laboratory settled the NG97 cell line derived from a human astrocytoma grade III, which started to develop and express important phenotypical characteristics of an astrocytoma grade IV after injection in the flank of nude mice. Astrocytomas are extremely aggressive malignancies of the Central Nervous System (CNS and account for 46% of all primary malignant brain tumors. Progression to worse prognosis occurs in 85% of the cases possibly due to changes in cell tumor microenvironment and through biological pathways that are still unclear. Methods This work focused on characterizing the NG97 cell line specifically after being recovered from the xenotransplant, who maintained their undifferentiated characteristics along the following 60th passages in vitro. These cells were subcultivated to evaluate the possible contribution of these undifferentiated characteristics to the malignant progression phenotype. These characteristics were the expression of molecules involved in the processes of migration, dedifferentiation and chromosomal instability. Results Results showed that NG97(ht had an decrease in doubling time through sub cultivation, which was characterized by a converse modulation between the expression of glial fibrillary acidic protein (GFAP and vimentin. In addition, β1 integrins were present in intermediate levels while α5 integrins had a high expression profile as well as fibronectin and laminin. Cytogenetic analysis of NG97(ht revealed several chromosomal abnormalities, 89% of the cells showed to be hyperdiploid and the modal number was assigned to be 63. Several acrocentric chromosomes were visualized and at least 30 figures were attributed to be murine. These findings suggest a possible fusion between the original NG97 cells

Characterization of cells recovered from the xenotransplanted NG97 human-derived glioma cell line subcultured in a long-term in vitro

International Nuclear Information System (INIS)

Machado, Camila ML; Boetcher-Luiz, Fátima; Verinaud, Liana; Ikemori, Rafael Y; Zorzeto, Tatiana Q; Nogueira, Ana CMA; Barbosa, Suse DS; Savino, Wilson; Schenka, André A; Vassallo, José; Heinrich, Juliana K

2008-01-01

In order to elucidate tumoral progression and drug resistance, cultured cell lines are valuable tools applied on tumor related assays provided they are well established and characterized. Our laboratory settled the NG97 cell line derived from a human astrocytoma grade III, which started to develop and express important phenotypical characteristics of an astrocytoma grade IV after injection in the flank of nude mice. Astrocytomas are extremely aggressive malignancies of the Central Nervous System (CNS) and account for 46% of all primary malignant brain tumors. Progression to worse prognosis occurs in 85% of the cases possibly due to changes in cell tumor microenvironment and through biological pathways that are still unclear. This work focused on characterizing the NG97 cell line specifically after being recovered from the xenotransplant, who maintained their undifferentiated characteristics along the following 60 th passages in vitro. These cells were subcultivated to evaluate the possible contribution of these undifferentiated characteristics to the malignant progression phenotype. These characteristics were the expression of molecules involved in the processes of migration, dedifferentiation and chromosomal instability. Results showed that NG97(ht) had an decrease in doubling time through sub cultivation, which was characterized by a converse modulation between the expression of glial fibrillary acidic protein (GFAP) and vimentin. In addition, β1 integrins were present in intermediate levels while α5 integrins had a high expression profile as well as fibronectin and laminin. Cytogenetic analysis of NG97(ht) revealed several chromosomal abnormalities, 89% of the cells showed to be hyperdiploid and the modal number was assigned to be 63. Several acrocentric chromosomes were visualized and at least 30 figures were attributed to be murine. These findings suggest a possible fusion between the original NG97 cells with stromal murine cells in the xenotransplant. In

Characterization of a continuous feline mammary epithelial cell line susceptible to feline epitheliotropic viruses.

Science.gov (United States)

Pesavento, Patricia; Liu, Hongwei; Ossiboff, Robert J; Stucker, Karla M; Heymer, Anna; Millon, Lee; Wood, Jason; van der List, Deborah; Parker, John S L

2009-04-01

Mucosal epithelial cells are the primary targets for many common viral pathogens of cats. Viral infection of epithelia can damage or disrupt the epithelial barrier that protects underlying tissues. In vitro cell culture systems are an effective means to study how viruses infect and disrupt epithelial barriers, however no true continuous or immortalized feline epithelial cell culture lines are available. A continuous cell culture of feline mammary epithelial cells (FMEC UCD-04-2) that forms tight junctions with high transepithelial electrical resistance (>2000Omegacm(-1)) 3-4 days after reaching confluence was characterized. In addition, it was shown that FMECs are susceptible to infection with feline calicivirus (FCV), feline herpesvirus (FHV-1), feline coronavirus (FeCoV), and feline panleukopenia virus (FPV). These cells will be useful for studies of feline viral disease and for in vitro studies of feline epithelia.

Biochemical changes in cuttings of Robinia pseudoacacia after treatment with naphthenate

Directory of Open Access Journals (Sweden)

SLAVKO KEVRESAN

2007-10-01

Full Text Available Naphthenic acids were isolated from gas oil fractions (distillation interval 168–290 °C of Vojvodina crude oil “Velebit”, characterized and their biological ac­tivity evaluated by the biochemical changes in cuttings of Robinia pseudoacacia after treatment with naphthenate. The activities of IAA peroxidase, total peroxi­da­ses and amylase, as well as the contents of reducing sugars and total proteins, were determined in the basal parts of soft wood cuttings of black locust after treat­ment with sodium naphthenate or the sodium salt of 1-naphthaleneacetic acid (NAA, con­­centration 10-7 mol dm-3 for 3 or 6 h. High activities of IAA oxidase and amy­lase, together with a low activity of peroxidase (which is known as being stimula­tory for the initiation and activation of primordia were obtained after the three-hour treatment with sodium naphthenate. Six-hour treatment had an inhibitory effect on the examined biochemical markers. The effects of three- and six-hour treatments with NAA were between those of the corresponding treatment with naphthenic acids.

Purification and Biochemical Characterization of a Neutral Serine Protease from Trichoderma harzianum. Use in Antibacterial Peptide Production from a Fish By-Product Hydrolysate.

Science.gov (United States)

Aissaoui, Neyssene; Chobert, Jean-Marc; Haertlé, Thomas; Marzouki, M Nejib; Abidi, Ferid

2017-06-01

This study reports the purification and biochemical characterization of an extracellular neutral protease from the fungus Trichoderma harzianum. The protease (Th-Protease) was purified from the culture supernatant to homogeneity by a three-step procedure with 14.2% recovery and 9.06-fold increase in specific activity. The purified enzyme appeared as a single protein band after sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with a molecular mass of about 20 kDa. The optimum pH and temperature for the proteolytic activity were pH 7.0 and 40 °C, respectively. The enzyme was then investigated for its potential application in the production of antibacterial peptides. Interestingly, Scorpaena notata viscera protein hydrolysate prepared using the purified serine protease (Th-Protease) showed remarkable in vitro antibacterial activities. A peptide with a high antibacterial activity was further purified by a three-step procedure, and its sequence was identified as FPIGMGHGSRPA. The result of this study offers a promising alternative to produce natural antibacterial peptides from fish protein hydrolysate.

Molecular and biochemical characterization of carbonic anhydrases of Paracoccidioides.

Science.gov (United States)

Tomazett, Mariana Vieira; Zanoelo, Fabiana Fonseca; Bailão, Elisa Flávia Cardoso; Bailão, Alexandre Melo; Borges, Clayton Luiz; Soares, Célia Maria de Almeida

2016-01-01

Carbonic anhydrases (CA) belong to the family of zinc metalloenzymes that catalyze the reversible hydration of carbon dioxide to bicarbonate. In the present work, we characterized the cDNAs of four Paracoccidioides CAs (CA1, CA2, CA3, and CA4). In the presence of CO2, there was not a significant increase in fungal ca1, ca2 and ca4 gene expression. The ca1 transcript was induced during the mycelium-to-yeast transition, while ca2 and ca4 gene expression was much higher in yeast cells, when compared to mycelium and mycelium-to-yeast transition. The ca1 transcript was induced in yeast cells recovered directly from liver and spleen of infected mice, while transcripts for ca2 and ca4 were down-regulated. Recombinant CA1 (rCA1) and CA4 (rCA4), with 33 kDa and 32 kDa respectively, were obtained from bacteria. The enzymes rCA1 (β-class) and rCA4 (α-class) were characterized regarding pH, temperature, ions and amino acids addition influence. Both enzymes were stable at pHs 7.5-8.5 and temperatures of 30-35 °C. The enzymes were dramatically inhibited by Hg+2 and activated by Zn+2, while only rCA4 was stimulated by Fe2+. Among the amino acids tested (all in L configuration), arginine, lysine, tryptophan and histidine enhanced residual activity of rCA1 and rCA4.

Extraction of lipase from Burkholderia cepacia by PEG/Phosphate ATPS and its biochemical characterization

Directory of Open Access Journals (Sweden)

Giovana da Silva Padilha

2012-02-01

Full Text Available This work aimed to study the partitioning of a lipase produced by Burkholderia cepacia in PEG/Phosphate aqueous two phase system (ATPS and its characterization. Lipase was produced by B. cepacia strains in a fermenter. Enzyme partitioning occurred at pH 6.0 and 8.0, using PEG 1500 and 6000 on two tie lines. Metal ions, pH and temperature effects on enzyme activity were evaluated. Five milliliter of 7.5% olive oil emulsion with 2.5% gumarabic in 0.1M sodium phosphate buffer at pH 8.0 and 37ºC were used for the activity determinations. Results showed that crude stratum from B. cepacia was partitioned by PEG1500/phosphate ATPS at pH 6.0 or 8.0 for, which the partitioning coefficients were 108-and 209-folds. Lipase presented optimal activity conditions at 37ºC and pH 8.0; it showed pH-stability for 4 h of incubation at different pH values at 37ºC. Metal ions such as Mn2+ , Co2+, I-and Ca2+ sustained enzymatic activities; however, it was inhibited by the presence of Fe2+, Hg2+ and Al3+ . Km and Vmax values were 0.258 U/mg and 43.90 g/L, respectively. A molecular weight of 33 kDa and an isoelectric point at pH 5.0 were determined by SDS-PAGE and IFS electrophoresis, respectively.

Biochemical characterization of a glucoamylase from Saccharomycopsis fibuligera R64

NARCIS (Netherlands)

Natalia, Dessy; Vidilaseris, Keni; Satrimafitrah, Pasjan; Ismaya Purkan, Wangsa T.; Permentier, Hjalmar; Fibriansah, Guntur; Puspasari, Fernita; Nurachman, Zeily; Dijkstra, Bauke; Soemitro, Soetijoso

Glucoamylase from the yeast Saccharomycopsis fibuligera R64 (GLL1) has successfully been purified and characterized. The molecular mass of the enzyme was 56,583 Da as determined by mass spectrometry. The purified enzyme demonstrated optimum activity in the pH range of 5.6-6.4 and at 50A degrees C.

Biochemical transport modeling, estimation, and detection in realistic environments

Science.gov (United States)

Ortner, Mathias; Nehorai, Arye

2006-05-01

Early detection and estimation of the spread of a biochemical contaminant are major issues for homeland security applications. We present an integrated approach combining the measurements given by an array of biochemical sensors with a physical model of the dispersion and statistical analysis to solve these problems and provide system performance measures. We approximate the dispersion model of the contaminant in a realistic environment through numerical simulations of reflected stochastic diffusions describing the microscopic transport phenomena due to wind and chemical diffusion using the Feynman-Kac formula. We consider arbitrary complex geometries and account for wind turbulence. Localizing the dispersive sources is useful for decontamination purposes and estimation of the cloud evolution. To solve the associated inverse problem, we propose a Bayesian framework based on a random field that is particularly powerful for localizing multiple sources with small amounts of measurements. We also develop a sequential detector using the numerical transport model we propose. Sequential detection allows on-line analysis and detecting wether a change has occurred. We first focus on the formulation of a suitable sequential detector that overcomes the presence of unknown parameters (e.g. release time, intensity and location). We compute a bound on the expected delay before false detection in order to decide the threshold of the test. For a fixed false-alarm rate, we obtain the detection probability of a substance release as a function of its location and initial concentration. Numerical examples are presented for two real-world scenarios: an urban area and an indoor ventilation duct.

Characterization of human lymphoid cell lines GM9947 and GM9948 as intra- and interlaboratory reference standards for DNA typing

Energy Technology Data Exchange (ETDEWEB)

Fregeau, C.J.; Elliott, J.C.; Fourney, R.M. [RCMP Central Forensic Laboratory, Ottawa, Ontario (Canada)] [and others

1995-07-20

The incorporation of reference DNA is crucial to the validation of any DNA typing protocol. Currently, reference DNA standards are restricted to molecular size DNA ladders and/or tumor cell line DNA. Either of these, however, presents some limitations. We have rigorously characterized two Epstein-Barr virus (EBV)-immortalized human lymphoid cell lines-GM9947 (female) and GM9948 (male)-to determine their suitability as alternative in-line standards for three widely employed allele profiling strategies. Twenty-one highly polymorphic VNTR-based allelic systems (7 RFLPs, 2 AmpFLPs, and 12 STRs) distributed over 12 chromosomes were scrutinized along with 3 gender-based discriminatory systems. The genetic stability of each locus was confirmed over a period of 225 in vitro population doublings. Allele size estimates and degree of informativeness for each of the 21 VNTR systems were compiled. The reproducibility of allele scoring by traditional RFLP analyses, using both cell lines as reference standards, was also verified by an interlaboratory validation study involving 13 analysts from two geographically distinct forensic laboratories. Taken together, our data indicate that GM9947 and GM9948 genomic DNAs could be adopted as reliable reference standards for DNA typing. 82 refs., 3 figs., 8 tabs.

Biochemical Characterization and Relative Expression Levels of Multiple Carbohydrate Esterases of the Xylanolytic Rumen Bacterium Prevotella ruminicola 23 Grown on an Ester-Enriched Substrate ▿ †

Science.gov (United States)

Kabel, Mirjam A.; Yeoman, Carl J.; Han, Yejun; Dodd, Dylan; Abbas, Charles A.; de Bont, Jan A. M.; Morrison, Mark; Cann, Isaac K. O.; Mackie, Roderick I.

2011-01-01

We measured expression and used biochemical characterization of multiple carbohydrate esterases by the xylanolytic rumen bacterium Prevotella ruminicola 23 grown on an ester-enriched substrate to gain insight into the carbohydrate esterase activities of this hemicellulolytic rumen bacterium. The P. ruminicola 23 genome contains 16 genes predicted to encode carbohydrate esterase activity, and based on microarray data, four of these were upregulated >2-fold at the transcriptional level during growth on an ester-enriched oligosaccharide (XOSFA,Ac) from corn relative to a nonesterified fraction of corn oligosaccharides (AXOS). Four of the 16 esterases (Xyn10D-Fae1A, Axe1-6A, AxeA1, and Axe7A), including the two most highly induced esterases (Xyn10D-Fae1A and Axe1-6A), were heterologously expressed in Escherichia coli, purified, and biochemically characterized. All four enzymes showed the highest activity at physiologically relevant pH (6 to 7) and temperature (30 to 40°C) ranges. The P. ruminicola 23 Xyn10D-Fae1A (a carbohydrate esterase [CE] family 1 enzyme) released ferulic acid from methylferulate, wheat bran, corn fiber, and XOSFA,Ac, a corn fiber-derived substrate enriched in O-acetyl and ferulic acid esters, but exhibited negligible activity on sugar acetates. As expected, the P. ruminicola Axe1-6A enzyme, which was predicted to possess two distinct esterase family domains (CE1 and CE6), released ferulic acid from the same substrates as Xyn10D-Fae1 and was also able to cleave O-acetyl ester bonds from various acetylated oligosaccharides (AcXOS). The P. ruminicola 23 AxeA1, which is not assigned to a CE family, and Axe7A (CE7) were found to be acetyl esterases that had activity toward a broad range of mostly nonpolymeric acetylated substrates along with AcXOS. All enzymes were inhibited by the proximal location of other side groups like 4-O-methylglucuronic acid, ferulic acid, or acetyl groups. The unique diversity of carbohydrate esterases in P. ruminicola 23

Biochemical and toxicological studies of aqueous extract of ...

African Journals Online (AJOL)

Biochemical and toxicological studies of aqueous extract of Syzigium ... tract diseases and also used as food spices), on some biochemical indices, such as ... liver functions and blood parameters were studied in adult albino rats of both sexes.

Characterization of stem-like cells in a new astroblastoma cell line

Energy Technology Data Exchange (ETDEWEB)

Coban, Esra Aydemir; Kasikci, Ezgi [Department of Genetics and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul (Turkey); Karatas, Omer Faruk [Molecular Biology and Genetics Department, Erzurum Technical University, Erzurum (Turkey); Suakar, Oznur; Kuskucu, Aysegul [Department of Medical Genetics, Yeditepe University Medical School and Yeditepe University Hospital, Istanbul (Turkey); Altunbek, Mine [Department of Genetics and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul (Turkey); Türe, Uğur [Department of Neurosurgery, Yeditepe University School of Medicine, Istanbul (Turkey); Sahin, Fikrettin [Department of Genetics and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul (Turkey); Bayrak, Omer Faruk, E-mail: ofbayrak@yeditepe.edu.tr [Department of Medical Genetics, Yeditepe University Medical School and Yeditepe University Hospital, Istanbul (Turkey)

2017-03-15

Cell lines established from tumors are the most commonly used models in cancer research, and their use in recent years has enabled a greater understanding of the biology of cancer and the means to develop effective treatment strategies. Astroblastomas are uncommon neuroepithelial tumors of glial origin, predominantly affecting young people, mainly teenagers and children, predominantly females. To date, only a single study has reported that astroblastomas contain a large number of neural stem-like cells, which had only a partial proliferation capacity and differentiation. Our objective was to establish an astroblastoma cell line to investigate the presence of astroblastic cells and cancer stem-like cells. The migratory and invasion abilities of the cells were quantified with invasion and migration assays and compared to a glioblastoma cell line. The presence of stem cells was detected with surface-marker analysis by using flow cytometry, and measuring the differentiation ability with a differentiation assay and the self-renewal capacity with a sphere-forming assay. These characteristics may determine whether this novel cell line is a model for astroblastomas that may have stem-cell characteristics. With this novel cell line, scientists can investigate the molecular pathways underlying astroblastomas and develop new therapeutic strategies for patients with these tumors. - Highlights: • An establishment of a novel astroblastoma cell line was proposed. • The presence of astroblastic cells and cancer stem-like cells was investigated. • The molecular pathways underlying astroblastomas may be investigated. • New therapeutic strategies for patients with astroblastoma may be developed.

Molecular and biochemical characterization of carbonic anhydrases of Paracoccidioides

Directory of Open Access Journals (Sweden)

Mariana Vieira Tomazett

Full Text Available Abstract Carbonic anhydrases (CA belong to the family of zinc metalloenzymes that catalyze the reversible hydration of carbon dioxide to bicarbonate. In the present work, we characterized the cDNAs of four Paracoccidioides CAs (CA1, CA2, CA3, and CA4. In the presence of CO2, there was not a significant increase in fungal ca1, ca2 and ca4 gene expression. The ca1 transcript was induced during the mycelium-to-yeast transition, while ca2 and ca4 gene expression was much higher in yeast cells, when compared to mycelium and mycelium-to-yeast transition. The ca1 transcript was induced in yeast cells recovered directly from liver and spleen of infected mice, while transcripts for ca2 and ca4 were down-regulated. Recombinant CA1 (rCA1 and CA4 (rCA4, with 33 kDa and 32 kDa respectively, were obtained from bacteria. The enzymes rCA1 (β-class and rCA4 (α-class were characterized regarding pH, temperature, ions and amino acids addition influence. Both enzymes were stable at pHs 7.5-8.5 and temperatures of 30-35 °C. The enzymes were dramatically inhibited by Hg+2 and activated by Zn+2, while only rCA4 was stimulated by Fe2+. Among the amino acids tested (all in L configuration, arginine, lysine, tryptophan and histidine enhanced residual activity of rCA1 and rCA4.

Functional Coding Variation in Recombinant Inbred Mouse Lines Reveals Novel Serotonin Transporter-Associated Phenotypes

Energy Technology Data Exchange (ETDEWEB)

Carneiro, Ana [Vanderbilt University; Airey, David [University of Tennessee Health Science Center, Memphis; Thompson, Brent [Vanderbilt University; Zhu, C [Vanderbilt University; Rinchik, Eugene M [ORNL; Lu, Lu [University of Tennessee Health Science Center, Memphis; Chesler, Elissa J [ORNL; Erikson, Keith [University of North Carolina; Blakely, Randy [Vanderbilt University

2009-01-01

The human serotonin (5-hydroxytryptamine, 5-HT) transporter (hSERT, SLC6A4) figures prominently in the etiology or treatment of many prevalent neurobehavioral disorders including anxiety, alcoholism, depression, autism and obsessive-compulsive disorder (OCD). Here we utilize naturally occurring polymorphisms in recombinant inbred (RI) lines to identify novel phenotypes associated with altered SERT function. The widely used mouse strain C57BL/6J, harbors a SERT haplotype defined by two nonsynonymous coding variants (Gly39 and Lys152 (GK)). At these positions, many other mouse lines, including DBA/2J, encode Glu39 and Arg152 (ER haplotype), assignments found also in hSERT. Synaptosomal 5-HT transport studies revealed reduced uptake associated with the GK variant. Heterologous expression studies confirmed a reduced SERT turnover rate for the GK variant. Experimental and in silico approaches using RI lines (C57Bl/6J X DBA/2J=BXD) identifies multiple anatomical, biochemical and behavioral phenotypes specifically impacted by GK/ER variation. Among our findings are multiple traits associated with anxiety and alcohol consumption, as well as of the control of dopamine (DA) signaling. Further bioinformatic analysis of BXD phenotypes, combined with biochemical evaluation of SERT knockout mice, nominates SERT-dependent 5-HT signaling as a major determinant of midbrain iron homeostasis that, in turn, dictates ironregulated DA phenotypes. Our studies provide a novel example of the power of coordinated in vitro, in vivo and in silico approaches using murine RI lines to elucidate and quantify the system-level impact of gene variation.

eQuilibrator--the biochemical thermodynamics calculator.

Science.gov (United States)

Flamholz, Avi; Noor, Elad; Bar-Even, Arren; Milo, Ron

2012-01-01

The laws of thermodynamics constrain the action of biochemical systems. However, thermodynamic data on biochemical compounds can be difficult to find and is cumbersome to perform calculations with manually. Even simple thermodynamic questions like 'how much Gibbs energy is released by ATP hydrolysis at pH 5?' are complicated excessively by the search for accurate data. To address this problem, eQuilibrator couples a comprehensive and accurate database of thermodynamic properties of biochemical compounds and reactions with a simple and powerful online search and calculation interface. The web interface to eQuilibrator (http://equilibrator.weizmann.ac.il) enables easy calculation of Gibbs energies of compounds and reactions given arbitrary pH, ionic strength and metabolite concentrations. The eQuilibrator code is open-source and all thermodynamic source data are freely downloadable in standard formats. Here we describe the database characteristics and implementation and demonstrate its use.

eQuilibrator—the biochemical thermodynamics calculator

Science.gov (United States)

Flamholz, Avi; Noor, Elad; Bar-Even, Arren; Milo, Ron

2012-01-01

The laws of thermodynamics constrain the action of biochemical systems. However, thermodynamic data on biochemical compounds can be difficult to find and is cumbersome to perform calculations with manually. Even simple thermodynamic questions like ‘how much Gibbs energy is released by ATP hydrolysis at pH 5?’ are complicated excessively by the search for accurate data. To address this problem, eQuilibrator couples a comprehensive and accurate database of thermodynamic properties of biochemical compounds and reactions with a simple and powerful online search and calculation interface. The web interface to eQuilibrator (http://equilibrator.weizmann.ac.il) enables easy calculation of Gibbs energies of compounds and reactions given arbitrary pH, ionic strength and metabolite concentrations. The eQuilibrator code is open-source and all thermodynamic source data are freely downloadable in standard formats. Here we describe the database characteristics and implementation and demonstrate its use. PMID:22064852

Isolation and characterization of a new cell line from spontaneous mouse mammary tumour, MBL-6, for in vivo cancer studies

Directory of Open Access Journals (Sweden)

Ladan Langroudi

2017-12-01

Full Text Available In search for treatments against breast cancer, cell lines are one of the basic resources, particularly as in vitro models. Additionally, animal models of cancer are used as the successive step in therapeutics research. In this regard, human breast cancer cell lines provide fundamental models in vitro. However, in vivo studies require immunodeficient mice, which lack the influence of other in vivo factors such as the native microenvironment and the immune system. There are few standard models to study the pathogenic mechanism at molecular level and cell signaling pathway of breast cancer. In this study, a new mouse breast cancer cell line, MBL-6, was successfully established and characterized from tissues of a spontaneous mammary tumor. The cell line had epithelial morphology, formed adherent monolayer, maintained continuously in vitro and was able to form new tumors when injected subcutaneously in syngeneic mice. The growth pattern and metastasis evaluations revealed a considerable in situ duration before invading distant organs. Real time polymerase chain reaction (PCR analysis showed the expression of ER-, PR- and Her-2 receptors. The chromosome analysis showed numerous chromosomal abnormalities. Aggressive tumorigenecity in tumorigenesis test and the IC50 to cyclophosphamide (CTX, celecoxib (CLX and cisplatin (CPN was also evaluated. The numerous tests performed on the new MBL-6 cell line suggest that it is in good quality and may be used in animal models of breast cancer studies.

Variations in seed protein content of cotton (Gossypium hirsutum L.) mutant lines by in vivo and in vitro mutagenesis.

Science.gov (United States)

Muthusamy, Annamalai; Jayabalan, Narayanasamy

2013-01-01

The present work describes the influence of gamma irradiation (GR), ethyl methane sulphonate (EMS) and sodium azide (SA) treatment on yield and protein content of selected mutant lines of cotton. Seeds of MCU 5 and MCU 11 were exposed to gamma rays (GR), ethyl methane sulphonate (EMS) and sodium azide (SA). Lower dose of gamma irradiation (100-500 Gy), 10-50 mM EMS and SA at lower concentration effectively influences in improving the yield and protein content. Significant increase in yield (258.9 g plant(-1)) and protein content (18.63 mg g(-1) d. wt.) as compared to parental lines was noted in M2 generations. During the subsequent field trials, number of mutant lines varied morphologically in terms of yield as well as biochemical characters such as protein. The selected mutant lines were bred true to their characters in M3 and M4 generations. The significant increase in protein content and profiles of the mutant lines with range of 10.21-18.63 mg g(-1). The SDS-PAGE analysis of mutant lines revealed 9 distinct bands of different intensities with range of 26-81 kDa. The difference in intensity of bands was more (41, 50 and 58 kDa) in the mutant lines obtained from in vitro mutation than in vivo mutation. Significance of such stimulation in protein content correlated with yielding ability of the mutant lines of cotton in terms of seed weight per plant. The results confirm that in cotton it is possible to enhance the both yield and biochemical characters by in vivo and in vitro mutagenic treatments.

Generation, genome edition and characterization of iPSC lines from a patient with coenzyme Q10 deficiency harboring a heterozygous mutation in COQ4 gene

Directory of Open Access Journals (Sweden)

Damià Romero-Moya

2017-10-01

Full Text Available We report the generation, CRISPR/Cas9-edition and characterization of induced pluripotent stem cell (iPSC lines from a patient with coenzyme Q10 deficiency harboring the heterozygous mutation c.483G > C in the COQ4 gene. iPSCs were generated using non-integrative Sendai Viruses containing the reprogramming factors OCT4, SOX2, KLF4 and C-MYC. The iPSC lines carried the c.483G > C COQ4 mutation, silenced the OKSM expression and were mycoplasma-free. They were bona fide pluripotent cells as characterized by morphology, immunophenotype/gene expression for pluripotent-associated markers/genes, NANOG and OCT4 promoter demethylation, karyotype and teratoma formation. The COQ4 mutation was CRISPR/Cas9 edited resulting in isogenic, diploid and off-target free COQ4-corrected iPSCs.

Biochemical characterization of nuclear receptors for vitamin D3 and glucocorticoids in prostate stroma cell microenvironment

International Nuclear Information System (INIS)

Hidalgo, Alejandro A.; Montecinos, Viviana P.; Paredes, Roberto; Godoy, Alejandro S.; McNerney, Eileen M.; Tovar, Heribelt; Pantoja, Diego; Johnson, Candace; Trump, Donald; Onate, Sergio A.

2011-01-01

Highlights: → Fibroblasts from benign and carcinoma-associated stroma were biochemically characterized for VDR and GR function as transcription factors in prostate stroma cell microenvironment. → Decreased SRC-1/CBP coactivators recruitment to VDR and GR may result in hormone resistance to 1,25D 3 in stromal cell microenvironment prostate cancer. → 1a,25-Dyhidroxyvitamin D 3 (1,25D 3 ) and glucocorticoids, either alone or in combination, may not be an alternative for 'some' advanced prostate cancers that fails androgen therapies. -- Abstract: The disruption of stromal cell signals in prostate tissue microenvironment influences the development of prostate cancer to androgen independence. 1α,25-Dihydroxyvitamin D 3 (1,25D 3 ) and glucocorticoids, either alone or in combination, have been investigated as alternatives for the treatment of advanced prostate cancers that fails androgen therapies. The effects of glucocorticoids are mediated by the intracellular glucocorticoid receptor (GR). Similarly, the effect of 1,25D 3 is mediated by the 1,25D 3 nuclear receptor (VDR). In this study, fibroblasts from benign- (BAS) and carcinoma-associated stroma (CAS) were isolated from human prostates to characterize VDR and GR function as transcription factors in prostate stroma. The VDR-mediated transcriptional activity assessed using the CYP24-luciferase reporter was limited to 3-fold induction by 1,25D 3 in 9 out of 13 CAS (70%), as compared to >10-fold induction in the BAS clinical sample pair. Expression of His-tagged VDR (Ad-his-VDR) failed to recover the low transcriptional activity of the luciferase reporter in 7 out of 9 CAS. Interestingly, expression of Ad-his-VDR successfully recovered receptor-mediated induction in 2 out of the 9 CAS analyzed, suggesting that changes in the receptor protein itself was responsible for decreased response and resistance to 1,25D 3 action. Conversely, VDR-mediated transcriptional activity was more efficient in 4 out of 13 CAS (30

Possible Biochemical Markers in Protein-Energy Malnutrition and ...

African Journals Online (AJOL)

This study was carried out to determine possible biochemical markers in children suffering from Plasmodium falciparum malaria and Protein-Energy Malnutrition in a Hospital setting in Western Kenya. Spectrophotometric assays of selected biochemical parameters namely, albumin, total proteins, glucose, glutamate ...

Phenotypic characterization of canine Malassezia spp., isolates

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Angélica Hurtado-Suárez

2016-09-01

Full Text Available Objective. To characterize and identify yeasts of the genus Malassezia by phenotypic features. Materials and methods. First, the macroscopic and microscopic morphological characteristics were described. In addition we performed biochemical and physiological assays as Tweens and Cremophor, including more. Results. Our results evidenced of 105 isolates obtained from dogs diagnosed with external otitis, it was possible to identify two distinct species from 46 isolates within the Malassezia genus: 36.19% (n=38 were identified as M. pachydermatis and 7.62% (n=8 as M. furfur. According to phenotypic patterns the remaining 56.19% (n=59 were reported as Malassezia spp., possibly corresponding to M. furfur and/or M. pachydermatis. Conclusions. Results emphasize the necessity to characterize according to species. It is not feasible to define Malassezia by species based on morphological, biochemical, and physiological findings. Therefore, molecular genotyping should be performed to identify markers allowing a more precise isolate identification. This would broaden our epidemiological knowledge regarding different species involved in canine otitis pathologies.

iPSCORE: A Resource of 222 iPSC Lines Enabling Functional Characterization of Genetic Variation across a Variety of Cell Types

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Athanasia D. Panopoulos

2017-04-01

Full Text Available Summary: Large-scale collections of induced pluripotent stem cells (iPSCs could serve as powerful model systems for examining how genetic variation affects biology and disease. Here we describe the iPSCORE resource: a collection of systematically derived and characterized iPSC lines from 222 ethnically diverse individuals that allows for both familial and association-based genetic studies. iPSCORE lines are pluripotent with high genomic integrity (no or low numbers of somatic copy-number variants as determined using high-throughput RNA-sequencing and genotyping arrays, respectively. Using iPSCs from a family of individuals, we show that iPSC-derived cardiomyocytes demonstrate gene expression patterns that cluster by genetic background, and can be used to examine variants associated with physiological and disease phenotypes. The iPSCORE collection contains representative individuals for risk and non-risk alleles for 95% of SNPs associated with human phenotypes through genome-wide association studies. Our study demonstrates the utility of iPSCORE for examining how genetic variants influence molecular and physiological traits in iPSCs and derived cell lines. : Working as part of the NHLBI NextGen consortium, Panopoulos and colleagues report the derivation and characterization of 222 publicly available iPSCs from ethnically diverse individuals with corresponding genomic data including SNP arrays, RNA-seq, and whole-genome sequencing. This collection provides a powerful resource to investigate the function of genetic variants. Keywords: iPSCORE, iPSC, GWAS, molecular traits, physiological traits, cardiac disease, NHLBI Next Gen, LQT2, KCNH2, iPSC-derived cardiomyocytes

The importance of biochemical and genetic findings in the diagnosis of atypical Norrie disease.

Science.gov (United States)

Rodríguez-Muñoz, Ana; García-García, Gema; Menor, Francisco; Millán, José M; Tomás-Vila, Miguel; Jaijo, Teresa

2018-01-26

Norrie disease (ND) is a rare X-linked disorder characterized by bilateral congenital blindness. ND is caused by a mutation in the Norrie disease pseudoglioma (NDP) gene, which encodes a 133-amino acid protein called norrin. Intragenic deletions including NDP and adjacent genes have been identified in ND patients with a more severe neurologic phenotype. We report the biochemical, molecular, clinical and radiological features of two unrelated affected males with a deletion including NDP and MAO genes. Biochemical and genetic analyses were performed to understand the atypical phenotype and radiological findings. Biogenic amines in cerebrospinal fluid (CSF) were measured by high-performance liquid chromatography. The coding exons of NDP gene were amplified by polymerase chain reaction. Multiplex ligation-dependent probe amplification and chromosomal microarray were carried out on both affected males. Computed tomography and magnetic resonance imaging were performed on the two patients. In one patient, the serotonin and catecholamine metabolite levels in CSF were virtually undetectable. In both patients, genetic studies revealed microdeletions in the Xp11.3 region, involving the NDP, MAOA and MAOB genes. Radiological examination demonstrated brain and cerebellar atrophy. We suggest that alterations caused by MAO deficit may remain during the first years of life. Clinical phenotype, biochemical findings and neuroimaging can guide the genetic study in patients with atypical ND and help us to a better understanding of this disease.

Possibilities and methods for biochemical assessment of radiation injury

Energy Technology Data Exchange (ETDEWEB)

Minkova, M [Meditsinska Akademiya, Sofia (Bulgaria). Nauchen Inst. po Rentgenologiya i Radiobiologiya

1986-01-01

An extensitive review (77 references) is made of the application of biochemical diagnostic methods for assessment of radiation diseases. A brief characteristics of several biochemical indicators is given: deoxycytidine, thymidine, rho-aminoisocarboxylic acid, DNA-ase, nucleic acids. Influence of such factors as age, sex, season etc. is studied by means of functional biochemical indicators as: creatine, triptophanic metabolites, 5-hydroxy-indolacetic acid, biogenic amines, serum proteins, enzymes, etc.

Clinical and biochemical manifestations of undifferentiated forms of connective tissue dysplasia in pregnant women with varicose veins of small pelvis

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N.M. Shibelgut

2010-03-01

Full Text Available Research objective is to define the pathogenesis of varicous veins of small pelvis in women. at Ultrasonic investigation of venous system of small pelvis has been carried out in 290 pregnant women. It revealed 190 patients with varicose veins of small pelvis (VVSP. By means of V.M. Jakovleva's technique phenotypic menifestation of connective tissue dysplasia was determined in all pregnant women. Biochemical manifestations of connective tissue dysplasia were identified by sialic acid level in blood serum, daily excretion of glycosaminoglycans and oxyproline. High frequency of clinical and biochemical manifestations of undifferentiated forms of connective tissue dysplasia was revealed in pregnant women with VVSP. Patients with VVSP developed tooth and jaw, facial and locomotor damages. Patients with VVSP characterized by visceral undifferentiated forms of connective tissue dysplasia demonstrated by refraction involvement, ventral hernias, flat feet, varicous veins of lower extremities, hypermobile syndrome, mitral valve prolapse of different degree. Biochemical manifestations of undifferentiated forms of connective tissue dysplasia in pregnant women with VVSP were insignificant

Isolation & characterization of Brucella melitensis isolated from patients suspected for human brucellosis in India

Science.gov (United States)

Barua, Anita; Kumar, Ashu; Thavaselvam, Duraipandian; Mangalgi, Smita; Prakash, Archana; Tiwari, Sapana; Arora, Sonia; Sathyaseelan, Kannusamy

2016-01-01

Background & objectives: Brucellosis is endemic in the southern part of India. A combination of biochemical, serological and molecular methods is required for identification and biotyping of Brucella. The present study describes the isolation and biochemical, molecular characterization of Brucella melitensis from patients suspected for human brucellosis. Methods: The blood samples were collected from febrile patients suspected to have brucellosis. A total of 18 isolates were obtained from 102 blood samples subjected to culture. The characterization of these 18 isolates was done by growth on Brucella specific medium, biochemical reactions, CO2 requirement, H2S production, agglutination with A and M mono-specific antiserum, dye sensitivity to basic fuchsin and thionin. Further, molecular characterization of the isolates was done by amplification of B. melitensis species specific IS711 repetitive DNA fragment and 16S (rRNA) sequence analysis. PCR-restriction fragment length polymorphism (RFLP) analysis of omp2 locus and IS711 gene was also done for molecular characterization. Results: All 102 suspected samples were subjected to bacteria isolation and of these, 18 isolates could be recovered on blood culture. The biochemical, PCR and PCR-RFLP and 16s rRNA sequencing revealed that all isolates were of B. melitensis and matched exactly with reference strain B. melitensis 16M. Interpretation & conclusions: The present study showed an overall isolation rate of 17.64 per cent for B. melitensis. There is a need to establish facilities for isolation and characterization of Brucella species for effective clinical management of the disease among patients as well as surveillance and control of infection in domestic animals. Further studies are needed from different geographical areas of the country with different level of endemicity to plan and execute control strategies against human brucellosis. PMID:27488010

Establishment and characterization of a new cell line, FPS-1, derived from human undifferentiated pleomorphic sarcoma, overexpressing epidermal growth factor receptor and cyclooxygenase-2.

Science.gov (United States)

Hakozaki, Michiyuki; Hojo, Hiroshi; Sato, Michiko; Tajino, Takahiro; Yamada, Hitoshi; Kikuchi, Shinichi; Abe, Masafumi

2006-01-01

Undifferentiated pleomorphic sarcoma (UPS) is among the most common soft tissue sarcomas in adults. In order to improve its aggressive course or prognosis and establish new therapeutic methods, molecular genetic and biological characterizations of UPS are required. A new human UPS cell line (FPS-1) was established from UPS of the upper arm of a 79-year-old man. The cell line has been maintained for over 14 months with more than 60 passages. FPS-1 cells were characterized using molecular biological methods. FPS-1 cells showed the same morphological and immunophenotypical characteristics as the primary tumor. Cytogenetic and molecular analyses revealed a nonsense mutation in exon 6 of the p53 gene. Epidermal growth factor receptor (EGFR) and cyclooxygenase-2 (COX-2) were expressed in FPS-1 cells. FPS-1 cells might be useful for investigating biological behavior and developing new molecular targeting antitumor drugs for UPS with EGFR or COX-2 expression.

[Biochemical markers of bone remodeling: pre-analytical variations and guidelines for their use. SFBC (Société Française de Biologie Clinique) Work Group. Biochemical markers of bone remodeling].

Science.gov (United States)

Garnero, P; Bianchi, F; Carlier, M C; Genty, V; Jacob, N; Kamel, S; Kindermans, C; Plouvier, E; Pressac, M; Souberbielle, J C

2000-01-01

Biochemical markers of bone turnover have been developed over the past 20 years that are more specific for bone tissue than conventional ones such as total alkaline phosphatase and urinary hydroxyproline. They have been widely used in clinical research and in clinical trials of new therapies as secondary end points of treatment efficacy. Most of the interest has been devoted to their use in postmenopausal osteoporosis, a condition characterized by subtle modifications of bone metabolism that cannot be detected readily by conventional markers of bone turnover. Although several recent studies have suggested that biochemical markers may be used for the management of the individual patient in routine clinical practice, this has not been clearly defined and is a matter of debate. Because of the crucial importance to clarify this issue, the Société Francaise de Biologie Clinique prompted an expert committee to summarize the available data and to make recommendations. The following paper includes a review on the biochemical and analytical aspects of the markers of bone formation and resorption and on the sources of variability such as sex, age, menstrual cycle, pregnancy and lactation, physical activity, seasonal variation and effects of diseases and treatments. We will also describe the effects of pre-analytical factors on the measurements of the different markers. Finally based on that review, we will make practical recommendations for the use of these markers in order to minimize the variability of the measurements and improve the clinical interpretation of the data.

Haematological and blood biochemical indices of West African ...

African Journals Online (AJOL)

Haematological and blood biochemical indices of West African dwarf goats vaccinated against Pestes des petit ruminants (PPR) ... blood biochemical indices of forty randomly selected West African dwarf (WAD) goats were studied. Packed cell volume ... neutrophil/lymphocyte ratio and white blood cells (WBC) than females.

BioNessie - a grid enabled biochemical networks simulation environment

OpenAIRE

Liu, X.; Jiang, J.; Ajayi, O.; Gu, X.; Gilbert, D.; Sinnott, R.O.

2008-01-01

The simulation of biochemical networks provides insight and understanding about the underlying biochemical processes and pathways used by cells and organisms. BioNessie is a biochemical network simulator which has been developed at the University of Glasgow. This paper describes the simulator and focuses in particular on how it has been extended to benefit from a wide variety of high performance compute resources across the UK through Grid technologies to support larger scale simulations.

Definitions of biochemical failure in prostate cancer following radiation therapy

International Nuclear Information System (INIS)

Taylor, Jeremy M.G.; Griffith, Kent A.; Sandler, Howard M.

2001-01-01

Purpose: The American Society for Therapeutic Radiology and Oncology (ASTRO) published a consensus panel definition of biochemical failure following radiation therapy for prostate cancer. In this paper, we develop a series of alternative definitions of biochemical failure. Using data from 688 patients, we evaluated the sensitivity and specificity of the various definitions, with respect to a defined 'clinically meaningful' outcome. Methods and Materials: The ASTRO definition of biochemical failure requires 3 consecutive rises in prostate-specific antigen (PSA). We considered several modifications to the standard definition: to require PSA rises of a certain magnitude, to consider 2 instead of 3 rises, to require the final PSA value to be greater than a fixed cutoff level, and to define biochemical failure based on the slope of PSA over 1, 1.5, or 2 years. A clinically meaningful failure is defined as local recurrence, distant metastases, initiation of unplanned hormonal therapy, unplanned radical prostatectomy, or a PSA>25 later than 6 months after radiation. Results: Requiring the final PSA in a series of consecutive rises to be larger than 1.5 ng/mL increased the specificity of biochemical failure. For a fixed specificity, defining biochemical failure based on 2 consecutive rises, or the slope over the last year, could increase the sensitivity by up to approximately 20%, compared to the ASTRO definition. Using a rule based on the slope over the previous year or 2 rises leads to a slightly earlier detection of biochemical failure than does the ASTRO definition. Even with the best rule, only approximately 20% of true failures are biochemically detected more than 1 year before the clinically meaningful event time. Conclusion: There is potential for improvement in the ASTRO consensus definition of biochemical failure. Further research is needed, in studies with long follow-up times, to evaluate the relationship between various definitions of biochemical failure and

Identification and characterization of SSE15206, a microtubule depolymerizing agent that overcomes multidrug resistance

KAUST Repository

Manzoor, Safia

2018-02-13

Microtubules are highly dynamic structures that form spindle fibres during mitosis and are one of the most validated cancer targets. The success of drugs targeting microtubules, however, is often limited by the development of multidrug resistance. Here we describe the discovery and characterization of SSE15206, a pyrazolinethioamide derivative [3-phenyl-5-(3,4,5-trimethoxyphenyl)-4,5-dihydro-1H-pyrazole-1-carbothioamide] that has potent antiproliferative activities in cancer cell lines of different origins and overcomes resistance to microtubule-targeting agents. Treatment of cells with SSE15206 causes aberrant mitosis resulting in G2/M arrest due to incomplete spindle formation, a phenotype often associated with drugs that interfere with microtubule dynamics. SSE15206 inhibits microtubule polymerization both in biochemical and cellular assays by binding to colchicine site in tubulin as shown by docking and competition studies. Prolonged treatment of cells with the compound results in apoptotic cell death [increased Poly (ADP-ribose) polymerase cleavage and Annexin V/PI staining] accompanied by p53 induction. More importantly, we demonstrate that SSE15206 is able to overcome resistance to chemotherapeutic drugs in different cancer cell lines including multidrug-resistant KB-V1 and A2780-Pac-Res cell lines overexpressing MDR-1, making it a promising hit for the lead optimization studies to target multidrug resistance.

Molecular Cytogenetic Characterization Identified the Murine B-Cell Lymphoma Cell Line A-20 as a Model for Sporadic Burkitt's Lymphoma.

Science.gov (United States)

Guja, Karolina; Liehr, Thomas; Rincic, Martina; Kosyakova, Nadezda; Hussein Azawi, Shaymaa S

2017-11-01

Here, we report the first molecular cytogenetic characterization of the BALB/cAnN mouse derived B-cell non-Hodgkin lymphoma (B-cell NHL) cell lines A-20. Even though previously used as a model for testing of, for example, dexametason, up to present, no data in the genetic properties of A-20 were available. The present study closed this gap and provides evidence that A-20 is a model for B-cell NHL subgroup sporadic Burkitt's lymphoma. C-myc oncogene is involved in a translocation and copy number alterations as gain of murine 14q material could be observed. Interestingly, the cell line showed the karyotype 39,X,-X or -Y,t(2;15)(qE5;qD2),del(6)(qB3qC3),del(9)(qA3qA4),dup(14)(qE1qE4) in ~95% of the cells, being exceptionally stable for cell lines being established 38 years ago. Still, ~5% of the cells showed polyploidization followed by chromothripsis. It remains to be determined if this can be observed also in other cell lines, just has not been reported yet, and/or if it is a unique feature of A-20. Overall, finally here, the necessary genetic data to identify A-20 as a model for human sporadic Burkitt's lymphoma are provided.

Characterization and diversity assessment in coconut collections of ...

African Journals Online (AJOL)

SUBDIC

conservation. Therefore, the present study was undertaken with the aim to characterize morphological, biochemical and nut component traits and to assess the diversity among 24 exotic and six indigenous in coconut accessions conserved at World Coconut Germplasm. Centre (WCGC) in Andaman and Nicobar Islands ...

Phytochemical analysis and In-vitro Biochemical Characterization of aqueous and methanolic extract of Triphala, a conventional herbal remedy

Directory of Open Access Journals (Sweden)

Romana Parveen

2018-03-01

Results revealed the presence of valuable bioactive compounds such as flavonoids, alkaloids, phenols, etc which might be responsible for biochemical activities. Extracts exhibited satisfactory radical-scavenging activity comparable with ascorbic acid. Methanolic extracts demonstrated higher antioxidant activity compared to aqueous extract. Extracts showed promising antibacterial potential against tested strain comparable to ampicillin. Hence, it can be concluded that triphala may be a promising candidate in pharmaceuticals and future medicine.

Microbiological and Biochemical Characterization of Cassava Retting, a Traditional Lactic Acid Fermentation for Foo-Foo (Cassava Flour) Production

OpenAIRE

Brauman, A.; Keleke, S.; Malonga, M.; Miambi, E.; Ampe, F.

1996-01-01

The overall kinetics of retting, a spontaneous fermentation of cassava roots performed in central Africa, was investigated in terms of microbial-population evolution and biochemical and physicochemical parameters. During the traditional process, endogenous cyanogens were almost totally degraded, plant cell walls were lysed by the simultaneous action of pectin methylesterase and pectate lysate, and organic acids (C2 to C4) were produced. Most microorganisms identified were found to be facultat...

40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides.

Science.gov (United States)

2010-07-01

... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections 158.2081...

CD147 expression predicts biochemical recurrence after prostatectomy independent of histologic and pathologic features

International Nuclear Information System (INIS)

Bauman, Tyler M.; Ewald, Jonathan A.; Huang, Wei; Ricke, William A.

2015-01-01

CD147 is an MMP-inducing protein often implicated in cancer progression. The purpose of this study was to investigate the expression of CD147 in prostate cancer (PCa) progression and the prognostic ability of CD147 in predicting biochemical recurrence after prostatectomy. Plasma membrane-localized CD147 protein expression was quantified in patient samples using immunohistochemistry and multispectral imaging, and expression was compared to clinico-pathological features (pathologic stage, Gleason score, tumor volume, preoperative PSA, lymph node status, surgical margins, biochemical recurrence status). CD147 specificity and expression were confirmed with immunoblotting of prostate cell lines, and CD147 mRNA expression was evaluated in public expression microarray datasets of patient prostate tumors. Expression of CD147 protein was significantly decreased in localized tumors (pT2; p = 0.02) and aggressive PCa (≥pT3; p = 0.004), and metastases (p = 0.001) compared to benign prostatic tissue. Decreased CD147 was associated with advanced pathologic stage (p = 0.009) and high Gleason score (p = 0.02), and low CD147 expression predicted biochemical recurrence (HR 0.55; 95 % CI 0.31–0.97; p = 0.04) independent of clinico-pathologic features. Immunoblot bands were detected at 44 kDa and 66 kDa, representing non-glycosylated and glycosylated forms of CD147 protein, and CD147 expression was lower in tumorigenic T10 cells than non-tumorigenic BPH-1 cells (p = 0.02). Decreased CD147 mRNA expression was associated with increased Gleason score and pathologic stage in patient tumors but is not associated with recurrence status. Membrane-associated CD147 expression is significantly decreased in PCa compared to non-malignant prostate tissue and is associated with tumor progression, and low CD147 expression predicts biochemical recurrence after prostatectomy independent of pathologic stage, Gleason score, lymph node status, surgical margins, and tumor volume in multivariable

Characterizing food waste substrates for co-digestion through biochemical methane potential (BMP) experiments.

Science.gov (United States)

Lisboa, Maria Sol; Lansing, Stephanie

2013-12-01

Co-digestion of food waste with dairy manure is increasingly utilized to increase energy production and make anaerobic digestion more affordable; however, there is a lack of information on appropriate co-digestion substrates. In this study, biochemical methane potential (BMP) tests were conducted to determine the suitability of four food waste substrates (meatball, chicken, cranberry and ice cream processing wastes) for co-digestion with flushed dairy manure at a ratio of 3.2% food waste and 96.8% manure (by volume), which equated to 14.7% (ice-cream) to 80.7% (chicken) of the VS being attributed to the food waste. All treatments led to increases in methane production, ranging from a 67.0% increase (ice cream waste) to a 2940% increase (chicken processing waste) compared to digesting manure alone, demonstrating the large potential methane production of food waste additions compared to relatively low methane production potential of the flushed dairy manure, even if the overall quantity of food waste added was minimal. Copyright © 2013 Elsevier Ltd. All rights reserved.

High-Throughput Flow Cytometric Method for the Simultaneous Measurement of CAR-T Cell Characterization and Cytotoxicity against Solid Tumor Cell Lines.

Science.gov (United States)

Martinez, Emily M; Klebanoff, Samuel D; Secrest, Stephanie; Romain, Gabrielle; Haile, Samuel T; Emtage, Peter C R; Gilbert, Amy E

2018-04-01

High-throughput flow cytometry is an attractive platform for the analysis of adoptive cellular therapies such as chimeric antigen receptor T cell therapy (CAR-T) because it allows for the concurrent measurement of T cell-dependent cellular cytotoxicity (TDCC) and the functional characterization of engineered T cells with respect to percentage of CAR transduction, T cell phenotype, and measurement of T cell function such as activation in a single assay. The use of adherent tumor cell lines can be challenging in these flow-based assays. Here, we present the development of a high-throughput flow-based assay to measure TDCC for a CAR-T construct co-cultured with multiple adherent tumor cell lines. We describe optimal assay conditions (such as adherent cell dissociation techniques to minimize impact on cell viability) that result in robust cytotoxicity assays. In addition, we report on the concurrent use of T cell transduction and activation antibody panels (CD25) that provide further dissection of engineered T cell function. In conclusion, we present the development of a high-throughput flow cytometry method allowing for in vitro interrogation of solid tumor, targeting CAR-T cell-mediated cytotoxicity, CAR transduction, and engineered T cell characterization in a single assay.

Atomic Structure and Biochemical Characterization of an RNA Endonuclease in the N Terminus of Andes Virus L Protein.

Directory of Open Access Journals (Sweden)

Yaiza Fernández-García

2016-06-01

Full Text Available Andes virus (ANDV is a human-pathogenic hantavirus. Hantaviruses presumably initiate their mRNA synthesis by using cap structures derived from host cell mRNAs, a mechanism called cap-snatching. A signature for a cap-snatching endonuclease is present in the N terminus of hantavirus L proteins. In this study, we aimed to solve the atomic structure of the ANDV endonuclease and characterize its biochemical features. However, the wild-type protein was refractory to expression in Escherichia coli, presumably due to toxic enzyme activity. To circumvent this problem, we introduced attenuating mutations in the domain that were previously shown to enhance L protein expression in mammalian cells. Using this approach, 13 mutant proteins encompassing ANDV L protein residues 1-200 were successfully expressed and purified. Protein stability and nuclease activity of the mutants was analyzed and the crystal structure of one mutant was solved to a resolution of 2.4 Å. Shape in solution was determined by small angle X-ray scattering. The ANDV endonuclease showed structural similarities to related enzymes of orthobunya-, arena-, and orthomyxoviruses, but also differences such as elongated shape and positively charged patches surrounding the active site. The enzyme was dependent on manganese, which is bound to the active site, most efficiently cleaved single-stranded RNA substrates, did not cleave DNA, and could be inhibited by known endonuclease inhibitors. The atomic structure in conjunction with stability and activity data for the 13 mutant enzymes facilitated inference of structure-function relationships in the protein. In conclusion, we solved the structure of a hantavirus cap-snatching endonuclease, elucidated its catalytic properties, and present a highly active mutant form, which allows for inhibitor screening.

Biochemical and morphological changes in rat lung tissue under the influence of external ionizing radiation

International Nuclear Information System (INIS)

Uzlenkova, N.Je.; Mamotyuk, Je.M.; Gusakova, V.A.; Kononenko, O.K.

2006-01-01

Single external x-ray exposure at minimum and mean lethal doses was established to cause a long activation of biochemical processes in the connective tissue of the rat lungs. Morphological and ultrastructure changes in the tissue of the lungs at early terms after x-ray and gamma-radiation exposure were due to development of destructive and degenerative reactions. The long-term changes were characterized by growth of connective tissue and formation of areas of fibrous changes in the structure of the lungs

Construction of a complete set of alien chromosome addition lines from Gossypium australe in Gossypium hirsutum: morphological, cytological, and genotypic characterization.

Science.gov (United States)

Chen, Yu; Wang, Yingying; Wang, Kai; Zhu, Xiefei; Guo, Wangzhen; Zhang, Tianzhen; Zhou, Baoliang

2014-05-01

We report the first complete set of alien addition lines of G. hirsutum . The characterized lines can be used to introduce valuable traits from G. australe into cultivated cotton. Gossypium australe is a diploid wild cotton species (2n = 26, GG) native to Australia that possesses valuable characteristics unavailable in the cultivated cotton gene pool, such as delayed pigment gland morphogenesis in the seed and resistances to pests and diseases. However, it is very difficult to directly transfer favorable traits into cultivated cotton through conventional gene recombination due to the absence of pairing and crossover between chromosomes of G. australe and Gossypium hirsutum (2n = 52, AADD). To enhance the transfer of favorable genes from wild species into cultivated cotton, we developed a set of hirsutum-australe monosomic alien chromosome addition lines (MAAL) using a combination of morphological survey, microsatellite marker-assisted selection, and molecular cytogenetic analysis. The amphidiploid (2n = 78, AADDGG) of G. australe and G. hirsutum was consecutively backcrossed with upland cotton to develop alien addition lines of individual G. australe chromosomes in G. hirsutum. From these backcross progeny, we generated the first complete set of chromosome addition lines in cotton; 11 of 13 lines are monosomic additions, and chromosomes 7G(a) and 13G(a) are multiple additions. MAALs of 1G(a) and 11G(a) were the first to be isolated. The chromosome addition lines can be employed as bridges for the transfer of desired genes from G. australe into G. hirsutum, as well as for gene assignment, isolation of chromosome-specific probes, flow sorting and microdissection of chromosome, development of chromosome-specific ''paints'' for fluorochrome-labeled DNA fragments, physical mapping, and selective isolation and mapping of cDNAs for a particular G. australe chromosome.

Accurate atom-mapping computation for biochemical reactions.

Science.gov (United States)

Latendresse, Mario; Malerich, Jeremiah P; Travers, Mike; Karp, Peter D

2012-11-26

The complete atom mapping of a chemical reaction is a bijection of the reactant atoms to the product atoms that specifies the terminus of each reactant atom. Atom mapping of biochemical reactions is useful for many applications of systems biology, in particular for metabolic engineering where synthesizing new biochemical pathways has to take into account for the number of carbon atoms from a source compound that are conserved in the synthesis of a target compound. Rapid, accurate computation of the atom mapping(s) of a biochemical reaction remains elusive despite significant work on this topic. In particular, past researchers did not validate the accuracy of mapping algorithms. We introduce a new method for computing atom mappings called the minimum weighted edit-distance (MWED) metric. The metric is based on bond propensity to react and computes biochemically valid atom mappings for a large percentage of biochemical reactions. MWED models can be formulated efficiently as Mixed-Integer Linear Programs (MILPs). We have demonstrated this approach on 7501 reactions of the MetaCyc database for which 87% of the models could be solved in less than 10 s. For 2.1% of the reactions, we found multiple optimal atom mappings. We show that the error rate is 0.9% (22 reactions) by comparing these atom mappings to 2446 atom mappings of the manually curated Kyoto Encyclopedia of Genes and Genomes (KEGG) RPAIR database. To our knowledge, our computational atom-mapping approach is the most accurate and among the fastest published to date. The atom-mapping data will be available in the MetaCyc database later in 2012; the atom-mapping software will be available within the Pathway Tools software later in 2012.

Biochemical Removal of HAP Precursors From Coal

Energy Technology Data Exchange (ETDEWEB)

Olson, G.; Tucker, L.; Richards, J.

1997-07-01

This project addresses DOE`s interest in advanced concepts for controlling emissions of air toxics from coal-fired utility boilers. We are determining the feasibility of developing a biochemical process for the precombustion removal of substantial percentages of 13 inorganic hazardous air pollutant (HAP) precursors from coal. These HAP precursors are Sb, As, Be, Cd, Cr, Cl, Co, F, Pb, Hg, Mn, Ni, and Se. Although rapid physical coal cleaning is done routinely in preparation plants, biochemical processes for removal of HAP precursors from coal potentially offer advantages of deeper cleaning, more specificity, and less coal loss. Compared to chemical processes for coal cleaning, biochemical processes potentially offer lower costs and milder process conditions. Pyrite oxidizing bacteria, most notably Thiobacillusferrooxidans, are being evaluated in this project for their ability to remove HAP precursors from U.S. coals.

Biochemical Removal of HAP Precursors From Coal

International Nuclear Information System (INIS)

Olson, G.; Tucker, L.; Richards, J.

1997-07-01

This project addresses DOE's interest in advanced concepts for controlling emissions of air toxics from coal-fired utility boilers. We are determining the feasibility of developing a biochemical process for the precombustion removal of substantial percentages of 13 inorganic hazardous air pollutant (HAP) precursors from coal. These HAP precursors are Sb, As, Be, Cd, Cr, Cl, Co, F, Pb, Hg, Mn, Ni, and Se. Although rapid physical coal cleaning is done routinely in preparation plants, biochemical processes for removal of HAP precursors from coal potentially offer advantages of deeper cleaning, more specificity, and less coal loss. Compared to chemical processes for coal cleaning, biochemical processes potentially offer lower costs and milder process conditions. Pyrite oxidizing bacteria, most notably Thiobacillusferrooxidans, are being evaluated in this project for their ability to remove HAP precursors from U.S. coals

Derivation, Characterization, and Neural Differentiation of Integration-Free Induced Pluripotent Stem Cell Lines from Parkinson's Disease Patients Carrying SNCA, LRRK2, PARK2, and GBA Mutations

DEFF Research Database (Denmark)

Momcilovic, Olga; Sivapatham, Renuka; Oron, Tal Ronnen

2016-01-01

We report generation of induced pluripotent stem cell (iPSC) lines from ten Parkinson's disease (PD) patients carrying SNCA, PARK2, LRRK2, and GBA mutations, and one age-matched control. After validation of pluripotency, long-term genome stability, and integration-free reprogramming, eight...... not be sufficient to determine the cause or mechanism of the disease, and highlights the need to use more focused strategies for large-scale data analysis........ We further examined gene expression in a stress model (MPTP-induced dopaminergic neuronal death) using two clones from the SNCA triplication line, and detected changes in genes associated with mitophagy. Our data suggested that even a well-characterized line of a monogenic disease may...

Biochemical and biophysical characterization of cell-free synthesized Rift Valley fever virus nucleoprotein capsids enables in vitro screening to identify novel antivirals.

Science.gov (United States)

Broce, Sean; Hensley, Lisa; Sato, Tomoharu; Lehrer-Graiwer, Joshua; Essrich, Christian; Edwards, Katie J; Pajda, Jacqueline; Davis, Christopher J; Bhadresh, Rami; Hurt, Clarence R; Freeman, Beverly; Lingappa, Vishwanath R; Kelleher, Colm A; Karpuj, Marcela V

2016-05-14

Viral capsid assembly involves the oligomerization of the capsid nucleoprotein (NP), which is an essential step in viral replication and may represent a potential antiviral target. An in vitro transcription-translation reaction using a wheat germ (WG) extract in combination with a sandwich ELISA assay has recently been used to identify small molecules with antiviral activity against the rabies virus. Here, we examined the application of this system to viruses with capsids with a different structure, such as the Rift Valley fever virus (RVFV), the etiological agent of a severe emerging infectious disease. The biochemical and immunological characterization of the in vitro-generated RVFV NP assembly products enabled the distinction between intermediately and highly ordered capsid structures. This distinction was used to establish a screening method for the identification of potential antiviral drugs for RVFV countermeasures. These results indicated that this unique analytical system, which combines nucleoprotein oligomerization with the specific immune recognition of a highly ordered capsid structure, can be extended to various viral families and used both to study the early stages of NP assembly and to assist in the identification of potential antiviral drugs in a cost-efficient manner. Reviewed by Jeffry Skolnick and Noah Isakov. For the full reviews please go to the Reviewers' comments section.

Linear rotary optical delay lines

Science.gov (United States)

Guerboukha, Hichem; Qu, Hang; Skorobogatiy, Maksim

2016-03-01

We present a semi-analytical solution for the design of a high-speed rotary optical delay line that use a combination of two rotating curvilinear reflectors. We demonstrate that it is possible to design an infinite variety of the optical delay lines featuring linear dependence of the optical delay on the rotation angle. This is achieved via shape optimization of the rotating reflector surfaces. Moreover, a convenient spatial separation of the incoming and outgoing beams is possible. For the sake of example, we present blades that fit into a circle of 10cm diameter. Finally, a prototype of a rotary delay line is fabricated using CNC machining, and its optical properties are characterized.

In vitro identification and characterization of CD133(pos cancer stem-like cells in anaplastic thyroid carcinoma cell lines.

Directory of Open Access Journals (Sweden)

Giovanni Zito

Full Text Available Recent publications suggest that neoplastic initiation and growth are dependent on a small subset of cells, termed cancer stem cells (CSCs. Anaplastic Thyroid Carcinoma (ATC is a very aggressive solid tumor with poor prognosis, characterized by high dedifferentiation. The existence of CSCs might account for the heterogeneity of ATC lesions. CD133 has been identified as a stem cell marker for normal and cancerous tissues, although its biological function remains unknown.ATC cell lines ARO, KAT-4, KAT-18 and FRO were analyzed for CD133 expression. Flow cytometry showed CD133(pos cells only in ARO and KAT-4 (64+/-9% and 57+/-12%, respectively. These data were confirmed by qRT-PCR and immunocytochemistry. ARO and KAT-4 were also positive for fetal marker oncofetal fibronectin and negative for thyrocyte-specific differentiating markers thyroglobulin, thyroperoxidase and sodium/iodide symporter. Sorted ARO/CD133(pos cells exhibited higher proliferation, self-renewal, colony-forming ability in comparison with ARO/CD133(neg. Furthermore, ARO/CD133(pos showed levels of thyroid transcription factor TTF-1 similar to the fetal thyroid cell line TAD-2, while the expression in ARO/CD133(neg was negligible. The expression of the stem cell marker OCT-4 detected by RT-PCR and flow cytometry was markedly higher in ARO/CD133(pos in comparison to ARO/CD133(neg cells. The stem cell markers c-KIT and THY-1 were negative. Sensitivity to chemotherapy agents was investigated, showing remarkable resistance to chemotherapy-induced apoptosis in ARO/CD133(pos when compared with ARO/CD133(neg cells.We describe CD133(pos cells in ATC cell lines. ARO/CD133(pos cells exhibit stem cell-like features--such as high proliferation, self-renewal ability, expression of OCT-4--and are characterized by higher resistance to chemotherapy. The simultaneous positivity for thyroid specific factor TTF-1 and onfFN suggest they might represent putative thyroid cancer stem-like cells. Our in

Structuring of conductive silver line by electrohydrodynamic jet printing and its electrical characterization

International Nuclear Information System (INIS)

Lee, Dae-Young; Lee, Jae-Chang; Shin, Yun-Soo; Park, Sung-Eun; Kim, Yong-Jun; Hwang, Jungho; Yu, Tae-U

2008-01-01

A set of silver lines with a few hundred nanometers in thickness and with a few hundred micrometers in width were obtained using the electrohydrodynamic jet printing. The lines exhibited about three times higher resistivity (4.8 μΩcm) than that of bulk silver after thermal sintering process. The characteristic impedance of the silver line was about 18 Ω while the value calculated was 20 Ω. This paper demonstrated the possibility of using electrohydrodynamic jet printing of silver nanoparticles to obtain conductive line onto circuit boards.

Purification and biochemical characterization of an Aspergillus niger phytase produced by solid-state fermentation using triticale residues as substrate.

Science.gov (United States)

Neira-Vielma, Alberto A; Aguilar, Cristóbal N; Ilyina, Anna; Contreras-Esquivel, Juan C; Carneiro-da-Cunha, María das Graça; Michelena-Álvarez, Georgina; Martínez-Hernández, José L

2018-03-01

In this study, an extracellular phytase produced by Aspergillus niger 7A-1, was biochemically characterized for possible industrial application. The enzyme was purified from a crude extract obtained by solid-state fermentation (SSF) of triticale waste. The extract was obtained by microfiltration, ultrafiltration (300, 100 and 30 kDa) and DEAE-Sepharose column chromatography. The molecular weight of the purified enzyme was estimated to be 89 kDa by SDS-PAGE. The purified enzyme was most active at pH 5.3 and 56 °C, and retained 50% activity over a wide pH range of 4 to 7. The enzymatic thermostability assay showed that the enzyme retained more than 70% activity at 80 °C for 60 s, 40% activity for 120 s and 9% after 300 s. The phytase showed broad substrate specificity, a K m value of 220 μM and V max of 25 μM/min. The purified phytase retained 50% of its activity with phosphorylated compounds such as phenyl phosphate, 1-Naphthyl phosphate, 2-Naphthyl phosphate, p-Nitrophenyl phosphate and Glycerol-2-phosphate. The inhibition of phytase activity by metal ions was observed to be drastically inhibited (50%) by Ca ++ and was slightly inhibited (10%) by Ni ++ , K + , and Na + , at 10 and 20 mM concentrations. A positive effect was obtained with Mg ++ , Mn ++ , Cu ++ , Cd ++ and Ba ++ at 25 and 35% with stimulatory effect on the phytase activity.

[Clinical, biochemical and allergological indices characterizing occupational diseases of the bronchial and pulmonary system in employees at aluminium production].

Science.gov (United States)

Kudaeva, I V; Dyakovich, O A; Beygel, E A; Masnavieva, L B; Naumova, O V; Budarina, L A

There are many harmful factors that possess a damaging impact on the body of employees at aluminum production. It leads to the development of bronchial asthma (BA), chronic nonobstructive bronchitis (CNB) and chronic obstructive pulmonary disease (COPD). The pathogenesis of these disorders, as well as sensitizing effect offluorine in the aluminum production is not fully understood. The purpose of this work was to study the characteristics of laboratory indices in patients with occupational diseases of the respiratory system. In workers of aluminum production with the diagnosis of occupational diseases of respiratory system (15 patients with a diagnosis of asthma, 30 CNB cases, 20 COPD patients) we evaluated the content of total protein, total cholesterol, high density lipoprotein cholesterol (HDLC), total calcium, phosphorus, ceruloplasmin, hematological indices and performed emigration of leukocytes braking test (TTEEL). Clinical and biochemical profile ofpersons with occupational asthma was characterized by a low level of total calcium and ceruloplasmin, a high concentration of phosphorus in the blood serum and inhibition of leukocyte emigration in the test with sodium fluoride. For aluminum production CNB workers characteristic active proatherogenic process was pronounced by a decrease in the HDLC level and an increase in atherogenic index; higher hematocrit value and concentration of erythrocytes, and more than 50% of cases of sensitization to the presence of sodium fluoride. COPD cases had occupational lower average concentration of hemoglobin in the erythrocyte, total protein in serum, as well as polymorphic variant response to sodium fluoride in the form of a depression and activation of leucocytes emigration.

Purification and biochemical characterization of an Aspergillus niger phytase produced by solid-state fermentation using triticale residues as substrate

Directory of Open Access Journals (Sweden)

Alberto A. Neira-Vielma

2018-03-01

Full Text Available In this study, an extracellular phytase produced by Aspergillus niger 7A-1, was biochemically characterized for possible industrial application. The enzyme was purified from a crude extract obtained by solid-state fermentation (SSF of triticale waste. The extract was obtained by microfiltration, ultrafiltration (300, 100 and 30 kDa and DEAE-Sepharose column chromatography. The molecular weight of the purified enzyme was estimated to be 89 kDa by SDS-PAGE. The purified enzyme was most active at pH 5.3 and 56 °C, and retained 50% activity over a wide pH range of 4 to 7. The enzymatic thermostability assay showed that the enzyme retained more than 70% activity at 80 °C for 60 s, 40% activity for 120 s and 9% after 300 s. The phytase showed broad substrate specificity, a Km value of 220 μM and Vmax of 25 μM/min. The purified phytase retained 50% of its activity with phosphorylated compounds such as phenyl phosphate, 1-Naphthyl phosphate, 2-Naphthyl phosphate, p-Nitrophenyl phosphate and Glycerol-2-phosphate. The inhibition of phytase activity by metal ions was observed to be drastically inhibited (50% by Ca++ and was slightly inhibited (10% by Ni++, K+, and Na+, at 10 and 20 mM concentrations. A positive effect was obtained with Mg++, Mn++, Cu++, Cd++ and Ba++ at 25 and 35% with stimulatory effect on the phytase activity.

Physiological response and productivity of safflower lines under water deficit and rehydration.

Science.gov (United States)

Bortolheiro, Fernanda P A P; Silva, Marcelo A

2017-01-01

Water deficit is one of the major stresses affecting plant growth and productivity worldwide. Plants induce various morphological, physiological, biochemical and molecular changes to adapt to the changing environment. Safflower (Carthamus tinctorius L.), a potential oil producer, is highly adaptable to various environmental conditions, such as lack of rainfall and temperatures. The objective of this work was to study the physiological and production characteristics of six safflower lines in response to water deficit followed by rehydration. The experiment was conducted in a protected environment and consisted of 30 days of water deficit followed by 18 days of rehydration. A differential response in terms of photosynthetic pigments, electrolyte leakage, water potential, relative water content, grain yield, oil content, oil yield and water use efficiency was observed in the six lines under water stress. Lines IMA 04, IMA 10, IMA 14 showed physiological characteristics of drought tolerance, with IMA 14 and IMA 16 being the most productive after water deficit. IMA 02 and IMA 21 lines displayed intermediate characteristics of drought tolerance. It was concluded that the lines responded differently to water deficit stress, showing considerable genetic variation and influence to the environment.

Physiological response and productivity of safflower lines under water deficit and rehydration

Directory of Open Access Journals (Sweden)

FERNANDA P.A.P. BORTOLHEIRO

2017-12-01

Full Text Available ABSTRACT Water deficit is one of the major stresses affecting plant growth and productivity worldwide. Plants induce various morphological, physiological, biochemical and molecular changes to adapt to the changing environment. Safflower (Carthamus tinctorius L., a potential oil producer, is highly adaptable to various environmental conditions, such as lack of rainfall and temperatures. The objective of this work was to study the physiological and production characteristics of six safflower lines in response to water deficit followed by rehydration. The experiment was conducted in a protected environment and consisted of 30 days of water deficit followed by 18 days of rehydration. A differential response in terms of photosynthetic pigments, electrolyte leakage, water potential, relative water content, grain yield, oil content, oil yield and water use efficiency was observed in the six lines under water stress. Lines IMA 04, IMA 10, IMA 14 showed physiological characteristics of drought tolerance, with IMA 14 and IMA 16 being the most productive after water deficit. IMA 02 and IMA 21 lines displayed intermediate characteristics of drought tolerance. It was concluded that the lines responded differently to water deficit stress, showing considerable genetic variation and influence to the environment.

Developments in commercially produced microbials at Biochem Products

Science.gov (United States)

John Lublinkhof; Douglas H. Ross

1985-01-01

Biochem Products is part of a large industrial and scientific family - the Solvay Group. Solvay, headquartered in Brussels, Belgium is a multinational company with 46,000 employees worldwide. In the U.S., our working partners include a large polymer manufacturer, a peroxygen producer and a leading poultry and animal health products company. Biochem Products is a...

Kombucha tea fermentation: Microbial and biochemical dynamics.

Science.gov (United States)

Chakravorty, Somnath; Bhattacharya, Semantee; Chatzinotas, Antonis; Chakraborty, Writachit; Bhattacharya, Debanjana; Gachhui, Ratan

2016-03-02

Kombucha tea, a non-alcoholic beverage, is acquiring significant interest due to its claimed beneficial properties. The microbial community of Kombucha tea consists of bacteria and yeast which thrive in two mutually non-exclusive compartments: the soup or the beverage and the biofilm floating on it. The microbial community and the biochemical properties of the beverage have so far mostly been described in separate studies. This, however, may prevent understanding the causal links between the microbial communities and the beneficial properties of Kombucha tea. Moreover, an extensive study into the microbial and biochemical dynamics has also been missing. In this study, we thus explored the structure and dynamics of the microbial community along with the biochemical properties of Kombucha tea at different time points up to 21 days of fermentation. We hypothesized that several biochemical properties will change during the course of fermentation along with the shifts in the yeast and bacterial communities. The yeast community of the biofilm did not show much variation over time and was dominated by Candida sp. (73.5-83%). The soup however, showed a significant shift in dominance from Candida sp. to Lachancea sp. on the 7th day of fermentation. This is the first report showing Candida as the most dominating yeast genus during Kombucha fermentation. Komagateibacter was identified as the single largest bacterial genus present in both the biofilm and the soup (~50%). The bacterial diversity was higher in the soup than in the biofilm with a peak on the seventh day of fermentation. The biochemical properties changed with the progression of the fermentation, i.e., beneficial properties of the beverage such as the radical scavenging ability increased significantly with a maximum increase at day 7. We further observed a significantly higher D-saccharic acid-1,4-lactone content and caffeine degradation property compared to previously described Kombucha tea fermentations. Our

Biochemical characterization of nuclear receptors for vitamin D{sub 3} and glucocorticoids in prostate stroma cell microenvironment

Energy Technology Data Exchange (ETDEWEB)

Hidalgo, Alejandro A. [Laboratory of Molecular Endocrinology, Department of Physiopathology, University of Concepcion, Concepcion (Chile); Department of Molecular Pharmacology and Therapeutics, NY (United States); Montecinos, Viviana P.; Paredes, Roberto; Godoy, Alejandro S.; McNerney, Eileen M.; Tovar, Heribelt; Pantoja, Diego [Laboratory of Molecular Endocrinology, Department of Physiopathology, University of Concepcion, Concepcion (Chile); Johnson, Candace [Department of Molecular Pharmacology and Therapeutics, NY (United States); Trump, Donald [Department of Medicine, Roswell Park Cancer Institute, Buffalo, NY (United States); Onate, Sergio A., E-mail: sergio.onate@udec.cl [Laboratory of Molecular Endocrinology, Department of Physiopathology, University of Concepcion, Concepcion (Chile); Department of Urology, State University of New York at Buffalo, NY (United States)

2011-08-19

Highlights: {yields} Fibroblasts from benign and carcinoma-associated stroma were biochemically characterized for VDR and GR function as transcription factors in prostate stroma cell microenvironment. {yields} Decreased SRC-1/CBP coactivators recruitment to VDR and GR may result in hormone resistance to 1,25D{sub 3} in stromal cell microenvironment prostate cancer. {yields} 1a,25-Dyhidroxyvitamin D{sub 3} (1,25D{sub 3}) and glucocorticoids, either alone or in combination, may not be an alternative for 'some' advanced prostate cancers that fails androgen therapies. -- Abstract: The disruption of stromal cell signals in prostate tissue microenvironment influences the development of prostate cancer to androgen independence. 1{alpha},25-Dihydroxyvitamin D{sub 3} (1,25D{sub 3}) and glucocorticoids, either alone or in combination, have been investigated as alternatives for the treatment of advanced prostate cancers that fails androgen therapies. The effects of glucocorticoids are mediated by the intracellular glucocorticoid receptor (GR). Similarly, the effect of 1,25D{sub 3} is mediated by the 1,25D{sub 3} nuclear receptor (VDR). In this study, fibroblasts from benign- (BAS) and carcinoma-associated stroma (CAS) were isolated from human prostates to characterize VDR and GR function as transcription factors in prostate stroma. The VDR-mediated transcriptional activity assessed using the CYP24-luciferase reporter was limited to 3-fold induction by 1,25D{sub 3} in 9 out of 13 CAS (70%), as compared to >10-fold induction in the BAS clinical sample pair. Expression of His-tagged VDR (Ad-his-VDR) failed to recover the low transcriptional activity of the luciferase reporter in 7 out of 9 CAS. Interestingly, expression of Ad-his-VDR successfully recovered receptor-mediated induction in 2 out of the 9 CAS analyzed, suggesting that changes in the receptor protein itself was responsible for decreased response and resistance to 1,25D{sub 3} action. Conversely, VDR

29-34 Yellow Rust Resistance in Advanced Lines and Commercial ...

African Journals Online (AJOL)

rust pathogen. The objective of this study was to characterize seedling yellow rust resistance in 21 advanced bread wheat lines and 20 cultivars from Ethiopia. Yellow rust infection types (ITs) produced on test wheat lines and cultivars from nine yellow rust races were compared with ITs produced on standard differential lines ...

Biochemical markers of bone turnover

International Nuclear Information System (INIS)

Kim, Deog Yoon

1999-01-01

Biochemical markers of bone turnover has received increasing attention over the past few years, because of the need for sensitivity and specific tool in the clinical investigation of osteoporosis. Bone markers should be unique to bone, reflect changes of bone less, and should be correlated with radiocalcium kinetics, histomorphometry, or changes in bone mass. The markers also should be useful in monitoring treatment efficacy. Although no bone marker has been established to meet all these criteria, currently osteocalcin and pyridinium crosslinks are the most efficient markers to assess the level of bone turnover in the menopausal and senile osteoporosis. Recently, N-terminal telopeptide (NTX), C-terminal telopeptide (CTX) and bone specific alkaline phosphatase are considered as new valid markers of bone turnover. Recent data suggest that CTX and free deoxypyridinoline could predict the subsequent risk of hip fracture of elderly women. Treatment of postmenopausal women with estrogen, calcitonin and bisphosphonates demonstrated rapid decrease of the levels of bone markers that correlated with the long-term increase of bone mass. Factors such as circadian rhythms, diet, age, sex, bone mass and renal function affect the results of biochemical markers and should be appropriately adjusted whenever possible. Each biochemical markers of bone turnover may have its own specific advantages and limitations. Recent advances in research will provide more sensitive and specific assays

Treatment of landfill leachate biochemical effluent using the nano-Fe3O4/Na2S2O8 system: Oxidation performance, wastewater spectral analysis, and activator characterization.

Science.gov (United States)

Liu, Zhanmeng; Li, Xian; Rao, Zhiwei; Hu, Fengping

2018-02-15

Nano-Fe 3 O 4 was used as heterogeneous catalyst to activate Na 2 S 2 O 8 for the generation of the sulfate radicals (SO 4 - ) to oxidize the residual pollutants in landfill leachate biochemical effluent. The oxidation performance, wastewater spectral analysis and activator characterization were discussed. Oxidation experimental result shows that nano-Fe 3 O 4 has obvious catalytic effect on Na 2 S 2 O 8 and can significantly enhance the oxidation efficiencies of Na 2 S 2 O 8 on landfill leachate biochemical effluent, with COD and color removals above 63% and 95%, respectively. Based on the analyses of three-dimensional excitation emission matrix fluorescence spectrum (3DEEM), ultraviolet-visible spectra (UV-vis), and Fourier Transform infrared spectroscopy (FTIR) of wastewater samples before and after treatment, it can be concluded that the pollution level of dissolved organic matter (DOM) declined and that the humic acid (HA) fractions were efficiently degraded into small molecules of fulvic acid (FA) fractions with less weight and stable structure. Compared to the raw wastewater sample, the aromaticity and substituent groups of the DOM were lessened in the treated wastewater sample. Moreover, the main structure of the organics and functional groups were changed by the Fe 3 O 4 /Na 2 S 2 O 8 system, with substantial decrease of conjugated double bonds. The micro morphology of nano-Fe 3 O 4 was characterized before and after reaction by the methods of scanning electron microscope spectra (SEM), X-ray diffraction pattern (XRD), and X-ray photoelectron spectroscopy (XPS). The XRD pattern analysis showed that nano-Fe 3 O 4 was oxidized into r-Fe 2 O 3 and that the particle size of it also became smaller after reaction. XPS was employed to analyze the content and iron valence on the nano-Fe 3 O 4 surface, and it can be found that the ratio of Fe 3+ /Fe 2+ decreased from 1.8 before reaction to 0.8 after reaction. From the SEM analysis after the treatment, it was

Improvement of fetus growth restriction diagnostics in pregnant women by means of biochemical markers that characterize the disorder of stress-adaptation

Directory of Open Access Journals (Sweden)

N. G. Kolokot

2018-04-01

Full Text Available Object of the work. Regulatory and adaptive processes of the system “mother –placenta – fetus” state determination and detection of pregnant women adaptive capabilities specific features in case of fetus growth restriction based on a number of the blood plasma biochemical indicators. Materials and methods. Markers of oxidative stress were detected spectrophotometrically in the blood plasma according to generally accepted methods: oxidative modification of proteins, stable metabolites of nitrogen oxide, L-arginine, malondialdehyde, thiol compounds and reduced glutathione. Statistical processing of data was made by methods of variational statistics with Microsoft Office Excel 2010, Statistica 6.0. Results. The use applicability of the number of biochemical markers which are predictors of perinatal complications has been scientifically substantiated. Using biochemical diagnostic technique it has been shown that fetus growth restriction syndrome is accompanied by disorders of regulatory and adaptive processes of the system “mother –placenta – fetus”. In particular, the level of proteins and lipids oxidative modification increases, nitric oxide synthase activity decreases and reserves of L-arginine and thiol compounds decrease in the blood plasma. Conclusions. Comparative analysis of a number of biochemical markers determination results of stress-realizing system activity in pregnant women with fetus growth restriction and in women with a physiological pregnancy has revealed a significant (P ≤ 0.05 increase in markers of protein oxidative modification blood levels (the level of dinitrophenylhydrazones of aliphatic aldehyd of basic amino acid residues and dinitrophenylhydrazones of carbonyl compounds of basic amino acid residues in the spontaneous sample increased by 14.2 % and 16.3 %, respectively, and in the stimulated sample by 46.6 % and 43.0 %, the level of malondialdehyde increased by 42.9 %. Reduction of the stress

Overview of the most commonly used methods in allergen characterization

Directory of Open Access Journals (Sweden)

TANJA CIRKOVIC VELICKOVIC

2005-03-01

Full Text Available The characterization of an allergen is a troublesome and difficult process, as it requires both the precise biochemical characterization of a (glycoprotein molecule and the establishment of its susceptibility to IgE antibodies, as they are themain link to histamine release in some hypersensitivity states (type I allergies. As the characterization of an allergen includes molecular weight determination of the allergenic molecule, its structure determination, physicochemical properties, IgE binding properties of the allergen molecule, and its allergenicity, an overal review of which biochemical and immunochemical methods are used in achieving this goal are presented in this paper. The information on the molecular level on the stuctures of allergens indicates that allergens are considerably heterogeneous protein structures, and that there is no particular aminoacid sequence which is responsible for the allergenicity. Therefore, information gained from detailed structural, functional and immunochemical studies of these intriguing molecules, which nowadaysmodulate a variety of pathophysiological conditions, would greatly improve our understanding of the underlying disease mechanisms, and the way to handle them.

Purification and partial biochemical characterization of polyphenol oxidase from mango (Mangifera indica cv. Manila).

Science.gov (United States)

Palma-Orozco, Gisela; Marrufo-Hernández, Norma A; Sampedro, José G; Nájera, Hugo

2014-10-08

Polyphenol oxidase (PPO) is an enzyme widely distributed in the plant kingdom that has been detected in most fruits and vegetables. PPO was extracted and purified from Manila mango (Mangifera indica), and its biochemical properties were studied. PPO was purified 216-fold by hydrophobic interaction and ion exchange chromatography. PPO was purified to homogeneity, and the estimated PPO molecular weight (MW) by SDS-PAGE was ≈31.5 kDa. However, a MW of 65 kDa was determined by gel filtration, indicating a dimeric structure for the native PPO. The isolated PPO showed the highest affinity to pyrogallol (Km = 2.77 mM) followed by 4-methylcatechol (Km = 3.14 mM) and catechol (Km = 15.14 mM). The optimum pH for activity was 6.0. PPO was stable in the temperature range of 20-70 °C. PPO activity was completely inhibited by tropolone, ascorbic acid, sodium metabisulfite, and kojic acid at 0.1 mM.

Cell lines authentication and mycoplasma detection as minimun quality control of cell lines in biobanking.

Science.gov (United States)

Corral-Vázquez, C; Aguilar-Quesada, R; Catalina, P; Lucena-Aguilar, G; Ligero, G; Miranda, B; Carrillo-Ávila, J A

2017-06-01

Establishment of continuous cell lines from human normal and tumor tissues is an extended and useful methodology for molecular characterization of cancer pathophysiology and drug development in research laboratories. The exchange of these cell lines between different labs is a common practice that can compromise assays reliability due to contamination with microorganism such as mycoplasma or cells from different flasks that compromise experiment reproducibility and reliability. Great proportions of cell lines are contaminated with mycoplasma and/or are replaced by cells derived for a different origin during processing or distribution process. The scientific community has underestimated this problem and thousand of research experiment has been done with cell lines that are incorrectly identified and wrong scientific conclusions have been published. Regular contamination and authentication tests are necessary in order to avoid negative consequences of widespread misidentified and contaminated cell lines. Cell banks generate, store and distribute cell lines for research, being mandatory a consistent and continuous quality program. Methods implementation for guaranteeing both, the absence of mycoplasma and authentication in the supplied cell lines, has been performed in the Andalusian Health System Biobank. Specifically, precise results were obtained using real time PCR detection for mycoplasma and 10 STRs identification by capillary electrophoresis for cell line authentication. Advantages and disadvantages of these protocols are discussed.

Phenotypic variability of the kyphoscoliotic type of Ehlers-Danlos syndrome (EDS VIA): clinical, molecular and biochemical delineation

Science.gov (United States)

2011-01-01

Background The kyphoscoliotic type of Ehlers-Danlos syndrome (EDS VIA) (OMIM 225400) is a rare inheritable connective tissue disorder characterized by a deficiency of collagen lysyl hydroxylase 1 (LH1; EC 1.14.11.4) due to mutations in PLOD1. Biochemically this results in underhydroxylation of collagen lysyl residues and, hence, an abnormal pattern of lysyl pyridinoline (LP) and hydroxylysyl pyridinoline (HP) crosslinks excreted in the urine. Clinically the disorder is characterized by hypotonia and kyphoscoliosis at birth, joint hypermobility, and skin hyperelasticity and fragility. Severe hypotonia usually leads to delay in gross motor development, whereas cognitive development is reported to be normal. Methods We describe the clinical, biochemical and molecular characterisation, as well as electron microscopy findings of skin, in 15 patients newly diagnosed with this rare type of Ehlers-Danlos syndrome. Results Age at diagnosis ranged from 5 months to 27 years, with only 1/3 of the patients been diagnosed correctly in the first year of life. A similar disease frequency was found in females and males, however a broad disease severity spectrum (intra- and interfamilial), independent of molecular background or biochemical phenotype, was observed. Kyphoscoliosis, one of the main clinical features was not present at birth in 4 patients. Importantly we also noted the occurrence of vascular rupture antenatally and postnatally, as well as developmental delay in 5 patients. Conclusion In view of these findings we propose that EDS VIA is a highly variable clinical entity, presenting with a broad clinical spectrum, which may also be associated with cognitive delay and an increased risk for vascular events. Genotype/phenotype association studies and additional molecular investigations in more extended EDS VIA populations will be necessary to further elucidate the cause of the variability of the disease severity. PMID:21699693

Phenotypic variability of the kyphoscoliotic type of Ehlers-Danlos syndrome (EDS VIA: clinical, molecular and biochemical delineation

Directory of Open Access Journals (Sweden)

Kariminejad Ariana

2011-06-01

Full Text Available Abstract Background The kyphoscoliotic type of Ehlers-Danlos syndrome (EDS VIA (OMIM 225400 is a rare inheritable connective tissue disorder characterized by a deficiency of collagen lysyl hydroxylase 1 (LH1; EC 1.14.11.4 due to mutations in PLOD1. Biochemically this results in underhydroxylation of collagen lysyl residues and, hence, an abnormal pattern of lysyl pyridinoline (LP and hydroxylysyl pyridinoline (HP crosslinks excreted in the urine. Clinically the disorder is characterized by hypotonia and kyphoscoliosis at birth, joint hypermobility, and skin hyperelasticity and fragility. Severe hypotonia usually leads to delay in gross motor development, whereas cognitive development is reported to be normal. Methods We describe the clinical, biochemical and molecular characterisation, as well as electron microscopy findings of skin, in 15 patients newly diagnosed with this rare type of Ehlers-Danlos syndrome. Results Age at diagnosis ranged from 5 months to 27 years, with only 1/3 of the patients been diagnosed correctly in the first year of life. A similar disease frequency was found in females and males, however a broad disease severity spectrum (intra- and interfamilial, independent of molecular background or biochemical phenotype, was observed. Kyphoscoliosis, one of the main clinical features was not present at birth in 4 patients. Importantly we also noted the occurrence of vascular rupture antenatally and postnatally, as well as developmental delay in 5 patients. Conclusion In view of these findings we propose that EDS VIA is a highly variable clinical entity, presenting with a broad clinical spectrum, which may also be associated with cognitive delay and an increased risk for vascular events. Genotype/phenotype association studies and additional molecular investigations in more extended EDS VIA populations will be necessary to further elucidate the cause of the variability of the disease severity.

Biochemical and functional characterization of recombinant fungal immunomodulatory proteins (rFIPs)

NARCIS (Netherlands)

Bastiaan-Net, S.; Chanput, W.; Hertz, A.; Zwittink, R.D.; Mes, J.J.; Wichers, H.J.

2013-01-01

In this study two novel FIPs have been identified and characterized. The first is FIP-nha, identified in the ascomycete Nectria haematococca, and as such, FIP-nha would be the first FIP to be identified outside the order of Basidiomycota. The second is LZ-9, an LZ-8 like protein identified in

Specification, construction, and exact reduction of state transition system models of biochemical processes.

Science.gov (United States)

Bugenhagen, Scott M; Beard, Daniel A

2012-10-21

Biochemical reaction systems may be viewed as discrete event processes characterized by a number of states and state transitions. These systems may be modeled as state transition systems with transitions representing individual reaction events. Since they often involve a large number of interactions, it can be difficult to construct such a model for a system, and since the resulting state-level model can involve a huge number of states, model analysis can be difficult or impossible. Here, we describe methods for the high-level specification of a system using hypergraphs, for the automated generation of a state-level model from a high-level model, and for the exact reduction of a state-level model using information from the high-level model. Exact reduction is achieved through the automated application to the high-level model of the symmetry reduction technique and reduction by decomposition by independent subsystems, allowing potentially significant reductions without the need to generate a full model. The application of the method to biochemical reaction systems is illustrated by models describing a hypothetical ion-channel at several levels of complexity. The method allows for the reduction of the otherwise intractable example models to a manageable size.

On line ultrasonic integrated backscatter

International Nuclear Information System (INIS)

Landini, L.; Picano, E.; Mazzarisi, A.; Santarelli, F.; Benassi, A.; De Pieri, G.

1988-01-01

A new equipment for on-line evaluation of index based on two-dimensional integrated backscatter from ultrasonic images is described. The new equipment is fully integrated into a B-mode ultrasonic apparatus which provides a simultaneous display of conventional information together with parameters of tissue characterization. The system has been tested with a backscattering model of microbubbles in polysaccharide solution, characterized by a physiological exponential time decay. An exponential fitting to the experimental data was performed which yielded r=0.95

Ultrastructural and biochemical studies on formation of calcium oxalate in plants

International Nuclear Information System (INIS)

Abdelmottaleb, A.M.

1989-01-01

Plant calcium oxalate crystals occur within cells called crystal idioblasts. Important aspects of this calcification phenomenon have not been characterized. This dissertation examines some of the aspects of this ubiquitous type of calcification including (1) characterization of ultrastructural features of developing crystal idioblasts, (2) determination of the relationship of specialized ultrastructural features of the idioblasts to transport of compounds and mechanisms of crystal deposition, and (3) the biochemical relationship between ascorbic acid metabolism and production of oxalic acid used for crystal formation. Structural and cytochemical studies revealed that crystal idioblasts have dense cytoplasm, modified plastids, enlarged nuclei, extensive endoplasmic reticulum, numerous dictyosomes and vesicles, and a bundle of raphide crystals in their vacuoles. A mechanism for Ca transport and crystal precipitation is proposed, based on these results. There is a strong and dynamic relationship between Ca concentration and oxalic acid produced for crystal formation, where increasing Ca level in the growth medium lead to increased total and insoluble oxalate in the plant. Calmodulin antagonists reduced oxalic acid production

Measures of Biochemical Sociology

Science.gov (United States)

Snell, Joel; Marsh, Mitchell

2008-01-01

In a previous article, the authors introduced a new sub field in sociology that we labeled "biochemical sociology." We introduced the definition of a sociology that encompasses sociological measures, psychological measures, and biological indicators Snell & Marsh (2003). In this article, we want to demonstrate a research strategy that would assess…

Characterization of fibroblast growth factor receptor 2 overexpression in the human breast cancer cell line SUM-52PE

International Nuclear Information System (INIS)

Tannheimer, Stacey L; Rehemtulla, Alnawaz; Ethier, Stephen P

2000-01-01

three times more transformed foci in soft agar than C1 variants (both IIIb), whereas full length FGFR2 and FGFR1 (both IIIc variants) showed no transforming activity [4]. Previous studies [5,6] have found amplification and overexpression of FGFR2 in 5-10% of primary breast cancer specimens. A recent study [7] done using a tissue array consisting of 372 primary breast cancer specimens found a 5% incidence of FGFR2 amplification. To our knowledge, none of the HBC cell lines studied thus far have an FGFR2 gene amplification, although overexpression of FGFR2 message and protein has been documented for some breast cancer cell lines [6,8,9]. SUM-52PE is a breast cancer cell line previously isolated in our laboratory that grows under serum-free and epidermal growth factor-free conditions, has high levels of tyrosine-phosphorylated membrane proteins, and has the capacity to invade and grow under anchorage-independent conditions [10,11,12]. This cell line exhibits all of the important hallmarks of transformed, highly malignant cells. Therefore, SUM-52PE was used as a model to study the diversity of FGFR2 expression in a breast cancer cell line that has true amplification and overexpression of FGFR2. This study was conducted to examine the degree of FGFR2 amplification and overexpression in the breast cancer cell line SUM-52PE. Subsequent sequencing and characterization of individual FGFR2 variants cloned from the SUM-52PE cell line was completed to determine the complexity of FGFR2 alternative splicing in the context of a highly metastatic breast cancer cell line. Southern, Northern and Western blot analyses were done in order to determine the degree of FGFR2 amplification and overexpression in the breast cancer cell line SUM-52PE. Individual FGFR2 variants were cloned out of SUM-52PE using FGFR2-specific primers in a reverse transcription (RT) polymerase chain reaction (PCR). FGFR2 cDNAs were characterized by restriction fragment analysis, sequencing and transient transfection

C IV LINE-WIDTH ANOMALIES

DEFF Research Database (Denmark)

Denney, Kelly Dianne; Pogge, R.W.; Assef, R.J.

2013-01-01

Comparison of six high-redshift quasar spectra obtained with the Large Binocular Telescope with previous observations from the Sloan Digital Sky Survey shows that failure to correctly identify absorption and other problems with accurate characterization of the CIV emission line profile in low S/N...

Biochemical and immunohistochemical characterization of proteins in Hürthle cell carcinoma.

Science.gov (United States)

De Keyser, L; Layfield, L; Van Herle, A; Costin, A; Lewin, K

1984-10-01

The present study reports the biochemical and immunohistochemical findings in the cytosol of a Hürthle cell carcinoma as compared with that of normal thyroid tissue. Sephadex G-200 chromatography of the extract derived from a Hürthle cell carcinoma and from normal thyroid tissue revealed three identical pools. Pool I consisted mainly of thyroglobulin (Tg), pool II corresponded to albumin, while pool III contained unidentified low molecular weight fragments which could not be studied further. Hürthle cell carcinoma, pool I, had a Tg content of 12.9 micrograms Tg/mg equivalent tissue and a 127I content of 5,6 mole/mole of Tg. Its sialic acid content was undetectable, however. In pool I of the normal thyroid gland, the respective values were 62.8 micrograms Tg/mg equivalent tissue, 21.3 +/mole 127I/mole Tg, and 15.4 mole sialic acid/mole Tg. The albumin contained in both pools II was shown to be ioidinated at the following levels: 0.025 mole 127I/mole albumin in Hürthle tumor pool II vs 1.28 mole 127I/mole albumin in normal thyroid pool II. Immunohistochemical studies confirmed the presence of Tg and albumin in the malignant Hürthle cells and acini and colloid. Thus, Hürthle cell carcinoma contained Tg and albumin. The Tg content was five times less compared with control tissue. Both proteins (Tg and albumin) were poorly iodinated in Hürthle carcinoma tissue, and the iodination of albumina seemed to be more severely impaired. The site of synthesis of both proteins could not be derived from the present studies.

Discovering Reliable Sources of Biochemical Thermodynamic Data to Aid Students' Understanding

Science.gov (United States)

Me´ndez, Eduardo; Cerda´, María F.

2016-01-01

Students of physical chemistry in biochemical disciplines need biochemical examples to capture the need, not always understood, of a difficult area in their studies. The use of thermodynamic data in the chemical reference state may lead to incorrect interpretations in the analysis of biochemical examples when the analysis does not include relevant…

Roughness of equipotential lines due to a self-affine boundary

International Nuclear Information System (INIS)

Assis, Thiago A de; Mota, Fernando de B; Miranda, Jose G V; Andrade, Roberto F S; Filho, Hugo de O Dias; Castilho, Caio M C de

2006-01-01

In this work, the characterization of the roughness of a set of equipotential lines l, due to a rough surface held at a nonzero voltage bias, is investigated. The roughness of the equipotential lines reflects the roughness of the profile, and causes a rapid variation in the electric field close to the surface. An ideal situation was considered, where a well known self-affine profile mimics the surface, while the equipotential lines are numerically evaluated using Liebmann's method. The use of an exact scale invariant profile helps to understand the dependency of the line roughness exponent α(l) on both the value of the potential (or on the average distance to the profile) and the profile's length. Results clearly support previous indications that: (a) for a system of fixed size, higher values of α characterize less corrugated lines far away from the profile; (b) for a fixed value of the potential, α decreases with the length of the profile towards the value of the boundary. This suggests that, for a system of infinite size, all equipotential lines share the same value of α

Genetic parameters for the prediction of abdominal fat traits using blood biochemical indicators in broilers.

Science.gov (United States)

Zhang, H L; Xu, Z Q; Yang, L L; Wang, Y X; Li, Y M; Dong, J Q; Zhang, X Y; Jiang, X Y; Jiang, X F; Li, H; Zhang, D X; Zhang, H

2018-02-01

1. Excessive deposition of body fat, especially abdominal fat, is detrimental in chickens and the prevention of excessive fat accumulation is an important problem. The aim of this study was to identify blood biochemical indicators that could be used as criteria to select lean Yellow-feathered chicken lines. 2. Levels of blood biochemical indicators in the fed and fasted states and the abdominal fat traits were measured in 332 Guangxi Yellow chickens. In the fed state, the genetic correlations (r g ) of triglycerides and very low density lipoprotein levels were positive for the abdominal fat traits (0.47 ≤ r g  ≤ 0.67), whereas total cholesterol, high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) showed higher negative correlations with abdominal fat traits (-0.59 ≤ r g  ≤ -0.33). Heritabilities of these blood biochemical parameters were high, varying from 0.26 to 0.60. 3. In the fasted state, HDL-C:LDL-C level was positively correlated with abdominal fat traits (0.35 ≤ r g  ≤ 0.38), but triglycerides, total cholesterol, HDL-C, LDL-C, total protein, albumin, aspartate transaminase, uric acid and creatinine levels were negatively correlated with abdominal fat traits (-0.79 ≤ r g  ≤ -0.35). The heritabilities of these 10 blood biochemical parameters were high (0.22 ≤ h 2  ≤ 0.59). 4. In the fed state, optimal multiple regression models were constructed to predict abdominal fat traits by using triglycerides and LDL-C. In the fasted state, triglycerides, total cholesterol, HDL-C, LDL-C, total protein, albumin and uric acid could be used to predict abdominal fat content. 5. It was concluded that these models in both nutritional states could be used to predict abdominal fat content in Guangxi Yellow broiler chickens.

'BioNessie(G) - a grid enabled biochemical networks simulation environment

OpenAIRE

Liu, X; Jiang, J; Ajayi, O; Gu, X; Gilbert, D; Sinnott, R

2008-01-01

The simulation of biochemical networks provides insight and understanding about the underlying biochemical processes and pathways used by cells and organisms. BioNessie is a biochemical network simulator which has been developed at the University of Glasgow. This paper describes the simulator and focuses in particular on how it has been extended to benefit from a wide variety of high performance compute resources across the UK through Grid technologies to support larger scal...

Dipeptidyl peptidase IV in two human glioma cell lines

Directory of Open Access Journals (Sweden)

A Sedo

2009-12-01

Full Text Available There is growing evidence that dipeptidyl peptidase IV [DPP-IV, EC 3.4.14.5] takes part in the metabolism of biologically active peptides participating in the regulation of growth and transformation of glial cells. However, the knowledge on the DPP-IV expression in human glial and glioma cells is still very limited. In this study, using histochemical and biochemical techniques, the DPP-IV activity was demonstrated in two commercially available human glioma cell lines of different transformation degree, as represented by U373 astrocytoma (Grade III and U87 glioblastoma multiforme (Grade IV lines. Higher total activity of the enzyme, as well as its preferential localisation in the plasma membrane, was observed in U87 cells. Compared to U373 population, U87 cells were morphologically more pleiomorphic, they were cycling at lower rate and expressing less Glial Fibrillary Acidic Protein. The data revealed positive correlation between the degree of transformation of cells and activity of DPP-IV. Great difference in expression of this enzyme, together with the phenotypic differences of cells, makes these lines a suitable standard model for further 57 studies of function of this enzyme in human glioma cells.

Isolation and characterization of β-glucosidase producing bacteria ...

African Journals Online (AJOL)

glucosidase activity. This activity was tested by growth in medium supplemented with esculin and ferric ammonium citrate. The esculin positive strains from both the sources were characterized biochemically and checked for their ability to transform ginsenoside Rb1. The growth medium and the pH for maximum growth were ...

Introgression of leginsulin, a cysteine-rich protein, and high-protein trait from an Asian soybean plant introduction genotype into a North American experimental soybean line.

Science.gov (United States)

Krishnan, Hari B; Kim, Won-Seok; Oehrle, Nathan W; Alaswad, Alaa A; Baxter, Ivan; Wiebold, William J; Nelson, Randall L

2015-03-25

Soybean is an important protein source for both humans and animals. However, soybean proteins are relatively poor in the sulfur-containing amino acids, cysteine and methionine. Improving the content of endogenous proteins rich in sulfur-containing amino acids could enhance the nutritive value of soybean meal. Leginsulin, a cysteine-rich peptide, predominantly accumulates in Asian soybean accessions but not in most North American cultivars. By screening diverse soybean accessions from the USDA Soybean Germplasm Collection, we were able to identify one plant introduction, PI 427138, as a high-protein line with relatively high amounts of both elemental sulfur and leginsulin. We introgressed these desirable traits from PI 427138 into an experimental line with the aim of improving the overall protein content and quality of seed proteins. Biochemical characterization of inbred progenies from the cross of LD00-3309 with PI 427138 grown at six locations revealed stable ingression of high protein, high elemental sulfur, and high leginsulin accumulation. Comparison of soybean seed proteins resolved by high-resolution 2-D gel electrophoresis in combination with Delta2D image analysis software revealed preferential accumulation of a few glycinin subunits contributed to the increased protein content in the introgressed lines. Amino acid analysis revealed that even though the leginsulin introgressed lines had higher protein, leginsulin, and elemental sulfur, the overall concentration of sulfur-containing amino acids was not significantly altered when compared with the parental lines. The experimental soybean lines developed during this study (Leg-3, Leg-7, and Leg-8) lack A5, A4, and B3 glycinin subunits and could be utilized in breeding programs to develop high-quality tofu cultivars.

Physicochemical and biochemical characterization of non-biodegradable cellulose in Miocene gymnosperm wood from the Entre-Sambre-et-Meuse, Southern Belgium

Energy Technology Data Exchange (ETDEWEB)

Lechien, Valerie; Rodriguez, Christian; Ongena, Marc; Hiligsmann, Serge; Thonart, Philippe [Liege Univ., Walloon Center of Industrial Biology, Gembloux (Belgium); Rulmont, Andre [Liege Univ., Chemistry Dept., Liege (Belgium)

2006-11-15

Specimens of Miocene fossil wood from the Entre-Sambre-et-Meuse karsts (southern Belgium) were examined using physicochemical and biochemical techniques in order to understand the reasons for the exceptional preservation of these fossilized remains after 15 million years. Structural and chemical changes were assessed by comparing the structural features of the fossil samples with those of their modern counterpart, Metasequoia. Solid state {sup 13} C nuclear magnetic resonance (NMR) and microscopic analysis showed good preservation of the cellulose structure in the fossil wood from the Florennes peat deposit. Despite the substantial cellulose fraction available in the fossil tissue, an enzymatic degradation test and a biochemical methane potential assay showed that the fossil cellulose could not be degraded by cellulases and anaerobic microorganisms usually involved in the biodegradation of organic matter. Moreover, the cellulose structure (crystallinity and surface area) seemed to have no effect on cellulose biodegradability in these Miocene fossil wood samples. On the basis of our observations, we suggest that the presence of a modified lignin structure could greatly influence cellulose preservation/biodegradability. (Author)

Exploring basic biochemical constituents in the body tissues of ...

African Journals Online (AJOL)

Feeding regime did not influence susceptibility to mass loss during export. Animal age influenced the biochemical composition and export performance of abalone. Keywords: abalone; aquaculture; feeds; Haliotis midae; live export; mass loss; tissue biochemical constituents. African Journal of Marine Science 2010, 32(1): ...

In-line quantification and characterization of membrane fouling

KAUST Repository

Bucs, Szilard; Hekkert, Sacco Te Lintel; Staal, Marc Jaap; Vrouwenvelder, Johannes S.

2016-01-01

Methods of detecting, quantifying and/or characterizing the fouling of a device from a combination of pressure and spectroscopic data are provided. The device can be any device containing components susceptible to fouling. Components can include

Genetic and biochemical basis of Gall Midge resistance in some cultivars of Indica Rice. Final report for the period 1 October 1980 - 30 November 1986

International Nuclear Information System (INIS)

Reddy, G.M.

1986-01-01

The stability of high productivity of modern rice varieties is greatly affected by insect pests. Rice gall midge is a serious insect pest of rice that is prevalent in several south eastern asian countries. Gall midge resistance has been mainly attributed to antibiosis. No progress has so far been made in identifying the exact biochemical nature of resistance. In Indica subspecies the understanding of chemical nature of disease would be helpful in the control of the disease and also in breeding programme aimed at developing resistance varieties. Studies were undertaken to establish the biochemical basis of resistance. Biochemical characterization of resistant and susceptible varieties were carried out. The parameters considered were: total sugar and reducing sugar content, total phenol content, amino acid profile, post infectional changes in sugar and phenol, isozyme studies. 2 figs, 6 tabs

Genetic and biochemical basis of Gall Midge resistance in some cultivars of Indica Rice. Final report for the period 1 October 1980 - 30 November 1986

Energy Technology Data Exchange (ETDEWEB)

Reddy, G M [Osmania Univ., Hyderabad (India). Dept. of Genetics

1987-12-31

The stability of high productivity of modern rice varieties is greatly affected by insect pests. Rice gall midge is a serious insect pest of rice that is prevalent in several south eastern asian countries. Gall midge resistance has been mainly attributed to antibiosis. No progress has so far been made in identifying the exact biochemical nature of resistance. In Indica subspecies the understanding of chemical nature of disease would be helpful in the control of the disease and also in breeding programme aimed at developing resistance varieties. Studies were undertaken to establish the biochemical basis of resistance. Biochemical characterization of resistant and susceptible varieties were carried out. The parameters considered were: total sugar and reducing sugar content, total phenol content, amino acid profile, post infectional changes in sugar and phenol, isozyme studies. 2 figs, 6 tabs.

Breeding and characterization of homokaryotic heteroplasmic male sterile lines in rice (oryza sativa)

International Nuclear Information System (INIS)

Mei, Q.; Zhou, X.; Liu, C.

2011-01-01

Twelve different Cytoplasmic Male-Sterile (CMS) lines were crossed with 18 rice varieties. From the hybrid with japonica rice Nongken 58, twelve homokaryotic-heteroplasmic male sterile lines were developed in B7F1 after successive back crossing and selection for stable male sterility and desirable agronomic traits such as flowering habit and high out crossing rate. The experimental results demonstrated that expression of the CMS factors were influenced by the corresponding nuclear genes. Three pollen abortion types, including the typical, the spherical and the stained abortion, were observed in the homokaryotic-heteroplasmic male sterile lines. Formation of the aborted pollen grains was influenced by the interaction among specific cytoplasmic and the corresponding nuclear genes. As the CMS carriers, these homokaryotic-heteroplasmic lines will have significant impact on the utilization of multiple types of CMS in hybrid rice breeding. What is more important is that these CMS lines are the invaluable materials for the investigation of the molecular mechanism of CMS formation in rice. (author)

Biochemical Stimulus-Based Strategies for Meniscus Tissue Engineering and Regeneration

Science.gov (United States)

Chen, Mingxue; Guo, Weimin; Gao, Shunag; Hao, Chunxiang; Shen, Shi; Zhang, Zengzeng; Wang, Zhenyong; Wang, Zehao; Li, Xu; Jing, Xiaoguang; Zhang, Xueliang; Yuan, Zhiguo; Wang, Mingjie; Zhang, Yu; Peng, Jiang; Wang, Aiyuan; Wang, Yu; Sui, Xiang

2018-01-01

Meniscus injuries are very common and still pose a challenge for the orthopedic surgeon. Meniscus injuries in the inner two-thirds of the meniscus remain incurable. Tissue-engineered meniscus strategies seem to offer a new approach for treating meniscus injuries with a combination of seed cells, scaffolds, and biochemical or biomechanical stimulation. Cell- or scaffold-based strategies play a pivotal role in meniscus regeneration. Similarly, biochemical and biomechanical stimulation are also important. Seed cells and scaffolds can be used to construct a tissue-engineered tissue; however, stimulation to enhance tissue maturation and remodeling is still needed. Such stimulation can be biomechanical or biochemical, but this review focuses only on biochemical stimulation. Growth factors (GFs) are one of the most important forms of biochemical stimulation. Frequently used GFs always play a critical role in normal limb development and growth. Further understanding of the functional mechanism of GFs will help scientists to design the best therapy strategies. In this review, we summarize some of the most important GFs in tissue-engineered menisci, as well as other types of biological stimulation. PMID:29581987

Thermodynamically consistent Bayesian analysis of closed biochemical reaction systems

Directory of Open Access Journals (Sweden)

Goutsias John

2010-11-01

Full Text Available Abstract Background Estimating the rate constants of a biochemical reaction system with known stoichiometry from noisy time series measurements of molecular concentrations is an important step for building predictive models of cellular function. Inference techniques currently available in the literature may produce rate constant values that defy necessary constraints imposed by the fundamental laws of thermodynamics. As a result, these techniques may lead to biochemical reaction systems whose concentration dynamics could not possibly occur in nature. Therefore, development of a thermodynamically consistent approach for estimating the rate constants of a biochemical reaction system is highly desirable. Results We introduce a Bayesian analysis approach for computing thermodynamically consistent estimates of the rate constants of a closed biochemical reaction system with known stoichiometry given experimental data. Our method employs an appropriately designed prior probability density function that effectively integrates fundamental biophysical and thermodynamic knowledge into the inference problem. Moreover, it takes into account experimental strategies for collecting informative observations of molecular concentrations through perturbations. The proposed method employs a maximization-expectation-maximization algorithm that provides thermodynamically feasible estimates of the rate constant values and computes appropriate measures of estimation accuracy. We demonstrate various aspects of the proposed method on synthetic data obtained by simulating a subset of a well-known model of the EGF/ERK signaling pathway, and examine its robustness under conditions that violate key assumptions. Software, coded in MATLAB®, which implements all Bayesian analysis techniques discussed in this paper, is available free of charge at http://www.cis.jhu.edu/~goutsias/CSS%20lab/software.html. Conclusions Our approach provides an attractive statistical methodology for

Uncertainty characterization of particle depth measurement using digital in-line holography and the hybrid method.

Science.gov (United States)

Gao, Jian; Guildenbecher, Daniel R; Reu, Phillip L; Chen, Jun

2013-11-04

In the detection of particles using digital in-line holography, measurement accuracy is substantially influenced by the hologram processing method. In particular, a number of methods have been proposed to determine the out-of-plane particle depth (z location). However, due to the lack of consistent uncertainty characterization, it has been unclear which method is best suited to a given measurement problem. In this work, depth determination accuracies of seven particle detection methods, including a recently proposed hybrid method, are systematically investigated in terms of relative depth measurement errors and uncertainties. Both synthetic and experimental holograms of particle fields are considered at conditions relevant to particle sizing and tracking. While all methods display a range of particle conditions where they are most accurate, in general the hybrid method is shown to be the most robust with depth uncertainty less than twice the particle diameter over a wide range of particle field conditions.

Biochemical toxicology of environmental agents

International Nuclear Information System (INIS)

Bruin, A. de

1976-01-01

A thorough and up-to-date account of the molecular-biological aspects of harmful agents - both chemical and physical - is given. This current treatise is principally intended to serve as an informative reference work for researchers in various areas of the field. In the pursuit of this aim, a devision of the entire field into 42 chapters has been made. Each chapter starts with a short introductory account dealing with the biochemical essentials of the particular subject. Radiation effects are discussed briefly at the end of each treatise. In order to make the treatise useful as a source book, a substantial collection of pertinent literature references is provided which are numbered in order of citation in the text. Initial chapters are devoted to the metabolic fate of the major classes of xenobiotic compounds. Peripheral topics, closely related to metabolism and dealing with modification of xenobiotic-metabolizing ability, as well as interaction phenomena follow (chs. 5-8). Subjects that draw heavily on the practical field of occupational hygiene are dealt with in chapters 9 and 10. The systematic treatment of how chemical and physical agents interact with the various biochemical and enzymatic systems they encounter during their passage through the organism occupies quantitatively the main part of the book (chs. 11-36). Finally, radiation biochemistry is discussed from the viewpoint of its high degree of scientific advancement, and secondly because the type of biochemical changes produced in vivo by X-rays closely parallel those evoked by chemical agents

Immunodominant role of CCHA subunit of Concholepas hemocyanin is associated with unique biochemical properties.

Science.gov (United States)

Becker, María Inés; Fuentes, Alejandra; Del Campo, Miguel; Manubens, Augusto; Nova, Esteban; Oliva, Harold; Faunes, Fernando; Valenzuela, María Antonieta; Campos-Vallette, Marcelo; Aliaga, Alvaro; Ferreira, Jorge; De Ioannes, Alfredo E; De Ioannes, Pablo; Moltedo, Bruno

2009-03-01

Hemocyanin, the oxygen transporter metallo-glycoprotein from mollusks, shows strong relationship between its notable structural features and intrinsic immunomodulatory effects. Here we investigated the individual contribution of CCHA and CCHB subunits from Concholepas hemocyanin (CCH) to in vivo humoral immune response and their pre-clinical evaluation as immunotherapeutic agent in a mice bladder cancer model, in relation to their biochemical properties. To this end, subunits were purified and well characterized. Homogeneous subunits were obtained by anionic exchange chromatography, and its purity assessed by electrophoretic and immunochemical methods. While each CCH subunit contains eight functional units showing partial cross reaction, the vibrational spectral analysis showed several spectral differences, suggesting structural differences between them. In addition, we demonstrated differences in the carbohydrate content: CCHA had a 3.6% w/w sugar with both N- and O-linked moieties. In turn, CCHB had a 2.5% w/w sugar with N-linked, while O-linked moieties were nearly absent. Considering these differences, it was not possible to predict a priori whether the immunogenic and immunotherapeutic properties of subunits might be similar. Surprisingly, both subunits by itself induced a humoral response, and showed an antitumor effect in the bladder carcinoma cell line MBT-2. However, when immunologic parameters were analyzed, CCHA showed better efficiency than CCHB. No allergic reactions or any toxic effects were observed in mice treated with CCHA, sustaining its potential therapeutic use. Our study supports that CCHA subunit accounts for the most important features involved in the immunogenicity of CCH, such as better hydrophilicity and higher content of carbohydrates.

Enzyme and biochemical producing fungi

DEFF Research Database (Denmark)

Lübeck, Peter Stephensen; Lübeck, Mette; Nilsson, Lena

2010-01-01

factories for sustainable production of important molecules. For developing fungi into efficient cell factories, the project includes identification of important factors that control the flux through the pathways using metabolic flux analysis and metabolic engineering of biochemical pathways....

On Line Characterization of SiC Nanoparticles Produced by Laser Pyrolysis

International Nuclear Information System (INIS)

Wattieaux, G; Lecerf, P; Meyer, L; Boufendi, L; Leconte, Y; Sublemontier, O; Herlin, N; Asimakopoulou, A; Tsakis, A; Daskalos, M; Konstandopoulos, A G; Ouf, F X

2011-01-01

On Line Measurements campaigns have been carried out for the first time in IRAMIS/SPAM CEA's laser gas-phase pyrolysis nanoparticles production facilities. The produced aerosol is composed of Argon and/or Helium laden with SiC nanoparticles concentrated up to ∼14 mg/l. Different commercial apparatus were used for sampling, and characterisation of size, and morphology of the particles. A series of experiment is performed by IRSN with a DMS500 from Cambustion Ltd TM , which gives electrical mobility equivalent diameter distribution in real time. Particles are sampled with an ejector-diluter VKL-10 from Palas TM . On the same sampling line, APTL performed aerodynamic equivalent diameter mass distribution using a Nanomoudi from MSP Corp. TM and TEM post analysis, as well as post elemental analysis (EDS) after collection on TEM grid with a Thermophoretic Precipitator (TP). A previously developed model is used to obtain the aggregate morphology and primary particles sizes. In addition to these commercial or well-known techniques, a new patent-pending technique called RFPM (Radio Frequency Plasma Metrology developed by CILAS + GREMI in the frame of NANOCARA program) is tested for the first time off the lab. The principle is based on levitation of particles in plasma RF. The entire set up is a prototype, including the sampling line. Experiments showed that this technique is very promising for on line gas phase monitoring, even though future improvements are needed, especially on the direct injection of the sample in the measuring chamber. First results of size distribution on pyrolysis line were obtained: three modes of different geometric mean diameter D50 were measured. Further analysis of TEM micrographs gave insight on these three modes, they can be interpreted as primary particles, aggregated particles, and a third one which could be composed of silica oxide nanoparticles issued from the combustion of the remaining silane initiated by an air leak in the exhaust of

Upgrading Laccase Production and Biochemical Properties: Strategies and Challenges.

Science.gov (United States)

Bertrand, Brandt; Martínez-Morales, Fernando; Trejo-Hernández, María R

2017-07-01

Improving laccases continues to be crucial in novel biotechnological developments and industrial applications, where they are concerned. This review breaks down and explores the potential of the strategies (conventional and modern) that can be used for laccase enhancement (increased production and upgraded biochemical properties such as stability and catalytic efficiency). The challenges faced with these approaches are briefly discussed. We also shed light on how these strategies merge and give rise to new options and advances in this field of work. Additionally, this article seeks to serve as a guide for students and academic researchers interested in laccases. This document not only gives basic information on laccases, but also provides updated information on the state of the art of various technologies that are used in this line of investigation. It also gives the readers an idea of the areas extensively studied and the areas where there is still much left to be done. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1015-1034, 2017. © 2017 American Institute of Chemical Engineers.

Polyphenol Oxidases in Crops: Biochemical, Physiological and Genetic Aspects

Directory of Open Access Journals (Sweden)

Francesca Taranto

2017-02-01

Full Text Available Enzymatic browning is a colour reaction occurring in plants, including cereals, fruit and horticultural crops, due to oxidation during postharvest processing and storage. This has a negative impact on the colour, flavour, nutritional properties and shelf life of food products. Browning is usually caused by polyphenol oxidases (PPOs, following cell damage caused by senescence, wounding and the attack of pests and pathogens. Several studies indicated that PPOs play a role in plant immunity, and emerging evidence suggested that PPOs might also be involved in other physiological processes. Genomic investigations ultimately led to the isolation of PPO homologs in several crops, which will be possibly characterized at the functional level in the near future. Here, focusing on the botanic families of Poaceae and Solanaceae, we provide an overview on available scientific literature on PPOs, resulting in useful information on biochemical, physiological and genetic aspects.

Biochemical and molecular characterization of the antioxidative system of Coffea sp. under cold conditions in genotypes with contrasting tolerance.

Science.gov (United States)

Fortunato, Ana S; Lidon, Fernando C; Batista-Santos, Paula; Leitão, António Eduardo; Pais, Isabel P; Ribeiro, Ana I; Ramalho, José Cochicho

2010-03-15

Low positive temperature (chilling) is frequently linked to the promotion of oxidative stress conditions, and is of particular importance in the coffee plant due to its severe impact on growth, development, photosynthesis and production. Nevertheless, some acclimation ability has been reported within the Coffea genus, and is possibly related to oxidative stress control. Using an integrated biochemical and molecular approach, the characterization of the antioxidative system of genotypes with different cold acclimation abilities was performed. Experiments were carried out using 1.5-year-old coffee seedlings of Coffea canephora cv. Apoatã, C. arabica cv. Catuaí, C. dewevrei and 2 hybrids, Icatu (C. arabicaxC. canephora) and Piatã (C. dewevreixC. arabica) subjected to a gradual cold treatment and a recovery period. Icatu showed the greatest ability to control oxidative stress, as reflected by the enhancement of several antioxidative components (Cu,Zn-SOD and APX activities; ascorbate, alpha-tocopherol and chlorogenic acids (CGAs) contents) and lower reactive oxygen species contents (H(2)O(2) and OH). Gene expression studies show that GRed, DHAR and class III and IV chitinases might also be involved in the cold acclimation ability of Icatu. Catuaí showed intermediate acclimation ability through the reinforcement of some antioxidative molecules, usually to a lesser extent than that observed in Icatu. On the other hand, C. dewevrei showed the poorest response in terms of antioxidant accumulation, and also showed the greatest increase in OH values. The difference in the triggering of antioxidative traits supports the hypothesis of its importance to cold (and photoinhibition) tolerance in Coffea sp. and could provide a useful probe to identify tolerant genotypes. Copyright 2009 Elsevier GmbH. All rights reserved.

Influence of process parameters on the weld lines of a micro injection molded component

DEFF Research Database (Denmark)

Tosello, Guido; Gava, Alberto; Hansen, Hans Nørgaard

2007-01-01

The insufficient entanglement of the molecular chains and the stress amplification at the v-notch of a weld line compromise the mechanical strength of a plastic product, also in the micro scale. To investigate the influence of process parameters on the weld lines formation, a special micro cavity...... was designed and manufactured by µEDM (Electro Discharge Machining). Weld lines were quantitatively characterized both in the two-dimensional (direction and position) and three-dimensional range (surface topography characterization). Results showed that shape and position of weld lines are mainly influenced...

Near-infrared Raman spectroscopy for assessing biochemical changes of cervical tissue associated with precarcinogenic transformation.

Science.gov (United States)

Duraipandian, Shiyamala; Mo, Jianhua; Zheng, Wei; Huang, Zhiwei

2014-11-07

Raman spectroscopy measures the inelastically scattered light from tissue that is capable of identifying native tissue biochemical constituents and their changes associated with disease transformation. This study aims to characterize the Raman spectroscopic properties of cervical tissue associated with the multi-stage progression of cervical precarcinogenic sequence. A rapid-acquisition fiber-optic near-infrared (NIR) Raman diagnostic system was employed for tissue Raman spectral measurements at 785 nm excitation. A total of 68 Raman spectra (23 benign, 29 low-grade squamous intraepithelial lesions (LSIL) and 16 high grade squamous intraepithelial lesions (HSIL)) were measured from 25 cervical tissue biopsy specimens, as confirmed by colposcopy-histopathology. The semi-quantitative biochemical modeling based on the major biochemicals (i.e., DNA, proteins (histone, collagen), lipid (triolein) and carbohydrates (glycogen)) in cervical tissue uncovers the stepwise accumulation of biomolecular changes associated with progressive cervical precarcinogenesis. Multi-class partial least squares-discriminant analysis (PLS-DA) together with leave-one tissue site-out, cross-validation yielded the diagnostic sensitivities of 95.7%, 82.8% and 81.3%; specificities of 100.0%, 92.3% and 88.5%,for discrimination among benign, LSIL and HSIL cervical tissues, respectively. This work suggests that the Raman spectral biomarkers have identified the potential to be used for monitoring the multi-stage cervical precarcinogenesis, forming the foundation of applying NIR Raman spectroscopy for the early diagnosis of cervical precancer in vivo at the molecular level.

Characterization of human warfarin reductase

OpenAIRE

Sokolová, Simona

2016-01-01

Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Simona Sokolová Supervisor: PharmDr. Petra Malátková, Ph.D. Title of diploma thesis: Characterization of human warfarin reductase Warfarin is widely used anticoagulant drug. Considering the narrow therapeutic window of warfarin, it is important to fully understand its metabolism in human body. Oxidative, reductive and conjugation reactions are involved in warfarin metabolism. Howev...

Probabilistic sensitivity analysis of biochemical reaction systems.

Science.gov (United States)

Zhang, Hong-Xuan; Dempsey, William P; Goutsias, John

2009-09-07

Sensitivity analysis is an indispensable tool for studying the robustness and fragility properties of biochemical reaction systems as well as for designing optimal approaches for selective perturbation and intervention. Deterministic sensitivity analysis techniques, using derivatives of the system response, have been extensively used in the literature. However, these techniques suffer from several drawbacks, which must be carefully considered before using them in problems of systems biology. We develop here a probabilistic approach to sensitivity analysis of biochemical reaction systems. The proposed technique employs a biophysically derived model for parameter fluctuations and, by using a recently suggested variance-based approach to sensitivity analysis [Saltelli et al., Chem. Rev. (Washington, D.C.) 105, 2811 (2005)], it leads to a powerful sensitivity analysis methodology for biochemical reaction systems. The approach presented in this paper addresses many problems associated with derivative-based sensitivity analysis techniques. Most importantly, it produces thermodynamically consistent sensitivity analysis results, can easily accommodate appreciable parameter variations, and allows for systematic investigation of high-order interaction effects. By employing a computational model of the mitogen-activated protein kinase signaling cascade, we demonstrate that our approach is well suited for sensitivity analysis of biochemical reaction systems and can produce a wealth of information about the sensitivity properties of such systems. The price to be paid, however, is a substantial increase in computational complexity over derivative-based techniques, which must be effectively addressed in order to make the proposed approach to sensitivity analysis more practical.

Advanced colorectal cancer, refractory to infusional fluorouracil treatment: efficacy of second line fluorouracil in combination with a different biochemical modulation

NARCIS (Netherlands)

van Halteren, H. K.; Wagener, D. J.; Vreugdenhil, G.; Punt, C. J.

1997-01-01

Currently there is no standard second line treatment for patients with advanced colorectal cancer (ACC). Previous reports have demonstrated that some patients may benefit from second line infusional 5-fluorouracil (5-FU) after failing 5-FU bolus treatment. We retrospectively studied the efficacy and

Expression analysis and biochemical characterization of beans plants biofortificated with zinc

Directory of Open Access Journals (Sweden)

Sandra Pérez Álvarez

2017-09-01

Full Text Available The present work was carried out in greenhouse conditions at the Centro de Investigación en Alimentación y Desarrollo AC in Delicias, Chihuahua, México. Four different concentrations (0, 25, 50 and 100 μM L−1 of Zn chelate and sulfate were used to study the antioxidant system of Phaseolus vulgaris L. Three genes related with antioxidant activity [superoxide dismutase (SOD, glutathione peroxidase (GSH-Px and catalase (CAT] were selected for expression study. Results showed that when Zn chelate at 50 and 100 μM L−1 were applied SOD was repressed and GSH-Px expression was low at 0, 25 and 100 μM L−1 while with sulfate form SOD expression was low and GSH-Px expression was strong in all treatment. CAT was highly expressed in all form and treatments. For a biochemical study the same enzymes were spectrophotometrically measured. SOD activity shows differences in both forms of Zn, chelate form was different at 25, 50 and 100 μM L−1 with less activity at 100 μM L−1 and sulfate treatment shows differences in all concentrations used. GSH-Px activity shows significant differences with sulfate form at 25, 50 μM L−1 where at 50 μM the activity was higher and low at 100 μM L−1, CAT does not exhibit significant differences but with chelate treatment at 50–100 μM L−1 the activity was higher compared to sulfate. Finally, to raise the Zn concentration in bean under biofortification program is a promising strategy in cropping systems in order to increase the ingestion of zinc and antioxidant capacity in the general population and provided the benefits that this element offered in human health.

Characterizing gene responses to drought stress in fourwing saltbush [Atriplex canescens (Pursh.) Nutt.)

Science.gov (United States)

Linda S. Adair; David L. Andrews; John Cairney; Edward A. Funkhouser; Ronald J. Newton; Earl F. Aldon

1992-01-01

New techniques in molecular biology can be used to characterize genes whose expression is induced by drought stress. These techniques can be used to understand responses of range plants to environmental stresses at the biochemical and molecular level. For example, they can be used to characterize genes that respond to drought stress conditions in the native shrub

Derivation and characterization of the NYSCFe003-A human embryonic stem cell line

Directory of Open Access Journals (Sweden)

Ana Sevilla

2017-12-01

Full Text Available The human embryonic stem cell line NYSCFe003-A was derived from a day 5 to day 6 blastocyst in feeder-free and antibiotic free conditions. The blastocyst was voluntarily donated for research as surplus after in vitro fertilization treatment following informed consent. The NYSCFe003-A line expresses all the pluripotency markers and has the potential to differentiate into all three germ layers in vitro. The line presents normal karyotype and is mycoplasma free.

In vivo and in vitro characterization of the biochemical and pathological changes induced by lionfish (pterios volitans) venom in mice.

Science.gov (United States)

Balasubashini, M Sri; Karthigayan, S; Somasundaram, S T; Balasubramanian, T; Viswanathan, P; Menon, Venugopal P

2006-01-01

Accidents caused by lionfish (Pterios volitans) envenomation are characterized by edema, intense pain, and necrosis at the site of sting. The mode of action and biochemistry of venoms are obviously complex and require a better knowledge and investigation to explore the toxic action and resulting biochemical changes. In the present study the LD(50) value of lionfish venom was found to be 42.5 mug/kg body weight (intraperitoneal injection) in Albino Swiss mice and was associated with reduced motor activity and asphyxiation followed by respiratory failure. The effect on vital organs revealed spongiosis in brain, vascular congestion in liver, cloudy swelling of renal tubules, congested blood vessels in renal tubules, and degeneration of myofibrils in heart. Whereas, the 10% of LD(50) (was 4.25 mug/kg b.w.), the sublethal dose showed reversible changes in the hematological (blood cell count, hematocrit, hemoglobin, mean corpuscular volume, mean corpuscular hemoglobin, and platelet count) parameters, serum enzymes (aspartate transaminase, alanine transaminase, alkaline phosphatase, lactate dehydrogenase, and creatinine phosphokinase), blood sugar, urea, creatinine, triglycerides, cholesterol, and total protein in mouse in vivo. The in vitro analysis of lionfish venom on mouse brain acetyl cholinesterase and Na(+), K(+), ATPase showed significant increased activity in a dose-dependent manner (10 to 40 mug). Moreover, the lionfish venom was observed to have a protease with a molecular weight of 45 kDa. Hence, the present study suggests the presence of bioactive proteins and peptides with excellent target specificity, which could be trapped for drug development in near future.

α-line systematics

International Nuclear Information System (INIS)

Caurier, E.; Zuker, A.P.

1991-01-01

A study of the α-line, defined as a set of differences (D α ) characterized binding energies of (N,Z) nuclei, is presented. The 2p2n-separation energies for even-even nuclei, odd-odd nuclei, even(Z)-odd(N) nuclei, odd(Z)-even(N) nuclei are presented. Trends and details in even-even nuclei, magic numbers of even-even and odd-odd nuclei are discussed. (G.P.) 1 ref.; 8 figs

Characterization of a Merkel Cell Polyomavirus-Positive Merkel Cell Carcinoma Cell Line CVG-1.

Science.gov (United States)

Velásquez, Celestino; Amako, Yutaka; Harold, Alexis; Toptan, Tuna; Chang, Yuan; Shuda, Masahiro

2018-01-01

Merkel cell polyomavirus (MCV) plays a causal role in ∼80% of Merkel cell carcinomas (MCC). MCV is clonally integrated into the MCC tumor genome, which results in persistent expression of large T (LT) and small T (sT) antigen oncoproteins encoded by the early locus. In MCV-positive MCC tumors, LT is truncated by premature stop codons or deletions that lead to loss of the C-terminal origin binding (OBD) and helicase domains important for replication. The N-terminal Rb binding domain remains intact. MCV-positive cell lines derived from MCC explants have been valuable tools to study the molecular mechanism of MCV-induced Merkel cell carcinogenesis. Although all cell lines have integrated MCV and express truncated LT antigens, the molecular sizes of the LT proteins differ between cell lines. The copy number of integrated viral genome also varies across cell lines, leading to significantly different levels of viral protein expression. Nevertheless, these cell lines share phenotypic similarities in cell morphology, growth characteristics, and neuroendocrine marker expression. Several low-passage MCV-positive MCC cell lines have been established since the identification of MCV. We describe a new MCV-positive MCV cell line, CVG-1, with features distinct from previously reported cell lines. CVG-1 tumor cells grow in more discohesive clusters in loose round cell suspension, and individual cells show dramatic size heterogeneity. It is the first cell line to encode an MCV sT polymorphism resulting in a unique leucine (L) to proline (P) substitution mutation at amino acid 144. CVG-1 possesses a LT truncation pattern near identical to that of MKL-1 cells differing by the last two C-terminal amino acids and also shows an LT protein expression level similar to MKL-1. Viral T antigen knockdown reveals that, like other MCV-positive MCC cell lines, CVG-1 requires T antigen expression for cell proliferation.

Molecular and biochemical characterization of a potato collection with contrasting tuber carotenoid content.

Directory of Open Access Journals (Sweden)

Maria Sulli

Full Text Available After wheat and rice, potato is the third most important staple food worldwide. A collection of ten tetraploid (Solanum tuberosum and diploid (S. phureja and S. chacoense genotypes with contrasting carotenoid content was subjected to molecular characterization with respect to candidate carotenoid loci and metabolic profiling using LC-HRMS. Irrespective of ploidy and taxonomy, tubers of these genotypes fell into three groups: yellow-fleshed, characterized by high levels of epoxy-xanthophylls and xanthophyll esters and by the presence of at least one copy of a dominant allele of the β-Carotene Hydroxylase 2 (CHY2 gene; white-fleshed, characterized by low carotenoid levels and by the presence of recessive chy2 alleles; and orange-fleshed, characterized by high levels of zeaxanthin but low levels of xanthophyll esters, and homozygosity for a Zeaxanthin Epoxidase (ZEP recessive allele. Novel CHY2 and ZEP alleles were identified in the collection. Multivariate analysis identified several groups of co-regulated non-polar compounds, and resulted in the grouping of the genotypes according to flesh color, suggesting that extensive cross-talk exists between the carotenoid pathway and other metabolite pathways in tubers. Postharvest traits like tuber dormancy and weight loss during storage showed little correlation with tuber carotenoid content, with the exception of zeaxanthin and its esters. Other tuber metabolites, such as glucose, monogalactosyldiacyglycerol (a glycolipid, or suberin precursors, showed instead significant correlations with both traits.

Molecular and biochemical characterization of a potato collection with contrasting tuber carotenoid content

Science.gov (United States)

Sulli, Maria; Mandolino, Giuseppe; Sturaro, Monica; Onofri, Chiara; Diretto, Gianfranco; Parisi, Bruno

2017-01-01

After wheat and rice, potato is the third most important staple food worldwide. A collection of ten tetraploid (Solanum tuberosum) and diploid (S. phureja and S. chacoense) genotypes with contrasting carotenoid content was subjected to molecular characterization with respect to candidate carotenoid loci and metabolic profiling using LC-HRMS. Irrespective of ploidy and taxonomy, tubers of these genotypes fell into three groups: yellow-fleshed, characterized by high levels of epoxy-xanthophylls and xanthophyll esters and by the presence of at least one copy of a dominant allele of the β-Carotene Hydroxylase 2 (CHY2) gene; white-fleshed, characterized by low carotenoid levels and by the presence of recessive chy2 alleles; and orange-fleshed, characterized by high levels of zeaxanthin but low levels of xanthophyll esters, and homozygosity for a Zeaxanthin Epoxidase (ZEP) recessive allele. Novel CHY2 and ZEP alleles were identified in the collection. Multivariate analysis identified several groups of co-regulated non-polar compounds, and resulted in the grouping of the genotypes according to flesh color, suggesting that extensive cross-talk exists between the carotenoid pathway and other metabolite pathways in tubers. Postharvest traits like tuber dormancy and weight loss during storage showed little correlation with tuber carotenoid content, with the exception of zeaxanthin and its esters. Other tuber metabolites, such as glucose, monogalactosyldiacyglycerol (a glycolipid), or suberin precursors, showed instead significant correlations with both traits. PMID:28898255

SABIO-RK: A data warehouse for biochemical reactions and their kinetics

Directory of Open Access Journals (Sweden)

Krebs Olga

2007-03-01

Full Text Available Systems biology is an emerging field that aims at obtaining a system-level understanding of biological processes. The modelling and simulation of networks of biochemical reactions have great and promising application potential but require reliable kinetic data. In order to support the systems biology community with such data we have developed SABIO-RK (System for the Analysis of Biochemical Pathways - Reaction Kinetics, a curated database with information about biochemical reactions and their kinetic properties, which allows researchers to obtain and compare kinetic data and to integrate them into models of biochemical networks. SABIO-RK is freely available for academic use at http://sabio.villa-bosch.de/SABIORK/.

Pin count-aware biochemical application compilation for mVLSI biochips

DEFF Research Database (Denmark)

Lander Raagaard, Michael; Pop, Paul

2015-01-01

Microfluidic biochips are replacing the conventional biochemical analyzers and are able to integrate the necessary functions for biochemical analysis on-chip. In this paper we are interested in flow-based biochips, in which the fluidic flow manipulated using integrated microvalves, which are cont...... a biochemical application. We focus on the compilation task, where the strategy is to delay operations, without missing their deadlines, such that the sharing of control signals is maximized. The evaluation shows a significant reduction in the number of control pins required....

Adding the s-Process Element Cerium to the APOGEE Survey: Identification and Characterization of Ce II Lines in the H-band Spectral Window

Science.gov (United States)

Cunha, Katia; Smith, Verne V.; Hasselquist, Sten; Souto, Diogo; Shetrone, Matthew D.; Allende Prieto, Carlos; Bizyaev, Dmitry; Frinchaboy, Peter; García-Hernández, D. Anibal; Holtzman, Jon; Johnson, Jennifer A.; Jőnsson, Henrik; Majewski, Steven R.; Mészáros, Szabolcs; Nidever, David; Pinsonneault, Mark; Schiavon, Ricardo P.; Sobeck, Jennifer; Skrutskie, Michael F.; Zamora, Olga; Zasowski, Gail; Fernández-Trincado, J. G.

2017-08-01

Nine Ce II lines have been identified and characterized within the spectral window observed by the Apache Point Observatory Galactic Evolution Experiment (APOGEE) survey (between λ1.51 and 1.69 μm). At solar metallicities, cerium is an element that is produced predominantly as a result of the slow capture of neutrons (the s-process) during asymptotic giant branch stellar evolution. The Ce II lines were identified using a combination of a high-resolution (R=λ /δ λ ={{100,000}}) Fourier Transform Spectrometer (FTS) spectrum of α Boo and an APOGEE spectrum (R = 22,400) of a metal-poor, but s-process enriched, red giant (2M16011638-1201525). Laboratory oscillator strengths are not available for these lines. Astrophysical gf-values were derived using α Boo as a standard star, with the absolute cerium abundance in α Boo set by using optical Ce II lines that have precise published laboratory gf-values. The near-infrared Ce II lines identified here are also analyzed, as consistency checks, in a small number of bright red giants using archival FTS spectra, as well as a small sample of APOGEE red giants, including two members of the open cluster NGC 6819, two field stars, and seven metal-poor N- and Al-rich stars. The conclusion is that this set of Ce II lines can be detected and analyzed in a large fraction of the APOGEE red giant sample and will be useful for probing chemical evolution of the s-process products in various populations of the Milky Way.

Vortex-line fluctuations in model high-temperature superconductors

International Nuclear Information System (INIS)

Li, Y.; Teitel, S.

1993-01-01

We carry out Monte Carlo simulations of the uniformly frustrated three-dimensional XY model, as a model for vortex-line fluctuations in a high-T c superconductor in an external magnetic field. A density of vortex lines of f=1/25 is considered. We find two sharp phase transitions. The low-T superconducting phase is an ordered vortex-line lattice. The high-T normal phase is a vortex-line liquid, with much entangling, cutting, and loop excitations. An intermediate phase is found, which is characterized as a vortex-line liquid of disentangled, approximately straight, lines. In this phase, the system displays superconducting properties in the direction parallel to the magnetic field, but normal behavior in planes perpendicular to the field. A detailed analysis of the vortex structure function is carried out

Development of a new first-aid biochemical detector

Science.gov (United States)

Hu, Jingfei; Liao, Haiyang; Su, Shilin; Ding, Hao; Liu, Suquan

2016-10-01

The traditional biochemical detector exhibits poor adaptability, inconvenient carrying and slow detection, which can't meet the needs of first-aid under field condition like natural or man-made disasters etc. Therefore a scheme of first-aid biochemical detector based on MOMES Micro Spectrometer, UV LED and Photodiode was proposed. An optical detection structure combined continuous spectrum sweep with fixed wavelength measurement was designed, which adopted mobile detection optical path consisting of Micro Spectrometer and Halogen Lamp to detect Chloride (Cl-), Creatinine (Cre), Glucose (Glu), Hemoglobin (Hb). The UV LED and Photodiode were designed to detect Potassium (K-), Carbon dioxide (CO2), Sodium (Na+). According to the field diagnosis and treatment requirements, we designed the embedded control hardware circuit and software system, the prototype of first-aid biochemical detector was developed and the clinical trials were conducted. Experimental results show that the sample's absorbance repeatability is less than 2%, the max coefficient of variation (CV) in the batch repeatability test of all 7 biochemical parameters in blood samples is 4.68%, less than the clinical requirements 10%, the correlation coefficient (R2) in the clinical contrast test with AU5800 is almost greater than 0.97. To sum up, the prototype meets the requirements of clinical application.

Biochemical failure after radical external beam radiotherapy for prostate cancer

International Nuclear Information System (INIS)

Nomoto, Satoshi; Imada, Hajime; Kato, Fumio; Yahara, Katsuya; Morioka, Tomoaki; Ohguri, Takayuki; Nakano, Keita; Korogi, Yukunori

2005-01-01

The purpose of this study was to evaluate biochemical failures after radical external beam radiotherapy for prostate cancer. A total of 143 patients with prostate cancer (5 cases in stage A2, 95 in stage B and 43 in stage C; 18 in low risk group, 37 in intermediate risk group, 67 in high risk group and 21 in unknown group) were included in this study. Patients of stage A2 and B underwent external irradiation of 46 Gy to the prostate gland and seminal vesicle and additional 20 Gy to the prostate gland, while patients of stage C underwent external irradiation of 66 Gy to the prostate gland and seminal vesicle including 46 Gy to the pelvis. Neoadjuvant hormonal therapy was done in 66 cases, and long-term hormonal therapy in 75 cases; two cases were treated with radiation therapy alone. The 3-year relapse free survival rates by stage A2, B and C were 100%, 96.7% and 88.1%, respectively. The 3-year relapse free survival rates by low, intermediate and high risk groups were 100%, 92.3% and 89.7%, respectively. Biochemical failure was noted in nine cases during the average observation term of 32.2 months; in this group the median of prostate specific antigen (PSA) value was 2.6 ng/ml, the doubling time was 8.6 months, and the term of biochemical failure was 33.2 months. Six of eight cases with biochemical failure were the neoadjuvant hormonal therapy group, but biochemical no evidence of disease (bNED) curve showed no significant difference between neoadjuvant and long-term hormonal groups. It is supposed that unnecessary hormonal therapies were performed based on the nonspecific diagnosis of biochemical failure after radical radiotherapy in our group of patients. A precise criterion of biochemical failure after radical radiotherapy for prostate cancer is necessary. (author)

Effect of earthworms on the biochemical characterization of biofilms in vermifiltration treatment of excess sludge.

Science.gov (United States)

Yang, Jian; Liu, Jing; Xing, Meiyan; Lu, Zhibo; Yan, Qiong

2013-09-01

In this study, the biofilms formed in a vermifilter (VF) with earthworms and a conventional biofilter (BF) without earthworms were compared to investigate the effects of earthworms on the characteristics of biofilms during an excess sludge treatment period of 4months. Typical macrographs and micrographs of the biofilms showed that the feeding and casting actions of earthworms remarkably modified the VF morphology. Elemental analysis and fluorescence spectra indicated that earthworms enhanced the stabilization of organic matter by accelerating the mineralization and humification of organic materials during vermiconversion. In addition, bacterial communities inhabiting the VF biofilm showed that earthworms increased both bacterial diversity and metabolic activities in the film, as revealed by automatic testing bacteriology (ATB) expression and sequencing data. These results demonstrate that earthworms influence the structure and biochemical characteristics of biofilms and enhance their bacterial diversity and functions for improved sludge stabilization. Copyright © 2013 Elsevier Ltd. All rights reserved.

Complexity of generic biochemical circuits: topology versus strength of interactions

Science.gov (United States)

Tikhonov, Mikhail; Bialek, William

2016-12-01

The historical focus on network topology as a determinant of biological function is still largely maintained today, illustrated by the rise of structure-only approaches to network analysis. However, biochemical circuits and genetic regulatory networks are defined both by their topology and by a multitude of continuously adjustable parameters, such as the strength of interactions between nodes, also recognized as important. Here we present a class of simple perceptron-based Boolean models within which comparing the relative importance of topology versus interaction strengths becomes a quantitatively well-posed problem. We quantify the intuition that for generic networks, optimization of interaction strengths is a crucial ingredient of achieving high complexity, defined here as the number of fixed points the network can accommodate. We propose a new methodology for characterizing the relative role of parameter optimization for topologies of a given class.

APOBEC3DE Inhibits LINE-1 Retrotransposition by Interacting with ORF1p and Influencing LINE Reverse Transcriptase Activity.

Directory of Open Access Journals (Sweden)

Weizi Liang

Full Text Available Human long interspersed elements 1 (LINE-1 or L1 is the only autonomous non-LTR retroelement in humans and has been associated with genome instability, inherited genetic diseases, and the development of cancer. Certain human APOBEC3 family proteins are known to have LINE-1 restriction activity. The mechanisms by which APOBEC3 affects LINE-1 retrotransposition are not all well characterized; here, we confirm that both A3B and A3DE have a strong ability to inhibit LINE-1 retrotransposition. A3DE interacts with LINE-1 ORF1p to target LINE-1 ribonucleoprotein particles in an RNA-dependent manner. Moreover, A3DE binds to LINE-1 RNA and ORF1 protein in cell culture system. Fluorescence microscopy demonstrated that A3DE co-localizes with ORF1p in cytoplasm. Furthermore, A3DE inhibits LINE-1 reverse transcriptase activity in LINE-1 ribonucleoprotein particles in a cytidine deaminase-independent manner. In contrast, A3B has less inhibitory effects on LINE-1 reverse transcriptase activity despite its strong inhibition of LINE-1 retrotransposition. This study demonstrates that different A3 proteins have been evolved to inhibit LINE-1 activity through distinct mechanisms.

Mutation breeding for downy mildew resistance in pearl millet. Nucleo-cytoplasmic interactions in disease-resistant lines

International Nuclear Information System (INIS)

Murty, B.R.; Thakur, S.R.; Prakash, N.; Mehta, S.L.; Bhakta, S.T.

1983-01-01

Under the need to rescue hybrid pearl millet cultivation in India from devastating damage by downy mildew, a mutation induction project was started in 1971 to make the commonly used male sterile parent Tift 23A resistant to the disease. Simultaneously sources of resistance from West Africa were used in crossbreeding by which climatic adaptation and male sterility had to be transferred. A number of mildew-resistant hybrids were developed, both from induced mutation and introduction. The resistant male sterile lines were further examined as to their common features and differences from susceptible lines. A strong evidence for nuclear-cytoplasmic interaction was obtained by biochemical and ultrastructural investigations. (author)

Derivation and characterization of the NIH registry human stem cell line NYSCF100 line under defined feeder-free conditions

Directory of Open Access Journals (Sweden)

Ana Sevilla

2018-05-01

Full Text Available The human embryonic stem cell line NYSCFe001-A was derived from a day 6 blastocyst in feeder-free and antibiotic free conditions. The blastocyst was voluntarily donated for research as surplus after in vitro fertilization treatment following informed consent. The NYSCFe001-A line, registered as NYSCF100 on the NIH registry, presents normal karyotype, is mycoplasma free, expresses all the pluripotency markers and has the potential to differentiate into all three germ layers in vitro.

Multidimensional biochemical information processing of dynamical patterns.

Science.gov (United States)

Hasegawa, Yoshihiko

2018-02-01

Cells receive signaling molecules by receptors and relay information via sensory networks so that they can respond properly depending on the type of signal. Recent studies have shown that cells can extract multidimensional information from dynamical concentration patterns of signaling molecules. We herein study how biochemical systems can process multidimensional information embedded in dynamical patterns. We model the decoding networks by linear response functions, and optimize the functions with the calculus of variations to maximize the mutual information between patterns and output. We find that, when the noise intensity is lower, decoders with different linear response functions, i.e., distinct decoders, can extract much information. However, when the noise intensity is higher, distinct decoders do not provide the maximum amount of information. This indicates that, when transmitting information by dynamical patterns, embedding information in multiple patterns is not optimal when the noise intensity is very large. Furthermore, we explore the biochemical implementations of these decoders using control theory and demonstrate that these decoders can be implemented biochemically through the modification of cascade-type networks, which are prevalent in actual signaling pathways.

THE PHOTOMETRIC AND KINEMATIC STRUCTURE OF FACE-ON DISK GALAXIES. II. INTEGRATED LINE PROFILE CHARACTERIZATION AND THE ORIGIN OF LINE PROFILE ASYMMETRY

International Nuclear Information System (INIS)

Andersen, David R.; Bershady, Matthew A.

2009-01-01

We perform a moments analysis of H I and H II global line profiles for 33 nearly face-on disk galaxies for the threefold purpose of rationalizing and interpreting line profile indices in the literature, presenting robust moment definitions with analytic error functions calibrated against Monte Carlo simulation, and probing the physical origin of line profile asymmetries. The first four profile moments serve as viable surrogates for the recession velocity, line width, asymmetry, and profile shape, respectively. The first three moments are superior, by a factor of ∼2 in precision, to related quantities defined in the literature. First and third profile moments are related; skew can be used to calculate more accurate recession velocities from global profiles. Second and fourth profile moments are linked, corresponding to the known trend that narrow (but well resolved) line widths tend to be more Gaussian. Hα kurtosis also appears correlated with the spatially resolved line width of the ionized gas. No systematics appear between various measures of line width and true rotation speed other than the wide range of normalizations, which we calibrate. This conclusion and calibration, however, is highly sample dependent. The ratio of H II to H I widths is consistent with unity, even at low projected line width. There may be a trend toward a decrease in the ratio of H II to H I widths consistent with previous studies, but we also observe greater scatter. While there is good agreement between H I and H II first, second, and fourth moments, we find no positive correlation between skew measured from H I and H II profiles. Detailed analysis of the spatially resolved Hα distribution demonstrates that H II global profile asymmetries are dominated by differences in the gas distribution, not kinematic asymmetries.

Biochemical markers in the follow-up of medullary thyroid cancer

NARCIS (Netherlands)

de Groot, Jan Willem B.; Kema, Ido P.; Breukelman, Henk; van der Veer, Eveline; Wiggers, Theo; Plukker, John T. M.; Wolffenbuttel, Bruce H. R.; Links, Thera P.

2006-01-01

Medullary thyroid cancer (MTC) shares biochemical features with other neuroendocrine tumors but the particular characteristics are largely unexplored. We investigated the biochemical neuroendocrine profile of MTC and whether specific markers could be useful in follow-up. In addition to the standard

Biogenic Synthesis, Characterization and Evaluation of Silver Nanoparticles from Aspergillus niger JX556221 Against Human Colon Cancer Cell Line HT-29.

Science.gov (United States)

Chengzheng, Wang; Jiazhi, Wen; Shuangjiang, Chen; Swamy, Mallappa Kumara; Sinniah, Uma Rani; Akhtar, Mohd Sayeed; Umar, Ahmad

2018-05-01

Nanobiotechnology has emerged as a promising technology to develop new therapeutically active nanomaterials. The present study was aimed to biosynthesize AgNPs extracellularly using Aspergillus niger JX556221 fungal extract and to evaluate their anticancer potential against colon cancer cell line, HT-29. UV-visible spectral characterization of the synthesized AgNPs showed higher absorption peak at 440 nm wavelength. Transmission Electron Microscopy (TEM) analysis revealed the monodispersed nature of synthesized AgNPs occurring in spherical shape with a size in the range of 20-25 nm. Further, characterization using Energy Dispersive Spectroscopy (EDX) confirmed the face-centred cubic crystalline structure of metallic AgNPs. FTIR data revealed the occurrence of various phytochemicals in the cell free fungal extract which substantiated the fungal extract mediated AgNPs synthesis. The cytotoxic effect of AgNPs was studied by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results evidenced the cytotoxic effect of AgNPs on HT-29 cell lines in a dose dependent manner. The highest activity was found at 100 μg/ml concentration after 24 h of incubation. Use of propidium iodide staining examination method confirmed the cytotoxic effect of AgNPs through inducing cell apoptosis. AgNPs cytotoxicity was found to be through elevating reactive oxygen species (ROS), and caspase-3 activation resulting in induced apoptosis. Therefore, this research finding provides an insight towards the development of novel anticancer agents using biological sources.

Bezafibrate induces a mitochondrial derangement in human cell lines: a PPAR-independent mechanism for a peroxisome proliferator.

Science.gov (United States)

Scatena, R; Bottoni, P; Vincenzoni, F; Messana, I; Martorana, G E; Nocca, G; De Sole, P; Maggiano, N; Castagnola, M; Giardina, B

2003-11-01

Bezafibrate is a hypolipidemic drug that belongs to the group of peroxisome proliferators because it binds to peroxisome proliferator-activated receptors type alpha (PPARs). Peroxisome proliferators produce a myriad of extraperoxisomal effects, which are not necessarily dependent on their interaction with PPARs. An investigation on the peculiar activities of bezafibrate could clarify some of the molecular events and the relationship with the biochemical and pharmacological properties of this class of compounds. In this view, the human acute promyelocytic leukemia HL-60 cell line and human rabdomiosarcoma TE-671 cell line were cultured in media containing bezafibrate and a number of observations such as spectrophotometric analysis of mitochondrial respiratory chain enzymes, NMR metabolite determinations, phosphofructokinase enzymatic analysis, and differentiation assays were carried on. Bezafibrate induced a derangement of NADH cytochrome c reductase activity accompanied by metabolic alterations, mainly a shift to anaerobic glycolysis and an increase of fatty acid oxidation, as shown by NMR analysis of culture supernatants where acetate, lactate, and alanine levels increased. On the whole, the present results suggest a biochemical profile and a therapeutic role of this class of PPARs ligands more complex than those previously proposed.

Biochemical evaluation of phenylketonuria (PKU: from diagnosis to treatment

Directory of Open Access Journals (Sweden)

Leticia Belmont-Martínez

2014-07-01

Besides periodical Phe and Tyr testing, biochemical follow-up includes the measurement of necessary elements that guarantee normal physical and intellectual development such as selenium, zinc, B12 vitamin, folates, iron and long chain fatty acids. Clinical context is as important as biochemical status so periodic evaluation of nutritional, medical, social and psychological aspects should be included.

Characterization of the solar climate in Malawi using NASA's surface ...

African Journals Online (AJOL)

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Characterization of the solar climate in Malawi using. NASA's surface meteorology and solar energy. (SSE) model. Senganimalunje, T. C.1 and Tenthani, C. M. 2*. 1Malawi Bureau of Standards, Metrology Services Department, Box 946, Blantyre, Malawi. 2Physics and Biochemical Sciences Department, Malawi Polytechnic, ...

Characterization of chlorinated solvent contamination in limestone using innovative FLUTe® technologies in combination with other methods in a line of evidence approach

DEFF Research Database (Denmark)

Broholm, Mette Martina; Janniche, Gry Sander; Mosthaf, Klaus

2016-01-01

Characterization of dense non-aqueous phase liquid (DNAPL) source zones in limestone aquifers/bedrock is essential to develop accurate site-specific conceptual models and perform risk assessment. Here innovative field methods were combined to improve determination of source zone architecture......, hydrogeology and contaminant distribution. The FACT™ is a new technology and it was applied and tested at a contaminated site with a limestone aquifer, together with a number of existing methods including wire-line coring with core subsampling, FLUTe® transmissivity profiling and multilevel water sampling...... groundwater sampling (under two flow conditions) and FACT™ sampling and analysis combined with FLUTe® transmissivity profiling and modeling were used to provide a line of evidence for the presence of DNAPL, dissolved and sorbed phase contamination in the limestone fractures and matrix. The combined methods...

Biochemical and Molecular Characterization of a Barley Seed ß-Glucosidase

DEFF Research Database (Denmark)

Leah, R.; Kigel, J.; Svendsen, I.

1995-01-01

blot analysis with the cDNA as probe indicated that BGQ60 is encoded by a single gene, and that BGQ60 mRNA only accumulates in the starchy endosperm tissue of late developing seeds. The bgq60 structural gene of approximately 5 kilobases contains an open reading frame encoding 485 amino acids...... during barley seed development and germination are discussed.......A 60-kDa ß-glucosidase (BGQ60) was purified and characterized from seeds of barley (Hordeum vulgare L.). BGQ60 catalytic activity was restricted to the cleavage of short-chain oligosaccharides composed of(1, 2) -,(1, 2, 3) -, and/or(1, 2, 3, 4) -ß-linked glucose or mannose units...

Establishment and Characterization of a Highly Tumourigenic and Cancer Stem Cell Enriched Pancreatic Cancer Cell Line as a Well Defined Model System

Science.gov (United States)

Fredebohm, Johannes; Boettcher, Michael; Eisen, Christian; Gaida, Matthias M.; Heller, Anette; Keleg, Shereen; Tost, Jörg; Greulich-Bode, Karin M.; Hotz-Wagenblatt, Agnes; Lathrop, Mark; Giese, Nathalia A.; Hoheisel, Jörg D.

2012-01-01

Standard cancer cell lines do not model the intratumoural heterogeneity situation sufficiently. Clonal selection leads to a homogeneous population of cells by genetic drift. Heterogeneity of tumour cells, however, is particularly critical for therapeutically relevant studies, since it is a prerequisite for acquiring drug resistance and reoccurrence of tumours. Here, we report the isolation of a highly tumourigenic primary pancreatic cancer cell line, called JoPaca-1 and its detailed characterization at multiple levels. Implantation of as few as 100 JoPaca-1 cells into immunodeficient mice gave rise to tumours that were histologically very similar to the primary tumour. The high heterogeneity of JoPaca-1 was reflected by diverse cell morphology and a substantial number of chromosomal aberrations. Comparative whole-genome sequencing of JoPaca-1 and BxPC-3 revealed mutations in genes frequently altered in pancreatic cancer. Exceptionally high expression of cancer stem cell markers and a high clonogenic potential in vitro and in vivo was observed. All of these attributes make this cell line an extremely valuable model to study the biology of and pharmaceutical effects on pancreatic cancer. PMID:23152778

Integrated cantilever-based flow sensors with tunable sensitivity for in-line monitoring of flow fluctuations in microfluidic systems

DEFF Research Database (Denmark)

Noeth, Nadine-Nicole; Keller, Stephan Sylvest; Boisen, Anja

2014-01-01

For devices such as bio-/chemical sensors in microfluidic systems, flow fluctuations result in noise in the sensor output. Here, we demonstrate in-line monitoring of flow fluctuations with a cantilever-like sensor integrated in a microfluidic channel. The cantilevers are fabricated in different...... is directly proportional to the flow rate fluctuations in the microfluidic channel. The SiN cantilevers show a detection limit below 1 nL/min and the thinnest SU-8 cantilevers a detection limit below 5 nL/min. Finally, the sensor is applied for in-line monitoring of flow fluctuations generated by external...

Characterization of T cell receptor assembly and expression in a Ti gamma delta-positive cell line

DEFF Research Database (Denmark)

Kuhlmann, J; Caspar-Bauguil, S; Geisler, C

1993-01-01

- variants of the T cell Lyon were induced and found to produce all of the Ti/CD3 components, with the exception of Ti-delta. Biochemical analysis indicated that: (1) Ti-gamma/CD3 gamma, delta, epsilon complexes were formed in the endoplasmic reticulum in the absence of Ti-delta; (2) the CD3-zeta chain did...

An improved solution of first order kinetics for biochemical oxygen ...

African Journals Online (AJOL)

This paper evaluated selected Biochemical Oxygen Demand first order kinetics methods. Domesticinstitutional wastewaters were collected twice in a month for three months from the Obafemi Awolowo University, Ile-Ife waste stabilization ponds. Biochemical Oxygen Demand concentrations at different days were determined ...

The effects of Islamic fasting on blood hematological-biochemical parameters

Directory of Open Access Journals (Sweden)

Mohamad Reza Sedaghat

2017-06-01

Conclusion:This study on healthy subjects suggests that fasting could affect some hematological-biochemical parameters but not all of them. Also, these changes in hematological-biochemical parameters were within the normal range and Ramadan fasting seems to be safe for healthy subjects.

Physiological and biochemical characterization of mango tree with paclobutrazol application via irrigation

Directory of Open Access Journals (Sweden)

Moisés Alves de Souza

2016-12-01

Full Text Available Paclobutrazol application in mango tree floral induction is followed by changes in the hormonal balance and carbohydrates production. This study aimed at evaluating the physiological and biochemical variables of mango tree (Palmer cultivar by applying paclobutrazol doses via irrigation. The experimental design was randomized blocks, with five paclobutrazol doses (0.7 g, 1.0 g, 1.3 g, 1.6 g and 1.9 g of a.i. per linear meter of canopy and one additional treatment, with one dose in the conventional form of application (1.9 g of a.i. per linear meter of canopy, and four replicates. The variables analyzed were: photosynthesis rate; stomatal conductance; transpiration; leaf temperature; CO2 ratio; total protein content; reducing, non-reducing and total soluble sugars in leaf tissue and fruit; and nitrate reductase enzyme. The lowest values for photosynthesis rate and stomatal conductance were obtained with paclobutrazol applied via irrigation. The highest values for gas exchange were obtained with the lowest paclobutrazol doses applied via irrigation, showing that this kind of application is efficient and that it is possible to reduce the application dose via irrigation system. The highest carbohydrate contents in the leaf tissue were observed for the doses of 1.06 g and 1.09 g of a.i. per linear meter of canopy, applied via irrigation. The carbohydrate contents in fruits decreased in response to the paclobutrazol dose. The application methods and doses applied via irrigation did not influence the nitrate reductase activity.

Rapid Discrimination Among Putative Mechanistic Models of Biochemical Systems.

Science.gov (United States)

Lomnitz, Jason G; Savageau, Michael A

2016-08-31

An overarching goal in molecular biology is to gain an understanding of the mechanistic basis underlying biochemical systems. Success is critical if we are to predict effectively the outcome of drug treatments and the development of abnormal phenotypes. However, data from most experimental studies is typically noisy and sparse. This allows multiple potential mechanisms to account for experimental observations, and often devising experiments to test each is not feasible. Here, we introduce a novel strategy that discriminates among putative models based on their repertoire of qualitatively distinct phenotypes, without relying on knowledge of specific values for rate constants and binding constants. As an illustration, we apply this strategy to two synthetic gene circuits exhibiting anomalous behaviors. Our results show that the conventional models, based on their well-characterized components, cannot account for the experimental observations. We examine a total of 40 alternative hypotheses and show that only 5 have the potential to reproduce the experimental data, and one can do so with biologically relevant parameter values.

Fluoroorotic acid-selected Nicotiana plumbaginifolia cell lines with a stable thymine starvation phenotype have lost the thymine-regulated transcriptional program.

Science.gov (United States)

Santoso, D; Thornburg, R

2000-08-01

We have selected 143 independent Nicotiana plumbaginifolia cell lines that survive in the presence of 5-fluoroorotic acid. These lines show several diverse phenotypes. The majority of these cell lines showed reduced levels of UMP synthase. However, one particular phenotype, which represents 14% of the total independent lines (20 cell lines), showed an unexpected, high level of UMP synthase and was therefore analyzed in detail. The selected cell lines showed no differences with wild-type cells with respect to uptake of orotic acid, affinity of UMP synthase for its substrates, or UMP synthase gene-copy number. Alternative detoxification mechanisms were also excluded. The elevated enzyme activity was correlated with elevated UMP synthase protein levels as well as elevated UMP synthase mRNA levels. In contrast to wild-type cell lines, the fluoroorotic acid-selected cell lines did not respond to thymine or to other biochemicals that affect thymine levels. In addition, there was also a concomitant up-regulation of aspartate transcarbamoylase, however, dihydroorotase and dihydroorotate dehydrogenase are not up-regulated in these cell lines.

Salvage conformal radiotherapy for biochemical recurrent prostate cancer after radical prostatectomy

Directory of Open Access Journals (Sweden)

Carlos R. Monti

2006-08-01

Full Text Available OBJECTIVE: Assess the results of salvage conformal radiotherapy in patients with biochemical failure after radical prostatectomy and identify prognostic factors for biochemical recurrence and toxicity of the treatment. MATERIALS AND METHODS: From June 1998 to November 2001, 35 patients were submitted to conformal radiotherapy for PSA > 0.2 ng/mL in progression after radical prostatectomy and were retrospectively analyzed. The mean dose of radiation in prostatic bed was of 77.4 Gy (68-81. Variables related to the treatment and to tumor were assessed to identify prognostic factors for biochemical recurrence after salvage radiotherapy. RESULTS: The median follow-up was of 55 months (17-83. The actuarial survival rates free of biochemical recurrence and free of metastasis at a distance of 5 years were 79.7% e 84.7%, respectively. The actuarial global survival rate in 5 years was 96.1%.The actuarial survival rate free of biochemical recurrence in 5 years was 83.3% with PSA pre-radiotherapy 1 and 2 (p = 0.023. Dose > 70 Gy in 30% of the bladder volume implied in more acute urinary toxicity (p = 0.035. The mean time for the development of late urinary toxicity was 21 months (12-51. Dose > 55 Gy in 50% bladder volume implied in more late urinary toxicity (p = 0.018. A patient presented late rectal toxicity of 2nd grade. CONCLUSIONS: Conformal radiotherapy showed to be effective for the control of biochemical recurrence after radical prostatectomy. Patients with pre-therapy PSA < 2 ng/mL have more biochemical control.

Modeling a multivariable reactor and on-line model predictive control.

Science.gov (United States)

Yu, D W; Yu, D L

2005-10-01

A nonlinear first principle model is developed for a laboratory-scaled multivariable chemical reactor rig in this paper and the on-line model predictive control (MPC) is implemented to the rig. The reactor has three variables-temperature, pH, and dissolved oxygen with nonlinear dynamics-and is therefore used as a pilot system for the biochemical industry. A nonlinear discrete-time model is derived for each of the three output variables and their model parameters are estimated from the real data using an adaptive optimization method. The developed model is used in a nonlinear MPC scheme. An accurate multistep-ahead prediction is obtained for MPC, where the extended Kalman filter is used to estimate system unknown states. The on-line control is implemented and a satisfactory tracking performance is achieved. The MPC is compared with three decentralized PID controllers and the advantage of the nonlinear MPC over the PID is clearly shown.