Characterization of cystathionine synthase as a selectable, liver-specific trait in rat hepatomas

1986 ◽  
Vol 82 (1) ◽  
pp. 309-320
Author(s):  
S.J. Goss
Keyword(s):  
Cell Growth ◽  
Serum Proteins ◽  
Selective Medium ◽  
Carbamoyl Phosphate ◽  
Cycle Enzyme ◽  
Wide Range ◽  
Specific Trait ◽  
Well Differentiated

Cell growth using homocysteine as a source of cysteine-sulphur requires two enzymes, cystathionine synthase (CS) and gamma-cystathionase (CT). The second of these enzymes, CT, is apparently present in most cell lines regardless of their tissues of origin, since most cells can grow in vitro in the absence of cystine if they are provided with cystathionine, the intermediate in the pathway. Likewise, homocysteine will support the growth of many human cells. However, of a wide range of rodent cells, only well-differentiated rat hepatoma cells were found to grow using homocysteine in place of cystine. It is shown that cell growth in homocysteine-medium correlates well with the presence in the cells of detectable levels of CS. Furthermore, in cells able to grow in homocysteine-medium, it is possible to demonstrate the homocysteine-dependent trans-sulphuration of serine to cysteine. Growth in homocysteine-medium is not dependent on the release of preformed cysteine from disulphide complexes with serum proteins. In cell hybrids, and in ‘dedifferentiated’ variants of rat hepatomas, CS, but not CT, is subject to extinction coordinately with well-characterized liver-specific traits. For rodent cells, homocysteine-medium thus acts as a selective medium requiring the expression of a single liver-specific trait, CS. In addition it is shown that, in certain hepatoma variants, CS is regulated co-ordinately with a urea-cycle enzyme (carbamoyl phosphate synthetase I) by glucocorticoids and cyclic-AMP. Cell death through cysteine starvation is briefly considered. The immediate cause of death is apparently an insufficient supply of reduced glutathione. Selenium and vitamin E assist cell growth when the supply of cysteine is limiting.

Arginine synthesis by hepatomas in vitro. I. The requirements for cell growth in medium containing ornithine in place of arginine and the isolation and characterization of variant hepatomas auxotrophic for arginine

1984 ◽  
Vol 68 (1) ◽  
pp. 285-303 ◽  
Author(s):  
S.J. Goss
Keyword(s):  
Cell Growth ◽  
The Other ◽  
Gene Products ◽  
Carbamoyl Phosphate Synthetase ◽  
Arginine Biosynthesis ◽  
Carbamoyl Phosphate ◽  
Isolation And Characterization ◽  
Ornithine Transcarbamoylase

Cell growth in ‘ornithine-medium’ requires the expression of two liver-specific genes, those for ornithine transcarbamoylase (OTC) and carbamoyl phosphate synthetase I (CPS-I). CPS-II appears unable to replace CPS-I in this system. The need for N-acetylglutamate (to activate CPS-I) can be met, at least in part, by providing it in the medium. The other gene products involved in arginine biosynthesis are probably all ubiquitous (i.e. not tissue-specific). In an attempt to study the factors responsible for the expression of liver-specific genes, variant hepatomas are isolated that have lost the ability to grow in ornithine-medium. Two classes of ‘orn-’ variants are identified: unstable variants that require dexamethasone for adequate CPS-I production, and ‘stable’ variants that have lost many liver-specific traits. Studies on one stable variant show that it can revert (though rarely), and that it regains its various liver-specific traits in a non-coordinate fashion.

scholarly journals In vitro and In Silico Approach For Characterization of Antimicrobial Peptide From Probiotics Against Staphylococcus Aureus and Escherichia Coli

2021 ◽  
Author(s):  
Amrutha Bindu ◽  
Lakshmi Devi
Keyword(s):  
Amino Acid ◽  
Antimicrobial Peptide ◽  
Lactobacillus Plantarum ◽  
Exchange Chromatography ◽  
Amino Acid Sequences ◽  
Proteinase K ◽  
Kinetic Assay ◽  
Wide Range

Abstract The focus of present study was to characterize antimicrobial peptide produced by probiotic cultures, Enterococcus durans DB-1aa (MCC4243), Lactobacillus plantarum Cu2-PM7 (MCC4246) and Lactobacillus fermentum Cu3-PM8 (MCC4233) against Staphylococus aureus and E. coli. The growth kinetic assay revealed 24 h of incubation to be optimum for bacteriocin production. The partially purified compound after ion-exchange chromatography was found to be thermoresistant and stable under wide range of pH. The compound was sensitive to proteinase-K, but resistant to trypsin, a-amylase and lipase. The apparent molecular weight of bacteriocin from MCC4243 and MCC4246 was found to be 3.5 KDa. Translated partial amino acid sequence of plnA gene in MCC4246 displayed 48 amino acid sequences showing 100% similarity with plantaricin A of Lactobacillus plantarum (WP_0036419). The sequence revealed 7 β sheets, 6 α sheets, 6 predicted coils and 9 predicted turns. The functions on cytoplasm show 10.82 isoelectric point and 48.6% hydrophobicity. The molecular approach of using Geneious Prime software and protein prediction data base for characterization of bacteriocin is novel and predicts “KSSAYSLQMGATAIKQVKKLFKKWGW” as peptide responsible for antimicrobial activity. The study provides information about broad spectrum bacteriocin in native probiotic culture and paves a way towards its application in functional foods as biopreservative agents.

Preparation and Characterization of a Novel γ-PGA/β-Tricalcium Phosphate Composite for Tissue Engineering

2014 ◽  
Vol 900 ◽  
pp. 306-311 ◽  
Author(s):  
Xiu Lin Shu ◽  
Qing Shan Shi ◽  
Xiao Bao Xie ◽  
Xiao Mo Huang ◽  
Yi Ben Chen
Keyword(s):  
Tissue Engineering ◽  
Cell Growth ◽  
Porous Structure ◽  
Water Absorption ◽  
Tricalcium Phosphate ◽  
Absorption Rate ◽  
Swelling Ratio ◽  
Composite Scaffolds

In order to improvedβ-TCP biocompatibility and cell growth, was chosen to modify β-TCP matrices to produce a γ-PGA/β-TCP composite biomaterial. Then, the morphology, water uptake and retention abilities,in vitrodegradation property in the simulated medium, cytotoxicity of this novel γ-PGA/β-TCP composite is investigated. SEM shows that the γ-PGA/β-TCP composite has a porous structure. By increasing the percentage ofγ-PGA from 0% to 50%, the swelling ratio of the composite s was enhanced from 9.0%to 297%. These data suggested that the surface hydrophilicity, water absorption rate, and swelling ratio were improved by adding γ-PGA to the composite. In the cytocompatibility test, the density of MC3T3-E1 preosteoblasts cells on the PTCP1:1 leachates was almost 110% higher than that on the controls on day 3. Therefore, the γ-PGA/β-TCP composite scaffolds, due to their better hydrophilicity, cytocompatibility, and porous structure, are very promising biomaterials for tissure engineering applications.

scholarly journals Identification and characterization of two amylase producing bacteria Cellulosimicrobium sp. and Demequina sp. isolated from marine organisms

2015 ◽  
Vol 20 ◽  
pp. 8 ◽  
Author(s):  
Laila S.H. Al-Naamani ◽  
Sergey Dobretsov ◽  
Jamal Al-Sabahi ◽  
Bassam Soussi
Keyword(s):  
Cell Growth ◽  
Bacterial Species ◽  
Marine Organisms ◽  
Ulva Rigida ◽  
Wide Range ◽  
Amylase Production ◽  
Maximum Cell ◽  
Commercial Applications ◽  
Identification And Characterization

Marine sources have been known to yield novel compounds with a wide range of bioactivity with various commercial applications. In this study, the abilities of bacteria isolated from eight marine organisms to produce α-amylase were examined. All eight organisms were found to harbor amylase producing bacteria. Two bacterial species isolated from the green alga Ulva rigida and the sponge Mycale sp. were further identified and their α-amylases were purified and characterized. The bacterial species isolated from U. rigida and Mycale sp. were identified by DNA sequencing as Cellulosimicrobium sp. and Demequina sp., respectively. Cellulosimicrobium sp. obtained maximum cell growth and amylase production at 29.C and in the presence of lactose as a carbon source. Optimal cell growth and amylase production by Demequina sp. was observed at 35.C. While lactose enhanced cell growth of Demequina sp., maximum amylase production was found when fructose and glycerol were the available sources of carbon. Both strains grew better in the presence of tryptone, whilst peptone stimulated amylase production. Maximal cell growth and amylase production by both of the strains was found at a medium salinity of 3% NaCl. 

scholarly journals Formulation and Characterization of Glucosamine Sulphate Loaded Carbopol Based Hydrogels for the Management of Osteoarthritis

2020 ◽  
Vol 11 (4) ◽  
pp. 7121-7127
Author(s):  
Sundareswara Kumar Chellaswamy ◽  
Satheesh Babu Natrajan
Keyword(s):  
Skin Irritation ◽  
Glucosamine Sulfate ◽  
Effective Dosage ◽  
Permeation Enhancers ◽  
Simple Method ◽  
Wide Range ◽  
Glucosamine Sulphate ◽  
Tween 40

Osteoarthritis is emerging as the most ordinary form of arthritis, affecting 22-39% of the Indian population. A wide range of medications and therapies are available for the treatment of osteoarthritis. With a desire to develop a therapeutically effective dosage form, the present study was carried out to formulate glucosamine sulfate loaded carbopol based hydrogel. Hydrogels H1 to H6 were formulated without permeation enhancers while formulations H7 to H12 were developed with a different class of permeation enhancers such as PEG400, oleic acid, Tween 40, DMSO and PG. Based on viscosity, it was detected that formulation H4 containing polymer 1% was ideal for incorporating drug. Considering H4 as a placebo, H6 was used for further evaluation. Drug content was found to be 99.2±0.64, with in vitro  drug release of 15±0.86, 22±1.59, 28±0.72, 35±0.68, 40±0.31, 47±0.83, 58±1.59, and 70±0.9 % at a duration of 1, 2, 3, 4, 5, 6, 7, 8 hours respectively. Skin irritation tests carried out on Wistar rats revealed that skin was intact with no inflammation or erythema detected, compared to untreated site. By diffusion disc method, it was evident that the levels of microbial load were relatively low, and no harmful microorganisms were identified. There were no significant changes in physicochemical properties on stability studies. Due to a simple method of preparation and effective drug delivery, glucosamine sulfate loaded hydrogels could be contemplated as a prominent formulation in the beneficiary treatment of osteoarthritis.

scholarly journals Characterization of Andean Blueberry in Bioactive Compounds, Evaluation of Biological Properties and in vitro Bioaccessibility

2020 ◽  
Author(s):  
Nieves Baenas ◽  
Jenny Ruales ◽  
Diego A. Moreno ◽  
Daniel Alejandro Barrio ◽  
Carla M. Stinco ◽  
...  
Keyword(s):  
Antioxidant Capacity ◽  
Bioactive Compounds ◽  
Biological Activities ◽  
In Vitro Digestion ◽  
Biological Properties ◽  
Wide Range ◽  
Zebrafish Embryogenesis

Andean blueberries are wild berries grown and consumed in Ecuador which contain high values of bioactive compounds, mainly anthocyanins, with powerful antioxidant activity. The aim of this study was to evaluate the profile and contents of (poly)phenols and carotenoids in Andean blueberry by HPLC-DAD-MSn and determine a wide range of its biological activities. The antioxidant capacity of this fruit was evaluated in vitro by three different methods and in vivo using the zebrafish animal model, also the toxicity effect was determined by the zebrafish embryogenesis test. Besides, the antimicrobial activity and the capacity of Andean blueberry to produce hemagglutination in blood cells were evaluated. Finally, the bioaccessibility of (poly)phenols and related antioxidant capacity were determined in the different phases of an in vitro digestion. The global results indicated no toxicity of Andean blueberry, weakly bacteriostatic activity, and high contents of anthocyanins and antioxidant capacity, which were partially bioaccesible in vitro (~ 50 % at the final intestinal step), contributing to the knowledge of its health benefits for consumers and its potential use in the food and pharmaceutical industry as functional ingredient.

scholarly journals Semi-automated high-throughput substrate screening assay for nucleoside kinases

2021 ◽  
Author(s):  
Katja Hellendahl ◽  
Maryke Fehlau ◽  
Sebastian Hans ◽  
Peter Neubauer ◽  
Anke Kurreck
Keyword(s):  
High Throughput ◽  
Viral Infections ◽  
Screening Assay ◽  
Liquid Handling ◽  
Wide Range ◽  
High Throughput Assay ◽  
Liquid Handling Robot

Nucleoside kinases (NKs) are key enzymes involved in the in vivo phosphorylation of nucleoside analogues used as drugs to treat cancer or viral infections. Having different specificities, the characterization of NKs is essential for drug design and the production of nucleotide analogues in an in vitro enzymatic process. Therefore, a fast and reliable substrate screening assay for NKs is of great importance. Here, we report the validation of a well-known luciferase-based assay for the detection of NK activity in 96-well plate format. The assay was semi-automated using a liquid handling robot. A good linearity was demonstrated (r² >0.98) in the range of 0 to 500 µM ATP, and it was shown that also alternative phosphate donors like dATP or CTP were accepted by the luciferase. The developed high-throughput assay revealed comparable results to HPLC analysis. The assay was exemplary used for the comparison of the substrate spectra of four nucleoside kinases using 20 (8 natural and 12 modified) substrates. The screening results correlated well with literature data and, additionally, previously unknown substrates were identified for three of the NKs studied. Our results demonstrate that the developed semi-automated high-throughput assay is suitable to identify best performing NKs for a wide range of substrates.

F.05 Characterization of NBCA glue polymerization for embolization of brain AVM’s

2016 ◽  
Vol 43 (S2) ◽  
pp. S19-S20
Author(s):  
BH Wang ◽  
D Pelz ◽  
D Lee ◽  
MR Boulton ◽  
SP Lownie
Keyword(s):  
High Speed ◽  
Contact Angles ◽  
Polymerization Rate ◽  
Physical Interaction ◽  
Wide Range ◽  
Glue Injection ◽  
Glue Embolization

Background: Brain arteriovenous malformations (AVM’s) are abnormal connections between arteries and veins. Endovascular glue embolization with N-butyl cyanoacrylate (NBCA) is an accepted form of treatment, with most complications related to timing of polymerization. Current literature reports a wide range of polymerization times with large discrepancies between in-vivo and in-vitro results. Methods: Polymerization time was measured for mixtures of lipiodol/NBCA of 50/50, 60/40, 70/30. The influence of pH, temperature and presence of biological catalysts on polymerization rate was investigated in-vivo using submerged droplet tests. PVA-C, silicone and endothelium surfaces were compared and contact angles were measured to assess physical interaction with NBCA. High-speed video of glue injection through a microcatheter was captured to characterize coaxial flow. Results: Polymerization rate increases with pH and temperature. A hydrophilic substrate such as PVA-C provides surface properties that are most similar to endothelium. Endothelium provides a catalytic surface that increases the rate of polymerization. Blood products further increase the polymerization rate with RBC’s providing almost instantaneous polymerization of NBCA upon contact. Characterization of coaxial flow shows dripping to jetting transition with significant wall effect. Conclusions: We have successfully deconstructed and characterized the dynamic behavior of NBCA embolization. A refined understanding of NBCA behavior could help reduce embolization-related complications.

scholarly journals Mesenchymal stromal cells orchestrate follicular lymphoma cell niche through the CCL2-dependent recruitment and polarization of monocytes

Blood ◽  
2012 ◽  
Vol 119 (11) ◽  
pp. 2556-2567 ◽  
Author(s):  
Fabien Guilloton ◽  
Gersende Caron ◽  
Cédric Ménard ◽  
Céline Pangault ◽  
Patricia Amé-Thomas ◽  
...  
Keyword(s):  
B Cells ◽  
Cell Growth ◽  
Follicular Lymphoma ◽  
B Cell ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Specific Gene ◽  
Wide Range ◽  
Cell Niche

Abstract Accumulating evidence indicates that infiltrating stromal cells contribute directly and indirectly to tumor growth in a wide range of cancers. In follicular lymphoma (FL), malignant B cells are found admixed with heterogeneous lymphoid-like stromal cells within invaded lymph nodes and BM. In addition, mesenchymal stromal cells (MSCs) support in vitro FL B-cell survival, in particular after their engagement toward lymphoid differentiation. We show here that BM-MSCs obtained from patients with FL (FL-MSCs) display a specific gene expression profile compared with MSCs obtained from healthy age-matched donors (HD-MSCs). This FL-MSC signature is significantly enriched for genes associated with a lymphoid-like commitment. Interestingly, CCL2 could be detected at a high level within the FL-cell niche, is up-regulated in HD-MSCs by coculture with malignant B cells, and is overexpressed by FL-MSCs, in agreement with their capacity to recruit monocytes more efficiently than HD-MSCs. Moreover, FL-MSCs and macrophages cooperate to sustain malignant B-cell growth, whereas FL-MSCs drive monocyte differentiation toward a proangiogenic and lipopolysaccharide-unresponsive phenotype close to that of tumor-associated macrophages. Altogether, these results highlight the complex role of FL stromal cells that promote direct tumor B-cell growth and orchestrate FL-cell niche, thus emerging as a potential therapeutic target in this disease.

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