Influence of the media ionic strength on the formation and in vitro biological performance of polycation-DNA complexes

2021 ◽  
pp. 117930
Author(s):  
Filipe Coelho ◽  
Claudia Botelho ◽  
Juan L. Paris ◽  
Eduardo F. Marques ◽  
Bruno F. B. Silva
Keyword(s):  
Ionic Strength ◽  
Dna Complexes ◽  
The Media ◽  
Biological Performance

scholarly journals Influence of Glucose Concentration on Colony-Forming Efficiency and Biological Performance of Primary Human Tissue–Derived Progenitor Cells

Cartilage ◽  
2020 ◽  
pp. 194760352090660 ◽  
Author(s):  
Venkata P. Mantripragada ◽  
Ryan Kaplevatsky ◽  
Wes A. Bova ◽  
Cynthia Boehm ◽  
Nancy A. Obuchowski ◽  
...  
Keyword(s):  
Glucose Concentration ◽  
Pairwise Comparison ◽  
Culture Methods ◽  
Glucose Levels ◽  
Colony Forming Efficiency ◽  
Forming Efficiency ◽  
The Media ◽  
Outerbridge Grade ◽  
Biological Performance

Objective Glucose concentrations used in current cell culture methods are a significant departure from physiological glucose levels. The study focuses on comparing the effects of glucose concentrations on primary human progenitors (connective tissue progenitors [CTPs]) used for cartilage repair. Design Cartilage- (Outerbridge grade 1, 2, 3; superficial and deep zone cartilage), infrapatellar fatpad-, synovium-, and periosteum-derived cells were obtained from 63 patients undergoing total knee arthroplasty and cultured simultaneously in fresh chondrogenic media containing 25 mM glucose (HGL) or 5 mM glucose (NGL) for pairwise comparison. Automated ASTM-based quantitative image analysis was used to determine colony-forming efficiency (CFE), effective proliferation rates (EPR), and sulfated-proteoglycan (GAG-ECM) staining of the CTPs across tissue sources. Results HGL resulted in increased cell cultures with CFE = 0 compared with NGL in all tissue sources ( P = 0.049). The CFE in NGL was higher than HGL for superficial cartilage ( P < 0.001), and contrary for synovium-derived CTPs ( P = 0.046) when CFE > 0. EPR of the CTPs did not differ between the media in the 6-day assay time period ( P = 0.082). The GAG-ECM area of the CTPs and their progeny was increased in presence of HGL ( P = 0.027). Conclusion Glucose concentration is critical to progenitor’s physiology and should be taken into account in the setting of protocols for clinical or in vitro cell expansion strategies.

A topoisomerase from chicken erythrocyte nuclei which does not assemble nucleosome core particles in vitro

1982 ◽  
Vol 60 (6) ◽  
pp. 651-658 ◽  
Author(s):  
Michael J. Ellison ◽  
David E. Pulleyblank
Keyword(s):  
Ionic Strength ◽  
Homogeneous Distribution ◽  
Chicken Erythrocyte ◽  
Nucleosome Assembly ◽  
Core Particle ◽  
Dna Complexes ◽  
Particle Assembly ◽  
Nucleosome Core ◽  
Nuclease Digestion

We have examined the ability of a topoisomerase purified from chicken erythrocyte nuclei to mediate nucleosome core assembly in vitro at physiological ionic strength (0.15 M NaCl). Although we have detected limited amounts of spontaneously assembled nucleosome cores at this salt concentration, the addition of this topoisomerase does not increase the amount of assembly observed. Nucleosome assembly was assayed by quantitating the amount of core particle length DNA accumulated with time upon the nuclease digestion of histone–DNA complexes. In addition, the amount of negative supercoils introduced into relaxed closed circular DNA upon nucleosome core particle assembly was determined. Correctly assembled complexes do not protect more DNA from nuclease digestion than random histone–DNA complexes but shift the heterogeneous size distribution of protected fragments to a more homogeneous distribution centred around 145 base pairs. Under our conditions of nucleosome assembly, a second histone–DNA complex which is distinct from the nucleosome core can be detected under physiological ionic strength conditions. This particle does not form in high salt assembly experiments. Similarly, the assembly of this particle is unaffected by the presence or absence of topoisomerase.

scholarly journals Enzymic oxidation of unconjugated bilirubin by rat liver

1986 ◽  
Vol 236 (3) ◽  
pp. 625-633 ◽  
Author(s):  
R Cardenas-Vazquez ◽  
O Yokosuka ◽  
B H Billing
Keyword(s):  
Ionic Strength ◽  
Rat Liver ◽  
Mitochondrial Protein ◽  
Subcellular Fractionation ◽  
Sprague Dawley ◽  
Bilirubin Oxidase ◽  
Outer Membranes ◽  
The Media ◽  
The Mean

The presence of the enzyme bilirubin oxidase, which degrades bilirubin in vitro, was demonstrated in the liver. Subcellular-fractionation experiments indicate that bilirubin oxidase is located in both the inner and outer membranes of the mitochondria. The mean rate of the reaction is 1.57 +/- 0.38 (S.D.) nmol of bilirubin degraded/min per mg of mitochondrial protein (munits/mg of protein). With respect to the overall breakdown of bilirubin, the enzyme has a Km' of 136 microM-bilirubin and a Vmax.' of 9.13 munits/mg of protein. Its activity is influenced by the ionic strength of the media and is inhibited by KCN, thiol reagents, NADH and albumin. The enzyme is aerobic, and between 1 and 1.5 mol of O2 are consumed per mol of bilirubin degraded. The products of the reaction include propentdyopents. The hepatic bilirubin oxidase activity of the jaundiced Gunn-rat liver is not significantly different from that of the Sprague-Dawley rat, and it is not induced by beta-naphthoflavone.

Torus Circumference and Thickness Parameters From a Linear DNA Size Series Suggest How Toruses Self-Assemble

1985 ◽  
Vol 43 ◽  
pp. 522-523
Author(s):  
George C. Ruben ◽  
Kenneth A. Marx
Keyword(s):  
Tertiary Structure ◽  
Dna Complexes ◽  
Tertiary Structures ◽  
Self Assembling ◽  
Double Stranded Dna ◽  
Calf Thymus ◽  
Dna Organization ◽  
Condensed Dna ◽  
Dna Size

Certain double stranded DNA bacteriophage and viruses are thought to have their DNA organized into large torus shaped structures. Morphologically, these poorly understood biological DNA tertiary structures resemble spermidine-condensed DNA complexes formed in vitro in the total absence of other macromolecules normally synthesized by the pathogens for the purpose of their own DNA packaging. Therefore, we have studied the tertiary structure of these self-assembling torus shaped spermidine- DNA complexes in a series of reports. Using freeze-etch, low Pt-C metal (10-15Å) replicas, we have visualized the microscopic DNA organization of both calf Thymus( CT) and linear 0X-174 RFII DNA toruses. In these structures DNA is circumferentially wound, continuously, around the torus into a semi-crystalline, hexagonal packed array of parallel DNA helix sections.

Efficient gene transfer to dispersed human pancreatic islet cells in vitro using adenovirus-polylysine/DNA complexes or polycationic liposomes

Diabetes ◽  
1996 ◽  
Vol 45 (9) ◽  
pp. 1197-1203 ◽  
Author(s):  
J. Saldeen ◽  
D. T. Curiel ◽  
D. L. Eizirik ◽  
A. Andersson ◽  
E. Strandell ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Pancreatic Islet ◽  
Islet Cells ◽  
Pancreatic Islet Cells ◽  
Dna Complexes ◽  
Human Pancreatic Islet ◽  
Efficient Gene

scholarly journals The in vitro assessment of the toxicity of volatile, oxidisable, redox-cycling compounds: phenols as an example

2021 ◽  
Author(s):  
Laia Tolosa ◽  
Teresa Martínez-Sena ◽  
Johannes P. Schimming ◽  
Erika Moro ◽  
Sylvia E. Escher ◽  
...  
Keyword(s):  
Cultured Cells ◽  
Degradation Products ◽  
Degradation Kinetics ◽  
Redox Cycling ◽  
In Vitro Toxicity ◽  
Cross Contamination ◽  
Physicochemical Changes ◽  
In Vitro Assessment ◽  
Biological Performance

AbstractPhenols are regarded as highly toxic chemicals. Their effects are difficult to study in in vitro systems because of their ambiguous fate (degradation, auto-oxidation and volatility). In the course of in vitro studies of a series of redox-cycling phenols, we found evidences of cross-contamination in several in vitro high-throughput test systems, in particular by trimethylbenzene-1, 4-diol/trimethylhydroquinone (TMHQ) and 2,6-di-tertbutyl-4-ethylphenol (DTBEP), and investigated in detail the physicochemical basis for such phenomenon and how to prevent it. TMHQ has fast degradation kinetics followed by significant diffusion rates of the resulting quinone to adjacent wells, other degradation products being able to air-diffuse as well. DTBEP showed lower degradation kinetics, but a higher diffusion rate. In both cases the in vitro toxicity was underestimated because of a decrease in concentration, in addition to cross-contamination to neighbouring wells. We identified four degradation products for TMHQ and five for DTBEP indicating that the current effects measured on cells are not only attributable to the parent phenolic compound. To overcome these drawbacks, we investigated in detail the physicochemical changes occurring in the course of the incubation and made use of gas-permeable and non-permeable plastic seals to prevent it. Diffusion was greatly prevented by the use of both plastic seals, as revealed by GC–MS analysis. Gas non-permeable plastic seals, reduced to a minimum compounds diffusion as well oxidation and did not affect the biological performance of cultured cells. Hence, no toxicological cross-contamination was observed in neighbouring wells, thus allowing a more reliable in vitro assessment of phenol-induced toxicity.

scholarly journals The effect of pH and ionic strength of dissolution media on in-vitro release of two model drugs of different solubilities from HPMC matrices

2013 ◽  
Vol 111 ◽  
pp. 384-391 ◽  
Author(s):  
Kofi Asare-Addo ◽  
Barbara R. Conway ◽  
Hassan Larhrib ◽  
Marina Levina ◽  
Ali R. Rajabi-Siahboomi ◽  
...  
Keyword(s):  
Ionic Strength ◽  
In Vitro Release ◽  
Effect Of Ph ◽  
Vitro Release

scholarly journals Viability of ‘Candidatus Liberibacter asiaticus’ Prolonged by Addition of Citrus Juice to Culture Medium

2014 ◽  
Vol 104 (1) ◽  
pp. 15-26 ◽  
Author(s):  
Jennifer K. Parker ◽  
Sarah R. Wisotsky ◽  
Evan G. Johnson ◽  
Faraj M. Hijaz ◽  
Nabil Killiny ◽  
...  
Keyword(s):  
Grapefruit Juice ◽  
Culture Medium ◽  
Quantitative Polymerase Chain Reaction ◽  
Chemical Characterization ◽  
Candidatus Liberibacter Asiaticus ◽  
Citrus Greening Disease ◽  
Candidatus Liberibacter ◽  
The Media ◽  
Liberibacter Asiaticus

Huanglongbing, or citrus greening disease, is associated with infection by the phloem-limited bacterium ‘Candidatus Liberibacter asiaticus’. Infection with ‘Ca. L. asiaticus’ is incurable; therefore, knowledge regarding ‘Ca. L. asiaticus’ biology and pathogenesis is essential to develop a treatment. However, ‘Ca. L. asiaticus’ cannot currently be successfully cultured, limiting its study. To gain insight into the conditions conducive for growth of ‘Ca. L. asiaticus’ in vitro, ‘Ca. L. asiaticus’ inoculum obtained from seed of fruit from infected pomelo trees (Citrus maxima ‘Mato Buntan’) was added to different media, and cell viability was monitored for up to 2 months using quantitative polymerase chain reaction in conjunction with ethidium monoazide. Media tested included one-third King's B (K), K with 50% juice from the infected fruit, K with 50% commercially available grapefruit juice, and 100% commercially available grapefruit juice. Results show that juice-containing media dramatically prolong viability compared with K in experiments reproduced during 2 years using different juice sources. Furthermore, biofilm formed at the air–liquid interface of juice cultures contained ‘Ca. L. asiaticus’ cells, though next-generation sequencing indicated that other bacterial genera were predominant. Chemical characterization of the media was conducted to discuss possible factors sustaining ‘Ca. L. asiaticus’ viability in vitro, which will contribute to future development of a culture medium for ‘Ca. L. asiaticus’.

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