scholarly journals Noninvasive Superoxide Monitoring ofIn VitroNeuronal Differentiation Using a Cell-Based Biosensor

2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Georgia Moschopoulou ◽  
Spyridon Kintzios
Keyword(s):  
Superoxide Dismutase ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Neuronal Differentiation ◽  
Superoxide Anion ◽  
Culture Medium ◽  
Ultrasensitive Detection ◽  
A Cell

Membrane-engineered cells bearing superoxide dismutase (SOD) molecules on their surface offer the capability of ultrarapid and ultrasensitive detection of the superoxide anion (O2-) through the measurement of changes of their cell membrane potential. We herewith report the application of this technology for the noninvasive determination of superoxide levels during thein vitrodifferentiation of PC12 cells. We were able to detect changes inO2-accumulation in the culture medium, which were closely associated with the progress of neuronal differentiation.

scholarly journals Imaging the transmembrane and transendothelial sodium gradients in gliomas

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Muhammad H. Khan ◽  
John J. Walsh ◽  
Jelena M. Mihailović ◽  
Sandeep K. Mishra ◽  
Daniel Coman ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Magnetic Resonance ◽  
Cell Membrane ◽  
Normal Tissue ◽  
Magnetic Resonance Spectroscopic Imaging ◽  
Biological Factors ◽  
High Sodium ◽  
Intracellular Milieu

AbstractUnder normal conditions, high sodium (Na+) in extracellular (Na+e) and blood (Na+b) compartments and low Na+ in intracellular milieu (Na+i) produce strong transmembrane (ΔNa+mem) and weak transendothelial (ΔNa+end) gradients respectively, and these manifest the cell membrane potential (Vm) as well as blood–brain barrier (BBB) integrity. We developed a sodium (23Na) magnetic resonance spectroscopic imaging (MRSI) method using an intravenously-administered paramagnetic polyanionic agent to measure ΔNa+mem and ΔNa+end. In vitro 23Na-MRSI established that the 23Na signal is intensely shifted by the agent compared to other biological factors (e.g., pH and temperature). In vivo 23Na-MRSI showed Na+i remained unshifted and Na+b was more shifted than Na+e, and these together revealed weakened ΔNa+mem and enhanced ΔNa+end in rat gliomas (vs. normal tissue). Compared to normal tissue, RG2 and U87 tumors maintained weakened ΔNa+mem (i.e., depolarized Vm) implying an aggressive state for proliferation, whereas RG2 tumors displayed elevated ∆Na+end suggesting altered BBB integrity. We anticipate that 23Na-MRSI will allow biomedical explorations of perturbed Na+ homeostasis in vivo.

Non-contact-based determination of membrane permeability to water and dimethyl sulfoxide of cells virtually trapped in a self-induced micro-vortex

Lab on a Chip ◽  
2021 ◽  
Author(s):  
Hsiu-Yang Tseng ◽  
Chiu-Jen Chen ◽  
Zong-Lin Wu ◽  
Yong-Ming Ye ◽  
Guo-Zhen Huang
Keyword(s):  
Cell Membrane ◽  
Dimethyl Sulfoxide ◽  
Membrane Permeability ◽  
Cell Membrane Permeability ◽  
Cell Type ◽  
Cryoprotective Agents ◽  
A Cell

Cell-membrane permeability to water (Lp) and cryoprotective agents (Ps) of a cell type is a crucial cellular information for achieving optimal cryopreservation in the biobanking industry. In this work, a...

In Vitro Removal of Ochratoxin A by Wine Lactic Acid Bacteria

2007 ◽  
Vol 70 (9) ◽  
pp. 2155-2160 ◽  
Author(s):  
VINCENZO DEL PRETE ◽  
HECTOR RODRIGUEZ ◽  
ALFONSO V. CARRASCOSA ◽  
BLANCA de las RIVAS ◽  
EMILIA GARCIA-MORUNO ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Ochratoxin A ◽  
High Performance ◽  
Culture Medium ◽  
Degradation Products ◽  
Cell Binding ◽  
A Cell ◽  
In Vitro Interaction

A study was carried out to determine the in vitro interaction between ochratoxin A (OTA) and wine lactic acid bacteria (LAB). Fifteen strains belonging to five relevant oenological LAB species were grown in liquid synthetic culture medium containing OTA. The portion of OTA removed during the bacterial growth was 8 to 28%. The OTA removed from the supernatants was partially recovered (31 to 57%) from the bacterial pellet. Cell-free extracts of three representative strains were produced by disrupting cells in a French pressure cell. The ability of crude cell-free extracts to degrade OTA was studied. OTA was not degraded by cell-free extracts of wine LAB strains, and no degradation products of OTA were detected in the high-performance liquid chromatograms of the methanol extract of the bacterial pellet. On the basis of these results, we conclude that OTA removal by wine LAB is a cell-binding phenomenon. The chemistry and the molecular basis of OTA binding to wine LAB remains unknown.

Studies on growth of the mink blastocyst

Development ◽  
1967 ◽  
Vol 17 (2) ◽  
pp. 293-302
Author(s):  
Joseph C. Daniel
Keyword(s):  
Culture Medium ◽  
Present Report ◽  
Blastocyst Stage ◽  
Pregnant Female ◽  
Culture Methods ◽  
Direct Treatment ◽  
Breeding Seasons

In those mammals in which implantation is delayed, the embryo enters a diapause at the blastocyst stage. The present report describes experiments with mink over the last three breeding seasons, attempting to define the factors that limit development at this stage. Four approaches to the problem were used: (1) determination of growth of mink blastocysts in vitro with specific modifications of media; (2) transplantation of mink embryos to rabbit uteri; (3) direct treatment of pregnant female mink with ergosterol; (4) growth of rabbit blastocysts in vitro in medium containing mink serum. (1) The culture methods used for mink embryos were those developed for the rabbit and described in earlier publications (Daniel, 1963, 1965). Mink blastocysts (Plate 1) were isolated in culture medium after being flushed from the uteri of mothers bred 9-20 days earlier. Various components were added to the medium, F10 (Ham, 1963), in concentrations that were previously tested against rabbit blastocysts and found to be non-toxic, and, in some cases, beneficial to growth.

scholarly journals Cortical Thinning and Hydrocephalus in Mice Lacking the Immunoglobulin Superfamily Member CDO

2006 ◽  
Vol 26 (10) ◽  
pp. 3764-3772 ◽  
Author(s):  
Wei Zhang ◽  
Min-Jeong Yi ◽  
Xiaoping Chen ◽  
Francesca Cole ◽  
Robert S. Krauss ◽  
...  
Keyword(s):  
Neuronal Differentiation ◽  
Myogenic Differentiation ◽  
Immunoglobulin Superfamily ◽  
Cortical Thinning ◽  
Helix Loop Helix ◽  
Neuronal Precursor Cells ◽  
A Cell ◽  
Multiple Cell

ABSTRACT CDO is a cell surface immunoglobulin superfamily member that positively regulates myogenic differentiation in vitro and in vivo and signals to posttranslationally activate myogenic basic helix-loop-helix (bHLH) transcription factors. The Cdo gene is also expressed in the dorsal aspect and midline structures of the developing central nervous system, and mice lacking CDO on the C57BL/6 background display holoprosencephaly with ∼80% penetrance, resulting in perinatal lethality. We report here that a fraction of Cdo −/− mice from this background have additional defects in brain development, including hydrocephalus and cortical thinning. Primary neural progenitor cultures from E14.5 Cdo −/− mutants display reduced proliferation, which may underlie the thinning. The cortical preplate and cortices of mutant animals also show reduced staining for β-tubulin III, indicating defective neuronal differentiation. CDO levels are strongly increased in cultured C17.2 neuronal precursor cells stimulated to differentiate; modulation of CDO levels in these cells by overexpression or interfering RNA approaches enhances or diminishes differentiation, respectively. Cotransfection of CDO enhances the activity of the neurogenic bHLH factor, neurogenin1, in reporter assays and enhances heterodimerization of neurogenin1 and E47. These results indicate that CDO promotes neuronal differentiation and support the hypothesis that CDO coordinates differentiation of multiple cell lineages by regulating the activity of tissue-specific bHLH factors.

scholarly journals Comparison of Extracellular and Intracellular Potency of Botulinum Neurotoxins

2006 ◽  
Vol 74 (10) ◽  
pp. 5617-5624 ◽  
Author(s):  
Fang Cai ◽  
Carrie B. Adrion ◽  
James E. Keller
Keyword(s):  
Culture Medium ◽  
Reaction Model ◽  
Cultured Neurons ◽  
Total Inhibition ◽  
Botulinum Neurotoxins ◽  
A Cell ◽  
Proteolytic Activities ◽  
Living Neurons ◽  
Subsequent Addition

ABSTRACT Levels of botulinum neurotoxin (BoNT) proteolytic activity were compared using a cell-free assay and living neurons to measure extracellular and intracellular enzymatic activity. Within the cell-free reaction model, BoNT serotypes A and E (BoNT/A and BoNT/E, respectively) were reversibly inhibited by chelating Zn2+ with N,N,N′,N′-tetrakis (2-pyridylmethyl) ethylenediamine (TPEN). BoNT/E required relatively long incubation with TPEN to achieve total inhibition, whereas BoNT/A was inhibited immediately upon mixing. When naïve Zn2+-containing BoNTs were applied to cultured neurons, the cellular action of each BoNT was rapidly inhibited by subsequent addition of TPEN, which is membrane permeable. Excess Zn2+ added to the culture medium several hours after poisoning fully restored intracellular toxin activity. Unlike TPEN, EDTA irreversibly inhibited both BoNT/A and -E within the cell-free in vitro reaction. Excess Zn2+ did not reactivate the EDTA-treated toxins. However, application of EDTA-treated BoNT/A or -E to cultured neurons demonstrated normal toxin action in terms of both blocking neurotransmission and SNAP-25 proteolysis. Different concentrations of EDTA produced toxin preparations with incrementally reduced in vitro proteolytic activities, which, when applied to living neurons showed undiminished cellular potency. This suggests that EDTA renders the BoNT proteolytic domain conformationally inactive when tested with the cell-free reaction, but this change is corrected during entry into neurons. The effect of EDTA is unrelated to Zn2+ because TPEN could be applied to living cells before or after poisoning to produce rapid and reversible inhibition of both BoNTs. Therefore, bound Zn2+ is not required for toxin entry into neurons, and removal of Zn2+ from cytosolic BoNTs does not irreversibly alter toxin structure or function. We conclude that EDTA directly alters both BoNTs in a manner that is independent of Zn2+.

scholarly journals In Vitro Antibacterial Activity and Mechanism of Vanillic Acid against Carbapenem-Resistant Enterobacter cloacae

Antibiotics ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 220 ◽  
Author(s):  
Weidong Qian ◽  
Yuting Fu ◽  
Miao Liu ◽  
Ting Wang ◽  
Jianing Zhang ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Crystal Violet ◽  
Vanillic Acid ◽  
Membrane Integrity ◽  
Enterobacter Cloacae ◽  
Carbapenem Resistant ◽  
Atp Concentration

Vanillic acid (VA) is a flavoring agent found in edible plants and fruits. Few recent studies exhibited robust antibacterial activity of VA against several pathogen microorganisms. However, little was reported about the effect of VA on carbapenem-resistant Enterobacter cloacae (CREC). The purpose of the current study was to assess in vitro antimicrobial and antibiofilm activities of VA against CREC. Here, minimum inhibitory concentrations (MIC) of VA against CREC was determined via gradient diffusion method. Furthermore, the antibacterial mode of VA against CREC was elucidated by measuring changes in intracellular adenosine triphosphate (ATP) concentration, intracellular pH (pHin), cell membrane potential and membrane integrity. In addition, antibiofilm formation of VA was measured by crystal violet assay and visualized with field emission scanning electron microscopy (FESEM) and confocal laser scanning microscopy (CLSM). The results showed that MIC of VA against E. cloacae was 600 μg/mL. VA was capable of inhibiting the growth of CREC and destroying the cell membrane integrity of CREC, as confirmed by the decrease of intracellular ATP concentration, pHin and membrane potential as well as distinctive variation in cellular morphology. Moreover, crystal violet staining, FESEM and CLSM results indicated that VA displayed robust inhibitory effects on biofilm formation of CREC and inactivated biofilm-related CREC cells. These findings revealed that VA exhibits potent antibacterial activity against CREC, and thus has potential to be exploited as a natural preservative to control the CREC associated infections.

scholarly journals BIOMIMETIC APPROACHES TO STUDY OF PROPERTIES OF MEDICINAL SUBSTANCES

2019 ◽  
Vol 62 (10) ◽  
pp. 4-29
Author(s):  
Nina B. Melnikova ◽  
Olga N. Solovyova ◽  
Evgeni N. Evgeni N. Kochetkov
Keyword(s):  
Antioxidant Activity ◽  
Superoxide Dismutase ◽  
Lipid Bilayers ◽  
Superoxide Anion ◽  
Molecular Form ◽  
Biologically Active ◽  
New Drugs ◽  
In Vitro Experiments ◽  
Medicinal Substances

The review is devoted to an assessment of the current level of use of biomimetic approaches to the study of the properties of known drugs and the development of new drugs. In this review, we consider the main biological functions of superoxide dismutase, namely the catalytic decomposition of toxic superoxide anion of oxygen to the molecular form of oxygen and protection against induced apoptosis. The biomimetic enzymes-Mn- and TEMPO-containing equivalents of superoxide dismutase SOD with antitumor and antioxidant activity were discussed more detail. The relationship between the properties and activity of SOD mimetics with their structure among them the nature of the anion and ligands, the coordination number, the geometry of the presence of conjugated bonds, and other parameters of the molecules. The study of the properties of Mn-SOD mimetics makes it possible to develop a new class of drugs successfully tested by in vivo and in vitro experiments and which are at the stages of clinical trials. Stable TEMPO radicals containing compounds are able to perform SOD functions, exhibiting antioxidant activity in relation not only to superoxide-anion, but also to peroxynitrile, and moreover to act as a spin label. The biomimetic membrane systems (monolayers, planar lipid bilayers, liposomes and other nano-sized objects) are discussed too for studying properties in in vitro experiments and for delivering potent and medicinal substances. The biomimetic approach combination allows to create the new promising drugs, including those based on SOD mimetics, and to develop the synthetic analogues of biologically active substances and methods of their delivery. The advantages of such dosage forms are lower toxicity of the preparations, lack of immunogenicity and a decrease in the dose of potent drugs.

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