scholarly journals Oxygen and Glucose Levels in Cell Culture Media Determine Resveratrol’s Effects on Growth, Hydrogen Peroxide Production, and Mitochondrial Dynamics

Antioxidants ◽  
2018 ◽  
Vol 7 (11) ◽  
pp. 157 ◽  
Author(s):  
Joao Fonseca ◽  
Fereshteh Moradi ◽  
Andrew Valente ◽  
Jeffrey Stuart
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Culture ◽  
Cell Growth ◽  
Cultured Cells ◽  
Culture Media ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Network ◽  
Hydrogen Peroxide Production ◽  
Network Characteristics ◽  
Glucose Levels

Resveratrol is a plant-derived polyphenol that has been widely studied for its putative health promoting effects. Many of those studies have been conducted in cell culture, in supra-physiological levels of oxygen and glucose. Resveratrol interacts with reactive oxygen species (ROS) as an antioxidant or pro-oxidant. Resveratrol affects the expression and activities of ROS-producing enzymes and organelles. It is therefore important to consider how cell culture conditions might determine the effects of resveratrol on cultured cells. We determined the effects of resveratrol on cell growth, hydrogen peroxide production, and mitochondrial network characteristics in C2C12 mouse myoblasts and PC3 human prostate cancer cells under conditions of physiological (5%) and supra-physiological (18%) oxygen, and normo- (5 mM) and hyper-glycemia (25 mM). Interestingly, most effects of resveratrol on the parameters measured here were dependent upon prevailing oxygen and glucose levels during the experiment. Many of the effects of resveratrol on cell growth, hydrogen peroxide production, and mitochondrial network characteristics that were seen in 25 mM glucose and/or 18% oxygen were absent under the physiologically relevant conditions of 5 mM glucose with 5% oxygen. These findings emphasize the importance of using physiologically meaningful starting conditions for cell-culture experiments with resveratrol and indeed any manipulation affecting ROS metabolism and mitochondria.

scholarly journals Reversible Bonding of Thermoplastic Elastomers for Cell Patterning Applications

Processes ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 54
Author(s):  
Byeong-Ui Moon ◽  
Keith Morton ◽  
Kebin Li ◽  
Caroline Miville-Godin ◽  
Teodor Veres
Keyword(s):  
Cell Culture ◽  
Cell Migration ◽  
Cell Growth ◽  
Cultured Cells ◽  
Culture Media ◽  
Fibroblast Cell ◽  
Thermoplastic Elastomers ◽  
Cell Patterning ◽  
Migration Studies ◽  
Cell Patterns

In this paper, we present a simple, versatile method that creates patterns for cell migration studies using thermoplastic elastomer (TPE). The TPE material used here can be robustly, but reversibly, bonded to a variety of plastic substrates, allowing patterning of cultured cells in a microenvironment. We first examine the bonding strength of TPE to glass and polystyrene substrates and com-pare it to thermoset silicone-based PDMS under various conditions and demonstrate that the TPE can be strongly and reversibly bonded on commercially available polystyrene culture plates. In cell migration studies, cell patterns are templated around TPE features cored from a thin TPE film. We show that the significance of fibroblast cell growth with fetal bovine serum (FBS)-cell culture media compared to the cells cultured without FBS, analyzed over two days of cell culture. This simple approach allows us to generate cell patterns without harsh manipulations like scratch assays and to avoid damaging the cells. We also confirm that the TPE material is non-toxic to cell growth and supports a high viability of fibroblasts and breast cancer cells. We anticipate this TPE-based patterning approach can be further utilized for many other cell patterning applications such as in cell-to-cell communication studies.

Change in mutation frequency at a TP53 hotspot during culture of ENU-mutagenised human lymphoblastoid cells

Mutagenesis ◽  
2019 ◽  
Author(s):  
Masahiko Watanabe ◽  
Masae Toudou ◽  
Taeko Uchida ◽  
Misato Yoshikawa ◽  
Hiroaki Aso ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Growth ◽  
Mutation Frequency ◽  
Cultured Cells ◽  
Tp53 Gene ◽  
Tumour Suppressor Genes ◽  
Restriction Sites ◽  
Mutation Spectra ◽  
Risk Of Cancer ◽  
Assay Conditions

Abstract Mutations in oncogenes or tumour suppressor genes cause increases in cell growth capacity. In some cases, fully malignant cancer cells develop after additional mutations occur in initially mutated cells. In such instances, the risk of cancer would increase in response to growth of these initially mutated cells. To ascertain whether such a situation might occur in cultured cells, three independent cultures of human lymphoblastoid GM00130 cells were treated with N-ethyl-N-nitrosourea to induce mutations, and the cells were maintained for 12 weeks. Mutant frequencies and spectra of the cells at the MspI and HaeIII restriction sites located at codons 247–250 of the TP53 gene were examined. Mutant frequencies at both sites in the gene exhibited a declining trend during cell culture and reached background levels after 12 weeks; this was also supported by mutation spectra findings. These results indicate that the mutations detected under our assay conditions are disadvantageous to cell growth.

Hydrogen peroxide production is affected by oxygen levels in mammalian cell culture

2017 ◽  
Vol 493 (1) ◽  
pp. 246-251 ◽  
Author(s):  
Lucas A. Maddalena ◽  
Shehab M. Selim ◽  
Joao Fonseca ◽  
Holt Messner ◽  
Shannon McGowan ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Culture ◽  
Mammalian Cell ◽  
Mammalian Cell Culture ◽  
Hydrogen Peroxide Production ◽  
Oxygen Levels

Formation of hydrogen peroxide in cell culture media by apple polyphenols and its effect on antioxidant biomarkers in the colon cell line HT-29

2009 ◽  
Vol 53 (10) ◽  
pp. 1226-1236 ◽  
Author(s):  
Phillip Bellion ◽  
Melanie Olk ◽  
Frank Will ◽  
Helmut Dietrich ◽  
Matthias Baum ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Culture ◽  
Cell Line ◽  
Culture Media ◽  
Cell Culture Media ◽  
Colon Cell ◽  
Colon Cell Line ◽  
Ht 29

scholarly journals Alkaloids Production and Cell Growth of Cinchona ledgeriana Moens: Effects of Fungal Filtrate and Methyl Jasmonate Elicitors

2021 ◽  
Vol 6 (1) ◽  
pp. 31-40
Author(s):  
Yustiny Andaliza Hasibuan ◽  
Diah Ratnadewi ◽  
Zainal Alim Mas’ud
Keyword(s):  
Cell Culture ◽  
Cell Growth ◽  
Secondary Metabolites ◽  
Methyl Jasmonate ◽  
Woody Plant ◽  
Cultured Cells ◽  
Cinchona Alkaloids ◽  
Cell Biomass ◽  
Cinchona Ledgeriana ◽  
Culture Technology

Cinchona alkaloids are known as antimalaria and anti-arrhythmic. Due to the long waiting time to harvest, cell culture technology is a challenge. This study aimed to determine the effects of elicitors, filtrate of two strains of endophytic fungi and methyl jasmonate (MeJA), in cell suspension culture of Cinchona ledgeriana on quinine and quinidine production. The cells were cultured for seven weeks in woody plant (WP) media treated with either of those elicitors in various concentrations. The cells growth was observed and the alkaloids were analyzed by HPLC. Cells treated with MeJA failed to grow that led to the cell biomass insufficiency for alkaloids determination.  It indicates that the cells are quite sensitive to even low concentration of MeJA that hampered the growth. Cells treated with the filtrate of Diaporthe sp. M13-Millipore filtered (S2M) gave the least cell biomass but presented the highest content of both alkaloids. Diaporthe sp. strain M-13 is stronger as elicitor than M-23 for this plant species. Filtrate of non-virulent fungi can elevate the biosynthesis of alkaloids. This reconfirms that cultured cells are capable to produce secondary metabolites and the productivity can be increased by using an appropriate elicitor.  

Using 3D perfusion bioreactor system to studying cell biology of ovarian cancer.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. e17558-e17558
Author(s):  
Alba Martínez ◽  
Molly Buckley ◽  
Joel Berry ◽  
Rebecca Christian Arend
Keyword(s):  
Ovarian Cancer ◽  
Cell Culture ◽  
Cell Growth ◽  
Cell Lines ◽  
Culture Media ◽  
Tumor Biology ◽  
Perfusion Bioreactor ◽  
Suitable Model ◽  
Cell Culture Media ◽  
Bioreactor System

e17558 Background: Epithelial Ovarian Cancer (EOC) is the most common cause of death among gynecological malignancies. This is a result of the high rate of recurrence and chemo-resistance in EOC patients. Therefore, the development of new therapeutics is crucial. A major factor contributing to this is the lack of therapeutic candidates is lack of translational accuracy in preclinical models. Recently, 3-dimensional (3-D) models have aided in accurately recreating tumor biology. We have developed an EOC 3-D perfused bioreactor system that recapitulates EOC tumor biology and incorporates tumor biomechanical regulation. This model allows for us to more accurately predict the clinical response of new drug candidates, which aids in elimination of ineffective candidates prior to clinical trials. Methods: EOC cell lines (luciferase-taggedSKOV-3 and OVCAR-8) were embedded in a relevant extracellular matrix (ECM) and injected into a perfused, polydimethylsiloxane (PDMS) bioreactor. Microchannels were embedded in matrigel so that the cell culture media with or without chemotherapy could flow through the perfused PDMS to provide nutrient delivery and gas exchange enhancing viability and function of surrounding cells. The bioreactors were connected to a peristaltic pump that allowed for the cell culture media to perfuse over a 7-day period. We monitored cell viability using bioluminescence imaging (BLI), immunohistochemistry (IHC), and lactate dehydrogenase (LDH) release in media. Results: BLI showed a linear increase in SKOV-3 and OVCAR-8 cell growth over 7 days. These results were confirmed by IHC measuring the number of nucleated cells per micron2. Graphical representation of the region of interest (ROI) showed a high correlation between IHC staining of nucleated cells and BLI score. IHC analysis of PAX8 staining was positive and proved that the perfusion bioreactor system maintains EOC biology over time. In addition, our results suggest that the bioreactor is a suitable model for drug preclinical testing in both cell lines as well as in patients’ samples. Conclusions: Our preliminary results using the 3D EOC perfused, PDMS bioreactor model showed increased EOC cell growth overtime, while maintaining original EOC histology. Moreover, our results suggest that this model could provide a novel platform to study therapeutic interventions in EOC. Our ultimate goal is to implement ovarian cancer microenvironment components (e.g. immune cells) into bioreactor system to study different drug treatments to better determine drug candidate’s translational efficacy.

scholarly journals An AlphaScreen Assay for the Discovery of Synthetic Chemical Inhibitors of Glucagon Production

2015 ◽  
Vol 21 (4) ◽  
pp. 325-332 ◽  
Author(s):  
Matthew R. Evans ◽  
Shuguang Wei ◽  
Bruce A. Posner ◽  
Roger H. Unger ◽  
Michael G. Roth
Keyword(s):  
Cell Culture ◽  
Small Molecules ◽  
Culture Media ◽  
Glucagon Secretion ◽  
Cell Culture Media ◽  
Glucagon Receptor ◽  
Synthetic Compounds ◽  
Glucose Levels ◽  
Blood Glucose Levels ◽  
Chemical Inhibitors

Glucose homeostasis is primarily controlled by two opposing hormones, insulin and glucagon, and diabetes results when insulin fails to inhibit glucagon action. Recent efforts to control glucagon in diabetes have focused on antagonizing the glucagon receptor, which is effective in lowering blood glucose levels but leads to hyperglucogonemia in rodents. An alternative strategy would be to control glucagon production with small molecules. In pursuit of this goal, we developed a homogeneous AlphaScreen assay for measuring glucagon in cell culture media and used this in a high-throughput screen to discover synthetic compounds that inhibited glucagon secretion from an alpha cell–like cell line. Some of these compounds inhibited transcription of the glucagon gene.

A Microfluidic Sensing System with a Multichannel Surface Plasmon Resonance Chip: Damage-free Characterization of Cells by Pattern Recognition

2020 ◽  
Author(s):  
Hiroka Sugai ◽  
Shunsuke Tomita ◽  
Sayaka Ishihara ◽  
Kyoko Yoshioka ◽  
Ryoji Kurita
Keyword(s):  
Cell Culture ◽  
Pattern Recognition ◽  
Surface Plasmon Resonance ◽  
Surface Plasmon ◽  
Plasmon Resonance ◽  
Cultured Cells ◽  
Culture Media ◽  
Biological Research ◽  
Cell Culture Media

<p>The development of a versatile sensing strategy for the damage-free characterization of cultured cells is of great importance for both fundamental biological research and industrial applications. Here, we present a pattern-recognition-based cell-sensing approach using a multichannel surface plasmon resonance (SPR) chip. The chip, in which five cysteine derivatives with different structures are immobilized on Au films, is capable of generating five unique SPR sensorgrams for the cell-secreted molecules that are contained in cell culture media. An automatic statistical program was built to acquire kinetic parameters from the SPR sensorgrams and to select optimal parameters as “pattern information” for subsequent multivariate analysis. Our system rapidly (~ 10 min) provides the complex information by merely depositing a small amount of cell culture media (~ 25 µL) onto the chip, and the amount of information obtained is comparable to that furnished by a combination of conventional laborious biochemical assays. This non-invasive pattern-recognition-based cell-sensing approach could potentially be employed as a versatile tool for characterizing cells. </p>

Sedimentation-Diffusion Behaviour of Nanoparticles

2020 ◽  
Author(s):  
francesco giorgi ◽  
Peter Macko ◽  
Judith M. Curran ◽  
Maurice Whelan ◽  
Andrew Worth ◽  
...  
Keyword(s):  
Cell Culture ◽  
Gold Nanoparticles ◽  
Theoretical Model ◽  
Cultured Cells ◽  
Culture Media ◽  
Biological Response ◽  
Optical Microscope ◽  
Local Concentration ◽  
Nominal Concentration

Abstract Background The biological response of organisms exposed to nanoparticles is often studied in vitro using adherent monolayers of cultured cells. In order to derive accurate concentration-response relationships, it is important to determine the local concentration of nanoparticles to which the cells are actually exposed rather than the nominal concentration of nanoparticles in the cell-culture medium. In this study, we investigated the sedimentation–diffusion process of different sized and charged gold nanoparticles in vitro by evaluating their settling dynamics and by developing a theoretical model to predict the concentration depth-profile of nanoparticles in solution over time.Results Experiments were carried out in water and in cell-culture media at a range of controlled temperatures. The optical phenomenon of caustics was exploited to track nanoparticles in real-time in a conventional optical microscope without any requirement for fluorescent labelling that potentially affects the dynamics of the nanoparticles. The results obtained demonstrate that size, temperature, and the stability of the nanoparticles play a pivotal role in regulating settling dynamics of nanoparticles. For gold nanoparticles larger than 60 nm in diameter, the initial nominal concentration did not accurately represent the concentration of nanoparticles local to the cells.Conclusion Nanoparticles dynamics in solution regulate the amount of material at the cellular level and must be taken into account when evaluating the biological response of organisms. the theoretical model proposed in this study accurately described the settling dynamics of the nanoparticles and thus represents a promising tool to support the design of in vitro experiments and the study of concentration-response relationships.

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