Rapid preparation of adherent mammalian cells for basic scanning electron microscopy (SEM) analysis

Osahor, Andrew; Deekonda, Karthik; Lee, Choon Weng; Sim, Edmund Ui Hang; Radu, Aurelian; Narayanan, K.

doi:10.1016/j.ab.2017.07.008

Cited by 14 publications

(7 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The samples were washed with sodium cacodylate buffer three times (each washing for 5 min); one drop of osmium tetraoxide was added to each sample and kept at 4°C for 1.5 h followed by dehydration through a series of alcohol concentrations (25%, 30%, 50%, 85%, 95% and 100%). The samples were then coated with gold and examined by SEM (Carl ZEISS, UK) (Osahor et al, 2017).…”

Section: Methodsmentioning

confidence: 99%

Enhanced steroidogenic and altered antioxidant response by ZnO nanoparticles in mouse testis Leydig cells

Bara

Kaul

2018

Toxicol Ind Health

View full text Add to dashboard Cite

Zinc oxide nanoparticles (ZnO NPs) are important nanomaterials with myriad applications and in widespread use. The main aim of this study was to evaluate the direct effect of ZnO NPs on steroidogenesis by considering mouse testicular Leydig cells (TM3) as an in vitro model system. The uptake, intracellular behaviour, cytotoxicity and morphological changes induced by ZnO NPs (0-200 µg/ml) in a time-dependent manner in the TM3 were assessed. A significant ( p < 0.05) decrease in TM3 viability was observed at 2 µg/ml ZnO NP after a 1-h incubation time period. Increased antioxidant enzyme activity, namely, superoxide dismutase (SOD) and catalase, was regularly observed. Not surprisingly, apoptosis also increased significantly after a 4-h exposure period. Transmission electron micrographs illustrated that ZnO NPs were taken up by Leydig cells and resulted in the formation of autophagosomes, autolysosomes and autophagic vacuoles. Concomitant real-time data indicated that ZnO NPs significantly increased the expression of steroidogenesis-related genes (steroidogenic acute regulatory protein and cytochrome P450 side-chain cleavage enzyme) and significantly ( p < 0.05) decreased antioxidant enzyme gene (SOD) expression after a 4-h incubation period. Moreover, ZnO NPs exposure significantly increased testosterone production at 2 µg/ml concentration after a 12-h incubation period. Our findings confirm the adverse effects of ZnO NPs by being cytotoxic, enhancing apoptosis, causing steroidogenic effect in Leydig cells and increasing autophagic vacuole formation possibly via alteration of antioxidant enzyme activity in TM3 cells.

show abstract

Section: Methodsmentioning

confidence: 99%

Enhanced steroidogenic and altered antioxidant response by ZnO nanoparticles in mouse testis Leydig cells

Bara

Kaul

2018

Toxicol Ind Health

View full text Add to dashboard Cite

show abstract

“…Passage three K5 iMSC and primary MSC were seeded on plastic coverslips, cultured for 2 days, and fixed with 2.5% glutaraldehyde. The samples were then subjected to ethanol gradient dehydration and dried in a desiccator, as described previously (Osahor et al, ). Subsequently, the samples were coated with a thin film of gold.…”

Section: Methodsmentioning

confidence: 99%

“…The samples were then subjected to ethanol gradient dehydration and dried in a desiccator, as described previously (Osahor et al, 2017). Subsequently, the samples were coated with a thin film of gold.…”

Section: Scanning Electron Microscopymentioning

confidence: 99%

Generation and characterization of a functional human adipose‐derived multipotent mesenchymal stromal cell line

Burk

Holland²,

Lauermann

et al. 2019

Biotech & Bioengineering

View full text Add to dashboard Cite

Multipotent mesenchymal stromal cells (MSC) and MSC-derived products have emerged as promising therapeutic tools. To fully exploit their potential, further mechanistic studies are still necessary and bioprocessing needs to be optimized, which requires an abundant supply of functional MSC for basic research. To address this need, here we used a novel technology to establish a human adipose-derived MSC line with functional characteristics representative of primary MSC. Primary MSC were isolated and subjected to lentiviral transduction with a library of expansion genes. Clonal cell lines were generated and evaluated on the basis of their morphology, immunophenotype, and proliferation potential. One clone (K5 iMSC) was then selected for further characterization. This clone had integrated a specific transgene combination including genes involved in stemness and maintenance of adult stem cells. Favorably, the K5 iMSC showed cell characteristics resembling juvenile MSC, as they displayed a shorter cell length and enhanced migration and proliferation compared with the nonimmortalized original primary MSC (p < 0.05). Still, their immunophenotype and differentiation potential corresponded to the original primary MSC and the MSC definition criteria, and cytogenetic analyses revealed no clonal aberrations. We conclude that the technology used is applicable to generate functional MSC lines for basic research and possible future bioprocessing applications. K E Y W O R D S bioprocessing, cell line, immortalization, mesenchymal stromal cells, multipotent mesenchymal stromal cell (MSC)

show abstract

“…Morphological evaluation of tissues was performed by means of SEM (JEOL JCM-6000, Tokyo, Japan). The samples were prepared according to Osahor et al [54] with some modifications. Plastic coverslips were sterilized with 70% ethanol and next by UV for 30 min.…”

Section: Semmentioning

confidence: 99%

Acrylamide Decreases Cell Viability, and Provides Oxidative Stress, DNA Damage, and Apoptosis in Human Colon Adenocarcinoma Cell Line Caco-2

et al. 2020

View full text Add to dashboard Cite

Acrylamide (AA) toxicity remains an interesting subject in toxicological research. The aim of the research performed in this paper was to determine mechanisms of cyto- and genotoxic effects of AA on the human colon adenocarcinoma cell line Caco-2, to estimate the inhibitory concentration (IC)50 values in cell viability assays, to measure the basal and oxidative DNA damage as well as the oxidative stress leading to apoptosis, and to assess the morphological changes in cells using microscopic methods. It has been proven that AA induces cytotoxic and genotoxic effects on Caco-2 cells. Higher cytotoxic activity was gained in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay compared with the PrestoBlue assay, with IC50 values of 5.9 and 8.9 mM after 24 h exposure, respectively. In the single-cell gel electrophoresis assay, the greatest DNA damage was caused by the highest concentration of acrylamide equal to 12.5 mM (89.1% ± 0.9%). AA also induced oxidative DNA damage and generated reactive oxygen species (ROS), which was concentration dependent and correlated with the depletion of mitochondrial membrane potential and apoptosis induction. In the microscopic staining of cells, AA in the dosage close to the IC50 induced morphological changes typical for apoptosis. Taken together, these results demonstrate that AA has a pro-oxidative effect on Caco-2 cells, leading to apoptotic cell death.

show abstract

Rapid preparation of adherent mammalian cells for basic scanning electron microscopy (SEM) analysis

Cited by 14 publications

References 19 publications

Enhanced steroidogenic and altered antioxidant response by ZnO nanoparticles in mouse testis Leydig cells

Enhanced steroidogenic and altered antioxidant response by ZnO nanoparticles in mouse testis Leydig cells

Generation and characterization of a functional human adipose‐derived multipotent mesenchymal stromal cell line

Acrylamide Decreases Cell Viability, and Provides Oxidative Stress, DNA Damage, and Apoptosis in Human Colon Adenocarcinoma Cell Line Caco-2

Contact Info

Product

Resources

About