scholarly journals In vitro determinants of e-cig aerosol condensate bioavailability and toxicity: Influence of cell culture parameters and utility of a normalized dose metric in e-cig studies

2021 ◽  
Author(s):  
Udochukwu C Obodo ◽  
Timothy R O'Connor
Keyword(s):  
Electronic Cigarettes ◽  
Seeding Density ◽  
Physical Factors ◽  
Culture Vessel ◽  
Cell Seeding ◽  
Health Issues ◽  
Cellular Survival ◽  
Cell Seeding Density ◽  
Species Production

Electronic cigarettes (e-cigs) have a strong foothold in the marketplace as a product to replace tobacco cigarette usage. Despite many researchers investigating the use of e-cigs and possible health issues, there is still controversy concerning how to evaluate and use e-cig condensates. Therefore, to identify factors that influence in vitro e-cig studies, we examined parameters that can impact experimental outcomes. We generated high wattage e-cig aerosol condensate (ECAC) to determine reproducible conditions to evaluate ECAC with respect to cellular survival. Cytotoxicity of ECAC was independent of serum conditions. However, cytotoxicity of ECAC is altered by treatment duration and by physical factors, including cell seeding density and volume of ECAC used. In addition, interactions between ECAC components and cells, as well as the culture vessel surface, diminish the bioavailability of ECAC components in vitro and altered the results obtained. Moreover, the cell seeding density changes reactive oxygen species production in response to ECAC exposure. Our data indicated that normalized ECAC doses (ECAC weight per cell) better reflect the toxicity of ECAC than nominal doses (ECAC percentage). These results provide factors for researchers to consider in the design of in vitro experiments using ECAC.

scholarly journals Stiff-to-Soft Transition from Glass to 3D Hydrogel Substrates in Neuronal Cell Culture

Micromachines ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 165
Author(s):  
Gulden Akcay ◽  
Regina Luttge
Keyword(s):  
Single Cells ◽  
Neuroblastoma Cells ◽  
Neuronal Cell ◽  
Seeding Density ◽  
Cell Aggregates ◽  
Cell Seeding ◽  
Glycol Diacrylate ◽  
Cell Seeding Density ◽  
Wide Range

Over the past decade, hydrogels have shown great potential for mimicking three- dimensional (3D) brain architectures in vitro due to their biocompatibility, biodegradability, and wide range of tunable mechanical properties. To better comprehend in vitro human brain models and the mechanotransduction processes, we generated a 3D hydrogel model by casting photo-polymerized gelatin methacryloyl (GelMA) in comparison to poly (ethylene glycol) diacrylate (PEGDA) atop of SH-SY5Y neuroblastoma cells seeded with 150,000 cells/cm2 according to our previous experience in a microliter-sized polydimethylsiloxane (PDMS) ring serving for confinement. 3D SH-SY5Y neuroblastoma cells in GelMA demonstrated an elongated, branched, and spreading morphology resembling neurons, while the cell survival in cast PEGDA was not supported. Confocal z-stack microscopy confirmed our hypothesis that stiff-to-soft material transitions promoted neuronal migration into the third dimension. Unfortunately, large cell aggregates were also observed. A subsequent cell seeding density study revealed a seeding cell density above 10,000 cells/cm2 started the formation of cell aggregates, and below 1500 cells/cm2 cells still appeared as single cells on day 6. These results allowed us to conclude that the optimum cell seeding density might be between 1500 and 5000 cells/cm2. This type of hydrogel construct is suitable to design a more advanced layered mechanotransduction model toward 3D microfluidic brain-on-a-chip applications.

Size Regulation of Chondrocyte Spheroids Using a PDMS-Based Cell Culture Chip

2013 ◽  
Vol 25 (4) ◽  
pp. 644-649 ◽  
Author(s):  
Takahisa Anada ◽  
◽  
Osamu Suzuki
Keyword(s):  
Tissue Engineering ◽  
Cell Differentiation ◽  
Chondrogenic Differentiation ◽  
Seeding Density ◽  
Cell Seeding ◽  
Initial Cell ◽  
Cell Seeding Density ◽  
Tissue Engineering Approach ◽  
Cellular Aggregates

Cartilage self-repair is limited due to a lack of blood supply and the low mitosis rate of chondrocytes. A tissue engineering approach using cells and biomaterials has the potential to treat cartilage injury. Threedimensional cellular aggregates are an excellent model for mimicking condensation and chondrogenic differentiation in vitro. We developed a technique for constructing spheroids utilizing a polydimethylsiloxane (PDMS)-based culture chip. The objective of this study is to determine how the initial cell density on a culture chip affects the chondrogenic ATDC5 cell differentiation. We demonstrate how culture chips having arrays of multicavities are able to generate high numbers of uniform spheroids rapidly and simultaneously with narrow size distribution. Spheroids are collected easily and noninvasively. Higher cell seeding density on the culture chip enhances chondrogenic cell differentiation. These results suggest the usefulness of this chip in engineering 3D cellular constructs with high functionality for tissue engineering.

scholarly journals Effect of Cell Seeding Density and Inflammatory Cytokines on Adipose Tissue-Derived Stem Cells: an in Vitro Study

2017 ◽  
Vol 13 (2) ◽  
pp. 267-277 ◽  
Author(s):  
Panithi Sukho ◽  
Jolle Kirpensteijn ◽  
Jan Willem Hesselink ◽  
Gerjo J. V. M. van Osch ◽  
Femke Verseijden ◽  
...  
Keyword(s):  
Stem Cells ◽  
Adipose Tissue ◽  
Inflammatory Cytokines ◽  
In Vitro Study ◽  
Vitro Study ◽  
Seeding Density ◽  
Cell Seeding ◽  
Cell Seeding Density

scholarly journals Identification and In Vitro Expansion of Buccal Epithelial Cells

2018 ◽  
Vol 27 (6) ◽  
pp. 957-966
Author(s):  
Soraya Rasi Ghaemi ◽  
Bahman Delalat ◽  
Frances J. Harding ◽  
Yazad D. Irani ◽  
Keryn A. Williams ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Transplantation ◽  
Progenitor Cell ◽  
Ex Vivo ◽  
Seeding Density ◽  
Cell Seeding ◽  
Cell Marker ◽  
Cell Seeding Density ◽  
Proliferation And Differentiation

Ex vivo-expanded buccal mucosal epithelial (BME) cell transplantation has been used to reconstruct the ocular surface. Methods for enrichment and maintenance of BME progenitor cells in ex vivo cultures may improve the outcome of BME cell transplantation. However, the parameter of cell seeding density in this context has largely been neglected. This study investigates how varying cell seeding density influences BME cell proliferation and differentiation on tissue culture polystyrene (TCPS). The highest cell proliferation activity was seen when cells were seeded at 5×104 cells/cm2. Both below and above this density, the cell proliferation rate decreased sharply. Differential immunofluorescence analysis of surface markers associated with the BME progenitor cell population (p63, CK19, and ABCG2), the differentiated cell marker CK10 and connexin 50 (Cx50) revealed that the initial cell seeding density also significantly affected the progenitor cell marker expression profile. Hence, this study demonstrates that seeding density has a profound effect on the proliferation and differentiation of BME stem cells in vitro, and this is relevant to downstream cell therapy applications.

scholarly journals Injectable, redox-polymerized carboxymethylcellulose hydrogels promote nucleus pulposus-like extracellular matrix elaboration by human MSCs in a cell density-dependent manner

2018 ◽  
Vol 33 (4) ◽  
pp. 576-589 ◽  
Author(s):  
Devika M Varma ◽  
Michelle S DiNicolas ◽  
Steven B Nicoll
Keyword(s):  
Extracellular Matrix ◽  
Minimally Invasive ◽  
Nucleus Pulposus ◽  
Seeding Density ◽  
Human Mscs ◽  
Dependent Manner ◽  
Cell Seeding ◽  
Swelling Properties ◽  
Matrix Deposition ◽  
Cell Seeding Density

Low back pain is a major cause for disability and is closely linked to intervertebral disc degeneration. Mechanical and biological dysfunction of the nucleus pulposus in the disc has been found to initiate intradiscal degenerative processes. Replacing or enriching the diseased nucleus pulposus with an injectable, stem cell-laden biomaterial that mimics its material properties can provide a minimally invasive strategy for biological and structural repair of the tissue. In this study, injectable, in situ-gelling carboxymethylcellulose hydrogels were developed for nucleus pulposus tissue engineering using encapsulated human marrow-derived mesenchymal stromal cells (hMSCs). With the goal of obtaining robust extracellular matrix deposition and faster construct maturation, two cell-seeding densities, 20 × 106 cells/ml and 40 × 106 cells/ml, were examined. The constructs were fabricated using a redox initiation system to yield covalently crosslinked, cell-seeded hydrogels via radical polymerization. Chondrogenic culture of the hydrogels over 35 days exhibited high cell viability along with deposition of proteoglycan and collagen-rich extracellular matrix, and mechanical and swelling properties similar to native human nucleus pulposus. Further, the matrix production and distribution in the carboxymethylcellulose hydrogels was found to be strongly influenced by hMSC-seeding density, with the lower cell-seeding density yielding a more favorable nucleus pulposus-specific matrix phenotype, while the rate of construct maturation was less dependent on the cell-seeding density. These findings are the first to demonstrate the utility of redox-polymerized carboxymethylcellulose hydrogels as hMSC carriers for potential minimally invasive treatment strategies for nucleus pulposus replacement.

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