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Biomolecules
Special Issues
Physiological Cell Culture
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Physiological Cell Culture

A special issue of Biomolecules (ISSN 2218-273X). This special issue belongs to the section "Cellular Biochemistry".

Deadline for manuscript submissions: closed (15 April 2023) | Viewed by 8916

Special Issue Editors

Dr. Jeffrey Stuart

E-Mail Website
Guest Editor
Department of Biological Sciences, Brock University, St. Catharines, ON L2S 3A1, Canada
Interests: mitochondrial bioenergetics; mitochondrial dynamics; aging; reactive oxygen/nitrogen species metabolism; oncometabolism; resveratrol
Special Issues, Collections and Topics in MDPI journals
Dr. Fereshteh Moradi

E-Mail Website
Guest Editor
Department of Biological Sciences, Brock University, St. Catharines, ON L2S 3A12, Canada
Interests: cancer biology; cell signaling; immunofluorescence; cell culture; PCR; western blot analysis; flowcytometry

Special Issue Information

Dear Colleagues,

Mammalian cell culture has been fundamental to gaining our current understanding of cell biology. First developed in the mid-20th century, cell culture has become a routine method for modeling and studying cellular functions and dysfunctions in health and disease. It is instrumental in the discovery and development of drugs. Yet, current cell culture practices are based on methods that promote cell proliferation, rather than those that recreate conditions similar to those experienced by cells in vivo. The concentrations of glucose, amino acids, and oxygen during cell culture are typically in excess to ensure adequate supplies of these nutrients, rather than keeping them within physiologically relevant ranges. Research over the past two decades has provided abundant evidence that non-physiological cell culture conditions can lead to experimental artifacts that limit the effectiveness with which cell culture models in vivo biology. It is increasingly clear that greater emphasis should be placed on adopting cell culture practices that more closely recreate the in vivo tissue environment.

This Special Issue welcomes original research and review articles focusing on the significance of physiological cell culture. This includes physioxia, growth medium formulation, 2- versus 3-dimensional cell culture models, serum versus serum-free media, and mono- versus co-culture. We are interested in contributions describing how the cell culture environment affects the behaviour of cellular disease models, the efficacy of in vitro fertilization approaches, cellular responses to drugs or other therapies, and other important applications of cell culture in biology and medicine.

Dr. Jeffrey Stuart
Dr. Fereshteh Moradi
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Biomolecules is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2700 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Published Papers (3 papers)

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Research

19 pages, 2650 KiB  
Article
Culture of Cancer Cells at Physiological Oxygen Levels Affects Gene Expression in a Cell-Type Specific Manner
by Ricardo Alva, Fereshteh Moradi, Ping Liang and Jeffrey A. Stuart
Biomolecules 2022, 12(11), 1684; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12111684 - 14 Nov 2022
Cited by 4 | Viewed by 2218
Abstract
Standard cell culture is routinely performed at supraphysiological oxygen levels (~18% O2). Conversely, O2 levels in most mammalian tissues range from 1–6% (physioxia). Such hyperoxic conditions in cell culture can alter reactive oxygen species (ROS) production, metabolism, mitochondrial networks, and [...] Read more.
Standard cell culture is routinely performed at supraphysiological oxygen levels (~18% O2). Conversely, O2 levels in most mammalian tissues range from 1–6% (physioxia). Such hyperoxic conditions in cell culture can alter reactive oxygen species (ROS) production, metabolism, mitochondrial networks, and response to drugs and hormones. The aim of this study was to investigate the transcriptional response to different O2 levels and determine whether it is similar across cell lines, or cell line-specific. Using RNA-seq, we performed differential gene expression and functional enrichment analyses in four human cancer cell lines, LNCaP, Huh-7, PC-3, and SH-SY5Y cultured at either 5% or 18% O2 for 14 days. We found that O2 levels affected transcript abundance of thousands of genes, with the affected genes having little overlap between cell lines. Functional enrichment analysis also revealed different processes and pathways being affected by O2 in each cell line. Interestingly, most of the top differentially expressed genes are involved in cancer biology, which highlights the importance of O2 levels in cancer cell research. Further, we observed several hypoxia-inducible factor (HIF) targets, HIF-2α targets particularly, upregulated at 5% O2, consistent with a role for HIFs in physioxia. O2 levels also differentially induced the transcription of mitochondria-encoded genes in most cell lines. Finally, by comparing our transcriptomic data from LNCaP and PC-3 with datasets from the Prostate Cancer Transcriptome Atlas, a correlation between genes upregulated at 5% O2 in LNCaP cells and the in vivo prostate cancer transcriptome was found. We conclude that the transcriptional response to O2 over the range from 5–18% is robust and highly cell-type specific. This latter finding indicates that the effects of O2 levels are difficult to predict and thus highlights the importance of regulating O2 in cell culture. Full article
(This article belongs to the Special Issue Physiological Cell Culture)
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23 pages, 2695 KiB  
Article
Cell Confluence Modulates TRPV4 Channel Activity in Response to Hypoxia
by Solène Barbeau, Alexandre Joushomme, Yann Chappe, Guillaume Cardouat, Isabelle Baudrimont, Véronique Freund-Michel, Christelle Guibert, Roger Marthan, Patrick Berger, Pierre Vacher, Yann Percherancier, Jean-François Quignard and Thomas Ducret
Biomolecules 2022, 12(7), 954; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12070954 - 07 Jul 2022
Cited by 3 | Viewed by 2228
Abstract
Transient receptor potential vanilloid 4 (TRPV4) is a polymodal Ca2+-permeable channel involved in various hypoxia-sensitive pathophysiological phenomena. Different tools are available to study channel activity, requiring cells to be cultured at specific optimal densities. In the present study, we examined if [...] Read more.
Transient receptor potential vanilloid 4 (TRPV4) is a polymodal Ca2+-permeable channel involved in various hypoxia-sensitive pathophysiological phenomena. Different tools are available to study channel activity, requiring cells to be cultured at specific optimal densities. In the present study, we examined if cell density may influence the effect of hypoxia on TRPV4 activity. Transiently TRPV4-transfected HEK293T cells were seeded at low or high densities corresponding to non-confluent or confluent cells, respectively, on the day of experiments, and cultured under in vitro normoxia or hypoxia. TRPV4-mediated cytosolic Ca2+ responses, single-channel currents, and Ca2+ influx through the channel were measured using Ca2+ imaging/microspectrofluorimetric assay, patch-clamp, and Bioluminescence Resonance Energy Transfer (BRET), respectively. TRPV4 plasma membrane translocation was studied using confocal microscopy, biotinylation of cell surface proteins, and BRET. Our results show that hypoxia exposure has a differential effect on TRPV4 activation depending on cell confluence. At low confluence levels, TRPV4 response is increased in hypoxia, whereas at high confluence levels, TRPV4 response is strongly inhibited, due to channel internalization. Thus, cell density appears to be a crucial parameter for TRPV4 channel activity. Full article
(This article belongs to the Special Issue Physiological Cell Culture)
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16 pages, 7883 KiB  
Article
Effects of Hypoxia on Proliferation and Differentiation in Belgian Blue and Hanwoo Muscle Satellite Cells for the Development of Cultured Meat
by Sanghun Park, Mick Gagliardi, Geertje Swennen, Arin Dogan, Yuna Kim, Yunhwan Park, Gyutae Park, Sehyuk Oh, Mark Post and Jungseok Choi
Biomolecules 2022, 12(6), 838; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12060838 - 16 Jun 2022
Cited by 8 | Viewed by 3492
Abstract
Among future food problems, the demand for meat is expected to increase rapidly, but the production efficiency of meat, which is a protein source, is very low compared to other foods. To address this problem, research on the development and production of cultured [...] Read more.
Among future food problems, the demand for meat is expected to increase rapidly, but the production efficiency of meat, which is a protein source, is very low compared to other foods. To address this problem, research on the development and production of cultured meat as an alternative meat source using muscle stem cells in vitro has recently been undertaken. Many studies have been conducted on myosatellite cells for medical purposes, but studies on alternative meat production are rare. In vitro cell culture mimics the in vivo environment for cell growth. The satellite cell niche is closer to hypoxic (2% O2) than normoxic (20% O2) conditions. The aim of this study was to investigate the efficient oxygen conditions of myosatellite cell cultures for the production of cultured meat. The bovine satellite cell counts and mRNA (Pax7, Myf5 and HIF1α) levels were higher in hypoxia than normoxia (p < 0.05). Through Hoechst-positive nuclei counts, and expression of Pax7, MyoD and myosin protein by immunofluorescence, it was confirmed that muscle cells performed normal proliferation and differentiation. Myoblast fusion was higher under hypoxic conditions (p < 0.05), and the myotube diameters were also thicker (p < 0.05). In the myotube, the number of cells was high in hypoxia, and the expression of the total protein amounts, differentiation marker mRNA (myogenin, myosin and TOM20), and protein markers (myosin and TOM20) was also high. The study results demonstrated that the proliferation and differentiation of bovine myosatellite cells were promoted more highly under hypoxic conditions than under normoxic conditions. Therefore, hypoxic cultures that promote the proliferation and differentiation of bovine myosatellite cells may be an important factor in the development of cultured meat. Full article
(This article belongs to the Special Issue Physiological Cell Culture)
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