Physiological Oxygen is Healthier for Cell Cultures

Physiological Oxygen is Healthier for Cell Cultures

Drs. Timpano and Uniacke, Hypoxystation users at University of Guelph in Ontario, have published a very thorough study examining the molecular basis of cells’ reactions to differing levels of hypoxia. In their paper “Human Cells Cultured Under Physiological Oxygen Utilize Two Cap-binding Proteins to Recruit Distinct mRNAs for Translation” (Journal of Biological Chemistry 291:20; 2016), they examine 2 different translation initiation proteins, eiF4E and eiF4E2, that are activated under either high (>8% O2) or low (<1% O2) oxygen levels, with the aid of mTORC1 or HIF-2α, respectively, and activated simultaneously in an area of low- to mid-level physioxia (1-8% O2). Timpano and Uniacke were able to stably and accurately create low oxygen in their Hypoxystation by Hypoxygen, which provides a closed workstation environment that enables researchers to culture and manipulate cells inside the chamber through gloveless sleeves, eliminating the negative consequences of spikes of higher oxygen and lower temperatures encountered in an incubator as cell cultures are growing. Their research into translational modulation of the proteome using the Hypoxystation gives seminal insights into physioxia as the natural condition for cells, both in vitro and in vivo.

“Culturing cells in ambient air could be far from physiological with respect to oxygen. Oxygen is a surprisingly neglected factor (in cell culture)” – Dr Timpano and Dr Uniacke, University of Guelph, Ontario, Canada

Through polysome association experiments with cells growing at ambient air versus lower oxygen levels of 1%, 3%, 5%, and 8%, RNA analysis, and m7-GTP cap-binding assays, Timpano and Uniacke were able to demonstrate that the oxygen concentration in the workstation was sufficient to either repress or increase the activity of eiF4E and eiF4E2, reflecting mechanisms that occur during development but also during tumor progression and in ischemic diseases. Cells can reversibly cycle between utilisation of the eiF4E protein, which preferentially binds to the 5′ TOP mRNA’s at >8% O2 and is impaired at hypoxia, and eiF4E2, which is active at <1% O2 and utilizes binding motifs in the 3′ UTR of the mRNA. The eIF4E type of mRNA’s code for housekeeping proteins while the eIF4E2-dependant mRNA’s encode signaling proteins needed to respond to environmental signals, allowing cells to control translation dynamically and giving cancer cells an edge during tumor progression, as hypoxia increases.

 

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Hypoxische Mikroumgebungen – Cell Press Nukleus

DWS Hypoxygen

Don Whitley Scientific und HypOxygen, der amerikanische Distributor von Don Whitley, haben sich mit Cell Press zusammengeschlossen, um ein forschungsbasiertes Nukleus mit Fokus auf Zellen in Umgebung mit geringem Sauerstoff zu fördern. Das Cell Press Webinar für „Hypoxische Mikroumgebungen“ war eine großartige Möglichkeit, um die Vorträge von Randall Johnson vom Karolinska Institut über die hypoxischen Reaktionen im Zusammenhang mit Metastasen sowie von Hannele Ruohola-Baker von der Universität Washington über die Rolle von hypoxischen Faktoren in der Reprogrammierung von somatischen Zellen zu hören.

 

Wir glauben, unser Engagement, das zuverlässigste, präziseste und robusteste Instrument für physiologische Zellkulturen anzubieten, hilft unseren Anwendern ihre Forschungen durchzuführen, und wir sind froh ein kleiner Teil davon zu sein. Whitley Hypoxystationen schaffen eine in vivo Umgebung für die Zellkultivierung und Manipulation über die Dauer der Inkubation einschließlich funktionale Studien über geringen Sauerstoff.

 

Wenn Sie das Webinar noch einmal aufrufen möchten, können Sie über diesen Link darauf zugreifen. Das Cell Press Nukleus, das ausgewählte Dokumente zu den Themen Krebs, Entzündungen, Stammzellen und Nervenentwicklung einschließt, bietet wertvolle Erkenntnisse der Hypoxie und stellt zudem Verbindungen zu Funktion und Krankheit her.

CellPress

Optimizing oxygen conditions enhances stem cell recovery

Two very recent papers, published in Cell and Regenerative Medicine, have described research into the advantages of reducing the oxygen exposure stem cells experience during isolation and cultivation, and the cellular mechanisms involved in translating hypoxia into increased stem cell survival and enhanced transplantation efficacy. The data presented by Mantel et al. on hematopoietic stem cells, and by Kay et al. on mesenchymal stem cells, caused quite a stir at the recent ISSCR meeting in Stockholm, as attendees discussed the implications for their own work.

Alasdair Kay and co-workers at Keele University evaluated the impact of various oxygen, glucose and serum levels on culturing human bone marrow-derived MSC’s. They found that the application of more physiological conditions, e.g. 2% oxygen instead of 21% O2 ambient, resulted in significantly increased cell yields; the highest yields were achieved in the stringently controlled hypoxic environment of a workstation. The authors attribute this to deoxygenation/reoxygenation cycles and very brief air exposure for cells cultured in an incubator, as compared to the controlled atmosphere in the workstation. Transcriptome analysis of hMSC’s cultured at 2% (incubator vs workstation) and 21% oxygen revealed scores of genes that were up or downregulated with very little overlap among the subsets. Kay et al. have identified downregulation of bone morphogenetic protein BMP2 expression, modulation of chemokine (C-X-C motif) receptor CXCR2 signaling, and LEPR upregulation as causative factors for enhanced hMSC isolation at hypoxia.

The term extraphysiologic oxygen shock/stress EPHOSS for hematopoietic stem cells experiencing “hyperoxia” outside of their natural niches is introduced by Mantel et al. from Indiana University. They delineated the detrimental effects of even brief exposure of bone marrow and cord blood to ambient conditions as compared to harvesting and processing HSC’s at constant 3% hypoxia. Up to 5-fold increased cell numbers and improved engraftment of transplanted HSC’s as a result of hypoxic conditions led the authors to question the relevance of the work currently being carried out at ambient oxygen. They attribute the beneficial effects of low oxygen on the yield and function of BM and CB stem cells to reduced production of reactive oxygen species ROS, implicating the mitochondrial permeability transition pore MPTP, cyclophilin CypD, and p53 in protection from EPHOSS. Mantel et al. predict that hypoxic processing will be significant for many other cell types and phenomena such as metabolism of cancer cells and stem cell aging.

This article was written by Burga Kalz Fuller, Product Manager at HypOxygen.

 

Alun Kitsell, Burga Kalz Fuller and Daniel Secker
Alun Kitsell, Burga Kalz Fuller and Daniel Secker with a Whitley HI35 Hypoxystation

 

  1. Kay et al. “BMP2 repression and optimized culture conditions promote human bone marrow-derived mesenchymal stem cell isolation” 2015 Med. 10(2), 109-125
  2. Mantel et al. “Enhancing hematopoietic stem cell transplantation efficacy by mitigating oxygen shock” 2015 Cell 161, 1553-1565

 

Hypoxia and stem cells

Later this month, the international stem cell research community will be travelling to Stockholm for the upcoming International Society for Stem Cell Research (ISSCR) meeting covering diverse topics from reprogramming and pluripotency of stem cells through tissue engineering and organ regeneration to therapy with stem cells.

Whitley H135 Hypoxystation
Whitley H135 Hypoxystation

Hypoxia is a crucial parameter determining the fate and development of stem cells, which leads Don Whitley Scientific to exhibit the Hypoxystation controlled environment workstation for low oxygen cell culture (see us on stand no. B15:33). Dr. Burga Kalz Fuller, Product Manager at our American distributor, HypOxygen, has summarized five recent papers delineating the role of hypoxia in stem cell research:

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Cell Press Webinar: Hypoxic Microenvironments

The hypoxic microenvironment triggers many of the events underlying cancer progression and stem cell maintenance and/or differentiation. In maintaining oxygen homeostasis, oxygen levels are sensed by mitochondria, communicated by reactive oxygen species (ROS), and conveyed by hypoxia-inducible factors HIF’s. Hypoxia signalling pathways have far-reaching consequences for cellular and tissue processes from metabolism through angiogenesis to metastasis.

Dr. Hannele Ruohola-Baker (University of Washington) and Dr. Randall Johnson (University of Cambridge) will be speaking at the upcoming Cell Press webinar on “Hypoxic Microenvironments”, which Don Whitley Scientific and HypOxygen are proud to sponsor. Dr. Ruohola-Baker’s lab investigates metabolic determinants of and miRNA function in stem cells, and Dr. Johnson examines the effects of hypoxia in cancer and inflammation.
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One City, Two Cultures

HypOxygen and Don Whitley Scientific are sponsoring a series of Keystone Symposia dealing with tumour micro environment (Vancouver), mitochondria and metabolism (Santa Fe), and immunity and inflammation (Olympic Valley). At the joint Keystone symposium on “Mitochondria, Metabolism and Heart Failure” and “Diabetes and Metabolic Dysfunction” in Santa Fe, research groups have come together to present their latest findings on metabolism.

Mitochondrial function is central to metabolism and energy sensing of the cells in all organs, and this joint meeting is bringing together researchers who gave talks on the regulation of metabolism under stress, mitochondrial quality control and dysregulated gluconeogenesis. Insight into the genes and the pathways controlling these processes at the cellular level is driving the search for more effective drugs.

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Whitley H35: Used in the Fight against Pancreatic Cancer

Whitley H35 Hypoxystation

Installation of H35 at MD Anderson

 

Widely regarded as the best cancer hospital in the USA, The MD Anderson Cancer Centre at the University of Texas currently uses a Whitley H35 Hypoxystation to contribute to their work focussed on pancreatic cancer.

 

 

Dr. Rafal Zielinski had this to say about their workstation:

“The hypoxia station we have purchased is being used for studies on STAT and HIF oncogenic transcription factors in tumours with major focus on pancreatic cancer. It is well-established that these tumours are highly hypoxic therefore experiments performed in oxygen-limited conditions better recapitulates the natural environment of the cancer, allows better understanding of the physiologic role of STATs and HIFs and enables more accurate evaluation of experimental therapeutic strategies. We also use chronic hypoxia to study metabolism of the cancer cells and to design optimal therapeutic approaches using new inhibitors of glycolysis.”

Fast Growing Hypoxia Technology Company Strengthens Resources

Burga Kalz Fuller

Don Whitley Scientific announce a key new appointment at their US distributor, HypOxygen (Frederick, MD, USA). Burga Kalz Fuller, PhD has joined the firm to further enhance their penetration of the Hypoxystation hypoxic workstation and related products into the growing field of mammalian cell based hypoxia research.

Burga comes with a wealth of experience within the life sciences industry, having spent the last ten years working with technology suppliers both in Europe and the United States where she was instrumental in the technical supervision of product and equipment development and marketing strategies.

Burga joins HypOxygen as Product Manager to continue HypOxygen’s recent success in providing state of the art hypoxic workstations that lead the way in advancing scientific research and innovation.

Burga comments, “I am excited to start a new challenge with HypOxygen and look forward to working with the team. The range of Whitley HEPA Hypoxystations will play a critical role in furthering advances in the fields of cancer and regenerative medicine research due to the wealth of special features for ultra-clean, precisely controlled physiological atmosphere that make it truly special. I am fortunate to be joining such a respected company that prides itself on providing industry leading technologies to the growing fields of tissue culture research and hypoxia.”

Jason Cadle, National Sales Manager at HypOxygen, said “Burga’s wealth of molecular cell biology experience and industry knowledge has already made her a key addition to the HypOxygen family. We view her appointment as a sign of our commitment to being the leading company in our industry. Our new innovations and the increasing demand from researchers led us to look for an addition to our team who will fit in with our ethos of innovation and exceptional service. It is very fortunate that we were able to find someone of Burga’s caliber to act in this capacity. I’m confident that Burga will play a key role in providing and implementing high quality technical solutions for our clients.”

Paul Walton, Managing Director of Don Whitley Scientific, added “Burga’s appointment is great news for the Whitley Hypoxystation range of products as we improve and expand the range. Having Burga on the team at HypOxygen will improve our international presence in terms of the service we can provide to all our distributors worldwide.”

Prize Winners Happy to Ski

Alison Janocha, Lerner Research Institute Ski Lesson Winner

Congratulations to Alison Janocha and colleagues from the Lerner Research Institute in Cleveland, Ohio. The ladies won the Hypoxygen prize at a recent event … a skiing lesson. Here are some photographs showing how very much they enjoyed their prize.

HypOxygen is our US distributor for the Hypoxystation range of workstations. HypOxygen have just returned from attending a successful meeting in Breckenridge, Colorado – The Keystone Symposia on Molecular and Cellular Biology meeting entitled Sensing and Signaling of Hypoxia: Interfaces with Biology and Medicine.Prize Winner, Lerner Research Institute Ski Lesson Winner 3

The event was concerned with hypoxia being an important component of most human diseases. The meeting focussed on advances in the understanding of the biochemistry and biology of hypoxia signaling and the impact on cell biology, physiology and medicine. Specific emphasis was placed on the emerging interfaces of hypoxia biology with new areas of scientific progress including metabolic regulation, epigenetic signaling, stem cell biology, inflammation, immunity, tissue repair and cancer.

Prize Winner, Lerner Research Institute Ski Lesson Winner 2