Low physiologic oxygen tensions reduce proliferation and differentiation of human multipotent mesenchymal stromal cells

BMC Cell Biology, Jan 2010

Background Human multipotent mesenchymal stromal cells (MSC) can be isolated from various tissues including bone marrow. Here, MSC participate as bone lining cells in the formation of the hematopoietic stem cell niche. In this compartment, the oxygen tension is low and oxygen partial pressure is estimated to range from 1% to 7%. We analyzed the effect of low oxygen tensions on human MSC cultured with platelet-lysate supplemented media and assessed proliferation, morphology, chromosomal stability, immunophenotype and plasticity. Results After transferring MSC from atmospheric oxygen levels of 21% to 1%, HIF-1α expression was induced, indicating efficient oxygen reduction. Simultaneously, MSC exhibited a significantly different morphology with shorter extensions and broader cell bodies. MSC did not proliferate as rapidly as under 21% oxygen and accumulated in G1 phase. The immunophenotype, however, was unaffected. Hypoxic stress as well as free oxygen radicals may affect chromosomal stability. However, no chromosomal abnormalities in human MSC under either culture condition were detected using high-resolution matrix-based comparative genomic hybridization. Reduced oxygen tension severely impaired adipogenic and osteogenic differentiation of human MSC. Elevation of oxygen from 1% to 3% restored osteogenic differentiation. Conclusion Physiologic oxygen tension during in vitro culture of human MSC slows down cell cycle progression and differentiation. Under physiological conditions this may keep a proportion of MSC in a resting state. Further studies are needed to analyze these aspects of MSC in tissue regeneration.

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Low physiologic oxygen tensions reduce proliferation and differentiation of human multipotent mesenchymal stromal cells

Christina Holzwarth 0 3 Martin Vaegler 0 3 Friederike Gieseke 0 3 Stefan M Pfister 2 Rupert Handgretinger 0 3 Gunter Kerst 1 Ingo Mller 0 3 0 University Children's Hospital, Department of General Pediatrics, Hematology and Oncology , Tubingen , Germany 1 University Children's Hospital, Department of Pediatric Cardiology, Pulmology and Intensive Care , Tubingen , Germany 2 German Cancer Research Center and University Children's Hospital , Heidelberg , Germany 3 University Children's Hospital, Department of General Pediatrics, Hematology and Oncology , Tubingen , Germany Background: Human multipotent mesenchymal stromal cells (MSC) can be isolated from various tissues including bone marrow. Here, MSC participate as bone lining cells in the formation of the hematopoietic stem cell niche. In this compartment, the oxygen tension is low and oxygen partial pressure is estimated to range from 1% to 7%. We analyzed the effect of low oxygen tensions on human MSC cultured with platelet-lysate supplemented media and assessed proliferation, morphology, chromosomal stability, immunophenotype and plasticity. Results: After transferring MSC from atmospheric oxygen levels of 21% to 1%, HIF-1a expression was induced, indicating efficient oxygen reduction. Simultaneously, MSC exhibited a significantly different morphology with shorter extensions and broader cell bodies. MSC did not proliferate as rapidly as under 21% oxygen and accumulated in G1 phase. The immunophenotype, however, was unaffected. Hypoxic stress as well as free oxygen radicals may affect chromosomal stability. However, no chromosomal abnormalities in human MSC under either culture condition were detected using high-resolution matrix-based comparative genomic hybridization. Reduced oxygen tension severely impaired adipogenic and osteogenic differentiation of human MSC. Elevation of oxygen from 1% to 3% restored osteogenic differentiation. Conclusion: Physiologic oxygen tension during in vitro culture of human MSC slows down cell cycle progression and differentiation. Under physiological conditions this may keep a proportion of MSC in a resting state. Further studies are needed to analyze these aspects of MSC in tissue regeneration. - Background Human multipotent mesenchymal stromal cells (MSC) obtained from bone marrow are characterized by a multilineage differentiation potential and a high proliferative capacity without losing their genetic stability [1]. By the mid of the last decade, the clinical potential was recognized [2]. Today, several clinical trials showed that the application of ex vivo expanded MSC is safe and feasible [3-5]. MSC have earned considerable attention as therapeutic tools in graft-versus-host disease after allogeneic hematopoietic stem cell transplantation [6]. The treatment of pediatric patients suffering from osteogenesis imperfecta was the first successful clinical application of MSC in regenerative medicine [3,7]. Another promising application of MSC has been explored in a pilot study where five children with steroid-induced osteonecrosis of the femur received MSC directly injected into the necrotic area of the bone [8]. Regenerative properties of MSC could also be demonstrated in the treatment of ischemic cardiovascular diseases [9]. These examples of tissue regeneration share the phenomenon of oxygen deprivation in the affected organs challenging the ability of MSC to differentiate into bone or other tissues [10]. Physiologically the cells are adapted to low oxygen levels with oxygen concentrations between 1% and 7%. Mathematical models of the pO2 distribution in human bone marrow suggest a gradient across the marrow from the relatively well oxygenated sinuses to the rather hypoxic endosteal region [11]. It is known that low oxygen tension is involved in keeping stem cells in a quiescent state retaining their plasticity [12]. Conversely, hypoxia may also serve as a danger signal and recruit MSC. In a rat model, Rochefort and colleagues showed that specifically MSC and not hematopoietic progenitor cells were mobilized out of the bone marrow into the peripheral blood by hypoxia [13]. However, it is an open issue if various oxygen concentrations over prolonged periods of time change the characteristic properties which define MSC in vitro as proposed by the International Society for Cellular Therapy [14]. Reports addressing this issue obtained conflicting results: In several in vitro studies, low oxygen concentrations have been found to be stimulating differentiation processes, exemplarily shown by inducing the cells toward the adipogenic, osteogenic or chondrogenic lineage [15-17]. In contrast, other groups reported suppressive effects of reduced oxygen tensions on MSC plasticity [10,18]. It is hypothesized that survival and proliferative capacity of MSC can be enhanced by maintaining the cells under low oxygen tensions. Standard culture of MSC at 21% O2 imply hyperoxic conditions compared to physiological O2 concentrations, i. (...truncated)


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Christina Holzwarth, Martin Vaegler, Friederike Gieseke, Stefan M Pfister, Rupert Handgretinger, Gunter Kerst, Ingo Müller. Low physiologic oxygen tensions reduce proliferation and differentiation of human multipotent mesenchymal stromal cells, BMC Cell Biology, 2010, pp. 11, 11, DOI: 10.1186/1471-2121-11-11