Hypoxia enhances proliferation and tissue formation of human mesenchymal stem cells.
06 Jul 2007-Biochemical and Biophysical Research Communications (Biochem Biophys Res Commun)-Vol. 358, Iss: 3, pp 948-953
TL;DR: It is demonstrated that oxygen concentrations affected many aspects of stem-cell physiology, including growth and in vitro development, and may be a critical parameter during expansion and differentiation.
About: This article is published in Biochemical and Biophysical Research Communications.The article was published on 2007-07-06. It has received 509 citations till now. The article focuses on the topics: Cellular differentiation & Progenitor cell.
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TL;DR: MSC derived from different adult and neonatal tissues are compared with respect to their cell biological properties, surface marker expression and proliferative capacities and several MSC functions including in vitro and in vivo differentiation capacities within a variety of lineages and immune-modulatory properties are highlighted.
Abstract: The mesenchymal stroma harbors an important population of cells that possess stem cell-like characteristics including self renewal and differentiation capacities and can be derived from a variety of different sources. These multipotent mesenchymal stem cells (MSC) can be found in nearly all tissues and are mostly located in perivascular niches. MSC have migratory abilities and can secrete protective factors and act as a primary matrix for tissue regeneration during inflammation, tissue injuries and certain cancers. These functions underlie the important physiological roles of MSC and underscore a significant potential for the clinical use of distinct populations from the various tissues. MSC derived from different adult (adipose tissue, peripheral blood, bone marrow) and neonatal tissues (particular parts of the placenta and umbilical cord) are therefore compared in this mini-review with respect to their cell biological properties, surface marker expression and proliferative capacities. In addition, several MSC functions including in vitro and in vivo differentiation capacities within a variety of lineages and immune-modulatory properties are highlighted. Differences in the extracellular milieu such as the presence of interacting neighbouring cell populations, exposure to proteases or a hypoxic microenvironment contribute to functional developments within MSC populations originating from different tissues, and intracellular conditions such as the expression levels of certain micro RNAs can additionally balance MSC function and fate.
1,369 citations
Cites background from "Hypoxia enhances proliferation and ..."
...Previous work has demonstrated that the culture of MSC under hypoxic conditions is accompanied by increased Oct4 expression and telomerase activity [81] which are involved in the maintenance of stemness....
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...Indeed, MSC cultured under normoxic conditions exhibit premature senescence and a reduction in population doublings in comparison to cells cultured under hypoxia [81,82] and may also show restricted cell division due to telomere shortening and replicative senescence [80,83]....
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TL;DR: This work has identified a broader spectrum of stem cells influenced by hypoxia that includes cancer stem cells and induced pluripotent stem cells, and elucidate an added dimension of stem cell control within the niche.
1,332 citations
Cites background from "Hypoxia enhances proliferation and ..."
...When cultured under hypoxic conditions, MSCs demonstrate increased Oct4 expression and telomerase activity (D’Ippolito et al., 2006; Grayson et al., 2007)....
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TL;DR: Intravenous autologous MSCs transplantation was safe for stroke patients during long‐term follow‐up and may improve recovery after stroke depending on the specific characteristics of the patients.
Abstract: We previously evaluated the short-term follow-up preliminary data of mesenchymal stem cells (MSCs) transplantation in patients with ischemic stroke. The present study was conducted to evaluate the long-term safety and efficacy of i.v. MSCs transplantation in a larger population. To accomplish this, we performed an open-label, observer-blinded clinical trial of 85 patients with severe middle cerebral artery territory infarct. Patients were randomly allocated to one of two groups, those who received i.v. autologous ex vivo cultured MSCs (MSC group) or those who did not (control group), and followed for up to 5 years. Mortality of any cause, long-term side effects, and new-onset comorbidities were monitored. Of the 52 patients who were finally included in this study, 16 were the MSC group and 36 were the control group. Four (25%) patients in the MSC group and 21 (58.3%) in the control group died during the follow-up period, and the cumulative surviving portion at 260 weeks was 0.72 in the MSC group and 0.34 in the control group (log-rank; p = .058). Significant side effects were not observed following MSC treatment. The occurrence of comorbidities including seizures and recurrent vascular episodes did not differ between groups. When compared with the control group, the follow-up modified Rankin Scale (mRS) score was decreased, whereas the number of patients with a mRS of 0-3 increased in the MSC group (p = .046). Clinical improvement in the MSC group was associated with serum levels of stromal cell-derived factor-1 and the degree of involvement of the subventricular region of the lateral ventricle. Intravenous autologous MSCs transplantation was safe for stroke patients during long-term follow-up. This therapy may improve recovery after stroke depending on the specific characteristics of the patients.
697 citations
TL;DR: The effects of preculturing human bone marrow‐derived MSC in hypoxic conditions (1%–3% oxygen) to elucidate the best conditions that enhance their tissue regenerative potential are examined, and it is demonstrated that MSC cultured in hypoxia activate the Akt signaling pathway while maintaining their viability and cell cycle rates.
Abstract: Mesenchymal stem cells (MSC) are adult multipotent cells found in bone marrow, adipose tissue, and other adult tissues. MSC have been shown to improve regeneration of injured tissues in vivo, but the mechanisms remain unclear. Typically, MSC are cultured under ambient, or normoxic, conditions (21% oxygen). However, the physiological niches for MSC in the bone marrow and other sites have much lower oxygen tension. When used as a therapeutic tool to repair tissue injuries, MSC cultured in standard conditions must adapt from 21% oxygen in culture to less than 1% oxygen in the ischemic tissue. We therefore examined the effects of preculturing human bone marrow-derived MSC in hypoxic conditions (1%–3% oxygen) to elucidate the best conditions that enhance their tissue regenerative potential. We demonstrated that MSC cultured in hypoxia activate the Akt signaling pathway while maintaining their viability and cell cycle rates. We also showed that MSC cultured in hypoxia induced expression of cMet, the major receptor for hepatocyte growth factor (HGF), and enhanced cMet signaling. MSC cultured in hypoxic conditions increased their migration rates. Since migration and HGF responsiveness are thought to be key mediators of MSC recruitment and/or activation in vivo, we next examined the tissue regenerative potential of MSC cultured under hypoxic conditions, using a murine hind limb ischemia model. We showed that local expression of HGF is increased in ischemic muscle in this model. Intra-arterial injection of MSC cultured in either normoxic or hypoxic conditions 24 hours after surgical induction of hind limb ischemia enhanced revascularization compared with saline controls. However, restoration of blood flow was observed significantly earlier in mice that had been injected with hypoxic preconditioned MSC. Collectively, these data suggest that preculturing MSC under hypoxic conditions prior to transplantation improves their tissue regenerative potential.
649 citations
TL;DR: It is suggested that possible ECM proteins and transcription factors affected by Mg(2+) that are responsible for the enhanced bone regeneration observed around degradable Mg orthopedic/craniofacial devices.
409 citations
Cites background from "Hypoxia enhances proliferation and ..."
...[47] reported that hypoxia induced proliferation and ECM production in mesenchymal stem cells....
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References
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TL;DR: Adult stem cells isolated from marrow aspirates of volunteer donors could be induced to differentiate exclusively into the adipocytic, chondrocytic, or osteocytic lineages.
Abstract: Human mesenchymal stem cells are thought to be multipotent cells, which are present in adult marrow, that can replicate as undifferentiated cells and that have the potential to differentiate to lineages of mesenchymal tissues, including bone, cartilage, fat, tendon, muscle, and marrow stroma. Cells that have the characteristics of human mesenchymal stem cells were isolated from marrow aspirates of volunteer donors. These cells displayed a stable phenotype and remained as a monolayer in vitro. These adult stem cells could be induced to differentiate exclusively into the adipocytic, chondrocytic, or osteocytic lineages. Individual stem cells were identified that, when expanded to colonies, retained their multilineage potential.
20,479 citations
TL;DR: The use of population doubling potential as a measure of biological age suggests that MSCs are intermediately between embryonic and adult tissues, and as such, may provide an in situ source for mesenchymal progenitor cells throughout an adult's lifetime.
Abstract: Recent studies have demonstrated the existence of a subset of cells in human bone marrow capable of differentiating along multiple mesenchymal lineages. Not only do these mesenchymal stem cells (MSCs) possess multilineage developmental potential, but they may be cultured ex vivo for many passages without overt expression of a differentiated phenotype. The goals of the current study were to determine the growth kinetics, self-renewing capacity and the osteogenic potential of purified MSCs during extensive subcultivation and following cryopreservation. Primary cultures of MSCs were established from normal iliac crest bone marrow aspirates, an aliquot was cryopreserved and thawed, and then both frozen and unfrozen populations were subcultivated in parallel for as many as 15 passages. Cells derived from each passage were assayed for their kinetics of growth and their osteogenic potential in response to an osteoinductive medium containing dexamethasone. Spindle-shaped human MSCs in primary culture exhibit a lag phase of growth, followed by a log phase, finally resulting in a growth plateau state. Passaged cultures proceed through the same stages, however, the rate of growth in log phase and the final number of cells after a fixed period in culture diminishes as a function of continued passaging. The average number of population doublings for marrow-derived adult human MSCs was determined to be 38 +/- 4, at which time the cells finally became very broad and flattened before degenerating. The osteogenic potential of cells was conserved throughout every passage as evidenced by the significant increase in APase activity and formation of mineralized nodular aggregates. Furthermore, the process of cryopreserving and thawing the cells had no effect on either their growth or osteogenic differentiation. Importantly, these studies demonstrate that replicative senescence of MSCs is not a state of terminal differentiation since these cells remain capable of progressing through the osteogenic lineage. The use of population doubling potential as a measure of biological age suggests that MSCs are intermediately between embryonic and adult tissues, and as such, may provide an in situ source for mesenchymal progenitor cells throughout an adult's lifetime.
1,540 citations
TL;DR: Assays for adipogenesis demonstrated that the adipogenic potential of cultures was directly related to their ability to generate single‐cell‐derived colonies and their enrichment for RS‐1A cells, and cultures enriched for RS-1B cells showed the greatest potential to differentiate into cartilage in a serum‐free system.
Abstract: There is considerable interest in the biology and therapeutic potential of adult stem cells from bone marrow stroma, variously referred to as mesenchymal stem cells or marrow stromal cells (MSCs). Human MSCs can expand rapidly in culture, but the rate of expansion and the yields of multipotential progenitors are inversely related to the plating density and incubation time of each passage. We have defined conditions for optimizing the yields of cultures enriched for early progenitors. Also, we developed a simple method for assessing the quality of the cultures by phase-contrast microscopy and image analysis or by forward light scatter in a flow cytometer. The cells expanded most rapidly on day 4 after plating, with a minimum average doubling time of about 10 hours for cells initially plated at 10 or 50 cells/cm(2). After plating the cells at 1 to 1000 cells/cm(2), the cultures underwent a time-dependent transition from early progenitors, defined as thin, spindle-shaped cells (RS-1A), to wider, spindle-shaped cells (RS-1B), and to still wider, spindle-shaped cells (RS-1C). Assays for adipogenesis demonstrated that the adipogenic potential of cultures was directly related to their ability to generate single-cell-derived colonies and their enrichment for RS-1A cells. In contrast, cultures enriched for RS-1B cells showed the greatest potential to differentiate into cartilage in a serum-free system. The results indicate that, when preparing cultures of human MSCs, it is necessary to compromise between conditions that provide the highest overall yields and those that provide the highest content of early progenitor cells.
979 citations
TL;DR: The appearance of differentiated regions as assessed morphologically, biochemically, and immunohistochemically was markedly reduced under hypoxic conditions, and hES cell growth under hypoxia provided enhanced formation of embryoid bodies.
Abstract: Early-stage mammalian embryos develop in a low O2 environment (hypoxia). hES cells, however, are generally cultured under an atmosphere of 21% O2 (normoxia), under which conditions they tend to differentiate spontaneously. Such conditions may not be the most suitable, therefore, for hES cell propagation. Here we have tested two hypotheses. The first hypothesis was that hES cells would grow as well under hypoxic as under normoxic conditions. The second hypothesis was that hypoxic culture would reduce the amount of spontaneous cell differentiation that occurs in hES colonies. Both hypotheses proved to be correct. Cells proliferated as well under 3% and 5% O2 as they did under 21% O2, and growth was only slightly reduced at 1% O2. The appearance of differentiated regions as assessed morphologically, biochemically (by the production of human chorionic gonadotropin and progesterone), and immunohistochemically (by the loss of stage-specific embryonic antigen-4 and Oct-4 and gain of stage-specific embryonic antigen-1 marker expression) was markedly reduced under hypoxic conditions. In addition, hES cell growth under hypoxia provided enhanced formation of embryoid bodies. Hypoxic culture would appear to be necessary to maintain full pluripotency of hES cells.
854 citations
TL;DR: Human marrow-isolated adult multilineage inducible cells, or MIAMI cells, proliferate extensively without evidence of senescence or loss of differentiation potential and thus may represent an ideal candidate for cellular therapies of inherited or degenerative diseases.
Abstract: We report here the isolation of a population of non-transformed pluripotent human cells from bone marrow after a unique expansion/selection procedure. This procedure was designed to provide conditions resembling the in vivo microenvironment that is home for the most-primitive stem cells. Marrow-adherent and -nonadherent cells were co-cultured on fibronectin, at low oxygen tension, for 14 days. Colonies of small adherent cells were isolated and further expanded on fibronectin at low density, low oxygen tension with 2% fetal bovine serum. They expressed high levels of CD29, CD63, CD81, CD122, CD164, hepatocyte growth factor receptor (cMet), bone morphogenetic protein receptor 1B (BMPR1B), and neurotrophic tyrosine kinase receptor 3 (NTRK3) and were negative for CD34, CD36, CD45, CD117 (cKit) and HLADR. The embryonic stem cell markers Oct-4 and Rex-1, and telomerase were expressed in all cultures examined. Cell-doubling time was 36 to 72 hours, and cells have been expanded in culture for more than 50 population doublings. This population of cells was consistently isolated from men and women of ages ranging from 3- to 72-years old. Colonies of cells expressed numerous markers found among embryonic stem cells as well as mesodermal-, endodermal- and ectodermal-derived lineages. They have been differentiated to bone-forming osteoblasts, cartilage-forming chondrocytes, fat-forming adipocytes and neural cells and to attachment-independent spherical clusters expressing genes associated with pancreatic islets. Based on their unique characteristics and properties, we refer to them as human marrow-isolated adult multilineage inducible cells, or MIAMI cells. MIAMI cells proliferate extensively without evidence of senescence or loss of differentiation potential and thus may represent an ideal candidate for cellular therapies of inherited or degenerative diseases.
719 citations