BACKGROUND
human mesenchymal stem cells  were first identified by friedenstein et al.
<cit>  in  <dig>  as non-hematopoetic, multipotent bone marrow stem cells, hmscs are more primitive and embryonic-like cells with the potential to differentiate into lineage-committed progenitors and mature cells, such as osteoblasts and fibroblasts  <cit> .
however, bone marrow is not the exclusive source of mscs; they have been isolated from virtually all post-natal and extra-embryonic tissues, including amniotic membrane, placenta, umbilical cord, and umbilical cord blood  <cit> .
in recent years, hmscs have been commonly used in tissue engineering, cell replacement therapy, gene therapy, and body organ/fluid transplantation.
oxygen is a potent signaling molecule, affecting the fundamental characteristics of various types of cells.
specifically, reduced oxygen levels or hypoxia influences blood-brain barrier permeability through influencing endothelial cell junctional complexes  <cit> .
furthermore, hypoxia induced proliferation of hematopoietic bone marrow stem cells  <cit> .
in addition, hypoxia-inducible factor- <dig> alpha , a critical transcription factor in the mammalian oxygen-sensing pathway, is activated in response to hypoxia, altering tumor xenograft gene expression, growth, and angiogenesis  <cit>  possibly through membrane type  <dig> metalloprotease  <cit> .
mscs exposed to hypoxic conditions exhibit greater colony-forming potential  <cit> , faster and prolonged proliferation  <cit> , and greater chemotaxis  <cit> .
exposure of mscs to hypoxia also prolonged their differentiation  <cit> .
hypoxia is often induced by decreasing oxygen concentrations  <cit> .
in addition, the hypoxia-mimetic agents, cobalt chloride  and desferrioxamine , an iron chelator, artificially induce hypoxia through blocking the degradation of hif-1α  <cit> .
the effects of hypoxia-mimetic agents are comparable to those resulting from reduced atmospheric oxygen levels  <cit> .
analysis of stem cell proliferation and differentiation is often carried out in 20% o <dig>  which is much greater than that found in vivo; arterial oxygen concentration is approximately 12% while bone marrow ranges from  <dig> to 7%  <cit> .
therefore, the effects of dfo and cocl <dig> on human umbilical cord-isolated hmscs were assessed.
the ultrastructure of hmscs in response to hypoxia was analyzed using atomic force microscopy  and transmission electron microscopy .
in addition, the effects of hypoxia mimetics on hmscs proliferation were analyzed.
determining the effects of hypoxia on hmscs morphology and proliferation may contribute to identifying the mechanisms by which hypoxia influences hmscs chemotaxis and migration as well as characterize their therapeutic application for hypoxia-related disease.
RESULTS
morphological and immunophenotypic characterization of umbilical cord-derived hmscs
both large and occasionally multi-nucleated hmscs as well as small, spindle-shaped mononucleated cells were present in the primary hmscs culture .
following the second passage, the cultures were primarily made up of the small, spindle-shaped cells, which continued to proliferate after numerous passages; the number of large hmscs diminished .
individual spindle-shaped cells appeared after 3- <dig> days in culture, while colonies formed as early as  <dig> days.
the primary hmscs expressed cd <dig> , cd <dig> , and cd <dig>  surface antigens whereas expression of the cd <dig> , cd <dig> , cd <dig> , cd <dig> , and hla-dr  surface molecules were below the detection limit .
hmscs differentiation
following adipogenic induction, hmscs morphology changed from elongated, confluent fibroblastic-like cells  to oval-shaped cells; a distinct ring of red coarse vacuoles around the cell periphery was observed upon oil red o staining after the fourth day, becoming larger and more numerous over time .
upon osteogenic differentiation, adherent monolayers of spindle-shaped cells  became multilayered cell clusters surrounded by a matrix-like substance that was visible after von kossa staining .
in addition, rapid mineralization and nodule formation characterized by the accumulation of overcrowded fibroblast-like cells in direct contact with one another was observed.
the cells bordering the nodules were fibroblastic, while those near the centers were polygonal .
effects of dfo and cocl <dig> on hmscs proliferation
hmscs proliferation rates were determined using the mtt assay; logarithmic growth was observed after  <dig> h. decreased hmscs proliferation was detected after dfo  and cocl <dig>  treatment.
specifically, significantly decreased cell growth was observed after treatment with  <dig> μm dfo and  <dig> and  <dig> μm cocl <dig> as compared to the untreated controls .
after  <dig> h, the inhibition rates were  <dig> ,  <dig> ,  <dig> , and  <dig> % for cells treated with  <dig>   <dig>   <dig> and  <dig> μm dfo, respectively .
similarly, the inhibition rates were  <dig> ,  <dig> ,  <dig> , and  <dig> % for hmscs treated for  <dig> h with  <dig>   <dig>   <dig>  and  <dig> μm cocl <dig>  respectively .
effects of hypoxia mimetics on hmscs cell cycle
as compared to the control group, the ratio of hmscs in the g0/g <dig> phase increased while those in the g2/m/s phase decreased after dfo and cocl <dig> treatment .
although no significant differences were observed among the various dfo or cocl <dig> concentrations, an increasing trend was observed in the number of cells in the g0/g <dig> phase while a decreasing trend was detected in the number of cells in the g2/s/m phase after treatment .
effects of hypoxia mimetics on hmscs morphology
after  <dig> h in normoxic conditions, afm revealed enlarged, spindle-shaped hmscs up to  <dig> μm in length and 30- <dig> μm wide .
in addition, a clear cytoskeleton, complex edge as well as synapse-like structure, and fishtail-like or cicada wing-like morphology were observed .
in addition, cells were in close contact with each other and presented a reticular structure, suggesting active cell-to-cell signaling .
hmscs were exposed to factors inducing adipogenic and osteogenic differentiation for  <dig> days in normoxic conditions after which the morphology was assessed.
the nuclei of adipogenic-induced hmscs moved to peripheral area while the cell membranes became more granular, which was suggestive of the s phase of the cell cylce .
osteogenic-induced hmscs appeared more elevated, square and rigid than undifferentiated cells .
altered hmscs morphology was observed after treatment with dfo  and cocl <dig> .
specifically, the spindle-like cells elongated up to nearly  <dig> μm.
in addition, cell-to-cell contacts diminished, and gaps appeared in between adjacent cells.
tem analysis of control hmscs revealed round cells with surface protrusions and microvilli .
the nuclei were irregularly shaped with ≥ one nucleoli, containing a large amount of euchromatin and a smaller amount of heterochromatin.
with the exception of a small amount of rough endoplasmic reticulum , ribosomes, and mitochondria  with clear and continuous cristae, relatively few organelles were found in the cytoplasm, suggesting that they had remained undifferentiated.
after treatment with dfo, more surface protrusions and microvilli, as well as a large number of intracellular vacuole-like structures were observed .
in addition, dfo treatment resulted in shrinkage or partial disintegration of the nuclear membrane, as well as chromatin condensation.
furthermore, an expanded rough endoplasmic reticulum, which was filled with a low density matrix, was detected.
although the number and volume of mitochondria were not significantly different from the control groups, mitochondria with an expanded ridge and containing a high density matrix were observed in dfo-treated hmscs .
significant changes in other organelles were not observed.
discussion
human umbilical cords represent a promising source of hmscs, which can be conveniently isolated and induce low immunogenicity.
in addition, as compared to other stem cell sources these present relatively few ethical issues.
the hmscs isolated from umbilical cords were positive for cd <dig>  cd <dig> and cd <dig>  and negative for cd <dig>  cd <dig>  cd <dig>  cd <dig> and hla-dr surface antigens, confirming their identity as mscs  <cit> .
in addition, the isolated hmscs were able to differentiate into adipogenic and osteogenic cells under specific culture conditions.
the intracellular cytoskeleton responds to external mechanical stimuli  <cit> .
tapping mode afm was used to observe the surface structures.
the position of the adipogenic cell nuclei and their granular quality of the membranes is suggestive of early s phase of the cell cycle.
osteogenic-induced hmscs appeared larger, squarer and more rigid as compared to undifferentiated cells, which is consistent with that reported by danti et al.
<cit> .
in addition, they were easily distinguishable from each other as the differentiation process prevented their proliferation, which also correlates with danti et al  <cit> .
differentiation of mscs is also influenced by hypoxia.
qu et al.
<cit>  reported that dfo increased the osteoblastic differentiation of bone morphogenetic protein-2-treated mscs.
valorani et al.
<cit>  and ren et al.
<cit>  observed that hypoxia  increased the adipogenic differentiation potential of mscs.
these studies suggest that pre-culturing mscs under hypoxic conditions prior to transplantation may enhance their efficacy.
as evidenced by clinical trials in hypoxic-ischemic diseases, mscs-based therapy has potential value in tissue replacement and regeneration  <cit> .
the effects of in vivo oxygen concentrations, which are much lower than common experimental conditions, on hmscs were largely unknown.
therefore, hmscs proliferation and morphology in response to hypoxia induced by dfo and cocl <dig> was assessed.
after treatment with dfo and cocl <dig>  hmscs were more elongated, and gaps appeared between adjacent cells.
thus, dfo and cocl <dig> may inhibit hmscs growth by weakening cell-to-cell signaling.
the reduction in cell-cell junctions may also mediate hmscs migration induced by hypoxia   <cit> .
further studies are necessary to determine the effects of dfo and cocl <dig> treatment on the signaling pathways that govern hmscs migration.
the influence of hypoxia on hmscs ultrastructure was also explored by tem.
after treatment with dfo, hmscs contained a large number of unidentifiable vacuoles that are early markers of apoptosis, which is consistent with ren et al.
<cit>  using 8% oxygen.
in addition, the observed shrinkage, disintegration, and dissolution of the nucleus, along with chromatin condensation indicated early apoptosis.
consistent with the signs of apoptosis, we found that hmscs proliferation decreased in a dose-dependent manner with increasing concentrations of cocl <dig> and dfo.
however, discrepant effects of physical hypoxia and hypoxia mimetics on mscs proliferation indicate differences.
for example, ren et al.
<cit>  reported that low oxygen levels  promoted mscs proliferation, whereas dfo  and cocl <dig>  inhibited their growth.
lavrentieva et al.
<cit>  reported that  <dig> -5% oxygen levels increased the proliferative capacity of hmscs.
whereas qu et al.
<cit>  found that dfo, ranging from  <dig> to  <dig> μm, inhibited cell growth in a dose-dependent manner.
hif-1α levels may differ under physically-induced hypoxia as opposed to cocl2-induced hypoxia  <cit>  which may account for these differences; however, further analysis is required.
the effects of cocl <dig> and dfo on hmscs proliferation may be mediated by cell cycle changes.
a larger percentage of dfo- and cocl2-treated cells in the g0/g <dig> phase was observed, while the ratio of those in the g2/m/s phase decreased.
similar results were reported by holzwarth et al.
<cit> , who reported hmscs accumulation in the g <dig> phase at 1% oxygen.
further studies are necessary to determine if the cell cycle effects of hypoxia mimetics can be recapitulated under low oxygen conditions.
there are several study limitations that warrant discussion.
firstly, the present study analyzed the effects of hypoxia mimics on hmscs; however, the effects of physical hypoxia were not assessed.
further studies will be carried out to compare the effects of physical hypoxia and hypoxia mimetics on hmscs morphology and growth.
in addition, the effects of dfo and cocl <dig> on hmsc hif-1α levels were not analyzed.
however, previous studies using the same concentrations of dfo and cocl <dig> used in the present study have reported upregulation of hif-1α expression  <cit> .
furthermore, the present study did not analyze the influence of dfo and cocl <dig> on mscs differentiation.
although the effects of these agents on mscs morphology is suggestive of greater self-renewal capacity as hmscs broaden and flatten with differentiation  <cit> , and differentiation was associated with changes in nuclear morphology  <cit> , determining their effects on mscs differentiation will be the focus of future studies.
finally, the effects of hypoxia-induced morphological changes on mscs function  was not explored in the present study, but will be analyzed in future studies.
although the present study has its limitations, determining the ways in which mscs respond to environments with lower than atmospheric oxygen concentrations, such as the blood, bone marrow, and cartilage, is crucial for their successful use regenerative medicine.
the present study advances our understanding of the influences of hypoxia on mscs morphology and proliferation.
CONCLUSIONS
we isolated and cultured hmscs from umbilical cords.
the proliferation of hmscs was inhibited by dfo and cocl <dig>  the cell surface and ultrastructure was viewed with atomic force microscopy and transmission electron microscopy after dfo- and cocl2-induced hypoxia, which is the first time this has been reported.
these data will provide a better understanding of the potential therapeutic applications of hmscs in hypoxic-ischemic disease.

 human mesenchymal stem cells were first identified by friedenstein et al. in as non-hematopoetic, multipotent bone marrow stem cells, hmscs are more primitive and embryonic-like cells with the potential to differentiate into lineage-committed progenitors and mature cells, such as osteoblasts and fibroblasts. however, bone marrow is not the exclusive source of mscs; they have been isolated from virtually all post-natal and extra-embryonic tissues, including amniotic membrane, placenta, umbilical cord, and umbilical cord blood. in recent years, hmscs have been commonly used in tissue engineering, cell replacement therapy, gene therapy, and body organ/fluid transplantation. oxygen is a potent signaling molecule, affecting the fundamental characteristics of various types of cells. specifically, reduced oxygen levels or hypoxia influences blood-brain barrier permeability through influencing endothelial cell junctional complexes. furthermore, hypoxia induced proliferation of hematopoietic bone marrow stem cells. in addition, hypoxia-inducible factor- alpha, a critical transcription factor in the mammalian oxygen-sensing pathway, is activated in response to hypoxia, altering tumor xenograft gene expression, growth, and angiogenesis possibly through membrane type metalloprotease. mscs exposed to hypoxic conditions exhibit greater colony-forming potential, faster and prolonged proliferation, and greater chemotaxis. exposure of mscs to hypoxia also prolonged their differentiation. hypoxia is often induced by decreasing oxygen concentrations. in addition, the hypoxia-mimetic agents, cobalt chloride and desferrioxamine, an iron chelator, artificially induce hypoxia through blocking the degradation of hif-1α. the effects of hypoxia-mimetic agents are comparable to those resulting from reduced atmospheric oxygen levels. analysis of stem cell proliferation and differentiation is often carried out in 20% o which is much greater than that found in vivo; arterial oxygen concentration is approximately 12% while bone marrow ranges from to 7%. therefore, the effects of dfo and cocl on human umbilical cord-isolated hmscs were assessed. the ultrastructure of hmscs in response to hypoxia was analyzed using atomic force microscopy and transmission electron microscopy. in addition, the effects of hypoxia mimetics on hmscs proliferation were analyzed. determining the effects of hypoxia on hmscs morphology and proliferation may contribute to identifying the mechanisms by which hypoxia influences hmscs chemotaxis and migration as well as characterize their therapeutic application for hypoxia-related disease.  morphological and immunophenotypic characterization of umbilical cord-derived hmscs both large and occasionally multi-nucleated hmscs as well as small, spindle-shaped mononucleated cells were present in the primary hmscs culture. following the second passage, the cultures were primarily made up of the small, spindle-shaped cells, which continued to proliferate after numerous passages; the number of large hmscs diminished. individual spindle-shaped cells appeared after 3- days in culture, while colonies formed as early as days. the primary hmscs expressed cd, cd, and cd surface antigens whereas expression of the cd, cd, cd, cd, and hla-dr surface molecules were below the detection limit. hmscs differentiation following adipogenic induction, hmscs morphology changed from elongated, confluent fibroblastic-like cells to oval-shaped cells; a distinct ring of red coarse vacuoles around the cell periphery was observed upon oil red o staining after the fourth day, becoming larger and more numerous over time. upon osteogenic differentiation, adherent monolayers of spindle-shaped cells became multilayered cell clusters surrounded by a matrix-like substance that was visible after von kossa staining. in addition, rapid mineralization and nodule formation characterized by the accumulation of overcrowded fibroblast-like cells in direct contact with one another was observed. the cells bordering the nodules were fibroblastic, while those near the centers were polygonal. effects of dfo and cocl on hmscs proliferation hmscs proliferation rates were determined using the mtt assay; logarithmic growth was observed after h. decreased hmscs proliferation was detected after dfo and cocl treatment. specifically, significantly decreased cell growth was observed after treatment with μm dfo and and μm cocl as compared to the untreated controls. after h, the inhibition rates were,,, and % for cells treated with and μm dfo, respectively. similarly, the inhibition rates were,,, and % for hmscs treated for h with and μm cocl respectively. effects of hypoxia mimetics on hmscs cell cycle as compared to the control group, the ratio of hmscs in the g0/g phase increased while those in the g2/m/s phase decreased after dfo and cocl treatment. although no significant differences were observed among the various dfo or cocl concentrations, an increasing trend was observed in the number of cells in the g0/g phase while a decreasing trend was detected in the number of cells in the g2/s/m phase after treatment. effects of hypoxia mimetics on hmscs morphology after h in normoxic conditions, afm revealed enlarged, spindle-shaped hmscs up to μm in length and 30- μm wide. in addition, a clear cytoskeleton, complex edge as well as synapse-like structure, and fishtail-like or cicada wing-like morphology were observed. in addition, cells were in close contact with each other and presented a reticular structure, suggesting active cell-to-cell signaling. hmscs were exposed to factors inducing adipogenic and osteogenic differentiation for days in normoxic conditions after which the morphology was assessed. the nuclei of adipogenic-induced hmscs moved to peripheral area while the cell membranes became more granular, which was suggestive of the s phase of the cell cylce. osteogenic-induced hmscs appeared more elevated, square and rigid than undifferentiated cells. altered hmscs morphology was observed after treatment with dfo and cocl. specifically, the spindle-like cells elongated up to nearly μm. in addition, cell-to-cell contacts diminished, and gaps appeared in between adjacent cells. tem analysis of control hmscs revealed round cells with surface protrusions and microvilli. the nuclei were irregularly shaped with ≥ one nucleoli, containing a large amount of euchromatin and a smaller amount of heterochromatin. with the exception of a small amount of rough endoplasmic reticulum, ribosomes, and mitochondria with clear and continuous cristae, relatively few organelles were found in the cytoplasm, suggesting that they had remained undifferentiated. after treatment with dfo, more surface protrusions and microvilli, as well as a large number of intracellular vacuole-like structures were observed. in addition, dfo treatment resulted in shrinkage or partial disintegration of the nuclear membrane, as well as chromatin condensation. furthermore, an expanded rough endoplasmic reticulum, which was filled with a low density matrix, was detected. although the number and volume of mitochondria were not significantly different from the control groups, mitochondria with an expanded ridge and containing a high density matrix were observed in dfo-treated hmscs. significant changes in other organelles were not observed. discussion human umbilical cords represent a promising source of hmscs, which can be conveniently isolated and induce low immunogenicity. in addition, as compared to other stem cell sources these present relatively few ethical issues. the hmscs isolated from umbilical cords were positive for cd cd and cd and negative for cd cd cd cd and hla-dr surface antigens, confirming their identity as mscs. in addition, the isolated hmscs were able to differentiate into adipogenic and osteogenic cells under specific culture conditions. the intracellular cytoskeleton responds to external mechanical stimuli. tapping mode afm was used to observe the surface structures. the position of the adipogenic cell nuclei and their granular quality of the membranes is suggestive of early s phase of the cell cycle. osteogenic-induced hmscs appeared larger, squarer and more rigid as compared to undifferentiated cells, which is consistent with that reported by danti et al.. in addition, they were easily distinguishable from each other as the differentiation process prevented their proliferation, which also correlates with danti et al. differentiation of mscs is also influenced by hypoxia. qu et al. reported that dfo increased the osteoblastic differentiation of bone morphogenetic protein-2-treated mscs. valorani et al. and ren et al. observed that hypoxia increased the adipogenic differentiation potential of mscs. these studies suggest that pre-culturing mscs under hypoxic conditions prior to transplantation may enhance their efficacy. as evidenced by clinical trials in hypoxic-ischemic diseases, mscs-based therapy has potential value in tissue replacement and regeneration. the effects of in vivo oxygen concentrations, which are much lower than common experimental conditions, on hmscs were largely unknown. therefore, hmscs proliferation and morphology in response to hypoxia induced by dfo and cocl was assessed. after treatment with dfo and cocl hmscs were more elongated, and gaps appeared between adjacent cells. thus, dfo and cocl may inhibit hmscs growth by weakening cell-to-cell signaling. the reduction in cell-cell junctions may also mediate hmscs migration induced by hypoxia. further studies are necessary to determine the effects of dfo and cocl treatment on the signaling pathways that govern hmscs migration. the influence of hypoxia on hmscs ultrastructure was also explored by tem. after treatment with dfo, hmscs contained a large number of unidentifiable vacuoles that are early markers of apoptosis, which is consistent with ren et al. using 8% oxygen. in addition, the observed shrinkage, disintegration, and dissolution of the nucleus, along with chromatin condensation indicated early apoptosis. consistent with the signs of apoptosis, we found that hmscs proliferation decreased in a dose-dependent manner with increasing concentrations of cocl and dfo. however, discrepant effects of physical hypoxia and hypoxia mimetics on mscs proliferation indicate differences. for example, ren et al. reported that low oxygen levels promoted mscs proliferation, whereas dfo and cocl inhibited their growth. lavrentieva et al. reported that -5% oxygen levels increased the proliferative capacity of hmscs. whereas qu et al. found that dfo, ranging from to μm, inhibited cell growth in a dose-dependent manner. hif-1α levels may differ under physically-induced hypoxia as opposed to cocl2-induced hypoxia which may account for these differences; however, further analysis is required. the effects of cocl and dfo on hmscs proliferation may be mediated by cell cycle changes. a larger percentage of dfo- and cocl2-treated cells in the g0/g phase was observed, while the ratio of those in the g2/m/s phase decreased. similar results were reported by holzwarth et al., who reported hmscs accumulation in the g phase at 1% oxygen. further studies are necessary to determine if the cell cycle effects of hypoxia mimetics can be recapitulated under low oxygen conditions. there are several study limitations that warrant discussion. firstly, the present study analyzed the effects of hypoxia mimics on hmscs; however, the effects of physical hypoxia were not assessed. further studies will be carried out to compare the effects of physical hypoxia and hypoxia mimetics on hmscs morphology and growth. in addition, the effects of dfo and cocl on hmsc hif-1α levels were not analyzed. however, previous studies using the same concentrations of dfo and cocl used in the present study have reported upregulation of hif-1α expression. furthermore, the present study did not analyze the influence of dfo and cocl on mscs differentiation. although the effects of these agents on mscs morphology is suggestive of greater self-renewal capacity as hmscs broaden and flatten with differentiation, and differentiation was associated with changes in nuclear morphology, determining their effects on mscs differentiation will be the focus of future studies. finally, the effects of hypoxia-induced morphological changes on mscs function was not explored in the present study, but will be analyzed in future studies. although the present study has its limitations, determining the ways in which mscs respond to environments with lower than atmospheric oxygen concentrations, such as the blood, bone marrow, and cartilage, is crucial for their successful use regenerative medicine. the present study advances our understanding of the influences of hypoxia on mscs morphology and proliferation.  we isolated and cultured hmscs from umbilical cords. the proliferation of hmscs was inhibited by dfo and cocl the cell surface and ultrastructure was viewed with atomic force microscopy and transmission electron microscopy after dfo- and cocl2-induced hypoxia, which is the first time this has been reported. these data will provide a better understanding of the potential therapeutic applications of hmscs in hypoxic-ischemic disease. 
the therapeutic efficacy of human mesenchymal stem cells for the treatment of hypoxic-ischemic diseases is closely related to level of hypoxia in the damaged tissues. the hypoxia-mimetic agents, dfo and cocl alter umbilical cord-derived hmscs morphology and inhibit their proliferation through influencing the cell cycle. after treatment with dfo and cocl hmscs were elongated, and adjacent cells were no longer in close contact. in addition, dfo and cocl treatments for h significantly inhibited hmscs proliferation in a concentration-dependent manner. in addition, vacuole-like structures were observed within the cytoplasm; the rough endoplasmic reticulum expanded, and expanded ridges were observed in mitochondria. to elucidate the potential therapeutic applications and limitations of hmscs derived from human umbilical cords, the effects of hypoxia on the morphology and proliferation of hmscs were analyzed. this treatment also increased the number of cells in g0/g phase and decreased those in g2/s/m phase.
 the therapeutic efficacy of human mesenchymal stem cells for the treatment of hypoxic-ischemic diseases is closely related to level of hypoxia in the damaged tissues. to elucidate the potential therapeutic applications and limitations of hmscs derived from human umbilical cords, the effects of hypoxia on the morphology and proliferation of hmscs were analyzed.  after treatment with dfo and cocl hmscs were elongated, and adjacent cells were no longer in close contact. in addition, vacuole-like structures were observed within the cytoplasm; the rough endoplasmic reticulum expanded, and expanded ridges were observed in mitochondria. in addition, dfo and cocl treatments for h significantly inhibited hmscs proliferation in a concentration-dependent manner. this treatment also increased the number of cells in g0/g phase and decreased those in g2/s/m phase.  the hypoxia-mimetic agents, dfo and cocl alter umbilical cord-derived hmscs morphology and inhibit their proliferation through influencing the cell cycle. 