International Journal of Cardiovascular ResearchISSN: 2324-8602

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Review Article, Int J Cardiovasc Res Vol: 1 Issue: 2

Molecular Aspects Contributing to Clinical Efficiency of Bone Marrow Stem Cell Transplantation

Fatemeh Pourrajab1,2*, Seyed Khalil Forouzannia1 and Seyed Hossain Hekmatimoghadam1,3
1Yazd Cardiovascular Research Center, Shahid Sadoughi University of Medical Sciences, Yazd, Iran
2Department of Clinical Biochemistry and Molecular Biology, Faculty of Medicine, Shahid Sadoughi University of Medical Sciences, Yazd, Iran
3School of paramedicine, Shahid Sadoughi University of Medical Sciences, Yazd, Iran
Corresponding author : Fatemeh Pourrajab, PhD
Yazd Cardiovascular Research Center, Shahid Sadoughi University of Medical Sciences, Yazd, Iran Postal Code: 8917945556
Tel: +98 351 5231421; Fax: +98 351 5253335
E-mail: [email protected]
Received: July 28, 2012 Accepted: August 08, 2012 Published: August 10, 2012
Citation: Pourrajab F, Forouzannia SK, Hekmatimoghadam SH (2012) Molecular Aspects Contributing to Clinical Efficiency of Bone Marrow Stem Cell Transplantation. Int J Cardiovasc Res 1:2. doi:10.4172/2324-8602.1000101


Molecular Aspects Contributing to Clinical Efficiency of Bone Marrow Stem Cell Transplantation

The multifunctional plasticity of bone marrow derived stem cells represents the most favorable for transplantation. The immunosuppressive properties of BMSCs have been demonstrated on a wide range of innate and adaptive immune cells. BMSC have emerged to be one of the most promising candidates for cellular immunotherapy in solid organ transplantation because the reduction of conventional immunosuppression is highly desirable. Particularly, BMSCs are able to migrate to site of inflammation/ stress to balance the cells through the PI3K/Akt pathway. As well as, BMSCs are exclusively recruited to tumor site and inhibit cancerous cells by down-regulation of Wnt signaling and the protein kinases.


Protective recovery; Switching gene expression; Signaling cascades

Protective Recovery and Anti-Oxidative Outcomes of Bmscs Behavior

Immunomodulatory activity of BMSCs to calm down inflammation
The protective properties of BMSCs have been demonstrated on a wide range of cells. The function includes inhibition of proliferation and activation of T lymphocytes, B lymphocytes, natural killer cells, dendritic cells and neutrophils [1]. Activated microglia is a common pathological hallmark of neurodegenerative diseases, whereby their continuous and chronic inflammatory responses are thought to exacerbate neuronal damage [2]. Cell proliferation is a consequence of microglia activation and has been implicated in Alzheimer’s disease, Parkinson’s disease, cerebral infarcts and other neuroinflammatory conditions. BMSCs inhibit microglia proliferation and modulate NO production [3]. The soluble factors released by activated microglia cells alone do stimulate BMSCs to produce high levels of NO [4]. Although NO has implicated as a mediator for T cell activation [5], but its boost level does appear to be one of the major factors in suppression of microglia proliferation by BMSCs [4]. Microglia is the resident macrophages of the central nervous system (CNS) and function as its main immune effector cell. Under normal conditions, they serve an important role in immune surveillance and homeostasis within the CNS [6]. Microglia are also highly responsive to stress and injury and become activated in response to these triggers. Inflammatory responses within the brain are initiated and maintained by a variety of inflammatory mediators secreted by microglia including nitric oxide (NO) and proinflammatory cytokines such as TNF-α and IL-1β [6,7]. The series of phenotypic changes that occur following microglia activation include cell proliferation, morphological changes, upregulation of activation markers such as CD80, CD86 and CD40 and gain of antigen-presenting function [2].
As the immunosuppressive properties of BMSCs, NO has been also suggested as cytoprotective with antioxidant mechanisms against reactive oxygen species, while its level is considered to be appropriate in the context of neurodegenerative disease therapy [8]. Accordingly in the LPS-activated microglia microenvironment, the low numbers of BMSCs significantly reduce NO levels, while the high number of BMSCs leads to an increase of NO production. In the presence of activated T cells, BMSCs are capable to produce NO for arresting the cell cycle of T cells [4,5,9].
It is noteworthy that additional factors expressed by BMSCs, are also contributing to the T cell suppression. Besides, BMSCs reduce expression of CD40 CD86 and MHC class II by microglia. Whereby, ligation of CD40 is crucial for complete activation and maturation of microglia. CD40 is a member of the tumor necrosis factor (TNF) receptor superfamily and its interactions with the ligand leading to exacerbation of multiple sclerosis [10,11], the fully activated microglia present antigen and stimulates T cells. As a result, the scenario of NO secretion by BMSCs limits T cell proliferation and CD40- down regulation of microglia not achieving full fledged activation, and appears ideal to manage inflammation within the CNS [4]. Accordingly, in the presence of BMSCs, LPS-induced proliferation of microglia is reduced, while being potent producers of NO when stimulated by soluble factors released by LPS-activated microglia.
More recently, BMSCs have also been found to primarily inhibit T cell proliferation but not its effector function. T cell responses consist of sequenced phases commencing from cellular activation, proliferation and effector functions. Cellular proliferation is being a vital phase whereby activated T cells are amplified into sufficient copies of antigen-specific T cells in order to perform effector function as cytotoxic cells. BMSCs show the ability to inhibit mitogen-activated T cell proliferation in a dose-dependent manner. However, BMSCs equally inhibit CD4+ and CD8+ subpopulations of T cells [11,12]. For this reason, they have the potential to be exploited in the control of unwanted immune responses such as graft versus host disease (GVHD) and autoimmunity. Data shows that BMSCs suppress T cell responses to polyclonal stimuli in a dose-dependent fashion and does not appear to be antigen-specific, whom target both primary and secondary responses [13,14].
Despite normal expression of MHC class I, studies implicate that BMSCs are unable to stimulate resting allogeneic T cells proliferation and IFN-γ production. Suggestively, the deficit may be related to the fact that MSCs do not express co-stimulatory molecules CD80 and CD86 which are crucial for providing the necessary second signal for T cell activation [12].
For the immunosuppressive effect, the latest studies suggest the importance of cell contact dependency, as well as soluble factors [15]. Depending on the immunologic environment, BMSCs inhibit or promote T cell proliferation, for example IFN-γ a key cytokine in “MSC-licensing”, on its own has no effect MSC-mediated suppression of T-cell proliferation [16], while in presence of IL-2, or TNF-α enhance BMSC-mediated suppression. The presence of IFN-γ, IL- 2, or TNF-α enhance BMSC-mediated suppression, although these cytokines promote T-cell proliferation according to their normal physiologic role. Thus, the suppression of T-cell proliferation is enhanced in a proinflammatory milieu, but attenuated or inhibited by the presence or absence of anti-inflammatory factors or inflammatory agents, respectively. In contrast, the presence of anti-inflammatory cytokines like IL-10 augments T-cell proliferation. However, cytokine IL-6 and IL-4 do not show suppression of BMSC-mediated T-cell proliferation. TNF-α and IL-2 can boost the inhibitory potential of BMSCs “licensed” by IFN-γ plus either IL-1α, or IL-1β [9,17,18]. In summary, the bimodal nature of BMSC function (inhibitory and stimulatory) has been subjected to pro inflammatory condition and depending on the degree of T-cell pre activation. BMSCs are able to switch from an “immunosuppressive MSC” to an “immunogeneic MSC”, depending on the frequency of immune stimulators.
BMSC antioxidative function by down/up regulation of oxidative stress
Recently, cell therapy has been demonstrated as a promising strategy to treat several oxidative stress-caused degenerative diseases. Neuron pretreatment with MSCs notably improves cell survival, prevents oxidative-associated apoptosis and abolishes the robust deterioration in oxidative status. Brain tissue is particularly vulnerable to oxidative damage, possibly due to its high consumption of oxygen, relatively low levels of antioxidant enzymes, high levels of free iron, and the consequent generation of high quantities of ROS [19,20]. Besides, physiological antioxidant defense mechanisms such as SOD (an antioxidant enzyme) and GSH (a cellular non-enzymatic antioxidant molecule) play an important role in protecting against free radicals by reducing ROS level. There is a large body of evidence indicating that oxidative stress plays a pivotal role in producing a large number of ROS and disrupt the antioxidant defense system the pathogenesis of oxidative-associated brain injury [20,21]. The chronic exposure to ROS results in an augmentation of lipid peroxidation in cortical neurons in the brain. It is well known that chronic exposure to high quantities of reactive oxygen species (ROS), results in alterations of DNA, lipids and proteins; and down regulation of endogenous antioxidants and subsequently cell apoptosis [20-22]. So far, the protective mechanism of BMSCs has been elucidated to be secretion of trophic factors, such as NGF, BDNF and bFGF. Also, the molecular signaling mechanism of BMSCs protection has been addressed to the PI3K/Akt pathway serving a pro-survival function in neurons exposed to various apoptosis-inducing stimuli. PI3K and its downstream effectors Akt is a well-known signaling pathway involved in cell protection under various stresses, including oxidative stress [23,24]. In contrast, ERK1/2 a family of MAPKs can be activated in response to various stress stimuli and has been proved to be involved in the neuronal apoptosis of degenerative diseases. As revealed, the inhibitors of ERK1/2 pathway, can rescue cell damage from various neurotoxins [25-27]. Accordingly, BMSCs by secreting several trophic factors including NGF, bFGF and BDNF, can restore the activities of the free radical scavenging enzymes, SOD, catalase, and glutathione peroxidase, which enormously protect cortical neurons against the deleterious effects [23,24,28]. Since ROS regulate cellular events, neuron chronic oxidative exposure leads to inactivation of PI3K/Akt and lasting activation of ERK1/2. Sustained activation of ERK1/2 phosphorylation contributes to neuronal cell death. Besides, inhibition of ERK1/2 signaling comprises activation of antioxidants such as copper/zinc SOD, catalase and SOD mimetics [19,29].
Recent findings indicate ERK1/2 as double-edged sward leading to apoptosis or cell survival at multiple points that include increasing p53 and BAX activity, phosphorylation of antiapoptotic protein Bcl- 2, increasing the Bax/Bcl-2 ratio, increasing caspase-3 and caspase-8 activities and increasing TNF-α production [26]. In spite of some discrepancies that can be related to the different treatments and conditions or the developmental state of the neurons, there is lots of evidence that transplanted BMSCs via protective factors, promote recovery function.

Switching Gene Expression and Cell Proliferation upon Residency

BMSCs down regulate collagen synthesis and fibroblast proliferation
MSC transplantation has been shown to decrease fibrosis in impaired organs such as lung, heart, liver and kidney through reparative activity. BMSCs exert anti-fibrotic effects at least in part through regulation of fibroblast proliferation and collagen synthesis [30-32].
Fibroblasts are predominantly involved in the maintenance of extracellular matrix by proliferation and collagen synthesis/ degradation. Collagen synthesis is regulated by fibrogenic factors, and its degradation is mediated by members of MMPs, which in turn are down regulated by metalloproteinase tissue inhibitors. Types I and III collagen are the major fibrillar collagen produced by fibroblasts. The expression of collagen genes is regulated at the transcriptional and post-transcriptional levels. After tissue impairment and at the fibrotic phase, an initial mesh of type III collagen forms the scaffold for subsequent deposition of large, highly aligned type I collagen fibers [33-35].
MSCs inactivate transcription of collagen types I and III, through down regulation of the expression of Col1a1 and Col3a1 in fibroblasts. The in vivo studies have demonstrated BMSC transplantation in rat models of myocardial infarction inhibited deposition of types I and III collagen [36]. Consequently, BMSCs have equivalent paracrine effects to attenuate excessive collagenase activity of activated fibroblasts. Collagenase (MMP-1) and gelatinase (MMP-2 and -9) activity are known to be elevated during the necrotic phase of infarct healing, and are involved in disruption of the collagen network [36,37] .
BMSCs induce anti-fibrotic effects through up regulation of Catna1 and Rarb, negatively regulator of cell proliferation, and down regulation of A2m and kit, positively regulator of cell proliferation. Catna1 encodes alpha-catenin which interacts with cadherin, a cell adhesion molecule, and targeted deletion of Catna1 in either the skin or in neuronal progenitor cells leads to hyper proliferation [38]. Rarb encodes a member of retinoic acid receptors, and regulate cell growth and differentiation in a variety of cells [39]. A2m encodes a plasma proteinase inhibitor and induces macrophage proliferation through cAMP-dependent signaling [40]. Kit encodes c-kit protein, a tyrosine kinase receptor for stem cell factor, and ectopic expression of c-kit in fibroblasts induces tumorigenesis [41].
Besides, three other negative regulators of cell proliferation have been reported to be also up regulated by paracrine action of BMSCs. Eln encodes a polymer of a precursor protein (tropoelastin), and impaired elastogenesis coincides with increased cell proliferation [42]. Mycd encodes a transcription factor important for smooth muscle and cardiac muscle development, and up regulation of Mycd in fibroblasts decreases their proliferative potential [43]. Ddit3 belongs to the CCAAT/enhancer binding protein family of transcription factors and its expression is capable of inducing growth arrest and apoptosis [44]. Whereas, fibroblasts have been reported to up regulate proliferation-related genes such as alpha- 2-macrogrobulin and v-kit, sarcoma viral oncogene homolog [36].
Taking together, BMSCs are able to mediate pleiotropic effects; negatively regulating fibroblast proliferation and exert anti-fibrotic effects, while capable to secret a large number of growth factors, anti-apoptotic factors and cytokines. The features of BMSCs can be beneficial for the treatment of impaired organs in which fibrotic changes are involved.
Wnt signaling as a double edged-sword by BMSCs to switch of tumor cells
BMSCs are recruited to tumor site and inhibits growth of tumor cells such as brain gliomas and Kaposis sarcoma 149 [6]. BMSCs can home to site of tumorogenesis and via direct cell–cell interactions potently inhibit tumor growth by down-regulation of the protein kinases such as Akt 149. Also via depression of Wnt- Dkk-1 signaling, BMSCs inhibit progression of tumor cells such as those in breast cancer [45,46]. Stem cells and tumor cells share many similarity and characteristics. In particular, they have similar active signaling pathways that regulate self-renewal and differentiation, such as the Wnt, Notch, Shh, and BMP pathways [47]. The canonical Wnt signaling pathway includes stabilization of cytosolic b-catenin, nuclear translocation, and gene regulation, whereby acts as a coactivator of T-cell factor (TCF) proteins. The Wnt ligands activate this canonical pathway by binding to the frizzled receptors and coreceptors, LRP5/6. Members of the proto-oncogene Wnt family can promote cellular proliferation and lead to tumor formation [48,49]. Genes regulated by Wnt signaling are involved in metabolism, proliferation, cell cycle, and apoptosis [50]. Wnt3a, for example, has been reported to increase expression of the anti-apoptotic protein Bcl-2 and PCNA. Initiation of Wnt signaling is modulated by soluble Wnt antagonists (sWAs), including soluble frizzled related proteins (sFRP), dickkopf proteins (Dkk) and Wnt inhibitory factor-1 (Wif1) [46,51]. The antagonist, Dkk-1, competes with Wnt for binding to LRP5/6, thus inhibits activation of the Wnt signaling pathway. Dkk-1 is a putative tumor suppressor and its deficiency implicates in tumor progression, while stimulating the proliferation and maintaining the undifferentiated phenotype of BMSCs [51,52]. Data analysis has shown that the expression level of Wnt inhibitor Dkk-1 in BMSCs is higher than that in cancerous cells, which may play a vital role in controlling Wnt signaling in cancer cells. The molecular mechanism of the inhibitory effect of BMSCs on the growth of human cancer cells can be related to the Dkk-1, a soluble inhibitor of Wnt signaling [46,53]. Data has also shown that b-catenin is down-regulated in cancerous cells by conditioned media from MSCs, and the expression level of Dkk-1 is higher in MSCs than that in cancerous cells. Neutralization of Dkk-1 and small interference RNA targeting Dkk-1 mRNA in MSCs attenuates the inhibitory effect of MSCs on cancer cells [45,46].

Signaling Cascades Participating in BMSC Life Span and Senescence

Interactions directly/ indirectly contributing to engineer BMSCs
The data showing that BMSCs fate are specifically influenced by direct contact with distinct differentiated cell types. In recovery of blood vessels and reparation of an impaired organ, endothelial cells (ECs) have been observed to display important implications for BMSC recruitment and differentiation. In direct cell interactions with ECs, BMSCs exhibit a smooth muscle cell- like synthetic phenotype, concurrently result in an increased smooth muscle a-actin expression, while soluble factors alone not cabling to induce such an effect [54,55].
Cell contacts between BMSCs and fibroblasts stimulate the appearance of myofibroblast-like cells with well-organized SM a-actin filaments in fibroblasts, indicating BMSCs induce fibroblast toward myofibroblast phenotype. It is tempting to speculate that BMSCs influence differentiation of adventitial fibroblasts to smooth muscle-like myofibroblasts causing vascular development and repair. Fibroblasts in turn, lead to a significant down regulation of the late SMC differentiation marker smoothelin-B and the MSC surface marker CD105 in BMSCs, indicating a modulation of the stem cell phenotype [54].
In other side, apoptosis of implanted BMSCs and poor survival weaken the transplantation and limits the efficiency of stem cell therapy [55]. Accordingly, H2O2 and serum deprivation induces apoptosis of BMSCs, while Lipopolysaccharide (LPS) preconditioning protects them from H2O2/SD-induced apoptosis and promoting their proliferation [56]. The ligand of Toll-like receptors (TLRs) can control the function of BMSCs and keep the survival capacity of them [57]. TLRs were thought to be expressed mainly on antigenpresenting cells, such as macrophages or dendritic cells, and their ligands can activate these cells to provoke innate immunity, as well as to establish adaptive immunity [56]. However, recent studies have demonstrated the presence and signaling function of TLRs in BMSCs [57]. LPS, an agonist of TLR4, in addition to myocytes and human dendritic cells, protects BMSCs from apoptosis through a PI3K/Akt-dependent pathways and NF-kB-dependent mechanism. Accordingly, via TLR-4, LPS can enhance phosphorylation of both Akt at Ser 473 and nuclear factor-kappa B (NF-kB) p65 at Ser 536, then provoke proliferation of BMSCs [56-58]. Hence, appropriate pre-treatments with LPS can protect BMSCs from stress-induced apoptosis and improve the survival of BMSCs via the TLR4 and PI3K/ Akt pathway.
Fully activated JAK-STAT pathway, essential for BMSC proliferation
Data also show that, whilst direct cell contact affects BMSC phenotype, ECs recruit BMSCs by growth factors such as PDGF-BB and TGF-β1 [55,59]. Here, we try to elucidate the findings serve to describe signaling events play pivotal roles in BMSC proliferation and differentiation. Accordingly, platelet-derived growth factor- BB (PDGF-BB) and basic fibroblast growth factor (bFGF), but not epithelial growth factor (EGF), induce BMSC proliferation through the Janus-activated kinase signal transducers and activators of transcription (JAK-STAT) cascade and the extracellular signalregulated kinase 1/2 (ERK1/2) pathway, respectively. Importantly, PDGF-BB and bFGF exhibited their proliferative effect on BMSCs without affecting their differentiating potential. Notably, both pathways are essential for the progression of BMSC from G1 phase to S phase, in response to the corresponding growth factors [59,60].
Following phosphorylation and activation by MEK, ERK1/2 is translocated into the nucleus where they activate various transcription factors. Similarly, the JAK-STAT pathway is activated by PDGFBB, bFGF and EGF in a variety of differentiated cells [61,62].
In BMSCs, ERK1/2 is mainly linked with TGF-β-induced proliferation and IGF-induced osteogenic differentiation and expansion, while PDGF-BB inducing proliferation via complete activation of the JAK-STAT pathway, including signal transducer and activator of transcription-3 (STAT-3) tyrosine and serine phosphorylation as well as JAK-2 tyrosine phosphorylation [60,62]. Data shows that PDGF-BB treatment leads to significant phosphorylation of STAT3 on both tyrosine and serine residues. Such dual phosphorylation is crucial for the complete activation of STAT3. The tyrosine phosphorylation is essential for dimerization and activation, while the serine phosphorylation required for maximal transcriptional activity of the activated STAT dimmer [59,60].
STAT proteins are a group of latent cytoplasmic transcription factors that are activated by tyrosine phosphorylation, usually mediated by members of the JAK tyrosine kinase family. STAT tyrosine phosphorylation promotes the formation of STAT dimers, followed by nuclear translocation of the activated dimmers. In the nucleus, the STAT dimers promote the transcription of several target genes, often associated with cell proliferation. Similarly, Serine phosphorylation of STAT proteins, is mediated by the MAPK family, and leads to the STAT-mediated transcriptional activity [60,62].
In response to PDGF-BB, both JAK-2 is required for STAT- 3 tyrosine phosphorylation, and the ERK1/2 pathway mediates STAT-3 serine phosphorylation. Whereas, bFGF stimulates BMSC proliferation via the ERK1/2 pathway without involvement the JAKSTAT cascade [60].
Nonetheless, a wide variety of GFs such as PDGF-BB, bFGF and EGF activate the BMSC ERK1/2 signaling pathway, while only PDGF-BB leads to a complete activation of the JAK-STAT pathway by directly and fully activating STAT3 tyrosine and serine phosphorylation, respectively by JAK2 and ERK1/2. PDGF-BB leads to BMSC proliferation via a complete activation of JAK-STAT pathway, but not by the ERK1/2 pathway on its own, while bFGFinduced ERK1/2 activation raising the possibility that the pathway is mainly responsible for other physiological processes rather proliferation of BMSCs [60,63]. Furthermore, some cytokines such as CXCL12 and CCL5 have also been reported to activate STAT isoforms in BMSCs [64]. Given the involvement of the JAK-STAT pathway in BMSC proliferation, it is reasonable to assume that the genes activated by STAT3 in response to PDGF-BB, are proliferationrelated and involved in the G1/S transition of the cells, as has been indicated by the cell cycle analysis elsewhere [60].

Strategies and In vitro Mechanisms for Efficient Transplantation

A preconditioning hypoxia helps stem cells to escape from senescence
Regarding the stem cell transplantation, significant number of transplanted cells undergoes cell death, which could hamper the potential benefits. Under in vivo hypoxic conditions, MSC may undergo significant stress which leads to cell death. A number of studies have addressed the effect of hypoxia on the growth and differentiation of human stem cells. It was reported that under normoxic conditions (20% pO2), MSCs cease to proliferate after 15–25 population doublings, while MSCs cultured under hypoxic conditions (1% pO2) retaining their ability to proliferate with an additional 8–20 population doublings [65]. Notable, the discrepancies existing are due to differences in species, the concentration of oxygen, and the length of culture periods. The discussion hereafter focuses on the data for human MSCs. As short-term effect, the hypoxia (2% pO2) increase the proliferation rate of MSCs by promoting progression of the cell cycle, or display no effect. But under the longer term effects of hypoxia, human MSCs display enhanced proliferation for seven passages over 6 weeks and a 30- fold increase in the cell number compared to normoxic condition. Another report shows that MSCs cultured under hypoxic condition (3% pO2) for up to 100 days, had 10- fold final population doublings (a high number), than those cultured under normoxic conditions [65,66].
The hypoxia preconditioning (HPC) preserve the cell survival by preparing BMSCs to confront the fallowing hypoxic challenge and prevent the deleterious effects [66]. Intriguingly, preconditioning of BMSCs with short periods of hypoxia/re-oxygenation, prior to the in vivo hypoxic challenge, led to a decrease in pro-oxidant enzymes, as well as an increase in anti-oxidant enzymes, what allows the cell to metabolize ROS (such as H2O2) into H2O and O2 [67]. Depending on the in vivo physiological state, an oxidative state can be characterized by either an increase in pro oxidants or a decrease in anti-oxidants, or a combination of both acting in concert. The HPC of stem cells also increases expression of pro-survival genes such as Akt and is associated with better stem cell survival [65]. The most the beneficial effect on cell maintenance is due to increased expression of “survival genes” like Akt, and bcl-2. Importantly, in vivo prolonged hypoxia condition causes an increase in the cell mobility to keep MSCs survive in a noxious environment [65,67]. Presumably, the process may cause BMSCs to leave the injury site and be washed away from the primary homing site. Then in the case of prolonged hypoxia condition, HPCBMSCs do not need to be stimulated and switched on prolonged hypoxia process. Generally, data exhibit that hypoxia dose extend life span than growth rate of the cells, being due to escape from cellular senescence.
Underlying mechanism of the hypoxia cause to expansion of stem cell life span
As indicated, stem cells when transplanted in vivo, undergo temporary exposure to hypoxia (≤1% O2). In an experiment, up to 14 days post temporary exposure to hypoxia (≤1% pO2) resulted in prevention of osteogenic differentiation but had no effect on MSC survival. Under hypoxia, down-regulation of cbfa-1/Runx2 (osteoblastic transcription factor), osteocalcin (osteogenic marker) and type I collagen (the main component of bone matrix) has been observed [68]. The ERK signaling is essential for the MSC differentiation, for example activation of the ERK pathway triggers osteogenic or adipogenic differentiation [69]. Therefore, inhibition of the ERK pathway can restrict the spontaneous differentiation and maintain BMSCs in an undifferentiated state. Although the complete molecular mechanisms underlying life expanded behavior of stem cells, are not yet known but reduction in oxygen cause downregulation of ERK, which help cells to escape from senescence.
Under the short-term hypoxia (24 h, 1% pO2), only chondrogenic differentiation is to be influenced by inducing of MAPK signals through up-regulation the expression of the Sox9 gene, transcription factor essential for chondrogenesis [65,70].
Intriguingly, temporary hypoxia lead to a 2-fold increase in angiogenic factor expression VEGF, but other growth factors and cytokines such as bFGF, TGFβ1 and IL-8 has been not affected [68].
Although, the ERK signaling promotes cell growth, but at the same time induces cellular senescence of MSCs. Further, hypoxia down-regulates the p16 gene expression (Cyclin-dependent kinase inhibitor 2A). The up-regulation of p16 gene expression has shown as a key player to induce human MSC senescence [69,71]. The late senescence of MSCs in hypoxic condition is related to downregulation of p16 gene. ROS induces p16 gene expression, and the p16 pathway in a positive loop increases the production of ROS, which in turn enhances cellular senescence. Inhibition of the ERK signal reduces the up-regulation of p16 gene expression. The temporary hypoxia induces down regulation of p38/MAPK and up-regulation of PTEN phosphatase, the inhibitor of Akt kinase [65,67,71].
However, acute (<1% pO2) prolonged hypoxia triggere a significant increase in the expression of p67phox and p47phox, subunits of pro-oxidant NAD(P)H oxidase, leading to an increase in ROS, associated with a decrease in cell survival [71]. Under acute prolonged hypoxic condition, there is a decrease in the expression of the anti-oxidant enzyme catalase, a major metabolizer of ROS, while the levels of SOD and its subunits can be unaltered by the hypoxic challenge. A decrease in the endogenous scavenger enzyme catalase will not allow for the metabolism of H2O2, leading to further intracellular accumulation of H2O2, with deleterious effects to the cell [65,67].
Generally, MSCs escape from senescence via down regulation of p38-MAPK-ERK pathway, down regulation of p16 gene expression, as well as down-regulation pro-oxidant NAD(P)H oxidase genes. Accordingly, different degrees of hypoxia and age of MSCs (senescence) have differential effects on the activation of the pathways.


Bone marrow stem cells (BMSCs) have been introduced as the most promising candidates for cellular transplantation. The regulatory effects of BMSCs on immune cell behavior offer insight into the potential moderating properties of BMSCs on inflammatory responses, particularly for immunotherapy in solid organ transplantation. More recently, BMSCs have also been found to primarily inhibit T cell proliferation but not its effector function. The paracrine effects of BMSCs on fibroblasts, imply their potential to attenuate fibrosis and impaired tissue remodeling at least in part through the down regulation of several genes involved in fibroblastic cell proliferation, inhibit type I and III collagen gene expression, and exhibit comparable collagenase activity. Cell contacts between BMSCs and fibroblasts stimulate the appearance of myofibroblast-like cells with well-organized SM α-actin filaments in fibroblasts, indicating BMSCs switch the fibroblast gene expression toward myofibroblast phenotype to reconstitute the organ. Together, we try to highlights the molecular mechanism and the in vivo efficiency of BMSCs. Also, understanding the mechanisms involved in BMSC beginning of senescence, is an important prerequisite to the design of future cellbased therapies with efficient out comes. This information is crucial for achieving a better control over the human BMSCs expansion and transplantation processes.


We would like to express special thanks to Yazd Shahid Sadouqi University of Medical Sciences for their support in stem cell research.
Disclosure statement for authors: On behalf of the authors, I (Fatemeh Pourrajab) should ensure that there are no potential conflicts of interest to disclose.


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