HIF-1α In Mesenchymal Stem Cells Research Articles

Mesenchymal Stem Cell Migration and Proliferation Are Mediated by Hypoxia-Inducible Factor-1α Upstream of Notch and SUMO Pathways.

Mesenchymal stem cells (MSCs) are effective in treating several pathologies. We and others have demonstrated that hypoxia or hypoxia-inducible factor 1 alpha (HIF-1α) stabilization improves several MSC functions, including cell adhesion, migration, and proliferation, thereby increasing their therapeutic potential. To further explore the mechanisms induced by HIF-1α in MSCs, we studied its relationship with Notch signaling and observed that overexpression of HIF-1α in MSCs increased protein levels of the Notch ligands Jagged 1–2 and Delta-like (Dll)1, Dll3, and Dll4 and potentiated Notch signaling only when this pathway was activated. Crosstalk between HIF and Notch resulted in Notch-dependent migration and spreading of MSCs, which was abolished by γ-secretase inhibition. However, the HIF-1-induced increase in MSC proliferation was independent of Notch signaling. The ubiquitin family member, small ubiquitin-like modifier (SUMO), has important functions in many cellular processes and increased SUMO1 protein levels have been reported in hypoxia. To investigate the potential involvement of SUMOylation in HIF/Notch crosstalk, we measured general SUMOylation levels and observed increased SUMOylation in HIF-1-expressing MSCs. Moreover, proliferation and migration of MSCs were reduced in the presence of a SUMOylation inhibitor, and this effect was particularly robust in HIF-MSCs. Immunoprecipitation studies demonstrated SUMOylation of the intracellular domain of Notch1 (N1ICD) in HIF-1-expressing MSCs, which contributed to Notch pathway activation and resulted in increased levels of N1ICD nuclear translocation as assessed by subcellular fractionation. SUMOylation of N1ICD was also observed in HEK293T cells with stabilized HIF-1α expression, suggesting that this is a common mechanism in eukaryotic cells. In summary, we describe, for the first time, SUMOylation of N1ICD, which is potentiated by HIF signaling. These phenomena could be relevant for the therapeutic effects of MSCs in hypoxia or under conditions of HIF stabilization.

Overexpression of HIF-1α in mesenchymal stem cells contributes to repairing hypoxic-ischemic brain damage in rats

Preclinical researches on mesenchymal stem cells (MSCs) transplantation, which is used to treat hypoxic-ischemic (HI) brain damage, have received inspiring achievements. However, the insufficient migration of active cells to damaged tissues has limited their potential therapeutic effects. There are some evidences that hypoxia inducible factor-1 alpha (HIF-1α) promotes the viability and migration of the cells. Here, we aim to investigate whether overexpression of HIF-1α in MSCs could improve the viability and migration capacity of cells, and its therapeutic efficiency on HI brain damage. In the study, MSCs with HIF-1α overexpression was achieved by recombinant lentiviral vector and transplanted to the rats subsequent to HI. Our data indicated that overexpression of HIF-1α promoted the viability and migration of MSCs, HIF-1α overexpressed MSCs also had a stronger therapeutic efficiency on HI brain damaged treatment by mitigating the injury on behavioral and histological changes evoked by HI insults, accompanied with more MSCs migrating to cerebral damaged area. This study demonstrated that HIF-1α overexpression could increase the MSCs’ therapeutic efficiency in HI and the promotion of the cells’ directional migration to cerebral HI area by overexpression may be responsible for it, which showed that transplantation of MSCs with HIF-1α overexpression is an attractive therapeutic option to treat HI-induced brain injury in the future.

Overexpression of HIF-1α in Mesenchymal Stem Cells Contributes to Repairing Hypoxic-Ischemic Brain Damage in Rats

Introduction: Preclinical researches on the use of mesenchymal stem cells (MSCs) transplantation to treat hypoxic-ischemic (HI) brain damage have received some encouraging results. However, the insufficient migration of active cells to damaged tissues has limited their potential therapeutic effects. There is a good evidence that hypoxia inducible factor-1 alpha (HIF-1α) promotes the viability and migration of cells. This study was designed to investigate whether overexpression of HIF-1α in MSCs could be effective to enhance the therapeutic efficiency.Methods: MSCs were obtained from rat femoral and tibial bone marrow, and cells with HIF-1α overexpression were gained by recombinant lentiviral vector. The proliferation and migration of MSCs were assessed by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and transwell culture system. Additionally, the therapeutic efficiency of MSCs after transplantion on HI brain damage in rats were investigated by Morris water maze test and histological examination to analyse the cognitive and pathological changes, and the recruitment in the hippocampus was also observed microscopically.Results: (1) As evidenced by phase contrast and fluorescence microscopy, more than 90% of the cells were GFP-positive when infected with the HIF-1α or GFP lentiviral, and the protein expression of HIF-1α in HIF-1α transduced MSCs was approximately 2-fold that in normal MSCs and GFP transduced MSCs (P<0.01). The results of MTT assay showed that overexpression of HIF-1α promoted the viability and proliferation of MSCs (P<0.01). (2) Meantime, we found MSCs were enhanced the migration both in vitro and in vivo experiments. According to transwell assay, the number of cells migrating to the lower compartment in HIF-1α overexpression group was more than other two groups (P<0.01). In vivo experiments in HI-induced rats, the number of MSCs in the hippocampus increased gradually in a time-dependent manner from day 7 to day 21 after transplantation. And a remarkably increased in the recruitment of HIF-1α transduced MSCs was observed in the hippocampus at every time point compared with GFP transduced MSCs (P<0.01). (3) HIF-1α overexpression also elevated the therapeutic efficiency of MSCs on behavioral and histological aspect in HI-induced brain damaged rats. Morris water maze test suggested that the rats suffered from cognitive dysfunction due to HI treatment. There were a significantly increased of time in the target quadrant in both MSCs transplanted groups compared with HI-vehicle group, but a more increase in the amount of time have been found in HIF-1α overexpressed MSCs group (P<0.01). In contrast with sham group, the pathological changes of the hippocampus in the HI-vehicle group revealed serious injury, and the cells were disordered and cavitation was visible by hematoxylin and eosin staining, but the damages were partly mitigated in HIF-1α overexpressed MSCs transplanted group as compared to other groups.Conclusion: Taken together, these data indicated that overexpression of HIF-1α could improve the therapeutic efficiency of MSCs, and transplantation of MSCs with HIF-1α overexpression may represent an attractive therapeutic option to treat HI brain damage in the future.Footnotes: This work was financially supported by National Natural Science Foundation of China (Gant No. 81270566, Gant No. 81500114). DisclosuresNo relevant conflicts of interest to declare.

Overexpression of Hif-1α in Mesenchymal Stem Cells Affects Cell-Autonomous Angiogenic and Osteogenic Parameters.

Reconstruction of large bone defects still represents a major medical challenge. In recent years tissue engineering has developed techniques based on adult mesenchymal stem cells (MSCs) that could represent an attractive therapeutical option to treat large bone defects in the future. It has been demonstrated in various animal models that ex vivo expanded MSCs are capable of promoting the regeneration of skeletal defects after implantation. However, for the efficient regeneration of bone in tissue engineering applications, a rapid vascularization of implanted grafts is essential to ensure the survival of cells in the early post-implantational phase. A promising strategy to enhance vascularization of MSC-containing implants could consist of overexpression of the angiogenic master transcription factor Hypoxia-inducible factor 1 (Hif-1) in the MSCs in order to induce angiogenesis and support osteogenesis. In the present study, we overexpressed Hif-1α in MSCs by using recombinant adenoviruses and investigated cell-autonomous effects. Overexpression of Hif-1α enhanced proliferation, migration, cell survival and expression of pro-angiogenic genes. Other parameters such as expression of the osteogenic markers BMP-2 and RunX2 were decreased. Hif-1α overexpression had no effect on invasion, senescence and osteogenic differentiation of MSCs. Our experiments revealed multifarious effects of Hif-1α overexpression on cell-autonomous parameters. Therefore, Hif-1α overexpression may represent a therapeutic option to improve cellular functions of MSCs to treat critical sized bone defects.

Over expression of HIF-1α in human mesenchymal stem cells increases their supportive functions for hematopoietic stem cells in an experimental co-culture model

IntroductionBone marrow transplantation is a critical approach for the treatment of many hematological disorders. Success of this approach is dependent on many factors the most important of which is the number of hematopoietic stem cells along with an efficient stroma. Co-transplantation of efficient mesenchymal stem cells can greatly improve the outcome of transplantations. Current researches assign a critical role for hypoxia inducible factor (HIF)-1α in protection of various cells and tissues probably through induction of cytokines. To make this feature applicable to human bone marrow-derived mesenchymal stem cells, we manipulated these cells to over express HIF-1α gene.Materials and methodsFull-length cDNA of human HIF-1α was inserted into human bone marrow mesenchymal stem cells by pcDNA.3.1 non-viral plasmid vector, and the effect of this over expression on production of some hematopoietic growth factors was explored. Moreover, using a co-culture system, the interactive impact of HIF-1α-overexpressed mesenchymal stem cells on hematopoietic stem cells was evaluated.ResultsOver expression of HIF-1α in mesenchymal stem cells in normoxia increased production of one of the most important hematopoietic growth factors, Stem cell factor (also known as Steel factor or c-kit ligand). HIF-1α overexpression had no effect on production of other hematopoietic growth factors. In co-culture of mesenchymal stem cells-HIF-1α with hematopoietic stem cells, enhancement of colony formation and reduced differentiation of hematopoietic stem cells were observed.ConclusionOver expression of HIF-1α in human bone marrow-derived mesenchymal stem cells can augment the production of some hematopoietic growth factors, and we suggest this response of mesenchymal stem cells could help to improve the outcome of bone marrow transplantation.

GW24-e1260 Overexpressing cellular repressor of E1A-stimulated genes promote hypoxia-induced VEGF paracrine via HIF-1 in mesenchymal stem cells

ObjectivesHypoxia inducible factor-1α (HIF-1α) is the key transcription regulator for multiple angiogenic factors, including vascular endothelial growth factor (VEGF). Cellular repressor of E1A-stimulated genes (CREG) has also been identified a...

ASSA13-10-9 Overexpressing Cellular Repressor of E1A-Stimulated Genes Promote Hypoxia-Induced VEGF Paracrine Via HIF-1α in Mesenchymal Stem Cells

BackgroundHypoxia inducible factor-1α (HIF-1α) is the key transcription regulator for multiple angiogenic factors, including vascular endothelial growth factor (VEGF). Cellular repressor of E1A-stimulated genes (CREG) has also been identified a...

HIF-1α Confers Resistance to Induced Stress in Bone Marrow-derived Mesenchymal Stem Cells

The major limiting factor in therapeutic application of mesenchymal stem cells (MSCs) is their high vulnerability during the early days of transplantation. Hence, researchers have been encouraged to find various strategies to make the cells resistant to different stresses before and after transplantation. Overexpression of HIF-1α in MSCs to confer resistance against harmful conditions was the aim of this study. Using an invitro approach, we engineered MSCs to overexpress HIF-1α and then evaluated their viability following exposure to hypoxic and oxidative stresses. The inherent expression of HIF-1α was downregulated by siRNA. Viability and apoptosis of the MSCs were then evaluated invitro following their exposure to hypoxic and oxidative stress conditions. Whereas overexpression of HIF-1α in MSCs was protective against cell death and apoptosis triggered by hypoxic and oxidative stress conditions, its downregulation increased apoptosis and death rate. Our study is the first to demonstrate how human MSCs can be manipulated to gain protection against stresses that potentially limit their clinical application.

Effects of Inflammatory Factors on Mesenchymal Stem Cells and Their Role in the Promotion of Tumor Angiogenesis in Colon Cancer

Mesenchymal stem cells (MSCs), which are modulated by cytokines present in the tumor microenvironment, play an important role in tumor progression. It is well documented that inflammation is an important part of the tumor microenvironment, so we investigated whether stimulation of MSCs by inflammatory cytokines would contribute to their ability to promote tumor growth. We first showed that MSCs could increase C26 colon cancer growth in mice. This growth-promoting effect was further accelerated when the MSCs were pre-stimulated by inflammatory factors IFN-γ and TNF-α. At the same time, we demonstrated that MSCs pre-stimulated by both inflammatory factors could promote tumor angiogenesis in vivo to a greater degree than untreated MSCs or MSCs pre-stimulated by either IFN-γ or TNF-α alone. A hen egg test-chorioallantoic membrane (HET-CAM) assay showed that treatment of MSC-conditioned medium can promote chorioallantoic membrane angiogenesis in vitro, especially treatment with conditioned medium of MSCs pretreated with IFN-γ and TNF-α together. This mechanism of promoting angiogenesis appears to take place via an increase in the expression of vascular endothelial growth factor (VEGF), which itself takes place through an increase in signaling in the hypoxia-inducible factor 1α (HIF-1α)-dependent pathway. Inhibition of HIF-1α in MSCs by siRNA was found to effectively reduce the ability of MSC to affect the growth of colon cancer in vivo in the inflammatory microenviroment. These results indicate that MSCs stimulated by inflammatory cytokines such as IFN-γ and TNF-α in the tumor microenvironment express higher levels of VEGF via the HIF-1α signaling pathway and that these MSCs then enhance tumor angiogenesis, finally leading to colon cancer growth in mice.