Transplantation Of Marrow Stromal Cells Research Articles

Endochondral Ossification Induced by Cell Transplantation of Endothelial Cells and Bone Marrow Stromal Cells with Copolymer Scaffold Using a Rat Calvarial Defect Model.

It has been recently reported that, in a rat calvarial defect model, adding endothelial cells (ECs) to a culture of bone marrow stromal cells (BMSCs) significantly enhanced bone formation. The aim of this study is to further investigate the ossification process of newly formed osteoid and host response to the poly(L-lactide-co-1,5-dioxepan-2-one) [poly(LLA-co-DXO)] scaffolds based on previous research. Several different histological methods and a PCR Array were applied to evaluate newly formed osteoid after 8 weeks after implantation. Histological results showed osteoid formed in rat calvarial defects and endochondral ossification-related genes, such as dentin matrix acidic phosphoprotein 1 (Dmp1) and collagen type II, and alpha 1 (Col2a1) exhibited greater expression in the CO (implantation with BMSC/EC/Scaffold constructs) than the BMSC group (implantation with BMSC/Scaffold constructs) as demonstrated by PCR Array. It was important to notice that cartilage-like tissue formed in the pores of the copolymer scaffolds. In addition, multinucleated giant cells (MNGCs) were observed surrounding the scaffold fragments. It was concluded that the mechanism of ossification might be an endochondral ossification process when the copolymer scaffolds loaded with co-cultured ECs/BMSCs were implanted into rat calvarial defects. MNGCs were induced by the poly(LLA-co-DXO) scaffolds after implantation, and more specific in vivo studies are needed to gain a better understanding of host response to copolymer scaffolds.

Bone marrow stromal cells-derived exosomes target DAB2IP to induce microglial cell autophagy, a new strategy for neural stem cell transplantation in brain injury.

Bone marrow stromal cells (MSCs) are a useful source of stem cells for the treatment of various brain injury diseases due to their abundant supply and fewer ethical problems compared with transplant treatment. However, the clinical application of MSCs is limited due to allograft rejection and immunosuppression in the process of MSCs transplantation. According to previous studies, microglial cell autophagy occurs following co-culture with MSCs. In the present study, exosomes were obtained from MSCs and subsequently characterized using transmission electron microscopy, atomic force microscopy and dynamic light scattering particle size analysis. The type of microRNAs (miRs) found in the exosomes was then analyzed via gene chip. The results demonstrated that microglial cell autophagy could be induced by exosomes. This mechanism was therefore investigated further via reverse transcription-quantitative PCR, western blotting and luciferase assays. These results demonstrated that exosomes from MSCs could induce microglial cell autophagy through the miR-32-mediated regulation of disabled homolog 2-interacting protein, thus providing a theoretical basis for the clinical application of miRs in MSCs.

Bone Marrow Stromal Cells Alleviate Secondary Damage in the Substantia Nigra After Focal Cerebral Infarction in Rats.

Transplantation of bone marrow stromal cells (BMSCs) is a promising therapy for ischemic stroke. Previously, we had reported that the secondary degeneration occurred in the ipsilateral substantia nigra (SN) after permanent distal branch of middle cerebral artery occlusion (dMCAO) in Sprague-Dawley rats. However, whether BMSCs have neurorestorative effects on the secondary damage in the SN after focal cerebral infarction has not known. In this study, rats were subjected to dMCAO followed by intravenous administration of BMSCs 1 day later. We found that transplanted BMSCs survived and migrated to cortical infarct areas and ipsilateral SN. Furthermore, BMSCs promoted neurogenesis through proliferation and differentiation in the SN after dMCAO. Rats implanted with BMSCs showed significant improvement in their performance of modified neurological severity scores and adhesive-removal test. Engrafted BMSCs enhanced survival of dopaminergic neuron, reduced gliosis in the ipsilateral SN, and increased contents of dopamine (DA) and its metabolites in the ipsilateral striatum after dMCAO. With pseudorabies virus-152 as a retrograde tracer, we also demonstrated that BMSCs could effectively enhance the cortico-striatum-nigral connections. These results suggest that BMSCs transplantation exerts neurorestorative effects after cortical infarction through promoting endogenous neurogenesis, increasing contents of DA and its metabolites, alleviating the secondary neuronal damage in the SN, enhancing the cortico-striatum-nigral projections pathway, and finally improving the neurological functional outcome.

A Study of Avascular Necrosis of the Femoral Head Using Danshen Combined with Marrow Stromal Cells In Vitro

As avascular necrosis of the femoral head (ANFH) has been one of the major problem plaguing the medical profession and Marrow Stromal Cells (MSCs) in orthopedic exploration for recent years, the aim of this study was to investigate the possible chemotactic effect in vitro treated by Danshen combined with transplantation of MSCs in ANFH. The MSCs were extracted from rabbit tibia and femur. The inhibition rate of Danshen in MSCs proliferation was assayed by MTT. Chemokine receptor expression was determined by RT-PCR. Chemotaxis migration was assayed by Boyden chamber. Results showed that high concentration of Danshen injection on the proliferation of MSCs have a certain inhibition rate. Less than 50 μl/ml did not substantially affect the proliferation of MSCs. After treatment with Danshen injection, it can promote chemokine receptors CCR2, CX3CR1 and CXCR4 expression in MSCs. Danshen could significantly promote cell migration after mixed with 50 ng/ml recombinant monocyte chemoattratant protein-1 (rMCP-1) and recombinant stromal cellderived factor-1 (rSDF-1). In conclusion, culture of MSCs can express chemokine receptors CCR2, CX3CR1 and CXCR4 in vitro. Danshen injection can increase the expression of chemokine receptor CCR2, CX3CR1, and CXCR4, which is beneficial to the migration of MSCs. In conclusion, the chemotactic effects of rMCP-1 and rSDF-1 on the chemotaxis of MSCs were positively correlated with the concentrations of rMCP-1 and rSDF-1 and significantly increased combined with Danshen.

Treadmill exercise enhances therapeutic potency of transplanted bone mesenchymal stem cells in cerebral ischemic rats via anti-apoptotic effects.

BackgroundThe transplantation of bone marrow stromal cells (MSCs) has proved to ameliorate ischemic brain injury in animals, but most transplanted MSCs undergo apoptosis in the ischemic penumbra, greatly compromising the therapeutic value of this treatment. Meanwhile, cell apoptosis can be inhibited by post-ischemia exercise which has been demonstrated to improve the expression of related anti-apoptotic proteins. The present study investigated whether treadmill exercise enhances the neuroprotective effects of transplanted MSCs in a rat experimental stroke model.ResultRats were subjected to 2-h middle cerebral artery occlusion (MCAO). Twenty-four hours after reperfusion, they were assigned randomly to receive no MSCs treatment and no exercise (control group), intravenous transplantation of MSCs and treadmill exercise (MSCs + Ex group), MSCs transplantation only (MSCs group) and treadmill exercise only (Ex group). Neurological assessment, TUNEL staining and western blot were performed. Compared with the MSCs group and Ex group, the MSCs + Ex group reported markedly improved neurological function, significantly decreased apoptotic cells, and increased expressions of survivin and bcl-2 (p < 0.05 or p < 0.01, respectively). Interestingly, the treadmill exercise significantly inhibited the apoptosis of transplanted MSCs. As a result, the number of engrafted MSCs in the MSCs + Ex group was significantly higher than that in the MSC group (p < 0.01).ConclusionsTreadmill exercise enhances the therapeutic potency of MSCs by improving neurological function and possibly inhibiting the apoptosis of neuron cells and transplanted MSCs. These effects may involve an increased expression of survivin and bcl-2.

骨髄間質細胞移植による脊髄損傷の再生

骨髄間質細胞移植による脊髄損傷の再生

Effect of combining exercise with transplantation of bobe marrow stromal cells on axonal sprouting, prevention of muscle atrophy and functional recovery after spinal cord contusion

Effect of combining exercise with transplantation of bobe marrow stromal cells on axonal sprouting, prevention of muscle atrophy and functional recovery after spinal cord contusion

Effects of transplanting bone marrow stromal cells on axonal and glial scarring after spinal cord injury

Objective To investigate the effects of bone marrow stromal cell (BMSC) transplantation on axonal and glial scarring after spinal cord injury (SCI).Methods Thirty New Zealand white rabbits were randomly assigned to a sham operation group (group A),a saline treatment group (group B) or a BMSC treatment group (group C).Group A served as controls,in which the canal was opened without damage to the spinal cord.In groups C and B SCI models were established with aneurysm clips and the rabbits of groups C and B were then given injections of BMSCs and saline solution respectively via the intra-intercostal artery at 1 week post injury.At 1 day,1 week,2 weeks and 4 weeks post injury,Basso Beattie-Bresnahan (BBB) scores were assessed to evaluate the recovery of locomotor function in the hind limbs.Spinal cord samples were harvested for HE and Nissl staining,and immunohistochemistry and image analysis were used to detect any changes in neurofilament (NF200) and glial fibrillary acidic protein (GFAP) in the injured spinal cords.Results The average BBB scores of group A were significantly higher those that of groups B and C at each time point,and those of group C were significantly better than those of group B at the 2nd and 4th week post injury.At the 4th week post injury,HE staining showed there was no glial scarring or cavities in group A,but that there was glial cellular proliferation,glial scarring and cavity formation at the injury site in groups B and C.In group C all were obviously less than in group B.Nissl staining indicated there were more typical neurons in group A,while there were a larger number of ruptured neurons,more degradation,and irregular remaining neurons in groups B and C.These abnormalities were again significantly more prevalent in group C.Immunohistochemical examination showed significant increases in NF200 positive neurons and GFAP in groups B and C compared with group A.The number of NF200 positive neurons was significantly higher in group C than in group B,but the GFAP positive area was significantly smaller in group C than in group B.Conclusion BMSC transplantation via the intercostal arteries can effectively improve axonal regeneration,attenuate glial cellular proliferation and reduce glial scar formation,promoting functional recovery after SCI,at least in rabbits. Key words: Spinal cord injury; Intercostal arteries; Bone marrow stromal cells; Neurofilaments; Glial fibrillary acidic protein; Rabbits

Spinal motor neurite outgrowth over glial scar inhibitors is enhanced by coculture with bone marrow stromal cells

Background contextTransplantation of bone marrow cells into spinal cord lesions promotes functional recovery in animal models, and recent clinical trials suggest possible recovery also in humans. The mechanisms responsible for these improvements are still unclear. PurposeTo characterize spinal cord motor neurite interactions with human bone marrow stromal cells (MSCs) in an in vitro model of spinal cord injury (SCI). Study design/settingPreviously, we have reported that human MSCs promote the growth of extending sensory neurites from dorsal root ganglia (DRG), in the presence of some of the molecules present in the glial scar, which are attributed with inhibiting axonal regeneration after SCI. We have adapted and optimized this system replacing the DRG with a spinal cord culture to produce a central nervous system (CNS) model, which is more relevant to the SCI situation. MethodsWe have developed and characterized a novel spinal cord culture system. Human MSCs were cocultured with spinal motor neurites in substrate choice assays containing glial scar–associated inhibitors of nerve growth. In separate experiments, MSC-conditioned media were analyzed and added to spinal motor neurites in substrate choice assays. ResultsAs has been reported previously with DRG, substrate-bound neurocan and Nogo-A repelled spinal neuronal adhesion and neurite outgrowth, but these inhibitory effects were abrogated in MSC/spinal cord cocultures. However, unlike DRG, spinal neuronal bodies and neurites showed no inhibition to substrates of myelin-associated glycoprotein. In addition, the MSC secretome contained numerous neurotrophic factors that stimulated spinal neurite outgrowth, but these were not sufficient stimuli to promote spinal neurite extension over inhibitory concentrations of neurocan or Nogo-A. ConclusionsThese findings provide novel insight into how MSC transplantation may promote regeneration and functional recovery in animal models of SCI and in the clinic, especially in the chronic situation in which glial scars (and associated neural inhibitors) are well established. In addition, we have confirmed that this CNS model predominantly comprises motor neurons via immunocytochemical characterization. We hope that this model may be used in future research to test various other potential interventions for spinal injury or disease states.

Transplantation of autologous angiogenin-overexpressing marrow stromal cells in treatment of a porcine chronic ischemia model

Objective To investigate the effect of autologous angiogenin(ANG)-overexpressing marrow stromal cells transplantation on the neoangiogenesis and cardiac function in a porcine model of chronic ischemia.Methods Porcine models of chronic ischemia were established and were randomized into the following 3groups:autologous marrow stromal cells transfected with Ad-ANG(GroupⅠ,n=11),autologous marrow stromal cells without transfection(GroupⅡ,n=10),and controls injected with serum-free DMEM medium(GroupⅢ,n=10).The autologous marrow stromal cells were cultured and were transfected with Ad-ANG,and the cells were labeled by CM-DiI.The cells were transplanted to the ischemia site of porcine heart.The parameters we observed included Rentrop score,ejection fraction,percentage of infracted area,CM-DiI labeled cells under fluorescence microscope,number of blood vessels and expression of ANG protein.Results Compared with groupⅡandⅢ,GroupⅠ had significantly increased Rentrop score,ejection fraction,numbers of blood vessels in the infarcted areas,CM-DiI labeled cells,and ANG protein expression,but significantly decreased infarcted area(P0.05or P0.01).Conclusion Transplantation of autologous angiogenin-overexpressing marrow stromal cells can yield a high survival rate of cells,greatly improving the blood supply of the infarcted cardiac area and cardiac function in porcine model.

발생 중인 포유류 망막으로 골수기질세포의 이식

발생 중인 포유류 망막으로 골수기질세포의 이식

Effect of transplantation of bone marrow stromal cells and treadmill exercise on axonal sprouting, prevention of muscle atrophy, and functional recovery after spinal cord contusion

Effect of transplantation of bone marrow stromal cells and treadmill exercise on axonal sprouting, prevention of muscle atrophy, and functional recovery after spinal cord contusion

Intracerebral Transplantation of Stromal Stem Cells: A Hope for Patients Suffering Neurological Discapacity after Hemorrhagic Stroke

At present, cell therapy with adult stem cells offers hope for treating neurological diseases considered incurable. In this paper, we review our current experience with delayed intracerebral transplantation of bone marrow stromal cells (BMSC) as a potential treatment of established neurological deficits occurring after intracerebral hemorrhage (ICH). The injection of collagenase IV into basal ganglia of adult Wistar rats causes intracerebral hemorrhage and subsequent neurological deficits that can be quantified by means of functional assessment tests. When the neurological deficit is stabilized, the intracerebral administration of BMSC is followed by a clear functional recovery of the treated animals compared to controls. Furthermore, histological studies show that transplanted stem cells can survive long-term in the injured brain, acquiring a phenotype of neurons and glial cells. These findings suggest that stem cell therapy represents an important field of research, with potential clinical application to treatment of established neurological deficits after brain hemorrhage.

Bone marrow stromal cells enhance the angiogenesis in ischaemic cortex after stroke: involvement of notch signalling

Angiogenesis takes place after brain ischaemia, and stroke-induced angiogenesis in ischaemic brain may be associated with improved neurological recovery. Bone MSCs (marrow stromal cells) transplantation can promote this vital angiogenesis in ischaemic zones, but the mechanisms by which MSCs promoting angiogenesis are unclear. The Notch signalling pathway may play an important role in embryonic blood vessels development and tumour angiogenesis, but whether it is also involved in angiogenesis after cerebral ischaemia is uncertain. We therefore investigated the Notch signalling pathway in angiogenesis after stroke. Rats were subjected to MCAo (middle cerebral artery occlusion) and treated intravenously with or without MSCs at 24 h after injury. On day 1, 3 and 7 after treatment with MSCs or PBS, immunofluorescent staining, Western blot and RT-PCR (reverse transcription-PCR) assays were carried out to evaluate angiogenesis, and expression of VEGF (vascular endothelial growth factor) and Notch signals in the ischaemic cortex. Immunofluorescent showed a significant increase in both new microvessels, VEGF-positive cells and Notch1-positive microvessels in the ischaemic cortex in MSCs-treated group. RT-PCR indicated that MSC transplantation significantly raised VEGF mRNA and Hes1 mRNA levels in the ischaemic cortex. The data suggest that treatment with MSCs enhances stroke-induced angiogenesis in ischaemic brain, and that the Notch signalling pathway is involved.

Intracerebroventricular transplanted bone marrow stem cells survive and migrate into the brain of rats with Parkinson's disease.

In this study, 6-hydroxydopamine was stereotaxically injected into the right substantia nigra compact and ventral tegmental area of rats to establish Parkinson's disease models. The rats then received a transplantation of bone marrow stromal cells that were previously isolated, cultured and labeled with 5-bromo-2’-deoxyuridine in vitro. Transplantation of the bone marrow stromal cells significantly decreased apomorphine-induced rotation time and the escape latency in the Morris water maze test as compared with rats with untreated Parkinson's disease. Immunohistochemical staining showed that, 5-bromo-2’-deoxyuridine-immunoreactive cells were present in the lateral ventricular wall and the choroid plexus 1 day after transplantation. These immunoreactive cells migrated to the surrounding areas of the lateral cerebral ventricle along the corpus callosum. The results indicated that bone marrow stromal cells could migrate to tissues surround the cerebral ventricle via the cerebrospinal fluid circulation and fuse with cells in the brain, thus altering the phenotype of cells or forming neuron-like cells or astrocytes capable of expressing neuron-specific proteins. Taken together, the present findings indicate that bone marrow stromal cells transplanted intracerebroventricularly could survive, migrate and significantly improve the rotational behavior and cognitive function of rats with experimentally induced Parkinson's disease.

骨髄間質細胞を用いた犬の脊髄損傷治療への臨床応用

骨髄間質細胞を用いた犬の脊髄損傷治療への臨床応用

Visualization of the Superparamagnetic Iron Oxide (SPIO)-Labeled Bone Marrow Stromal Cells Using a 3.0-T MRI—a Pilot Study for Clinical Testing of Neurotransplantation

Recent studies have elucidated that transplantation of the bone marrow stromal cells (BMSC) has therapeutic potential for the central nervous system (CNS) disorders. However, no imaging modalities have been established to track the engrafted cells in the CNS in clinical situation. This study aimed to investigate the ability of magnetic resonance imaging (MRI) to visualize the BMSC labeled with superparamagnetic iron oxide (SPIO). The BMSC of mice were labeled with SPIO. Various numbers of the cells were injected into the agar phantom and were visualized using a 3.0-T MR apparatus. The SPIO-labeled cells were injected into the temperature-sensitive gelation polymer (TGP) hydrogel and were cultured for 7days. They were also visualized just after the injection and at 7days postinjection. After a 7-day culture, they were stained with Turnbull blue technique. T2-, T2*-, and susceptibility-weighted imaging could identify minimally 1,000 cells in the agar or TGP hydrogel, although it was difficult to quantify their number on MRI. All of these sequences could track the SPIO-labeled BMSC for at least 7days when injected into the TGP. Turnbull blue staining revealed the survival and proliferation of the SPIO-labeled BMSC in the TGP for 7days. The findings strongly suggest that the SPIO labeling may enable to track minimally 1,000 cells engrafted in the CNS on clinical MR apparatus. These data would be valuable to consider the application of imaging technique into cell transplantation therapy for CNS disorders.

Expression of DeltaNp73 in hippocampus of APP/PS1 transgenic mice following GFP-BMSCs transplantation

Objective: To study the effect of hippocampal bone marrow stromal cells (GFP-BMSCs) transplantation on spatial memory and DeltaNp73 expression in APP/PS1 transgenic mice.Methods: Twelve APP/PS1 transgenic mice randomly received either 10 μl GFP-BMSCs suspension in medium (GFP-BMSCs transplantation group) or 10 μl complete medium (sham-operated group). Learning and memory function of mice in both groups were observed and tested in Morris water maze experiment at 2 weeks after surgery. Senile plaques and DeltaNp73 protein in hippocampuses were determined by immunohistochemistry and western blot at 3 weeks after surgery, respectively.Results: APP/PS1 mice treated with BMSCs performed significantly better on the water maze test than those in sham-operated group (P<0·05). Immunohistochemistry showed that GFP-BMSCs distributed uniformly and the number of Alzheimer’s senile plaques reduced after transplantation. Western blot showed that quantified DeltaNp73 protein expression was significantly higher in BMSCs transplantation group when compared with sham-operated group (P<0·01).Conclusions: Our results suggest that BMSCs transplatation could retard Alzheimer’s disease (AD) like pathology and upregulate DeltaNp73 expression in hippocampuses of APP/PS1 transgenic mice. GFP-BMSCs transplantation will be a potential treatment for AD.

Combination therapy with bone marrow stromal cells and FK506 enhanced amelioration of ischemic brain damage in rats

AimsTransplantation of bone marrow stromal cells (MSCs) has been shown to ameliorate ischemic brain injury in animals. In the present study, we investigated whether the transplantation of MSCs combined with FK506, a clinically used immunosuppressant, enhanced neuroprotective effects in rat experimental stroke. Main methodsMale Sprague–Dawley rats underwent transient 90min middle cerebral artery occlusion (MCAO). Two or 6h after ischemia onset, the rats were randomly assigned to receive intravenous administration of MSCs plus FK506, MSCs alone, FK506 alone, or vehicle. Infarct volume, and neurological and immunohistological assessments were performed to examine the effects of these therapies. Key findingsIn 2-hour post-ischemia treatment groups, significant improvement of infarct volume and neurological scores were observed 1day after combination therapy compared with monotherapy, and this neuroprotection continued for 7days. Combination therapy significantly reduced the number of TUNEL-positive apoptotic cells, increased Bcl-2 expression, decreased Bax expression, and suppressed neutrophil infiltration and microglia/macrophage activation compared to monotherapy. In 6-hour post-ischemia treatment groups, a significant reduction of infarct volume, edema index, and neurological score was observed only in the combination therapy group. Moreover, the number of engrafted MSCs on day 7 with combination therapy was significantly higher than with MSCs alone. SignificanceCombination therapy using FK506 enhanced the anti-apoptotic and anti-inflammatory effects of MSCs and increased the survival of transplanted cells, leading to expansion of the therapeutic time window for MSCs.

Marrow Stromal Cells Differentiate Into Vasculature After Allogeneic Transplantation Into Ischemic Myocardium

Marrow stromal cells (MSCs) are reportedly able to improve ventricular function after myocardial infarction through the paracrine effect or regenerating myocytes. However, the evidence to prove that is scant. In this animal study, we employed MSCs isolated from transgenic pigs designed to express enhanced green fluorescent proteins as the donor to study the fate of the cells after allogeneic transplantation. Green MSCs prepared from transgenic pigs were allogeneically transplanted into chronic ischemic myocardium of 8 Yorkshire pigs by direct intramyocardial injection (total 1.2 × 10(8) cells in 2.5 mL saline, with 25 injection sites). Cohorts of 2 animals were sacrificed at 1, 2, 4, and 6 weeks, and 3 months after injection to study the fate of the injected cells. Allogeneic injection of the green MSCs is safe; no observable side effects or signs of graft versus host disease were observed. By 4',6-diamidino-2-phenylindole (DAPI) counterstained frozen sections, the green cells were found migrating from the injected area into deeper layers of myocardium over the course of 1 to 6 weeks. By immunofluorescent staining, the green cells were associated with smooth muscle actin or von Willebrand factor positive cells, suggesting that the transplanted cells were contributing to the formation of new vessels. We found no evidence that these cells were associated with the new generation of cardiac myocytes. Three months after injection, clusters of MSCs still can be found in the middle layer of ischemic myocardium; however, no unlimited cell growth was found. Allogeneic transplantation of green MSCs can be safely used to elucidate the mechanisms of cell-based therapy. The benefits of this therapy appear mainly due to the angiogenesis, not the regeneration, of cardiac myocytes.