Bone Marrow Stromal Cells Treatment Research Articles

Role of Serum Inflammatory Cytokines in Sepsis Rats Following BMSCs Transplantation: Protein Microarray Analysis.

Bone marrow stromal cells (BMSCs) have emerged as a potential therapy for sepsis, yet the underlying mechanisms remain unclear. In this study, we investigated the effects of BMSCs on serum inflammatory cytokines in a rat model of lipopolysaccharide (LPS)-induced sepsis. Sepsis was induced by intravenous injection of LPS, followed by transplantation of BMSCs. We monitored survival rates for 72 h and evaluated organ functions, histopathological changes, and cytokines expression. Sepsis rats showed decreased levels of white blood cells, platelets, lymphocyte ratio, and oxygen partial pressure, along with increased levels of neutrophil ratio, carbon dioxide partial pressure, lactic acid, alanine aminotransferase, and aspartate aminotransferase. Histologically, lung, intestine, and liver tissues exhibited congestion, edema, and infiltration of inflammatory cells. However, after BMSCs treatment, there was improvement in organ functions, histopathological injuries, and survival rates. Protein microarray analysis revealed significant changes in the expression of 12 out of 34 inflammatory cytokines. These findings were confirmed by enzyme-linked immunosorbent assay. Pro-inflammatory factors, such as interleukin-1β (IL-1β), IL-1α, tumor necrosis factor-α (TNF-α), tissue inhibitor of metal protease 1 (TIMP-1), matrix metalloproteinase 8 (MMP-8), Leptin, and L-selectin were upregulated in sepsis, whereas anti-inflammatory and growth factors, including IL-4, β-nerve growth factor (β-NGF), ciliary neurotrophic factor (CNTF), interferon γ (IFN-γ), and Activin A were downregulated. BMSCs transplantation led to a decrease in pro-inflammatory cytokines and an increase in anti-inflammatory and growth factors. We summarized relevant molecular signaling pathways that resulted from cytokines in BMSCs for treating sepsis. Our results illustrated that BMSCs could promote tissue repair and improve organ functions and survival rates in sepsis through modulating cytokine networks.

Bone Marrow Stromal Cells (BMSCs) Inhibit Retinal Ganglion Cell Apoptosis and Alleviate Inflammation Through Up-Regulation of MicroRNA-43 and Brain-Derived Neurotrophic Factor in Glaucoma

Glaucoma is a leading cause of vision loss mainly due to retinal ganglion cells (RGC) loss. MicroRNAs (miRNAs) are highlighted as potential biomarkers in diseases. This study aims to investigate the role of miR-43 and BMSCs in the RGC apoptosis and glaucoma.RGCs were transfected with miR-43 inhibitors and mimics, and then co-cultured with BMSCs. RT-qPCR analysis was conducted to determine miR-43 expression, whilst Western blot, and flow cytometry were carried out to assess the role of miR-43 in apoptosis and inflammation. The interaction between miR-43 and BDNF, a neurotrophic factor, was detected by dual-luciferase reporter gene assay. Overexpression of miR-43 promoted RGC proliferation and decreased apoptosis. Furthermore, miR-43 overexpression diminished the contents of apoptosis- and inflammatory-related factors, and elevated the expression of BDNF. Down-regulation of BDNF exerted similar effect as down-regulation of miR-43, enhancing apoptosis and aggravating inflammation. Importantly, BMSC treatment reversed the in vitro inhibitory effect of si-BDNF on RGC with enhancement of miR-43 expression. Mechanically, miR-43 was indicated to target BDNF in glaucoma. Collectively, miR-43 delivered by BMSCs plays an important role in the inflammatory injury and abnormal apoptosis of RGC by regulating the expression of BDNF. These findings might help development of new treatment for glaucoma and provide a promising biomarker for diagnosis and treatment.

Intranasal Delivery of Bone Marrow Stromal Cells Preconditioned with Fasudil to Treat a Mouse Model of Parkinson's Disease.

ObjectiveStem cell transplantation is a promising strategy with great potential to treat Parkinson’s disease (PD). Nevertheless, improving the cell delivery route and optimising implanted cells are necessary to increase the therapeutic effect. Herein, we investigated whether intranasal delivery of bone marrow stromal cells (BMSCs) has beneficial effects in a PD mouse model and whether the therapeutic potential of BMSCs could be enhanced by preconditioning with fasudil.MethodsA PD mouse model was developed by intraperitoneally administering 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Mice were treated intranasally with phosphate buffered saline (PBS), BMSCs, or BMSCs preconditioned with fasudil. One month later, the effects of BMSC treatment were analysed.ResultsOur study showed that fasudil could accelerate the proliferation of BMSCs and promote brain-derived neurotrophic factor (BDNF) secretion in vitro. Intranasally administered BMSCs were capable of surviving and migrating in the brain. Intranasal delivery of BMSCs preconditioned with fasudil significantly improved motor function and reduced dopaminergic neuron loss in substantia nigra; treatment with BMSCs and PBS resulted in similar outcomes. Preconditioning with fasudil inhibited the activation and aggregation of microglia, suppressed immune response, and reinforced BDNF secretion in MPTP-PD mice significantly more than treatment with BMSCs alone.ConclusionThe present study demonstrates that intranasally administering BMSCs preconditioned with fasudil is a promising cell-based therapy for PD.

Bone marrow stromal cells promote neuromotor functional recovery, via upregulation of neurotrophic factors and synapse proteins following traumatic brain injury in rats.

It has previously been demonstrated that bone marrow stromal cells (BMSCs) exhibit great therapeutic potential in neuronal injuries; however, there is limited understanding of the precise underlying mechanisms that contribute to functional improvement following brain injury. The aim of the present study was to assess the effect of BMSC treatment on traumatic brain injury (TBI) in rats, and investigate if they migrate to injured areas and promote neuromotor functional recovery via upregulation of neurotrophic factors and synaptic proteins. BMSCs were cultured in vitro from female Sprague Dawley (SD) rat bone marrow and were subsequently infused into male adult SD rats via the tail vein, following induction of TBI. The results demonstrated that treatment with BMSCs significantly reduced TBI-induced neuromotor impairment and neuronal loss, as assessed by rota rod testing, western blot analysis, modified neurological severity score and immunohistochemistry. The distribution of transplanted BMSCs was tracked by monitoring the expression of sex determining region Y (SRY) in rats. The number of cells double-positive for SRY/neuronal nuclear antigen or SRY/glial fibrillary acidic protein was increased in the BMSC group, which demonstrated that BMSCs migrated to injured areas and differentiated into neurons and astrocytes, following TBI. Furthermore, administration of BMSCs increased expression of vascular endothelial growth factor and brain derived neurotrophic factor. Protein expression levels of synaptophysin were downregulated following TBI and this was reversed in part by treatment with BMSCs. These findings uncovered some underlying mechanisms of action of BMSCs, and may lead to their potential use as a future effective therapeutic agent for the treatment of TBI.

Bone marrow stromal cells promote neuroplasticity of cerebral ischemic rats via a phosphorylated CRMP2-mediated mechanism

Collapsin response mediator protein 2 (CRMP2), an important protein involved in axonal growth and the maintenance of neuronal membrane integrity, has proved to be altered in nervous system diseases. This study was aimed to investigate the role of CRMP2 in bone marrow stromal cells (BMSCs) treating rats with cerebral ischemia. BMSCs were isolated from shaft of the femurs, tibiae, and humeri and were intra-carotid administrated immediately after middle cerebral artery occlusion (MCAO). Modified Neurological Severity Scores (mNSS) was conducted at 3, 7, 14dpo and the electrophysiologic evaluation was evaluated at 14dpo. Then all rats were sacrificed and brain tissues were harvested for RT-PCR, Western blot and Immunohistochemistry analysis. We found BMSCs treatment significantly improved the neurobehavioral performance impaired by ischemic brain injury, accompanied with the notably increasing levels of Synaptophysin (SYP) and Growth associated protein 43 (GAP43). We also found the protein level of phosphorylated CRMP2 (p-CRMP2) and phosphorylation-mediated protein including Glycogen synthase kinase 3 Beta (GSK3β), Cyclin-dependent kinase 5 (CDK5) were dramatically downregulated in ischemic rats following BMSCs transplant. Furthermore, the GSK3β-mediated factors including neurotrophic and signaling factors were all significantly upregulated in BMSCs-treated group. On the basis of these findings, we suggest that the neuroplasticity effect of BMSCs on cerebral ischemia may be associated with the phosphorylated modulation of CRMP2.

Sodium ferulate and n-butylidenephthalate combined with bone marrow stromal cells (BMSCs) improve the therapeutic effects of angiogenesis and neurogenesis after rat focal cerebral ischemia

BackgroundStudies have indicated that bone marrow stromal cell (BMSC) administration is a promising approach for stroke treatment. For our study, we chose sodium ferulate (SF) and n-butylidenephthalide (BP) combined with BMSC, and observed if the combination treatment possessed more significant effects on angiogenesis and neurogenesis post-stroke.MethodsWe established rat permanent middle cerebral artery occlusion (MCAo) model and evaluated ischemic volumes of MCAo, BMSC, SF + BP, Simvastatin + BMSC and SF + BP + BMSC groups with TTC staining on the 7th day after ischemia. Immunofluorescence staining of vascular endothelial growth factor (VEGF) and brain derived neurotrophic factor (BDNF), as well as immunohistochemistry staining of von Willebrand factor (vWF) and neuronal class III β-tubulin (Tuj1) were performed in ischemic boundary zone (IBZ), furthermore, to understand the mechanism, western blot was used to investigate AKT/mammalian target of rapamycin (mTOR) signal pathway in ischemic cortex. We also tested BMSC derived-VEGF and BDNF expressions by western blot assay in vitro.ResultsSF + BP + BMSC group obviously decreased infarction zone, and elevated the expression of VEGF and the density and perimeter of vWF-vessels as same as Simvastatin + BMSC administration; moreover, its effects on BDNF and Tuj1 expressions were superior to Simvastatin + BMSC treatment in IBZ. Meanwhile, it showed that SF and BP combined with BMSC treatment notably up-regulated AKT/mTOR signal pathway compared with SF + BP group and BMSC alone post-stroke. Western blot results showed that SF and BP treatment could promote BMSCs to synthesize VEGF and BDNF in vitro.ConclusionsWe firstly demonstrate that SF and BP combined with BMSC can significantly improve angiogenesis and neurogenesis in IBZ following stroke. The therapeutic effects are associated with the enhancement of VEGF and BDNF expressions via activation of AKT/mTOR signal pathway. Furthermore, triggering BMSC paracrine function of SF and BP might contribute to amplifying the synergic effects of the combination treatment.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-016-0979-5) contains supplementary material, which is available to authorized users.

Bone marrow stromal cells inhibits HMGB1-mediated inflammation after stroke in type 2 diabetic rats

High-mobility group box 1 (HMGB1), a ligand of receptor for advanced glycation endproducts (RAGE), functions as a proinflammatory factor. It is mainly involved in inflammatory activation and contributes to the initiation and progression of stroke. By using a model of transient middle cerebral artery occlusion (MCAo) in type 2 diabetic rats, we investigated the changes of pro-inflammation mediators, blood–brain barrier (BBB) leakage and functional outcome after stroke. Type 2 diabetic rats did not show an increased lesion volume, but exhibited significantly increased expression of HMGB1 and RAGE, BBB leakage, as well as decreased functional outcome after stroke compared with control rats. Injection of bone marrow stromal cells (BMSCs) into type 2 diabetic rats significantly reduced the expression of HMGB1 and RAGE, attenuated BBB leakage, and improved functional outcome after stroke. BMSCs-treated type 2 diabetic rats inhibited inflammation and improved functional outcome after stroke. Furthermore, in vitro data support the hypothesis that BMSCs-induced reduction of HMGB1 and RAGE in T2DM-MCAo rats contributed to attenuated inflammatory response in the ischemic brain, which may lead to the beneficial effects of BMSCs treatment. Further investigation of BMSCs treatment in type 2 diabetic stroke is warranted.

Cell Treatment for Stroke in Type Two Diabetic Rats Improves Vascular Permeability Measured by MRI.

Treatment of stroke with bone marrow stromal cells (BMSC) significantly enhances brain remodeling and improves neurological function in non-diabetic stroke rats. Diabetes is a major risk factor for stroke and induces neurovascular changes which may impact stroke therapy. Thus, it is necessary to test our hypothesis that the treatment of stroke with BMSC has therapeutic efficacy in the most common form of diabetes, type 2 diabetes mellitus (T2DM). T2DM was induced in adult male Wistar rats by administration of a high fat diet in combination with a single intraperitoneal injection (35mg/kg) of streptozotocin. These rats were then subjected to 2h of middle cerebral artery occlusion (MCAo). T2DM rats received BMSC (5x106, n = 8) or an equal volume of phosphate-buffered saline (PBS) (n = 8) via tail-vein injection at 3 days after MCAo. MRI was performed one day and then weekly for 5 weeks post MCAo for all rats. Compared with vehicle treated control T2DM rats, BMSC treatment of stroke in T2DM rats significantly (p<0.05) decreased blood-brain barrier disruption starting at 1 week post stroke measured using contrast enhanced T1-weighted imaging with gadopentetate, and reduced cerebral hemorrhagic spots starting at 3 weeks post stroke measured using susceptibility weighted imaging, although BMSC treatment did not reduce the ischemic lesion volumes as demarcated by T2 maps. These MRI measurements were consistent with histological data. Thus, BMSC treatment of stroke in T2DM rats initiated at 3 days after stroke significantly reduced ischemic vascular damage, although BMSC treatment did not change infarction volume in T2DM rats, measured by MRI.

PS1/γ-Secretase-Mediated Cadherin Cleavage Induces β-Catenin Nuclear Translocation and Osteogenic Differentiation of Human Bone Marrow Stromal Cells.

Bone marrow stromal cells (BMSCs) are considered a promising tool for bone bioengineering. However, the mechanisms controlling osteoblastic commitment are still unclear. Osteogenic differentiation of BMSCs requires the activation of β-catenin signaling, classically known to be regulated by the canonical Wnt pathway. However, BMSCs treatment with canonical Wnts in vitro does not always result in osteogenic differentiation and evidence indicates that a more complex signaling pathway, involving cadherins, would be required to induce β-catenin signaling in these cells. Here we showed that Wnt3a alone did not induce TCF activation in BMSCs, maintaining the cells at a proliferative state. On the other hand, we verified that, upon BMSCs osteoinduction with dexamethasone, cadherins were cleaved by the PS1/γ-secretase complex at the plasma membrane, and this event was associated with an enhanced β-catenin translocation to the nucleus and signaling. When PS1/γ-secretase activity was inhibited, the osteogenic process was impaired. Altogether, we provide evidence that PS1/γ-secretase-mediated cadherin cleavage has as an important role in controlling β-catenin signaling during the onset of BMSCs osteogenic differentiation, as part of a complex signaling pathway responsible for cell fate decision. A comprehensive map of these pathways might contribute to the development of strategies to improve bone repair.

Simvastatin combined with bone marrow stromal cells treatment activatesastrocytes to ameliorate neurological function after ischemic stroke in rats

Evidence indicates that reactive astrocytes play an important role in neurological functional amelioration after stroke. Bone marrow stromal cell (BMSC) transplantation-promoted astrocyte-derived neurotrophic factors' expressions resulted in decreasing the number of apoptotic neurons and improving neurogenesis and angiogenesis in the ischemic boundary zone. Recent studies found that simvastatin had neuroprotective activity in acute stroke. However, it is still ambiguous as to whether simvastatin combined with BMSCs can further activate astrocytes, which contribute to neurological functional recovery after ischemic stroke. In our present study, we established a middle cerebral artery occlusion model in rats and found that combination treatment of simvastatin and BMSCs could significantly increase neurological functional scores and induce activated astrocytes to enhance stromal cell-derived factor-1 α, vascular endothelial growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor expressions compared with BMSC alone (P < 0.05) by immunofluorescence staining on the 7th day after ischemia. Furthermore, western blot results showed that combination treatment could notably activate the AKT/mammalian target of rapamycin signaling pathway in oxygen-glucose deprivation-induced astrocytes. It was superior to BMSC treatment at the 5-min time point (P < 0.01). Therefore, we think that combination treatment should be a more effective therapy after stroke due to synergic functions of supplementary approaches.

Therapeutic effects of intravenous administration of bone marrow stromal cells on sevoflurane-induced neuronal apoptosis and neuroinflammation in neonatal rats.

BackgroundSevoflurane exposure during the early postnatal period causes neuroinflammation and neuronal apoptosis in rodents. Bone marrow stromal cells (BMSCs) have been shown to protect and repair the damaged central nervous system, for example in ischemic stroke models. In this study, we investigated whether intravenous administration of BMSCs ameliorated neurodegeneration, induced by sevoflurane exposure, in neonatal rats.MethodsSprague-Dawley rat pups (postnatal day 7) were exposed to 2% sevoflurane for 6 h (vehicle group, n = 7). BMSCs were administered 30 min after induction of sevoflurane anesthesia (BMSCs group, n = 7). The pups were exposed to carrier gas only, as a negative control (mock anesthesia group, n = 4). We assessed the therapeutic effects of BMSC treatment by measuring expression of the pro-inflammatory cytokine interleukin-6 (IL-6), and levels of cleaved caspase-3, in brain tissues immediately following sevoflurane anesthesia.ResultsAnalysis of the cleaved caspase-3 bands revealed that levels of activated caspase-3 were elevated in the vehicle group compared with the mock anesthesia group, indicating that a single exposure to sevoflurane at subclinical concentrations can precipitate neuronal apoptosis. BMSC treatment did not suppress apoptosis induced by sevoflurane exposure (compared with the vehicle group). The vehicle group had higher proinflammatory cytokine IL-6 protein levels compared with the mock anesthesia group, indicating that sevoflurane exposure induces IL-6 expression. BMSC treatment suppressed sevoflurane-induced increases in IL-6 expression, indicating that these cells can inhibit the neuroinflammation induced by sevoflurane exposure (vehicle group vs. BMSC group).ConclusionsIntravenous administration of BMSCs reduces neuroinflammation, but does not attenuate apoptosis induced by sevoflurane exposure.

Bone Marrow Stromal Cells Combined with Oxiracetam Influences the Expression of B-cell Lymphoma 2 in Rats with Ischemic Stroke

This study aimed to investigate the combination effects of bone marrow stromal cells (BMSCs) and oxiracetam for ischemic stroke. Forty Sprague Dawley female rats (220 ± 20 g) were subjected to a 2-hour ischemic middle cerebral artery occlusion (MCAO)-24 hours reperfusion model. The rats were randomly divided into 4 groups. Rats from BMSCs group, oxiracetam group, and BMSCs + oxiracetam group accepted injection of BMSCs (3 × 10(6) cells), oxiracetam (800 mg/kg), and BMSCs + oxiracetam, respectively. Rats from control group did not receive any interventions after ischemia reperfusion. The neurologic function was examined by modified neurological severity scores (mNSS). B-cell lymphoma 2 (Bcl-2) expression and apoptosis were detected by immunohistochemistry and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. The mNSS was decreased in all treatment groups and that in BMSCs + oxiracetam group was lower than BMSCs group and oxiracetam group (P < .05). The expression of Bcl-2 was unregulated in all treatment groups (P < .05), and similarly, the expression of Bcl-2 in BMSCs + oxiracetam group was higher than BMSCs group and oxiracetam group (P < .05). Control group displayed more TUNEL-positive cells than the treatment groups, and BMSCs + oxiracetam group displayed less apoptotic cells than BMSCs group or oxiracetam group (P < .05). Transplantation of BMSCs can promote the recovery of neurologic function in MCAO rats, and the effect of BMSCs combined with oxiracetam was better than the either one. Upregulation of Bcl-2 resulting in a decrease of apoptosis may be one of the mechanisms of BMSCs treatment for cerebral ischemic stroke.

Canine Bone Marrow Stromal Cells Promote Functional Recovery in Mice with Spinal Cord Injury

ABSTRACTRegenerative therapy has begun to be clinically applied in humans and dogs to treat neurological disorders, such as spinal cord injury (SCI). Here, we show the therapeutic potential of transplantation of cultured canine bone marrow stromal cells (BMSCs) into mice with SCI. Canine BMSC transplantation therapy was performed, immediately after the spinal cord was injured. Canine BMSC therapy enhanced functional recovery of the hind limbs in mice with SCI. Nestin-positive cells were observed only in the lesion of mice with SCI that received BMSCs. These results suggest that canine BMSCs promote functional recovery in mice with SCI and that migration of nestin-positive cells may contribute to the efficacy of the BMSC treatment.

The effects of BMSCs transplantation on autophagy by CX43 in the hippocampus following traumatic brain injury in rats

Traumatic brain injury (TBI) can initiate a series of complicated pathological events, and induce various types of neuronal cell death including autophagy and apoptosis. Currently, the treatment of TBI is one of the main challenges in neurobiology. In this regard, the administration of bone marrow stromal cells (BMSCs) represents a novel treatment modality for TBI. However, the protective mechanism of BMSCs was unknown in the TBI. The aim of the present study was to assess the effects of BMSCs on connexin 43(CX43) and autophagy in the hippocampus following TBI in rats. A rat model of TBI was created using a modified weight-drop device. Double-membrane structures in the process of autophagy formation were frequently observed in injured brain by electron microscopy. The levels of autophagic pathway associated proteins and CX43 were also detected by western blot analysis. Specifically, immunoblotting results showed that BMSCs treatment after TBI could down-regulate light chain 3 (LC3), Beclin-1 and CX43 expression in the hippocampus. Taken together, our results demonstrated that BMSCs were able to significantly suppress TBI-induced autophagy activity, and the potential mechanism by regulating CX43 levels.

Combination BMSC and Niaspan Treatment of Stroke Enhances White Matter Remodeling and Synaptic Protein Expression in Diabetic Rats

ObjectiveWhite matter remodeling plays an important role in neurological recovery after stroke. Bone marrow stromal cells (BMSCs) and Niaspan, an agent which increases high density lipoprotein (HDL), each induces neurorestorative effects and promotes white matter remodeling after stroke in non-diabetic rats. In this study, we test whether combination of BMSCs with Niaspan induces an enhanced white matter remodeling in the ischemic brain of diabetic rats.Research design and methodsType-1 diabetes (T1DM) rats were subjected to transient middle cerebral artery occlusion (MCAo) and treated with or without BMSCs; Niaspan; and the combination of BMSCs + Niaspan daily for 14 days after MCAo. Immunostaining for white matter remodeling and synaptic protein expression including NG2; CNPase; BS (Bielschowsky silver); LFB (luxol fast blue); Synaptophysin and SMI-31 immunostaining were performed.ResultsBMSC monotherapy did not regulate NG2 and CNPase expression compared to T1DM control rats. Both, combination of BMSCs + Niaspan treatment, and Niaspan monotherapy significantly increase NG2 and CNPase expression compared to T1DM control. While combination BMSC+Niaspan, BMSC monotherapy and Niaspan monotherapy groups all increase BS, LFB, synaptophysin, and SMI-31 expression in the ischemic brain compared to T1DM-MCAo control. In addition, the combination treatment significantly enhances LFB, SMI-31, and Synaptophysin expression compared to BMSC monotherapy.ConclusionsCombination treatment of stroke with BMSCs and Niaspan in T1DM rats increases white matter remodeling and additively increases BMSC monotherapy induced myelination and synaptic plasticity after stroke in T1DM rats.

Exogenous and endogenous therapeutic effects of combination Sodium Ferulate and bone marrow stromal cells (BMSCs) treatment enhance neurogenesis after rat focal cerebral ischemia

Combining bone marrow stromal cells (BMSCs) with pharmacological therapy is an attractive approach for neurological function recovery of stroke. Our previous reports demonstrated that Sodium Ferulate (SF) combined with BMSCs administration could facilitate BMSCs migration into the ischemic brain by up-regulation of stromal cell-derived factor-1 alpha (SDF-1α)/chemokine (CXC motif) receptor-4 axis after stroke. To further investigate whether combination treatment could enhance neurogenesis through exogenous and endogenous therapeutic effects, we established rat permanent middle cerebral artery occlusion (pMCAo) model and measured ischemic infarct size by magnetic resonance imaging (MRI) scanning in the present study. The results showed that combination treatment could dramatically reduce ischemic infarction size which may be attributed to the effects on decreasing brain edema and enhancing cerebral tissue perfusion at 3days after stroke. Immunofluorescence staining results indicated that combination treatment could not only promote expression of Glucose transporter 1(Glut1) and Neuron-specific class III beta-tubulin (Tuj1) in the periinfarct area, but also improve BMSCs expression of Glut1, GFAP and Tuj1. Moreover, it showed combination treatment could enhance the endogenous expression of Tuj-1 in ischemic boundary zone. These results perhaps associated with combination treatment up-regulating bone morphogenetic proteins (BMP)2/4 expressions and down-regulating Notch-1, Hes1 and Hes5 expressions as detected by Western Blot analysis. Our study firstly demonstrated in vivo that combination treatment could facilitate exogenous BMSCs differentiation into neural-and astrocytic-like cells, as well as enhance repair capacity of brain parenchymal cells by promoting glucose metabolism and endogenous neurogenesis after stroke. These results illustrate that administration of SF and BMSCs is a potential pathway of cell-based pharmacological treatment towards ischemic stroke.

Synergistic effects of ultrashort wave and bone marrow stromal cells on nerve regeneration with acellular nerve allografts

Acellular nerve allografts (ANA) possess bioactivity and neurite promoting factors in nerve tissue engineering. Previously we reported that low dose ultrashort wave (USW) radiation could enhance the rate and quality of peripheral nerve regeneration with ANA repairing sciatic nerve defects. Meanwhile, ANA implanted with bone marrow stromal cells (BMSCs) exhibited a similar result. Thus, it is interesting to know whether it might yield a synergistic effect when USW radiation is combined with BMSCs-laden ANA. Here we investigated the effectiveness of ANA seeded with BMSCs, combined with USW therapy on repairing peripheral nerve injuries. Adult male Wistar rats were randomly divided into four groups: Dulbecco's modified Eagle's medium (DMEM) control group, BMSCs-laden group, ultrashort wave (USW) group and BMSC + USW group. The regenerated nerves were assayed morphologically and functionally, and growth-promoting factors in the regenerated tissues following USW administration or BMSCs integration were also detected. The results indicated that the combination therapy caused much better beneficial effects evidenced by increased myelinated nerve fiber number, myelin sheath thickness, axon diameter, sciatic function index, nerve conduction velocity, and restoration rate of tibialis anterior wet weight. Moreover, the mRNA levels of brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) in the spinal cord and muscles were elevated significantly. In conclusion, we found a synergistic effect of USW radiation and BMSCs treatment on peripheral nerve regeneration, which may help establish novel strategies for repairing peripheral nerve defects.

Intrathecal transplantation of bone marrow stromal cells attenuates blood-spinal cord barrier disruption induced by spinal cord ischemia-reperfusion injury in rabbits

Intrathecal administration of bone marrow stromal cells has been found to produce beneficial effects on ischemia-reperfusion injury to the spinal cord. The blood-spinal cord barrier is critical to maintain spinal cord homeostasis and neurologic function. However, the effects of bone marrow stromal cells on the blood-spinal cord barrier after spinal cord ischemia-reperfusion injury are not well understood. This study investigated the effects and possible mechanisms of bone marrow stromal cells on blood-spinal cord barrier disruption induced by spinal cord ischemia-reperfusion injury. This was a prospective animal study conducted at the Central Laboratory of the First Affiliated Hospital, China Medical University. The study used 81 Japanese white rabbits (weight, 1.8-2.6 kg). Spinal cord ischemia-reperfusion injury was induced in rabbits by infrarenal aortic occlusion for 30 minutes. Two days before the injury was induced, bone marrow stromal cells (1 × 10(8) in 0.2-mL phosphate-buffered saline) were transplanted by intrathecal injection. Hind-limb motor function was assessed using Tarlov criteria, and motor neurons in the ventral gray matter were counted by histologic examination. The permeability of the blood-spinal cord barrier was examined using Evans blue (EB) and lanthanum nitrate as vascular tracers. The expression and localization of tight junction protein occludin were assessed by Western blot, real-time polymerase chain reaction, and immunofluorescence analysis. Matrix metalloproteinase-9 (MMP-9) and tumor necrosis factor-α (TNF-α) expression were also measured. Intrathecal transplantation of bone marrow stromal cells minimized the neuromotor dysfunction and histopathologic deficits (P < .01) and attenuated EB extravasation at 4 hours (5.41 ± 0.40 vs 7.94 ± 0.36 μg/g; P < .01) and 24 hours (9.03 ± 0.44 vs 15.77 ± 0.89 μg/g; P < .01) after spinal cord ischemia-reperfusion injury. In addition, bone marrow stromal cells treatment suppressed spinal cord ischemia-reperfusion injury-induced decreases in occludin (P < .01). Finally, bone marrow stromal cells reduced the excessive expression of MMP-9 and TNF-α (P < .01). Pre-emptive intrathecal transplantation of bone marrow stromal cells stabilized the blood-spinal cord barrier integrity after spinal cord ischemia-reperfusion injury in a rabbit model of transient aortic occlusion. This beneficial effect was partly mediated by inhibition of MMP-9 and TNF-α and represents a potential therapeutic approach to mitigating spinal cord injury after aortic occlusion. Clinical thoracoabdominal aorta surgery may trigger spinal cord ischemia-reperfusion injury, resulting in paraplegia as well as bladder, bowel, and sexual dysfunction. Transplantation of bone marrow stromal cells has attracted increasing attention in the field of nervous system protection, but its mechanisms have not been elucidated completely. The blood-spinal cord barrier plays a crucial role to maintain normal spinal cord function. This study suggested that intrathecal transplantation of bone marrow stromal cells stabilized blood-spinal cord barrier integrity through inhibiting the upregulation of matrix metalloproteinase-9 and tumor necrosis factor-a and ameliorated spinal cord ischemia-reperfusion injury. This may provide a novel train of thought to enhance the protective effects of bone marrow stromal cells on spinal cord injury.

The immunomodulatory effect of bone marrow stromal cells (BMSCs) on interleukin (IL)-23/IL-17-mediated ischemic stroke in mice

Ischemic cerebral infarction is a leading cause of death and disability, but the key inflammatory cytokines were not fully understood and no successful therapy has been established. We have used the methods of reverse transcriptase-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), western blot, and immunohistochemical staining to detect the dynamic changes of IL-23/IL-17 axis in brain infarction and BMSC treatment on the sixth day. BMSC transplantation could reduce the infarct size (P<0.05) and improve functional deficits (P<0.001) in brain infarction. The level of IL-23/IL-17 axis expression is higher (P<0.05) in pMCAO-operated group than that in sham operated group. BMSC treatment could reduce IL-23 expression (P<0.05) and attenuate IL-17 expression (P<0.01) both in serum and around infarct lesion. So we have drawn the conclusion that IL-23/IL-17 axis induces inflammation in the pathophysiological process of cerebral infarction. BMSC treatment plays therapeutic role by immunomodulating the expression of IL-23/IL-17 axis. These findings may help to understand the cytokines in cerebral infarction and open up a possible new way of immunological treatment.

Abstract 191: Neurorestorative Therapy For Stroke In Type 2 Diabetic Rats Using Bone Marrow Stromal Cells

Objective: Treatment of stroke may differ between the diabetic (DM) and non-DM stroke populations. Cell therapy with bone-marrow-stromal cells (BMSC) initiated at 24h after stroke improves functional recovery in non-DM rats, but not in type one DM rats. However, the effect of BMSC treatment of stroke in type two DM (T2DM) is unknown and is the subject of this investigation. Methods: T2DM was induced with high fat diet and low dose of streptozotocin. T2DM rats were subjected to 2h of middle cerebral artery occlusion (MCAo), then treated with either BMSC (5X106) or vehicle (n=8/group) initiated at 3 days after MCAo and monitored for 28 days. A battery functional tests, and vascular and white matter changes and protein expression were measured in the ischemic brain. Results: BMSC treatment significantly improved functional outcome at 14 and 21 days after MCAo in T2DM rats. However, the functional benefit was lost at 28 days after MCAo. BMSC significantly increased vascular and white matter remodeling identified by an increase of vascular density (vWF: 403±35/mm2 vs 303±17/mm2), cerebral arteriole number (aSMA 65.0±6.9/mm2 vs 38.6±4.4/mm2), Bielschowsky silver (axon marker, 26±2.4% vs 18±1.8%) and Lucifer fast blue (myelin marker, 39±3.1% vs 25±3.5%) expression in the ischemic boundary zone (IBZ). To test the mechanisms underlying the BMSC induced neurorestorative effects, decreased RAGE and Angiogenin expression were observed in the IBZ (RAGE: 3.1±0.4% vs 13.2±3%; Angiogenin: 9.5±2.2% vs 27.7±5.2%) of BMSC treated rats. However, BMSC treatment significantly increased internal carotid wall thickness (23.4±1.5μm vs 17.9±0.6μm), artery intimae thickness (11.1±0.6μm vs 7.6±0.5μm), and cerebral artery wall thickness (10.5±0.8μm vs 6.4±0.2μm); and significantly decreased cerebral artery diameter (26.5±1.7μm vs 36.3±2.8μm) compared to vehicle control. Conclusion: BMSC therapy in T2DM-MCAo significantly improved early functional outcome, but did not improve long term functional outcome. BMSC promote vascular and white matter remodeling, but increase cerebral artery neointimal hyperplasia. The increased cerebral artery neointimal hyperplasia may contribute to the failed improvement long term functional outcome after stroke in T2DM rats.