Glial Cell Line-derived Neurotrophic Factor-overexpressing Human Neural Stem/Progenitor Cells Enhance Therapeutic Efficiency in Rat with Traumatic Spinal Cord Injury

Spinal cord injury (SCI) causes axonal damage and demyelination, neural cell death, and comprehensive tissue loss, resulting in devastating neurological dysfunction. Neural stem/progenitor cell (NSPCs) transplantation provides therapeutic benefits for neural repair in SCI, and glial cell line-derived neurotrophic factor (GDNF) has been uncovered to have capability of stimulating axonal regeneration and remyelination after SCI. In this study, to evaluate whether GDNF would augment therapeutic effects of NSPCs for SCI, GDNF-encoding or mock adenoviral vector-transduced human NSPCs (GDNF-or Mock-hNSPCs) were transplanted into the injured thoracic spinal cords of rats at 7 days after SCI. Grafted GDNF-hNSPCs showed robust engraftment, long-term survival, an extensive distribution, and increased differentiation into neurons and oligodendroglial cells. Compared with Mock-hNSPC- and vehicle-injected groups, transplantation of GDNF-hNSPCs significantly reduced lesion volume and glial scar formation, promoted neurite outgrowth, axonal regeneration and myelination, increased Schwann cell migration that contributed to the myelin repair, and improved locomotor recovery. In addition, tract tracing demonstrated that transplantation of GDNF-hNSPCs reduced significantly axonal dieback of the dorsal corticospinal tract (dCST), and increased the levels of dCST collaterals, propriospinal neurons (PSNs), and contacts between dCST collaterals and PSNs in the cervical enlargement over that of the controls. Finally grafted GDNF-hNSPCs substantially reversed the increased expression of voltage-gated sodium channels and neuropeptide Y, and elevated expression of GABA in the injured spinal cord, which are involved in the attenuation of neuropathic pain after SCI. These findings suggest that implantation of GDNF-hNSPCs enhances therapeutic efficiency of hNSPCs-based cell therapy for SCI.

Effect of oxygen glucose deprivation-reperfusion in astrocytes overexpressing endothelin-1 on the proliferation of neural stem/progenitor cells / 器官移植

Objective To investigate the effect of oxygen glucose deprivation-reperfusion (OGD-R) in astrocytes overexpressing endothelin (ET)-1 on the proliferation of neural stem/progenitor cells (NSPCs). Methods OGD-R models of negative control astrocytes (C6-Mock) and astrocytes over-expressing ET-1 (C6-ET-1) were constructed. Transwell co-culture system of astrocytes and NSPCs was established. Morphologic observation and identification of the astrocytes and primary NSPCs were performed. The cells were divided into four groups: C6-Mock+NSPCs, OGD-R+C6-Mock+NSPCs, C6-ET-1+NSPCs and OGD-R+C6-ET-1+NSPCs groups and co-cultured for 0, 24, 48 and 72 h respectively. The diameter of neurosphere was measured in each group. Results In the C6-Mock and C6-ET-1 cells, type Ⅰ astrocytes in fibrous morphology were observed. Glial fibrillary acidic protein (GFAP) was expressed in the cytoplasm of these two types of cells. Primary NSPCs were positive for nestin staining. After co-culture for 48 and 72 h, the neurosphere diameter in the OGD-R+C6-Mock+NSPCs group was significantly greater than that in the C6-Mock+NSPCs group. The neurosphere diameter in the OGD-R+C6-ET-1+NSPCs group was considerably greater than that in the C6-ET-1+NSPCs group. The neurosphere diameter in the OGD-R+C6-ET-1+NSPCs group was significantly greater compared with that in the OGD-R+C6-Mock+NSPCs group (all P<0.05). Conclusions OGD-R astrocytes can promote the proliferation of NSPCs. ET-1 over-expression further accelerates the proliferation of NSPCs.

Curcumin Increase the Expression of Neural Stem/Progenitor Cells and Improves Functional Recovery after Spinal Cord Injury

OBJECTIVE: To investigates the effect of curcumin on proliferation of spinal cord neural stem/progenitor cells (SC-NSPCs) and functional outcome in a rat spinal cord injury (SCI) model. METHODS: Sixty adult male Sprague-Dawley rats were randomly and blindly allocated into three groups (sham control group; curcumin treated group after SCI; vehicle treated group after SCI). Functional recovery was evaluated by the Basso, Beattie, and Bresnahan (BBB) scale during 6 weeks after SCI. The expression of SC-NSPC proliferation and astrogliosis were analyzed by nestin/Bromodeoxyuridine (BrdU) and Glial fibrillary acidic protein (GFAP) staining. The injured spinal cord was then examined histologically, including quantification of cavitation. RESULTS: The BBB score of the SCI-curcumin group was better than that of SCI-vehicle group up to 14 days (p < 0.05). The co-immunoreactivity of nestin/BrdU in the SCI-curcumin group was much higher than that of the SCI-vehicle group 1 week after surgery (p < 0.05). The GFAP immunoreactivity of the SCI-curcumin group was remarkably lower than that of the SCI-vehicle group 4 weeks after surgery (p < 0.05). The lesion cavity was significantly reduced in the curcumin group as compared to the control group (p < 0.05). CONCLUSION: These results indicate that curcumin could increase the expression of SC-NSPCs, and reduce the activity of reactive astrogliosis and lesion cavity. Consequently curcumin could improve the functional recovery after SCI via SC-NSPC properties.

Curcumin Increase the Expression of Neural Stem/Progenitor Cells and Improves Functional Recovery after Spinal Cord Injury

OBJECTIVE: To investigates the effect of curcumin on proliferation of spinal cord neural stem/progenitor cells (SC-NSPCs) and functional outcome in a rat spinal cord injury (SCI) model.METHODS: Sixty adult male Sprague-Dawley rats were randomly and blindly allocated into three groups (sham control group; curcumin treated group after SCI; vehicle treated group after SCI). Functional recovery was evaluated by the Basso, Beattie, and Bresnahan (BBB) scale during 6 weeks after SCI. The expression of SC-NSPC proliferation and astrogliosis were analyzed by nestin/Bromodeoxyuridine (BrdU) and Glial fibrillary acidic protein (GFAP) staining. The injured spinal cord was then examined histologically, including quantification of cavitation.RESULTS: The BBB score of the SCI-curcumin group was better than that of SCI-vehicle group up to 14 days (p < 0.05). The co-immunoreactivity of nestin/BrdU in the SCI-curcumin group was much higher than that of the SCI-vehicle group 1 week after surgery (p < 0.05). The GFAP immunoreactivity of the SCI-curcumin group was remarkably lower than that of the SCI-vehicle group 4 weeks after surgery (p < 0.05). The lesion cavity was significantly reduced in the curcumin group as compared to the control group (p < 0.05).CONCLUSION: These results indicate that curcumin could increase the expression of SC-NSPCs, and reduce the activity of reactive astrogliosis and lesion cavity. Consequently curcumin could improve the functional recovery after SCI via SC-NSPC properties.

M2 Phenotype Microglia-derived Cytokine Stimulates Proliferation and Neuronal Differentiation of Endogenous Stem Cells in Ischemic Brain

Microglia play a key role in the immune response and inflammatory reaction that occurs in response to ischemic stroke. Activated microglia promote neuronal damage or protection in injured brain tissue. Extracellular signals polarize the microglia towards the M1/M2 phenotype. The M1/M2 phenotype microglia released pro- and anti-inflammatory cytokines which induce the activation of neural stem/progenitor cells (NSPCs). In this study, we investigated how the cytokines released by microglia affect the activation of NSPCs. First, we treated BV2 cells with a lipopolysaccharide (LPS; 20 ng/ml) for M1 phenotype microglia and interleukin-4 (IL-4; 20 ng/ml) for M2 phenotype microglia in BV2 cells. Mice were subjected to transient middle cerebral artery occlusion (tMCAO) for 1 h. In ex vivo, brain sections containing the subventricular zone (SVZ) were cultured in conditioned media of M1 and M2 phenotype-conditioned media for 3 d. We measured the expression of cytokines in the conditioned media by RT-PCR and ELISA. The M2 phenotype microglia-conditioned media led to the proliferation and neural differentiation of NSPCs in the ipsilateral SVZ after ischemic stroke. The RT-PCR and ELISA results showed that the expression of TGF-α mRNA was significantly higher in the M2 phenotype microglia-conditioned media. These data support that M2 phenotype microglia-derived TGF-α is one of the key factors to enhance proliferation and neural differntiation of NSPCs after ischemic stroke.

Generation of Induced Pluripotent Stem Cells and Neural Stem/Progenitor Cells from Newborns with Spina Bifida Aperta

STUDY DESIGN: We established induced pluripotent stem cells (iPSCs) and neural stem/progenitor cells (NSPCs) from three newborns with spina bifida aperta (SBa) using clinically practical methods. PURPOSE: We aimed to develop stem cell lines derived from newborns with SBa for future therapeutic use. OVERVIEW OF LITERATURE: SBa is a common congenital spinal cord abnormality that causes defects in neurological and urological functions. Stem cell transplantation therapies are predicted to provide beneficial effects for patients with SBa. However, the availability of appropriate cell sources is inadequate for clinical use because of their limited accessibility and expandability, as well as ethical issues. METHODS: Fibroblast cultures were established from small fragments of skin obtained from newborns with SBa during SBa repair surgery. The cultured cells were transfected with episomal plasmid vectors encoding reprogramming factors necessary for generating iPSCs. These cells were then differentiated into NSPCs by chemical compound treatment, and NSPCs were expanded using neurosphere technology. RESULTS: We successfully generated iPSC lines from the neonatal dermal fibroblasts of three newborns with SBa. We confirmed that these lines exhibited the characteristics of human pluripotent stem cells. We successfully generated NSPCs from all SBa newborn-derived iPSCs with a combination of neural induction and neurosphere technology. CONCLUSIONS: We successfully generated iPSCs and iPSC-NSPCs from surgical samples obtained from newborns with SBa with the goal of future clinical use in patients with SBa.

Induction of Angiogenesis and Neurogenesis by Serum from Rats Treated with Shunaoxin Dropping Pills / 中草药·英文版

Objective Shunaoxin Dropping Pills (SDPs), a Chinese patent medicine, has been used widely in China for the treatment of headache, amnesia, and insomnia. The aim of the present study is to observe the effect of SDPs on inducing angiogenesis and neurogenesis in vitro. Methods The present testing system using the serum obtained from animals ig treated with SDPs and a co-culture system in vitro was used to investigate if SDPs promotes brain microvascular endothelial cells (BMECs) tube formation and neural differentiation of neural stem/progenitor cells (NSPCs), which plays important roles in angiogenesis and neurogenesis. Results The SDPs serum sampled from rats ig treated with SDPs for 3 d dose-dependently promoted the tube like structure formation of cultured BMECs, and enhanced the fraction of MAP-2 positive cells of NSPCs, which co-cultured with the BMECs and astrocyte. In addition, there was no significant change in the percentage of glial fibrillary acidic protein positive cells. Conclusion Our results show that SDPs serum can induce neural differentiation and BMECs tube formation in vitro.

Advances of Cell Transplantation for Stroke (review) / 中国康复理论与实践

@#A variety kinds of cells were transplanted for brain and spinal cord repair, and some types of cells have been used for stroke, which including bone marrow stromal cells, neural stem/progenitor cells, olfactory ensheathing cells, human neuronal cell and so on. The complementary advantages of different types of cells and the integrated application of varied strategies of nerve restoration is an important direction for future exploration.

Differentiation of Rat Neural Stem Cells Following Transplantation in the Brain of Huntington's Disease Rat Model

Stem cells provide an important means for regenerative medicine due to the capacity to generate multiple types of differentiated cells and at the same time to maintain self-renewal. To identify the therapeutic effect of the transplantation of neural stem cells, differentiation and migration capacity of the neural stem cells that were isolated from E14 rat embryo and maintained in culture were examined after transplantation to the striatum of the quinolinic acid (QA)-induced Huntington's disease rat model. in vitro co-culture of the neural stem cells with the mixture of primary neurons and astrocytes promoted the maturation and the synapse formation of neuronal progenies of neural stem cells. Following the implantation, the neural stem cells survived, differentiated, and migrated in the damaged striatum region, exhibiting immunoreactivities against nestin, Tuj-1, GFAP, GAD(67) and synapsin 1 to a varying degree. These data provide clear evidence supporting that the neural stem cells isolated from the rat embryo and maintained in the primary culture have a multiple capacity to differentiate into neurons or glial cells both in vitro and in vivo.