Neural Stem Cells Transplantation Group Research Articles

Mechanism of tetramethylpyrazine combined with neural stem cells transplantation on cerebral ischemia-reperfusion rats

This study aimed to investigate the intervention effect of tetramethylpyrazine(TMP) combined with transplantation of neural stem cells(NSCs) on middle cerebral artery occlusion(MCAO) rat model and to explore the mechanism of TMP combined with NSCs transplantation on ischemic stroke based on the regulation of stem cell biological behavior. MCAO rats were randomly divided into a model group, a TMP group, an NSCs transplantation group, and a TMP combined with NSCs transplantation group according to neurological function scores. A sham group was set up at the same time. The neurological function score was used to evaluate the improvement of neurological function in MCAO rats after TMP combined with NSCs transplantation. The proliferation, migration, and differentiation of NSCs were evaluated by BrdU, BrdU/DCX, BrdU/NeuN, and BrdU/GFAP immunofluorescence labeling. The protein expression of stromal cell-derived factor 1(SDF-1), C-X-C motif chemokine receptor 4(CXCR4), as well as oxidative stress pathway proteins nuclear factor erythroid 2-related factor 2(Nrf2), Kelch-like ECH-associated protein 1(KEAP1), heme oxygenase 1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1) was detected by Western blot to study the migration mechanism of TMP combined with NSCs. The results showed that TMP combined with NSCs transplantation significantly improved the neurological function score in MCAO rats. Immunofluorescence staining showed a significant increase in the number of BrdU~+, BrdU~+/DCX~+, BrdU~+/NeuN~+, and BrdU~+/GFAP~+ cells in the TMP, NSCs transplantation, and combined treatment groups, with the combined treatment group showing the most significant increase. Further Western blot analysis revealed significantly elevated expression of CXCR4 protein in the TMP, NSCs transplantation, and combined treatment groups, along with up-regulated protein expression of Nrf2, HO-1, and NQO1, and decreased KEAP1 protein expression. This study showed that both TMP and NSCs transplantation can promote the recovery of neurological function by promoting the proliferation, migration, and differentiation of NSCs, and the effect of TMP combined with NSCs transplantation is superior. The mechanism of action may be related to the activation of the Nrf2/HO-1/CXCR4 pathway.

Linc-NSC affects cell differentiation, apoptosis and proliferation in mouse neural stem cells and embryonic stem cells in vitro and in vivo

BackgroundStem cell therapy is a promising therapeutic strategy. In a previous study, we evaluated tumorigenicity by the stereotactic transplantation of neural stem cells (NSCs) and embryonic stem cells (ESCs) from experimental mice. Twenty-eight days later, there was no evidence of tumor formation or long-term engraftment in the NSCs transplantation group. In contrast, the transplantation of ESCs caused tumor formation; this was due to their high proliferative capacity. Based on transcriptome sequencing, we found that a long intergenic non-coding RNA (named linc-NSC) with unknown structure and function was expressed at 1100-fold higher levels in NSCs than in ESCs. This finding suggested that linc-NSC is negatively correlated with stem cell pluripotency and tumor development, but positively correlated with neurogenesis. In the present study, we investigated the specific role of linc-NSC in NSCs/ESCs in tumor formation and neurogenesis.MethodsWhole transcriptome profiling by RNA sequencing and bioinformatics was used to predict lncRNAs that are widely associated with enhanced tumorigenicity. The expression of linc-NSC was assessed by quantitative real-time PCR. We also performed a number of in vitro methods, including cell proliferation assays, differentiation assays, immunofluorescence assays, flow cytometry, along with in vivo survival and immunofluorescence assays to investigate the impacts of linc-NSC on tumor formation and neurogenesis in NSCs and ESCs.ResultsFollowing the knockdown of linc-NSC in NSCs, NSCs cultured in vitro and those transplanted into the cortex of mice showed stronger survival ability (P < 0.0001), enhanced proliferation(P < 0.001), and reduced apoptosis (P < 0.05); the opposite results were observed when linc-NSC was overexpressed in ESCs. Furthermore, the overexpression of linc-NSC in ECSs induced enhanced apoptosis (P < 0.001) and differentiation (P < 0.01), inhibited tumorigenesis (P < 0.05) in vivo, and led to a reduction in tumor weight (P < 0.0001).ConclusionsOur analyses demonstrated that linc-NSC, a promising gene-edited target, may promote the differentiation of mouse NSCs and inhibit tumorigenesis in mouse ESCs. The knockdown of linc-NSC inhibited the apoptosis in NSCs both in vitro and in vivo, and prevented tumor formation, revealing a new dimension into the effect of lncRNA on low survival NSCs and providing a prospective gene manipulation target prior to transplantation. In parallel, the overexpression of linc-NSC induced apoptosis in ESCs both in vitro and in vivo and attenuated the tumorigenicity of ESCs in vivo, but did not completely prevent tumor formation.

Restorative effects and mechanisms of neural stem cell transplantation on ischemic brain injury based on the Wnt signaling pathway

Objective To explore the restorative effects and mechanisms of neural stem cell (NSC) transplantation on ischemic brain injury based on the Wnt signaling pathway. Methods Out of 102 male KM mice, 15 were randomly selected as the control group without any intervention, while the remaining 87 underwent middle cerebral artery occlusion (MCAO) using the Zea-Longa suture method. Seven mice that did not successfully model MCAO were excluded, leaving 80 mice that successfully underwent MCAO, randomized into two groups: the Ischemic Brain Injury group (n = 40) receiving 10 μL of sterile PBS solution injected into the lateral ventricle, and the Ischemic Brain Injury + NSCs Transplantation group (n = 40) receiving 10 μL of NSCs suspension injected into the lateral ventricle. Results Compared to the ischemic brain injury group, mice in the Ischemic Brain Injury + NSCs Transplantation group exhibited significantly alleviated edema in the middle cerebral artery supply area, with neurons displaying more normal morphological characteristics and fewer signs of degeneration and necrosis. The mice with NSC transplantation had significantly smaller infarct volume than those in the ischemic brain injury group (p < 0.05). The mice with NSC transplantation showed significantly lower Zea-Longa scores and a lower proportion of TUNEL-positive cells compared to those in the ischemic brain injury group (p < 0.05). Conclusion NSC transplantation can significantly inhibit neuronal apoptosis in the ischemic region of mice with ischemic brain injury, alleviate brain tissue edema, reduce infarct volume, and improve neurological function. The mechanism may be related to Wnt signaling pathway activation.

Effects of neural stem cells in combination with nerve neurotrophic factor 3 gene modified olfactory ensheathing cell co-transplants on functional recovery after traumatic brain injury in the adult rats

Objective To observe the effects of neural stem cells (NSCs) combined with neurotrophin-3 gene modified olfactory ensheathing cells (OECs-NT-3) co-transplants on functional recovery after traumatic brain injury in the adult rats. Methods NSCs and OECs were cultured from hippocampus and olfactory bulb of neonatal rats and then identified. OECs-NT-3 was constructed by lentiviral vector containing NT-3 gene. Male SD rats weighing (200±20) g were randomly divided into sham group, model group, OECs-NT-3 transplantation group, NSCs transplantation group and OECs-NT-3 combined NSCs transplantation group. The rat brain injury model was established. The corresponding cells were transplanted. At day 1, 7, 14 and 28 after transplantation, the modified neurological severity score (mNSS) was used to evaluate neurological function, and brain tissues were taken at each time point for paraffin sections. Hematoxylin-eosin staining was performed. Cell apoptosis was detected by terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) method. Immunohistochemical staining of BrdU and p75 was done. Results At day 28 of transplantation, the mNSS scores in rats in the combined group (4.00±1.05) were significantly different from those in the model group (8.50±1.43), the OECs-NT-3 group (5.50±1.35) and the NSCs group (6.00±1.49) (t=-7.997, -2.764, -3.464, P 0.05). Hematoxylin and eosin (HE) staining revealed that the cell density in the sham group, model group, OECs-NT-3 group, NSCs group and the combined group was (220±22), (299±47), (347±35), (390±72) and (523±55) cells, respectively. As compared with the model group, OECs-NT-3 group and NSCs group, the cell density in the combined group was significantly increased (t=7.246, 5.996, 3.373, P<0.05), and the tissue structure was relatively regular. The p75 and BrdU immunohistochemistry showed that the two kinds of transplanted cells could survive continuously and were widely distributed around the lesion. At day 7 after transplantation, the apoptotic index in the sham group, model group, OECs-NT-3 group, NSCs group and combined group was (1.67±0.17)%, (47.49±7.92)%, (31.22±3.97)%, (34.48±4.69)% and (18.17±3.13)% respectively. The apoptosis rate in the combined transplantation group was significantly lower than that in the model group, OECs-NT-3 group and NSCs group (t=5.963, 4.470, 5.012, P<0.05). Conclusion Transplantation of NSCs combined with OECs-NT-3 can alleviate neuronal apoptosis after brain injury in rats, repair tissue defects, and promote the recovery of nerve function. Key words: Neural stem cell; Neurotrophin-3 gene; Gene modify; Olfactory ensheathing cells; Combined transplantation; Traumatic brain injury

Anti-Inflammatory Mechanism of Neural Stem Cell Transplantation in Spinal Cord Injury.

Neural stem cell (NSC) transplantation has been proposed to promote functional recovery after spinal cord injury. However, a detailed understanding of the mechanisms of how NSCs exert their therapeutic plasticity is lacking. We transplanted mouse NSCs into the injured spinal cord seven days after SCI, and the Basso Mouse Scale (BMS) score was performed to assess locomotor function. The anti-inflammatory effects of NSC transplantation was analyzed by immunofluorescence staining of neutrophil and macrophages and the detection of mRNA levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6) and interleukin-12 (IL-12). Furthermore, bone marrow-derived macrophages (BMDMs) were co-cultured with NSCs and followed by analyzing the mRNA levels of inducible nitric oxide synthase (iNOS), TNF-α, IL-1β, IL-6 and IL-10 with quantitative real-time PCR. The production of TNF-α and IL-1β by BMDMs was examined using the enzyme-linked immunosorbent assay (ELISA). Transplanted NSCs had significantly increased BMS scores (p < 0.05). Histological results showed that the grafted NSCs migrated from the injection site toward the injured area. NSCs transplantation significantly reduced the number of neutrophils and iNOS+/Mac-2+ cells at the epicenter of the injured area (p < 0.05). Meanwhile, mRNA levels of TNF-α, IL-1β, IL-6 and IL-12 in the NSCs transplantation group were significantly decreased compared to the control group. Furthermore, NSCs inhibited the iNOS expression of BMDMs and the release of inflammatory factors by macrophages in vitro (p < 0.05). These results suggest that NSC transplantation could modulate SCI-induced inflammatory responses and enhance neurological function after SCI via reducing M1 macrophage activation and infiltrating neutrophils. Thus, this study provides a new insight into the mechanisms responsible for the anti-inflammatory effect of NSC transplantation after SCI.

Effectiveness of muscle basal lamina carrying neural stem cells and olfactory ensheathing cells in spinal cord repair.

We examined the effect of muscle basal lamina (MBL) with neural stem cells (NSCs) and olfactory ensheathing cells (OECs) on spinal cord injury repair. Seventy-two Sprague-Dawley rats were subjected to spinal cord hemisection and divided into 6 groups. In blank control group (group A), the ends of the spinal cord hemisection model were flushed with physiological saline. In NSC transplantation group (B), OEC transplantation group (C), MBL with NSC transplantation group (D), MBL with OEC transplantation group (E), and MBL with NSC and OEC transplantation group (F), NSCs, OECs, MBL with NSCs, MBL with OECs, and MBL with NSCs and OECs were implanted into the ends of the hemisection model. Survival and migration of transplanted cells were detected by immunohistochemistry and immunofluorescence after 4 and 8 weeks. Hind limb function repair was evaluated by Bundle branch block score at various time points before and after surgery. MBL could promote NSC growth along its lumen and promote host cell advancement in the lumen, reducing local inflammatory responses. Using MBL with NSCs and/or OECs for spinal cord repair shows advantages over simple cell transplantation. Group F contained more nerve cells in muscle basal lamina than group E. This method is useful for forming more axons, synaptic connections, and signal transduction pathways. However, these new axons showed nerve demyelination, which may greatly limit nerve signal conduction. In group F, OECs could induce neural stem cells, axonal growth, and synaptic connection formation, but its role is limited.

Effects of hypothermia combined with neural stem cell transplantation on recovery of neurological function in rats with spinal cord injury.

The microenvironment of the injured spinal cord is hypothesized to be involved in driving the differentiation and survival of engrafted neural stem cells (NSCs). Hypothermia is known to improve the microenvironment of the injured spinal cord in a number of ways. To investigate the effect of NSC transplantation in combination with hypothermia on the recovery of rat spinal cord injury, 60 Sprague-Dawley female rats were used to establish a spinal cord hemisection model. They were divided randomly into three groups: A, spinal cord injury group; B, NSC transplantation group; and C, NSC transplantation + hypothermia group. At 1, 2, 4, 6 and 8 weeks post-injury, the motor function of all animals was evaluated using the Basso, Beattie and Besnaham locomotor scoring system and the inclined plane test. At 4 weeks post-transplantation, histological analysis and immunocytochemistry were performed. At 8 weeks post-transplantation, horseradish peroxidase nerve tracing and transmission electron microscopy were conducted to observe axonal regeneration. The outcome of hind limb motor function recovery in group C significantly surpassed that in group B at 4 weeks post-injury (P<0.05). Recovery was also observed in group A, but to a lesser degree. For the pathological sections no neural axonal were observed in group A. A few axon-like structures were observed in group B and more in group C. Horseradish peroxidase-labeled neurofibers and bromodeoxyuridine-positive cells were observed in the spinal cords of group C. Fewer of these cells were found in group B and fewer still in group A. The differences among the three groups were significant (P<0.05). Using transmission electron microscopy, newly formed nerve fibers and myelinated nerve fibers were observed in the central transverse plane in groups B and C, although these nerve fibers were not evident in group A. In conclusion, NSC transplantation promoted the recovery of hind limb function in rats, and combination treatment with hypothermia produced synergistic effects.

The role of exogenous neural stem cells transplantation in cerebral ischemic stroke.

To observe the effects of neural stem cells (NSCs) transplantation in rats' striatum and subventricular zone (SVZ) in rat models of focal cerebral ischemia and reperfusion. Hippocampus was extracted from fetal rats with 14 days of gestation. Suspension culture was used to isolate and culture the rat's NSCs. A cerebral ischemia and reperfusion rat's model was made on the left side of the brain through occlusion of the left middle cerebral artery. Neurological signs were assessed by Zea Longa's five-grade scale, with scores 1, 2, and 3 used to determine the successful establishment of the rat's model. The NSCs were stereotaxically injected into the left striatum 24 hours after the successful rat's model was built. Rats were then randomly divided into 5 groups, namely, normal group, sham operation group, ischemia group, PBS transplantation group, and NSCs transplantation group, each of which was observed on day 3, day 7, and day 14. The ischemia-related neurological deficits were assessed by using a 7-point evaluation criterion. Forelimb injuries were evaluated in all rats using the foot-fault approach. Infarct size changes were observed through TTC staining and cell morphology and structure in the infarct region were investigated by Nissl staining. Apoptosis and apoptosis-positive cell counts were studied by Tunel assay. Expressions of double-labeling positive cells in the striatum and subventricular zone (SVZ) were observed by BrdU/NeuN and BrdU/GFAP fluorescent double-labeling method and the number of positive cells in the striatum and SVZ was counted. Results from the differently treated groups showed that right hemiplegia occurred in the ischemia group, PBS transplantation group, and NSCs transplantation group in varying degrees. Compared with the former two groups, there was least hemiplegia in the NSCs transplantation group. The TTC staining assay showed that rats in the NSCs transplantation group had smaller infarct volume than those from the PBS transplantation group. The Nissl dyeing showed that there was a large area of neuronal necrosis and apoptosis in the ischemia and PBS transplantation groups, and damage was mainly focused in the striatum. Degeneration and damage of nerve cells were significantly reduced in the NSCs transplantation group. The Tunel assay showed that the number of apoptosis-positive cells in the NSCs transplantation group was less than that in the PBS transplantation group at each time point. Double immunofluorescent labeling showed that the proliferation of endogenous neural stem cells began at the third day, reaching the peak at the 7th day, and was significantly reduced at the 14th day in the SVZ. The number of BrdU/NeuN increased significantly in the NSCs transplantation group compared to that in the PBS transplantation group (P < 0.05). The number of BrdU/GFAP decreased significantly in the NSCs transplantation group compared to that of PBS transplantation group (P < 0.05). The number of BrdU/GFAP-positive cells in the striatum was observed to be much more in the PBS transplantation group than in the NSCs transplantation group. Both neurological deficits and coordination capacity of rats with cerebral ischemia were significantly improved via transplantation of the neural stem cells. In conclusion, transplantation of neural stem cells can therefore possibly promote the differentiation of endogenous NSCs into neurons and reduce their differentiation towards glial cells. Transplantation of the neural stem cells may also change the ischemic microenvironment of striatum, possibly inhibiting the proliferation of glial cells.

Neural stem cells transplantation combined with monosialotetrahexosyl ganglioside for treatment of spinal cord injury in rats

Objective To investigate the curative effect of neural stem cells (NSCs) transplantation combined with monosialotetrahexosyl ganglioside (GMi) in treatment of acute spinal cord injury in rats. Methods Compressive spinal cord injury model at T8 segment was established in the adult SD rats that were then randomly divided into three groups, ie, control group, NSCs transplantation group and NSCs + GM1 group.Continuous observation was performed at 1,2, 4 and 8 weeks.Functional neurological recovery of the injured spinal cord was evaluated with motor function scale, pathology, transmission electron microscopy and somatosensory evoked potential (SEP). Results The motor function of the lower extremities was recovered at different degrees in three groups.While the motor function recovery level of the animals and the positive staining cells of the calcitonin gene-related peptide (CGRP) in the NSCs + GM1 group were higher than those in the other two groups at 4 and 8 weeks (P ＜ 0.01).Compared with control group and NSCs group, focal necrosis and small vessel regeneration were observed only in the center of the injured segment in the NSCs + GM1 group at 8 weeks.Electron microscope scan showed edema under the membrane of the large myelin sheath in the control group, much intact myelin sheath, well-differentiated neurons and many kinds of synapse vesicles in the NSCs + GM1 group.The latent period of SEP was shortened markedly in the NSCs + GM1 group two weeks after transplantation (P ＜0.05).The latent period shortening was apparent in the NSCs group at 4 and 8 weeks after transplantation but was still longer than that in the control group. Conclusions Transplantation of neural stem cells combined with use of GM1 can protect the nervous tissues after spinal cord injury, when GM1 reconstructs the spinal cord through promoting differentiation of the transplanted stem cells and linking with the host cells. Key words: Spinal cord injuries; GM1 ganglioside; Neural stem cells

The effect of neural stem cell transplantation on the hindlimb motor function and caspase-3 expression of motor cortex in spinal cord transected rats

Objective To observe the effect of neural stem cell (NSC)transplantation on the hindlimb motor function and caspase-3 expression of motor cortex (MC) in spinal cord transected (SCT) rats. Methods S prague-Dawley (SD) rats were randomly divided into sham operated group, operation group( T9 transection), and subacute NSC transplantation group. The MC of each group( n= 8 )was harvested 7 days post operation (dpo), then western blot was employed to detect the level of caspase-3( β-actin was used as control ). The left 5 animals of each group were subjected to BBB score evaluation at 16th week,then the animals were sacrificed and the MC was harvested and performed immunostain by using caspase-3 rabbit antibody. Results Following NSC transplanta tion, BBB scores( ( 7.58 ± 0. 99 ) ) increased significantly than seen in the SCT animals( ( 5.16 ± 1.19) ). The expressional level of caspase-3 at 7day post operation was( (0.89 + 0.12)) in MC of SCT rats,while it decreased significantly to( (0.76 + 0.11 ) ) in NSC transplantation rats(P＜0.05). The immunoreactive stain of caspase-3 was seen in the cytoplasm of pyramidal neuron in the cortex. Conclusion NSC engraft can downregulate the expression of caspase-3, corresponding to a significant improvement in hindlimb motor function. These findings indicate that NSC transplantation probably regulate the expression of apoptosis genes in MC to promote neurological function recovery in rats subjected SCT. Key words: Neural stem cells; Spinal cord transection; Transplantation; Apoptosis gene; Rat

Transplantation of neurotrophin-3 modified neural stem cell for spinal cord injury treatment in rats

Objective To investigate the survival and differentiation of enhanced green fluorescent protein (EGFP)-labeled neurotrophin-3 (NT-3) modified fetal spinal cord-derived neural stem cells (NSCs) after transplanted into injured spinal cord.Methods EGFP labeled NT-3 modified NSCs were transplanted into the lesioned site of T9 hemisected rats (n = 10).The survival of grafted cells, expressions of NT-3 and differentiation into oligodendrocytes were detected by immunohistochemical assessment, and compared with the rats in unmodified NSCs transplantation group (n = 10).Results Cell counting presented more surviving cells in NT-3-NSCs group (304 ± 40) than NSCs group (258 ± 41) (P 0.05).The percentage of NT-3 positive cells in NT-3-NSCs group [(74.2 ± 14.1)%] was higher than NSCs group [(22.9 ± 11.1) %] (P < 0.01).More grafted cells in NT-3-NSCs group [(53.8 ± 12.4) %] differentiated into oligedendrocytes than NSCs group [(39.7 ± 13.7) %] (P 0.05).Conclusion NT-3 modification improves the survival of grafted NSCs, and promotes the expression of NT-3 and the differentiation into oligodendrocytes, which can increase the therapeutic effect of NSCs transplantation for spinal cord injury. Key words: Spinal cord injury; Transplantation; Neural stem cells; Neurotrophin3

Experimental study on treatment of glioma by embyonic neural stem cell transplnation in rats

The neural stem cells in Wistar rats were cultured in vitro, purified, and transplanted into C6 glioma model in order to observe their biological characters and provide a basic foundation for treatment of neurological diseases by neural stem cell transplantation. The cells at hippocampal area from gestation 15-day rats were cultured in vitro, and frozen and preserved in liquid nitrogen. C6 tumor-bearing models (n=25) and neural stem cells transplantation models (n=35) were established. When the tumor grew to 3 to 4 weeks, 5 rats in each group were randomly selected for MRI examination. At different intervals, the rats were perfused and sampled for HE staining, GFAP and BrdU immunohistochemical staining. The results showed that after resuscitation of neural stem cells at 1-4 passages, the cell viability was 40%-63% with the difference being not significant. The cells could proliferate, passage, and most cells transplanted into glioma model survived. The mean survival time in neural stem cell transplantation group and control was 4.28 and 3.88 weeks respectively, and the average tumor size in the former was smaller than in the latter. It was concluded that embryonic neural stem cells in rats could proliferate and differentiate, and after resuscitation the biological characteristic and viability of the cells were not influenced. Neural stem cells had inhibitory effects on the growth of glioma cells and could prolong the survival of rat model.

Transplantation of gene-transfected neural stem cells for transient cerebral ischemia in rats

To detect the expression and the role of vascular endothelial growth factor (VEGF)-transfected neural stem cells (NSCs) in rat brain subjected to ischemia. Fetal NSCs were cultured from E14 days SD rats and transfected with VEGF121 gene by using lipofectAMINE technique. The gene expression of transfected cells was detected by RT-PCR and immunofluorescent staining in vitro. Temporary middle cerebral artery occlusion (tMCAO) model was established in 40 SD rats and then rate were randomly divided into (1) control group (n = 10), (2) PBS transplantation group (n = 10), (3) neural stem cells transplantation group (n = 10), and (4) VEGF-secreting neural stem cells transplantation group (n = 10). BrdU-labelled NSCs and VEGF-secreting NSCs were transplanted into the penumbra zones respectively 3 days after the tMCAO model was established. Neurological Severity Score (NSS) was checked in all groups 2, 4, 6, 8, 10, 12 weeks respectively after transplantation. One and 12 weeks after the transplantation, 10 rats in the group (4) were killed and then brains taken out respectively. By using immunofluorescent staining, the VEGF expression of transplanted cells 1 week after transplantation, differentiation and migration of transplanted neural stem cells 12 week after transplantation were detected respectively. VEGF-transfected neural stem cells could continuously express gene products during the first 2 weeks. Both transfected NSCs and their progeny expressed VEGF gene products, which was demonstrated by fluorescence study. There were no significant differences in NSS in groups 4 when tMCAO models were just established. However, the values of NSS in (4) group were 5.8 +/- 1.5, 5.0 +/- 1.0, 4.6 +/- 1.0, 4.0 +/- 0.7, 4.0 +/- 1.0, 3.8 +/- 0.4 from 2 approximately 12 weeks after transplantation, significantly lower than those in groups (1) and (2) 8 weeks (P = 0.008) and those in groups (1), (2) and (3) 12 weeks (P = 0.000) after transplantation. NSS in group (3) was also lower than that in groups (1) and (2) 8 and 12 weeks after transplantation. One week after transplantation, immunofluorescent staining showed that VEGF-transfected NSCs migrated and expressed VEGF into hosts' brains. Twelve weeks after transplantation, transplanted NSCs survived and migrated, some of them differentiated to neurons and integrated well with hosts' cytoarchitectural components. VEGF-transfected NSCs express gene products during the early time after transplantation, which reduce brain injury through protecting the vascular system against ischemia and reperfusion injury. Transplantation of VEGF-transfected NSCs might be a novel method for treatment of cerebral ischemia.

Transplanted neural stem cells promote regeneration of peripheral nerve

Purpose of study: The purposes of this study are to investigate the characters of neural stem cells in collagen medium and to achieve the peripheral nerve regeneration after neural stem cell transplantation.Methods used: Neural stem cells were harvested from hippocampus of embryo rat. The cells were floated and cultured in medium containing bFGF (20 ng/ml) and collagen gel (1%). To investigate whether these cells in collagen medium could maintain the undifferentiated condition, cells were stained with anti-nestin antibody, which is specific for undifferentiated neural cells. Thereafter we removed bFGF to investigate whether these cells could differentiate into neurons and whether the axon could extend in the collagen medium. A 15-mm defect was made of the sciatic nerves of inbred Fischer 344 adult rats. The defects were bridged by silicone tubes (20 mm in length, 1.5 mm in diameter) filled inside with neural stem cells in collagen gel. In the control rats, silicone tubes were filled with collagen gel without neural stem cells. We counted the numbers and diameters of axons at the regenerative nerves in the silicone tubes 8 weeks after transplantation. The regenerative nerves were stimulated electrically to investigate whether the descending nerve action potential (NAP) could be recognized.of findings: 1) Neural stem cells started to proliferate within 48 hours in the collagen medium, and neurospheres were recognized after 8 days. On immunostains with anti-nestin antibody, positive findings were observed in the cytoplasm. In medium without bFGF, these cells differentiated into neurons and gradually extended axons. 2) Eight weeks after neural stem cells were transplanted, regenerative nerves were visible in the silicone tube. The mean number of myelinated fibers was 7,064 and the mean diameter of myelinated fibers was 3.51 μm in the neural stem cell transplantation group, but these values were 293 and 1.51 μm in control group. NAPs were recognized in all regenerative nerves. Mean latency was 7.92 ms, and mean amplitude was 1.38 mV. The mean latency was almost the same as the normal control nerve, but mean amplitude was 38% of the normal control nerve.Relationship between findings and existing knowledge: For the peripheral nerve regeneration, several kinds of cells were used for the source of transplantation. This is the first report of neural stem cells being transplanted to a nerve injury site, and definite regeneration was recognized.Overall significance of findings: Neural stem cells from the hippocampus of fetal rats maintained the potentials of proliferate and their undifferentiated condition and ability to differentiate into neurons. Transplanted neural stem cells could supply regenerative myelinated fiber at nerve injury site.Disclosures: No disclosures.Conflict of interest: No conflicts.