BACKGROUND The development of regenerative therapy for human spinal cord injury(SCI)is dramatically restricted by two main challenges:the need for a safe source of functionally active and reproducible neural stem cell...BACKGROUND The development of regenerative therapy for human spinal cord injury(SCI)is dramatically restricted by two main challenges:the need for a safe source of functionally active and reproducible neural stem cells and the need of adequate animal models for preclinical testing.Direct reprogramming of somatic cells into neuronal and glial precursors might be a promising solution to the first challenge.The use of non-human primates for preclinical studies exploring new treatment paradigms in SCI results in data with more translational relevance to human SCI.AIM To investigate the safety and efficacy of intraspinal transplantation of directly reprogrammed neural precursor cells(drNPCs).METHODS Seven non-human primates with verified complete thoracic SCI were divided into two groups:drNPC group(n=4)was subjected to intraspinal transplantation of 5 million drNPCs rostral and caudal to the lesion site 2 wk post injury,and lesion control(n=3)was injected identically with the equivalent volume of vehicle.RESULTS Follow-up for 12 wk revealed that animals in the drNPC group demonstrated a significant recovery of the paralyzed hindlimb as well as recovery of somatosensory evoked potential and motor evoked potential of injured pathways.Magnetic resonance diffusion tensor imaging data confirmed the intraspinal transplantation of drNPCs did not adversely affect the morphology of the central nervous system or cerebrospinal fluid circulation.Subsequent immunohistochemical analysis showed that drNPCs maintained SOX2 expression characteristic of multipotency in the transplanted spinal cord for at least 12 wk,migrating to areas of axon growth cones.CONCLUSION Our data demonstrated that drNPC transplantation was safe and contributed to improvement of spinal cord function after acute SCI,based on neurological status assessment and neurophysiological recovery within 12 wk after transplantation.The functional improvement described was not associated with neuronal differentiation of the allogeneic drNPCs.Instead,directed drNPCs migration to the areas of active growth cone formation may provide exosome and paracrine trophic support,thereby further supporting the regeneration processes.展开更多
Previous studies have shown that proliferation of endogenous neural precursor cells cannot alone compensate for the damage to neurons and axons. From the perspective of neural plastici- ty, we observed the effects of ...Previous studies have shown that proliferation of endogenous neural precursor cells cannot alone compensate for the damage to neurons and axons. From the perspective of neural plastici- ty, we observed the effects of functional electrical stimulation treatment on endogenous neural precursor cell proliferation and expression of basic fibroblast growth factor and epidermal growth factor in the rat brain on the infarct side. Functional electrical stimulation was performed in rat models of acute middle cerebral artery occlusion. Simultaneously, we set up a placebo stimulation group and a sham-operated group. Immunohistochemical staining showed that, at 7 and 14 days, compared with the placebo group, the numbers of nestin （a neural precursor cell marker）-positive cells in the subgranular zone and subventricular zone were increased in the functional electrical stimulation treatment group. Western blot assays and reverse-transcription PCR showed that total protein levels and gene expression of epidermal growth factor and basic fibroblast growth factor were also upregulated on the infarct side. Prehensile traction test results showed that, at 14 days, prehension function of rats in the functional electrical stimulation group was significantly better than in the placebo group. These results suggest that functional electrical stimulation can promote endogenous neural precursor cell proliferation in the brains of acute cerebral infarction rats, enhance expression of basic fibroblast growth factor and epidermal growth factor, and improve the motor function of rats.展开更多
BACKGROUND： Choosing proper donor cells is one of keys in experimental and clinical studies on cell replacement therapy （CRT） for treating Parkinson disease （PD）. Embryonic mesencephalic precursor cells （MPCs） ...BACKGROUND： Choosing proper donor cells is one of keys in experimental and clinical studies on cell replacement therapy （CRT） for treating Parkinson disease （PD）. Embryonic mesencephalic precursor cells （MPCs） can stably differentiate into dopaminergic neuron after in vitro proliferated culture. As compared with embryonic stem cell and neural stem cell strains, cell composition of embryonic MPCs after primary culture is also the most close to that of embryonic mesencephalic ventral cell suspension without proliferated culture. Successful experience accumulated in the latter suggests that primary cultured embryonic MPCs might be the most potential donor cells in clinical application with CRT for treating PD so far. OBJECTIVE： To investigate the feasibility of primary cultured embryonic precursor cells cultured primarily as donor cells in CRT for treating PD in rats. DESIGN ： A randomized and controlled trial taking SD rats as experimental animals.SETTING： Department of Neurosurgery, Huashan Hospital Affiliated to Fudan University.MATERIALS： This experiment was carried out at the Institute of Neuroscience, Shanghai Institute for Biological Science, Chinese Academy of Sciences from July 2003 to June 2004. Totally 26 female SD rats, with body mass of 200 to 220 g, were provided by Shanghai Experimental Animal Center of Chinese Academy of Sciences. METHODS ： Stereotaxic injection of 6-hydroxydopamine into the medial forebrain bundle were perfored to develop PD model rat. Among 26 SD rats, 20 rats achieved a more than 5 turns/min in apomorphine induced rotation test, reaching the standard of PD model rats. Immunohistochemical detection was performed on 1 out of 20 model rats after execution, and the other 19 rats were randomly divided into control group （n=5）, sham transplantation group （n=5）and cell grafted group （n=9）. Primary cultured E12 MPC cell suspension （1.2×10^11 L^-1）were used as donor cells. 4μL primary cultured E12 MPC cell suspension prepared freshly was injected into the lesioned corpus striatum of rats in cell grafted group, and 4μL D-Hank＇s solution was injected in sham transplantation group in the same way. There was no injection in control group. Apomorphine-induced rotation rate of PD rats were recorded respectively in cell grafted group and sham transplantation group pre-operation （initial value） and at postoperative 2, 4, 6 and 16 weeks. Apomorphine-induced rotation rate of PD rats was recorded in control group at postoperative 2 months （initial value） and following 2,4,6 and 16 weeks. To determine TH antigen with immunohistological ABC method （DAB developing） at 6 months post-transplantation to investigate the differentiation and survival of donor cells in the host body.MAIN OUTCOME MEASURES： Apomorphine-induced rotation behavior before and after transplantation and the survival and differentiation of implanted cells in the host body at 6 months post-transplantation. RESULTS： Among 19 model rats, one rat died after transplantation respectively in the cell grafted group and sham transplantation group; finally 17 model rats entered the stage of result analysis. Relative apomorphine-induced rotation rate was significantly decreased in the cell grafted group as compared with that before transplantation , with significant difference （P 〈 0.01 .P 〈 0.05）;the mean value of relative apomorphine-induced rotation rate was significantly decreased at postoperative 16 weeks in cell grafted group as compared with that of corresponding relative rotation rate in control group , also with significant difference （P 〈 0.05）.Immunohistological results showed that donor cells could differentiate into large and multi-polar dopaminergic neurons in the host body. CONCLUSION ： Primary cultured embryonic MPCs can be used as the donor cells in CRT for treating PD.展开更多
BACKGROUND： Cerebral injury in adult mammals can induce neural precursor cells （NPCs） to proliferate and migrate towards the focal zone, but it is unclear whether endogenous NPCs can migrate towards regions distal ...BACKGROUND： Cerebral injury in adult mammals can induce neural precursor cells （NPCs） to proliferate and migrate towards the focal zone, but it is unclear whether endogenous NPCs can migrate towards regions distal to the hemorrhagic focus or whether NPCs differentiate in the peripheral hemorrhagic region. OBJECTIVE： To investigate the distribution of endogenous NPCs in different brain regions of rats with experimental cerebral hemorrhage, as well as NPC proliferation and differentiation with time. DESIGN, TIME AND SE＇B＇ING： A randomized, controlled animal experiment was performed at the Department of Neurobiology, Luzhou Medical College, between January 2007 and October 2008. MATERIALS： Bromodeoxyuridine （BrdU） was purchased from Roche, Germany. Mouse anti-rat BrdU monoclonal antibody, rabbit anti-nestin polyclonal antibody, rabbit anti-neuron specific enolase （NSE） polyclonal antibody were purchased from Wuhan Boster, China. Rabbit anti-glial fibrillary acidic protein （GFAP） polyclonal antibody was purchased from Sigma, USA. METHODS： Thirty-five adult Sprague Dawley rats were randomly divided into three groups： （1） cerebral hemorrhage group （n = 25）, rats were stereotaxically administered 50 p L autologous arterial blood via the dorsal caudate putamen to induce cerebral hemorrhage; （2） sham-surgery group （n = 5）, rats underwent surgery but did not receive blood injection; （3） blank control group （n = 5）, rats received no surgery and blood administration. At 2 hours after surgery, all rats were intraperitoneally administered BrdU. MAIN OUTCOME MEASURES： Distribution and proliferation of BrdU-positive cells were observed by immunohistochemical staining. BrdU-positive cell differentiation into neurons and glial cells in the peripheral hemorrhagic region was detected by double-label immunofluorescence. RESULTS： Immunohistochemistry results revealed that BrdU-positive cells existed not only in the peripheral hemorrhagic region, such as the subependymal layer and hippocampal dentate gyrus, but also in the lateral septal nucleus, diagonal band, habenular nucleus, and cerebral cortex. Following cerebral hemorrhage, BrdU-positive cells in the peripheral hemorrhagic region gradually increased （P 〈 0.05）, and peaked at 7 14 days. Double-label immunofluorescence showed that with time after cerebral hemorrhage, BrdU/nestin-positive cells decreased, but BrdU/GFAP- and BrdU/NSE-positive cells increased in the peripheral cerebral hemorrhagic region （P 〈 0.05）. CONCLUSION： Cerebral hemorrhage can induce the proliferation of endogenous NPCs, which peaks at 1-2 weeks after hemorrhage. NPCs can also migrate towards the regions distal to the hemorrhagic focus, such as a diagonal band or lateral septal nucleus. NPCs can gradually differentiate with increasing time after hemorrhage.展开更多
We recently reported that targeted deletion of Pannexin 1 in neural precursor cells of the ventricular zone impairs the maintenance of these cells in healthy and stroke-injured brain. Here we frame this exciting new f...We recently reported that targeted deletion of Pannexin 1 in neural precursor cells of the ventricular zone impairs the maintenance of these cells in healthy and stroke-injured brain. Here we frame this exciting new finding in the context of our previous studies on Pannexin 1 in neural precursors as well as the close rela- tionship between Pannexin 1 and purinergic receptors established by other groups. Moreover, we identify important gaps in our understanding of Pannexin 1 in neural precursor cell biology in terms of the under- lying molecular mechanisms and functional/behavioural outcomes.展开更多
Objective To investigate whether buffalo rat liver cell conditioned medium (BRL CM) can be used as the culture medium of embryonic stem (ES) cells, and to get relatively pure neural precursor cells (NPCs) for treat...Objective To investigate whether buffalo rat liver cell conditioned medium (BRL CM) can be used as the culture medium of embryonic stem (ES) cells, and to get relatively pure neural precursor cells (NPCs) for treatment aim. Methods Mouse ES cells were cultured in BRL CM and medium contain leukemia inhibitory factor (LIF), respectively. NPCs were selectively cultured in serum free medium. Alkaline phosphatase activity was visualized with NBT/BCIP and nestin antigen was detected with immunocytochemical methods. Results BRL CM could be used as an efficiency culture condition instead of LIF in ES cells culture. About 86% of cells derived from ES cells in the serum free culture were NPCs. Conclusion BRL CM can replace LIF to use in ES cell culture. High purity of NPC can be induced from ES cells with serum free culture method.展开更多
Recent research has shown that defined sets of exogenous factors are sufficient to convert rodent and human somatic cells directly into induced neural stem cells or neural precursor cells（iNSCs/iNPCs）.The process of...Recent research has shown that defined sets of exogenous factors are sufficient to convert rodent and human somatic cells directly into induced neural stem cells or neural precursor cells（iNSCs/iNPCs）.The process of transdifferentiation bypasses the step of a pluripotent state and reduces the risk of tumorigenesis and genetic instability while retaining the self-renewing capacity.This iNSC/iNPC technology has fueled much excitement in regenerative medicine,as these cells can be differentiated into target cells for replacement therapy for neurodegenerative diseases.Patients＇ somatic cell-derived iNSCs/iNPCs have also been proposed to serve as disease models with potential value in both fundamental studies and clinical applications.This review focuses on the mechanisms,techniques,and applications of iNSCs/iNPCs from a series of related studies,as well as further efforts in designing novel strategies using iNSC/iNPC technology and its potential applications in neurodegenerative diseases.展开更多
Human embryonic stem (hES) cells provide a promising supply of specific cell types for transplantation therapy. We presented here the method to induce differentiation of purified neural precursors from hES cells. hES ...Human embryonic stem (hES) cells provide a promising supply of specific cell types for transplantation therapy. We presented here the method to induce differentiation of purified neural precursors from hES cells. hES cells (Line PKU-1 and Line PKU-2) were cultured in sus- pension in bacteriological Petri dishes, which differentiated into cystic embryoid bodies (EBs). The EBs were then cultured in N2 medium containing bFGF in poly-L-lysine-coated tissue culture dishes for two weeks. The central, small cells with 2―3 short processes of the spreading out- growth were isolated mechanically and replated. The resulting neurospheres were cultured in suspension for 10 days, then dissociated into single cell suspension with a Pasteur pipette and plated. Cells grew vigorously in an attached way and were passed every 4―5 days. Almost all the cells were proved nestin positive by immunostaining. Following withdrawal of bFGF, they differentiated into neurons expressing β-tubulin isotypeIII, GABA, serotonin and synaptophysin. Through induction of PDGF-AA, they differentiated into astrocytes expressing GFAP and oli- godendrocytes expressing O4. The results showed that hES cells can differentiate into typical neural precursors expressing the specific marker nestin and capable of generating all three cell types of the central nervous system (CNS) in vitro.展开更多
There are few studies on the membrane protein Ankfyl. We have found Ankfyl is specifically expressed in neural stem/precursor cells during early development in mice （murine）. To further explore Ankfyl function in ne...There are few studies on the membrane protein Ankfyl. We have found Ankfyl is specifically expressed in neural stem/precursor cells during early development in mice （murine）. To further explore Ankfyl function in neural development, we developed a gene knockout mouse with a mixed Balb/C and C57/BL6 genetic background. Using immunofluorescence and in situ hybridization, neural defects were absent in mixed genetic Ankfyl null mice during development and in adults up to 2 months old. However, Ankfyl gene knockout mice with a pure genetic background were found to be lethal in the C57/BL6 inbred mice embryos, even after seven generations of backcrossing. Polymerase chain reaction confirmed homozygotes were unattainable as early as embryonic day 11.5. We conclude that Ankfyl protein is dispensable in neural stem/precursor ceils, but could be critical for early embryonic murine development, depending on the genetic background.展开更多
Artificial neural network (NN) is such a model as to imitate the structure and intelligence feature of human brain. It has strong nonlinear mapping function. To introduce NN into the study of earthquake prediction is ...Artificial neural network (NN) is such a model as to imitate the structure and intelligence feature of human brain. It has strong nonlinear mapping function. To introduce NN into the study of earthquake prediction is not only an extension of the application of artificial neural network model but also a new try for precursor observation to serve the earthquake prediction. In this paper, we analyzed the predictability of time series and gave a method of application of artificial neural network in forecasting earthquake precursor chaotic time series. Besides, taking the ground tilt observation of Jiangning and Xuzhou Station, the bulk strain observation of Liyang station as examples, we analyzed and forecasted their time series respectively. It is indicated that the precision of this method can meet the needs of practical task and therefore of great value in the application to the future practical earthquake analysis and prediction.展开更多
Hippocampal neuronal loss causes cognitive dysfunction in Alzheimer’s disease.Adult hippocampal neurogenesis is reduced in patients with Alzheimer’s disease.Exercise stimulates adult hippocampal neurogenesis in rode...Hippocampal neuronal loss causes cognitive dysfunction in Alzheimer’s disease.Adult hippocampal neurogenesis is reduced in patients with Alzheimer’s disease.Exercise stimulates adult hippocampal neurogenesis in rodents and improves memory and slows cognitive decline in patients with Alzheimer’s disease.However,the molecular pathways for exercise-induced adult hippocampal neurogenesis and improved cognition in Alzheimer’s disease are poorly understood.Recently,regulator of G protein signaling 6(RGS6)was identified as the mediator of voluntary running-induced adult hippocampal neurogenesis in mice.Here,we generated novel RGS6fl/fl;APP_(SWE) mice and used retroviral approaches to examine the impact of RGS6 deletion from dentate gyrus neuronal progenitor cells on voluntary running-induced adult hippocampal neurogenesis and cognition in an amyloid-based Alzheimer’s disease mouse model.We found that voluntary running in APP_(SWE) mice restored their hippocampal cognitive impairments to that of control mice.This cognitive rescue was abolished by RGS6 deletion in dentate gyrus neuronal progenitor cells,which also abolished running-mediated increases in adult hippocampal neurogenesis.Adult hippocampal neurogenesis was reduced in sedentary APP_(SWE) mice versus control mice,with basal adult hippocampal neurogenesis reduced by RGS6 deletion in dentate gyrus neural precursor cells.RGS6 was expressed in neurons within the dentate gyrus of patients with Alzheimer’s disease with significant loss of these RGS6-expressing neurons.Thus,RGS6 mediated voluntary running-induced rescue of impaired cognition and adult hippocampal neurogenesis in APP_(SWE) mice,identifying RGS6 in dentate gyrus neural precursor cells as a possible therapeutic target in Alzheimer’s disease.展开更多
Objective:Diabetes-induced gastrointestinal(GI)motility disorders are increasingly prevalent.Damage to the enteric nervous system(ENS),composed primarily of enteric neurons and glial cells,is an essential mechanism in...Objective:Diabetes-induced gastrointestinal(GI)motility disorders are increasingly prevalent.Damage to the enteric nervous system(ENS),composed primarily of enteric neurons and glial cells,is an essential mechanism involved in these disorders.Although electroacupuncture(EA)has shown the potential to mitigate enteric neuronal loss,its mechanism is not fully understood.Additionally,the effects of EA on enteric glial cells have not been investigated.Enteric neural precursor cells(ENPCs)contribute to the structural and functional integrity of the ENS,yet whether EA enhances their differentiation into enteric neurons and glial cells remains unexplored.This study investigates whether EA promotes ENS repair through enhancing ENPC-derived neurogenesis and gliogenesis and elucidates the potential molecular mechanisms involved.Methods:Transgenic mice were used to trace Nestin+/nerve growth factor receptor(Ngfr)+ENPCs labeled with green fluorescent protein(GFP)in vivo.Mice were randomly divided into four groups:control,diabetes mellitus(DM),DM+sham EA,and DM+EA.The effects of EA on diabetic mice were evaluated by GI motility,ENS structure,and ENPC differentiation.Glial cell line-derived neurotrophic factor(GDNF)/Ret signaling was detected to clarify the underlying molecular mechanisms.Results:EA alleviated diabetes-induced GI motility disorders,as indicated by reduced whole gut transit time,shortened colonic bead expulsion time,and enhanced smooth muscle contractility.Furthermore,EA attenuated diabetes-induced losses of enteric neurons and glial cells,thereby restoring ENS integrity.Notably,EA reversed the diabetes-induced decrease in ENPCs and significantly increased the absolute number and the proportion of ENPC-derived enteric neurons.However,immunofluorescence analyses revealed no colocalization between EA-induced glial fibrillary acidic protein+glial cells and GFPlabeled ENPCs.Mechanistically,GDNF/Ret signaling was elevated in intestinal tissues and upregulated in ENPCs in EA-treated diabetic mice.Conclusion:EA facilitates ENS repair by promoting Nestin+/Ngfr+ENPC differentiation into enteric neurons via upregulation of GDNF/Ret signaling,and driving enteric gliogenesis from non-Nestin+/Ngfr+ENPCs.These findings highlight EA's role in ameliorating diabetes-induced GI dysmotility through ENPC-derived ENS restoration.展开更多
Endogenous neural stem cells become ＂activated＂ after neuronal injury, but the activation sequence and fate of endogenous neural stem cells in focal cerebral ischemia model are little known. We evaluated the relatio...Endogenous neural stem cells become ＂activated＂ after neuronal injury, but the activation sequence and fate of endogenous neural stem cells in focal cerebral ischemia model are little known. We evaluated the relationships between neural stem cells and hypoxia-inducible factor-1α and vascular endothelial growth factor expression in a photothromobotic rat stroke model using immunohistochemistry and western blot analysis. We also evaluated the chronological changes of neural stem cells by 5-bromo-2′-deoxyuridine（BrdU） incorporation. Hypoxia-inducible factor-1α expression was initially increased from 1 hour after ischemic injury, followed by vascular endothelial growth factor expression. Hypoxia-inducible factor-1α immunoreactivity was detected in the ipsilateral cortical neurons of the infarct core and peri-infarct area. Vascular endothelial growth factor immunoreactivity was detected in bilateral cortex, but ipsilateral cortex staining intensity and numbers were greater than the contralateral cortex. Vascular endothelial growth factor immunoreactive cells were easily found along the peri-infarct area 12 hours after focal cerebral ischemia. The expression of nestin increased throughout the microvasculature in the ischemic core and the peri-infarct area in all experimental rats after 24 hours of ischemic injury. Nestin immunoreactivity increased in the subventricular zone during 12 hours to 3 days, and prominently increased in the ipsilateral cortex between 3–7 days. Nestin-labeled cells showed dual differentiation with microvessels near the infarct core and reactive astrocytes in the peri-infarct area. BrdU-labeled cells were increased gradually from day 1 in the ipsilateral subventricular zone and cortex, and numerous BrdU-labeled cells were observed in the peri-infarct area and non-lesioned cortex at 3 days. BrdU-labeled cells rather than neurons, were mainly co-labeled with nestin and GFAP. Early expressions of hypoxia-inducible factor-1α and vascular endothelial growth factor after ischemia made up the microenvironment to increase the neuronal plasticity of activated endogenous neural stem cells. Moreover, neural precursor cells after large-scale cortical injury could be recruited from the cortex nearby infarct core and subventricular zone.展开更多
基金Supported by Russian Science Foundation,No.16-15-10432。
文摘BACKGROUND The development of regenerative therapy for human spinal cord injury(SCI)is dramatically restricted by two main challenges:the need for a safe source of functionally active and reproducible neural stem cells and the need of adequate animal models for preclinical testing.Direct reprogramming of somatic cells into neuronal and glial precursors might be a promising solution to the first challenge.The use of non-human primates for preclinical studies exploring new treatment paradigms in SCI results in data with more translational relevance to human SCI.AIM To investigate the safety and efficacy of intraspinal transplantation of directly reprogrammed neural precursor cells(drNPCs).METHODS Seven non-human primates with verified complete thoracic SCI were divided into two groups:drNPC group(n=4)was subjected to intraspinal transplantation of 5 million drNPCs rostral and caudal to the lesion site 2 wk post injury,and lesion control(n=3)was injected identically with the equivalent volume of vehicle.RESULTS Follow-up for 12 wk revealed that animals in the drNPC group demonstrated a significant recovery of the paralyzed hindlimb as well as recovery of somatosensory evoked potential and motor evoked potential of injured pathways.Magnetic resonance diffusion tensor imaging data confirmed the intraspinal transplantation of drNPCs did not adversely affect the morphology of the central nervous system or cerebrospinal fluid circulation.Subsequent immunohistochemical analysis showed that drNPCs maintained SOX2 expression characteristic of multipotency in the transplanted spinal cord for at least 12 wk,migrating to areas of axon growth cones.CONCLUSION Our data demonstrated that drNPC transplantation was safe and contributed to improvement of spinal cord function after acute SCI,based on neurological status assessment and neurophysiological recovery within 12 wk after transplantation.The functional improvement described was not associated with neuronal differentiation of the allogeneic drNPCs.Instead,directed drNPCs migration to the areas of active growth cone formation may provide exosome and paracrine trophic support,thereby further supporting the regeneration processes.
基金the National Natural Science Foundation of China,grants No.30772304,30973166,and 81171863
文摘Previous studies have shown that proliferation of endogenous neural precursor cells cannot alone compensate for the damage to neurons and axons. From the perspective of neural plastici- ty, we observed the effects of functional electrical stimulation treatment on endogenous neural precursor cell proliferation and expression of basic fibroblast growth factor and epidermal growth factor in the rat brain on the infarct side. Functional electrical stimulation was performed in rat models of acute middle cerebral artery occlusion. Simultaneously, we set up a placebo stimulation group and a sham-operated group. Immunohistochemical staining showed that, at 7 and 14 days, compared with the placebo group, the numbers of nestin （a neural precursor cell marker）-positive cells in the subgranular zone and subventricular zone were increased in the functional electrical stimulation treatment group. Western blot assays and reverse-transcription PCR showed that total protein levels and gene expression of epidermal growth factor and basic fibroblast growth factor were also upregulated on the infarct side. Prehensile traction test results showed that, at 14 days, prehension function of rats in the functional electrical stimulation group was significantly better than in the placebo group. These results suggest that functional electrical stimulation can promote endogenous neural precursor cell proliferation in the brains of acute cerebral infarction rats, enhance expression of basic fibroblast growth factor and epidermal growth factor, and improve the motor function of rats.
文摘BACKGROUND： Choosing proper donor cells is one of keys in experimental and clinical studies on cell replacement therapy （CRT） for treating Parkinson disease （PD）. Embryonic mesencephalic precursor cells （MPCs） can stably differentiate into dopaminergic neuron after in vitro proliferated culture. As compared with embryonic stem cell and neural stem cell strains, cell composition of embryonic MPCs after primary culture is also the most close to that of embryonic mesencephalic ventral cell suspension without proliferated culture. Successful experience accumulated in the latter suggests that primary cultured embryonic MPCs might be the most potential donor cells in clinical application with CRT for treating PD so far. OBJECTIVE： To investigate the feasibility of primary cultured embryonic precursor cells cultured primarily as donor cells in CRT for treating PD in rats. DESIGN ： A randomized and controlled trial taking SD rats as experimental animals.SETTING： Department of Neurosurgery, Huashan Hospital Affiliated to Fudan University.MATERIALS： This experiment was carried out at the Institute of Neuroscience, Shanghai Institute for Biological Science, Chinese Academy of Sciences from July 2003 to June 2004. Totally 26 female SD rats, with body mass of 200 to 220 g, were provided by Shanghai Experimental Animal Center of Chinese Academy of Sciences. METHODS ： Stereotaxic injection of 6-hydroxydopamine into the medial forebrain bundle were perfored to develop PD model rat. Among 26 SD rats, 20 rats achieved a more than 5 turns/min in apomorphine induced rotation test, reaching the standard of PD model rats. Immunohistochemical detection was performed on 1 out of 20 model rats after execution, and the other 19 rats were randomly divided into control group （n=5）, sham transplantation group （n=5）and cell grafted group （n=9）. Primary cultured E12 MPC cell suspension （1.2×10^11 L^-1）were used as donor cells. 4μL primary cultured E12 MPC cell suspension prepared freshly was injected into the lesioned corpus striatum of rats in cell grafted group, and 4μL D-Hank＇s solution was injected in sham transplantation group in the same way. There was no injection in control group. Apomorphine-induced rotation rate of PD rats were recorded respectively in cell grafted group and sham transplantation group pre-operation （initial value） and at postoperative 2, 4, 6 and 16 weeks. Apomorphine-induced rotation rate of PD rats was recorded in control group at postoperative 2 months （initial value） and following 2,4,6 and 16 weeks. To determine TH antigen with immunohistological ABC method （DAB developing） at 6 months post-transplantation to investigate the differentiation and survival of donor cells in the host body.MAIN OUTCOME MEASURES： Apomorphine-induced rotation behavior before and after transplantation and the survival and differentiation of implanted cells in the host body at 6 months post-transplantation. RESULTS： Among 19 model rats, one rat died after transplantation respectively in the cell grafted group and sham transplantation group; finally 17 model rats entered the stage of result analysis. Relative apomorphine-induced rotation rate was significantly decreased in the cell grafted group as compared with that before transplantation , with significant difference （P 〈 0.01 .P 〈 0.05）;the mean value of relative apomorphine-induced rotation rate was significantly decreased at postoperative 16 weeks in cell grafted group as compared with that of corresponding relative rotation rate in control group , also with significant difference （P 〈 0.05）.Immunohistological results showed that donor cells could differentiate into large and multi-polar dopaminergic neurons in the host body. CONCLUSION ： Primary cultured embryonic MPCs can be used as the donor cells in CRT for treating PD.
文摘BACKGROUND： Cerebral injury in adult mammals can induce neural precursor cells （NPCs） to proliferate and migrate towards the focal zone, but it is unclear whether endogenous NPCs can migrate towards regions distal to the hemorrhagic focus or whether NPCs differentiate in the peripheral hemorrhagic region. OBJECTIVE： To investigate the distribution of endogenous NPCs in different brain regions of rats with experimental cerebral hemorrhage, as well as NPC proliferation and differentiation with time. DESIGN, TIME AND SE＇B＇ING： A randomized, controlled animal experiment was performed at the Department of Neurobiology, Luzhou Medical College, between January 2007 and October 2008. MATERIALS： Bromodeoxyuridine （BrdU） was purchased from Roche, Germany. Mouse anti-rat BrdU monoclonal antibody, rabbit anti-nestin polyclonal antibody, rabbit anti-neuron specific enolase （NSE） polyclonal antibody were purchased from Wuhan Boster, China. Rabbit anti-glial fibrillary acidic protein （GFAP） polyclonal antibody was purchased from Sigma, USA. METHODS： Thirty-five adult Sprague Dawley rats were randomly divided into three groups： （1） cerebral hemorrhage group （n = 25）, rats were stereotaxically administered 50 p L autologous arterial blood via the dorsal caudate putamen to induce cerebral hemorrhage; （2） sham-surgery group （n = 5）, rats underwent surgery but did not receive blood injection; （3） blank control group （n = 5）, rats received no surgery and blood administration. At 2 hours after surgery, all rats were intraperitoneally administered BrdU. MAIN OUTCOME MEASURES： Distribution and proliferation of BrdU-positive cells were observed by immunohistochemical staining. BrdU-positive cell differentiation into neurons and glial cells in the peripheral hemorrhagic region was detected by double-label immunofluorescence. RESULTS： Immunohistochemistry results revealed that BrdU-positive cells existed not only in the peripheral hemorrhagic region, such as the subependymal layer and hippocampal dentate gyrus, but also in the lateral septal nucleus, diagonal band, habenular nucleus, and cerebral cortex. Following cerebral hemorrhage, BrdU-positive cells in the peripheral hemorrhagic region gradually increased （P 〈 0.05）, and peaked at 7 14 days. Double-label immunofluorescence showed that with time after cerebral hemorrhage, BrdU/nestin-positive cells decreased, but BrdU/GFAP- and BrdU/NSE-positive cells increased in the peripheral cerebral hemorrhagic region （P 〈 0.05）. CONCLUSION： Cerebral hemorrhage can induce the proliferation of endogenous NPCs, which peaks at 1-2 weeks after hemorrhage. NPCs can also migrate towards the regions distal to the hemorrhagic focus, such as a diagonal band or lateral septal nucleus. NPCs can gradually differentiate with increasing time after hemorrhage.
基金Research in the Swayne lab was supported by operating grants to LAS from the Natural Sciences and Engineering Research Council of Canada(NSERC Discovery Grant)the Canadian Institutes of Health Research(CIHR Grant MOP142215)+5 种基金The Scottish Rite Charitable Foundation of Canada and the University of Victoria Division of Medical Sciencesby infrastructure grants from the Canadian Foundation for Innovation(CFI)the British Columbia Knowledge Development Fund(BCKDF)supported by a Michael Smith Foundation for Health Research and British Columbia Schizophrenia Society Foundation Scholar Awardsupported by a University of Victoria Fellowship Graduate Awardsupported by a Vanier Canada Graduate Scholarship(NSERC)
文摘We recently reported that targeted deletion of Pannexin 1 in neural precursor cells of the ventricular zone impairs the maintenance of these cells in healthy and stroke-injured brain. Here we frame this exciting new finding in the context of our previous studies on Pannexin 1 in neural precursors as well as the close rela- tionship between Pannexin 1 and purinergic receptors established by other groups. Moreover, we identify important gaps in our understanding of Pannexin 1 in neural precursor cell biology in terms of the under- lying molecular mechanisms and functional/behavioural outcomes.
文摘Objective To investigate whether buffalo rat liver cell conditioned medium (BRL CM) can be used as the culture medium of embryonic stem (ES) cells, and to get relatively pure neural precursor cells (NPCs) for treatment aim. Methods Mouse ES cells were cultured in BRL CM and medium contain leukemia inhibitory factor (LIF), respectively. NPCs were selectively cultured in serum free medium. Alkaline phosphatase activity was visualized with NBT/BCIP and nestin antigen was detected with immunocytochemical methods. Results BRL CM could be used as an efficiency culture condition instead of LIF in ES cells culture. About 86% of cells derived from ES cells in the serum free culture were NPCs. Conclusion BRL CM can replace LIF to use in ES cell culture. High purity of NPC can be induced from ES cells with serum free culture method.
基金supported by the National Natural Science Foundation of China (81271248 and 81400933)
文摘Recent research has shown that defined sets of exogenous factors are sufficient to convert rodent and human somatic cells directly into induced neural stem cells or neural precursor cells（iNSCs/iNPCs）.The process of transdifferentiation bypasses the step of a pluripotent state and reduces the risk of tumorigenesis and genetic instability while retaining the self-renewing capacity.This iNSC/iNPC technology has fueled much excitement in regenerative medicine,as these cells can be differentiated into target cells for replacement therapy for neurodegenerative diseases.Patients＇ somatic cell-derived iNSCs/iNPCs have also been proposed to serve as disease models with potential value in both fundamental studies and clinical applications.This review focuses on the mechanisms,techniques,and applications of iNSCs/iNPCs from a series of related studies,as well as further efforts in designing novel strategies using iNSC/iNPC technology and its potential applications in neurodegenerative diseases.
文摘Human embryonic stem (hES) cells provide a promising supply of specific cell types for transplantation therapy. We presented here the method to induce differentiation of purified neural precursors from hES cells. hES cells (Line PKU-1 and Line PKU-2) were cultured in sus- pension in bacteriological Petri dishes, which differentiated into cystic embryoid bodies (EBs). The EBs were then cultured in N2 medium containing bFGF in poly-L-lysine-coated tissue culture dishes for two weeks. The central, small cells with 2―3 short processes of the spreading out- growth were isolated mechanically and replated. The resulting neurospheres were cultured in suspension for 10 days, then dissociated into single cell suspension with a Pasteur pipette and plated. Cells grew vigorously in an attached way and were passed every 4―5 days. Almost all the cells were proved nestin positive by immunostaining. Following withdrawal of bFGF, they differentiated into neurons expressing β-tubulin isotypeIII, GABA, serotonin and synaptophysin. Through induction of PDGF-AA, they differentiated into astrocytes expressing GFAP and oli- godendrocytes expressing O4. The results showed that hES cells can differentiate into typical neural precursors expressing the specific marker nestin and capable of generating all three cell types of the central nervous system (CNS) in vitro.
基金Dr.Hui Fu was supported by the National Natural Science Foundation of China,No.81371338by Open Research Fund Program of Hubei-MOST KLOS & KLOBMEDr.Zu-neng Lu was supported by grants from Health and Family Planning Commission of Hubei Province scientific research project,No.WJ2015MA007
文摘There are few studies on the membrane protein Ankfyl. We have found Ankfyl is specifically expressed in neural stem/precursor cells during early development in mice （murine）. To further explore Ankfyl function in neural development, we developed a gene knockout mouse with a mixed Balb/C and C57/BL6 genetic background. Using immunofluorescence and in situ hybridization, neural defects were absent in mixed genetic Ankfyl null mice during development and in adults up to 2 months old. However, Ankfyl gene knockout mice with a pure genetic background were found to be lethal in the C57/BL6 inbred mice embryos, even after seven generations of backcrossing. Polymerase chain reaction confirmed homozygotes were unattainable as early as embryonic day 11.5. We conclude that Ankfyl protein is dispensable in neural stem/precursor ceils, but could be critical for early embryonic murine development, depending on the genetic background.
文摘Artificial neural network (NN) is such a model as to imitate the structure and intelligence feature of human brain. It has strong nonlinear mapping function. To introduce NN into the study of earthquake prediction is not only an extension of the application of artificial neural network model but also a new try for precursor observation to serve the earthquake prediction. In this paper, we analyzed the predictability of time series and gave a method of application of artificial neural network in forecasting earthquake precursor chaotic time series. Besides, taking the ground tilt observation of Jiangning and Xuzhou Station, the bulk strain observation of Liyang station as examples, we analyzed and forecasted their time series respectively. It is indicated that the precision of this method can meet the needs of practical task and therefore of great value in the application to the future practical earthquake analysis and prediction.
基金supported by the National Institutes of Health,Nos.AA025919,AA025919-03S1,and AA025919-05S1(all to RAF).
文摘Hippocampal neuronal loss causes cognitive dysfunction in Alzheimer’s disease.Adult hippocampal neurogenesis is reduced in patients with Alzheimer’s disease.Exercise stimulates adult hippocampal neurogenesis in rodents and improves memory and slows cognitive decline in patients with Alzheimer’s disease.However,the molecular pathways for exercise-induced adult hippocampal neurogenesis and improved cognition in Alzheimer’s disease are poorly understood.Recently,regulator of G protein signaling 6(RGS6)was identified as the mediator of voluntary running-induced adult hippocampal neurogenesis in mice.Here,we generated novel RGS6fl/fl;APP_(SWE) mice and used retroviral approaches to examine the impact of RGS6 deletion from dentate gyrus neuronal progenitor cells on voluntary running-induced adult hippocampal neurogenesis and cognition in an amyloid-based Alzheimer’s disease mouse model.We found that voluntary running in APP_(SWE) mice restored their hippocampal cognitive impairments to that of control mice.This cognitive rescue was abolished by RGS6 deletion in dentate gyrus neuronal progenitor cells,which also abolished running-mediated increases in adult hippocampal neurogenesis.Adult hippocampal neurogenesis was reduced in sedentary APP_(SWE) mice versus control mice,with basal adult hippocampal neurogenesis reduced by RGS6 deletion in dentate gyrus neural precursor cells.RGS6 was expressed in neurons within the dentate gyrus of patients with Alzheimer’s disease with significant loss of these RGS6-expressing neurons.Thus,RGS6 mediated voluntary running-induced rescue of impaired cognition and adult hippocampal neurogenesis in APP_(SWE) mice,identifying RGS6 in dentate gyrus neural precursor cells as a possible therapeutic target in Alzheimer’s disease.
基金supported by the National Natural Science Foundation of China(No.81700471 and No.82270583)the National Key Research and Development Program of China(No.2022YFC2504005)。
文摘Objective:Diabetes-induced gastrointestinal(GI)motility disorders are increasingly prevalent.Damage to the enteric nervous system(ENS),composed primarily of enteric neurons and glial cells,is an essential mechanism involved in these disorders.Although electroacupuncture(EA)has shown the potential to mitigate enteric neuronal loss,its mechanism is not fully understood.Additionally,the effects of EA on enteric glial cells have not been investigated.Enteric neural precursor cells(ENPCs)contribute to the structural and functional integrity of the ENS,yet whether EA enhances their differentiation into enteric neurons and glial cells remains unexplored.This study investigates whether EA promotes ENS repair through enhancing ENPC-derived neurogenesis and gliogenesis and elucidates the potential molecular mechanisms involved.Methods:Transgenic mice were used to trace Nestin+/nerve growth factor receptor(Ngfr)+ENPCs labeled with green fluorescent protein(GFP)in vivo.Mice were randomly divided into four groups:control,diabetes mellitus(DM),DM+sham EA,and DM+EA.The effects of EA on diabetic mice were evaluated by GI motility,ENS structure,and ENPC differentiation.Glial cell line-derived neurotrophic factor(GDNF)/Ret signaling was detected to clarify the underlying molecular mechanisms.Results:EA alleviated diabetes-induced GI motility disorders,as indicated by reduced whole gut transit time,shortened colonic bead expulsion time,and enhanced smooth muscle contractility.Furthermore,EA attenuated diabetes-induced losses of enteric neurons and glial cells,thereby restoring ENS integrity.Notably,EA reversed the diabetes-induced decrease in ENPCs and significantly increased the absolute number and the proportion of ENPC-derived enteric neurons.However,immunofluorescence analyses revealed no colocalization between EA-induced glial fibrillary acidic protein+glial cells and GFPlabeled ENPCs.Mechanistically,GDNF/Ret signaling was elevated in intestinal tissues and upregulated in ENPCs in EA-treated diabetic mice.Conclusion:EA facilitates ENS repair by promoting Nestin+/Ngfr+ENPC differentiation into enteric neurons via upregulation of GDNF/Ret signaling,and driving enteric gliogenesis from non-Nestin+/Ngfr+ENPCs.These findings highlight EA's role in ameliorating diabetes-induced GI dysmotility through ENPC-derived ENS restoration.
基金supported by the National Research Foundation of Korea Grant funded by the Korean Government,No.NRF-013-2011-1-E00045
文摘Endogenous neural stem cells become ＂activated＂ after neuronal injury, but the activation sequence and fate of endogenous neural stem cells in focal cerebral ischemia model are little known. We evaluated the relationships between neural stem cells and hypoxia-inducible factor-1α and vascular endothelial growth factor expression in a photothromobotic rat stroke model using immunohistochemistry and western blot analysis. We also evaluated the chronological changes of neural stem cells by 5-bromo-2′-deoxyuridine（BrdU） incorporation. Hypoxia-inducible factor-1α expression was initially increased from 1 hour after ischemic injury, followed by vascular endothelial growth factor expression. Hypoxia-inducible factor-1α immunoreactivity was detected in the ipsilateral cortical neurons of the infarct core and peri-infarct area. Vascular endothelial growth factor immunoreactivity was detected in bilateral cortex, but ipsilateral cortex staining intensity and numbers were greater than the contralateral cortex. Vascular endothelial growth factor immunoreactive cells were easily found along the peri-infarct area 12 hours after focal cerebral ischemia. The expression of nestin increased throughout the microvasculature in the ischemic core and the peri-infarct area in all experimental rats after 24 hours of ischemic injury. Nestin immunoreactivity increased in the subventricular zone during 12 hours to 3 days, and prominently increased in the ipsilateral cortex between 3–7 days. Nestin-labeled cells showed dual differentiation with microvessels near the infarct core and reactive astrocytes in the peri-infarct area. BrdU-labeled cells were increased gradually from day 1 in the ipsilateral subventricular zone and cortex, and numerous BrdU-labeled cells were observed in the peri-infarct area and non-lesioned cortex at 3 days. BrdU-labeled cells rather than neurons, were mainly co-labeled with nestin and GFAP. Early expressions of hypoxia-inducible factor-1α and vascular endothelial growth factor after ischemia made up the microenvironment to increase the neuronal plasticity of activated endogenous neural stem cells. Moreover, neural precursor cells after large-scale cortical injury could be recruited from the cortex nearby infarct core and subventricular zone.
