Characterization of SH-SY5Y neurons subject to 92kHz ultrasound stimulation / Caracterización de neuronas SH-SY5Y sujetas a estimulación por ultrasonido de 92 kHz

SUMMARY: Cellular microstructural changes due to ultrasound exposure are critical to understand and characterize in order to further the establishment of ultrasonics in cell and tissue engineering and medicine. In this study, neurite length, nuclear morphology, and cellular toxicity are assessed at varying intensities of 92 kHz ultrasound provided by a piezoceramic disk element and incident upon SH- SY5Y neurons in vitro. Findings suggest that stimulation increases neurite length up to 2.73 fold tested at α = 0.05 in an intensity dependent manner. Additionally, stimulation causes a statistically significant (α = 0.05) decrease in nuclear area and less elongated nuclei, by 1.78 fold and 1.38 fold respectively, also in an intensity dependent manner. For maximum transducer surface intensities ranging from 0 to 39.11 W/cm2, the toxicity of 92 kHz ultrasound is assessed and a nontoxic range is determined using Caspase-3 and Annexin V staining, in addition to Calcium imaging via Calcein-AM staining. Intensities of up to 1.6 W/cm2 are found to be nontoxic for the cells under the parameters used in this study.

RESUMEN: Los cambios micro estructurales celulares debidos a la exposición a los ultrasonidos son fundamentales para comprender y caracterizar el establecimiento de los ultrasonidos en la ingeniería y la medicina de células y tejidos. En este estudio, la longitud de las neuritas, la morfología nuclear y la toxicidad celular se evalúan a intensidades variables de ultrasonido de 92 kHz proporcionado por un elemento de disco piezocerámico e incidente sobre las neuronas SH-SY5Y in vitro. Los resultados sugieren que la estimulación aumenta la longitud de las neuritas hasta 2,73 veces probada a α = 0,05 de una manera dependiente de la intensidad. Además, la estimulación provoca una disminución estadísticamente significativa (α = 0,05) en el área nuclear y núcleos menos alargados, en 1,78 veces y 1,38 veces, respectivamente y también de una manera dependiente de la intensidad. Para intensidades máximas de la superficie del transductor que oscilan entre 0 y 39,11 W / cm2, se evaluó la toxicidad del ultrasonido de 92 kHz y se determinó un rango no tóxico mediante tinción con Caspasa-3 y Anexina V, además de imágenes de calcio mediante tinción con Calceína-AM. Se encontró que las intensidades de hasta 1.6 W / cm2 no son tóxicas para las células bajo los parámetros usados en este estudio.

Graded and pan-neural disease phenotypes of Rett Syndrome linked with dosage of functional MeCP2

Rett syndrome (RTT) is a progressive neurodevelopmental disorder, mainly caused by mutations in MeCP2 and currently with no cure. We report here that neurons from R106W MeCP2 RTT human iPSCs as well as human embryonic stem cells after MeCP2 knockdown exhibit consistent and long-lasting impairment in maturation as indicated by impaired action potentials and passive membrane properties as well as reduced soma size and spine density. Moreover, RTT-inherent defects in neuronal maturation could be pan-neuronal and occurred in neurons with both dorsal and ventral forebrain features. Knockdown of MeCP2 led to more severe neuronal deficits as compared to RTT iPSC-derived neurons, which appeared to retain partial function. Strikingly, consistent deficits in nuclear size, dendritic complexity and circuitry-dependent spontaneous postsynaptic currents could only be observed in MeCP2 knockdown neurons but not RTT iPSC-derived neurons. Both neuron-intrinsic and circuitry-dependent deficits of MeCP2-deficient neurons could be fully or partially rescued by re-expression of wild type or T158M MeCP2, strengthening the dosage dependency of MeCP2 on disease phenotypes and also the partial function of the mutant. Our findings thus reveal stable neuronal maturation deficits and unexpectedly, graded sensitivities of neuron-inherent and neural transmission phenotypes towards the extent of MeCP2 deficiency, which is informative for future therapeutic development.

Restorative Mechanism of Neural Progenitor Cells Overexpressing Arginine Decarboxylase Genes Following Ischemic Injury

Cell replacement therapy using neural progenitor cells (NPCs) following ischemic stroke is a promising potential therapeutic strategy, but lacks efficacy for human central nervous system (CNS) therapeutics. In a previous in vitro study, we reported that the overexpression of human arginine decarboxylase (ADC) genes by a retroviral plasmid vector promoted the neuronal differentiation of mouse NPCs. In the present study, we focused on the cellular mechanism underlying cell proliferation and differentiation following ischemic injury, and the therapeutic feasibility of NPCs overexpressing ADC genes (ADC-NPCs) following ischemic stroke. To mimic cerebral ischemia in vitro , we subjected the NPCs to oxygen-glucose deprivation (OGD). The overexpressing ADC-NPCs were differentiated by neural lineage, which was related to excessive intracellular calcium-mediated cell cycle arrest and phosphorylation in the ERK1/2, CREB, and STAT1 signaling cascade following ischemic injury. Moreover, the ADC-NPCs were able to resist mitochondrial membrane potential collapse in the increasingly excessive intracellular calcium environment. Subsequently, transplanted ADC-NPCs suppressed infarct volume, and promoted neural differentiation, synapse formation, and motor behavior performance in an in vivo tMCAO rat model. The results suggest that ADC-NPCs are potentially useful for cell replacement therapy following ischemic stroke.

LncRNA Neat1 is upregulated in P19 cell line processed with all-trans-retinoic acid / 基础医学与临床

Objective To investigate the expression of Neat1 in the differentiation of P19 cells induced by all-trans-retinoic acid(atRA) and to explore the effect of histone modification on its expression. Methods The differentiation of P19 cells was induced with the α-MEM culture media containing 0.5 μmol/L all-trans-retinoic( atRA) acid and the expression Mash1 which is the neural differentiation maker gene was measured. The expression of Neat1 was measured with RT-qPCR. The cells were treated with TSA, NAM or AdOx respectively to investigate the effect of the histone modifier inhibitor on the expression of Neat1. Results The model of differentiation of P19 cells induced by atRA was successfully constructed. Mash1 was significantly upregulated in the process of P19 cells differentiation.Neat1 was significantly upregulated with the induction of P19 cells treated with atRA(P＜0.01). The TSA but not the NAM or AdOx could induce the expression of the Neat1.Conclusions The expression of Neat1 is significantly upregulated in the process of P19 differentiation induced by atRA and the the high level of histone acetylation maintained by TSA potentially induce the expression of Neat1.

Comparison of Inducing Pluripotent Stem Cells to Differentiate into Motor Neuron Precursor in Two Kinds of System / 中国康复理论与实践

Objective To induce human-induced pluripotent stem cells(iPSCs)to differentiate into spinal motor neuron precursor (MNP)and compare the induction efficiency in systems of feeder and feeder-free. Methods iPSCs cultured on mouse feeder cells or in feeder-free condition were induced into neuroepithelial progenitors (NEP) on the sixth day and MNP on the twelveth day.Their morphology was observed under inverted micro-scope,and the markers of iPSCs,NEP,MNP were detected with immunofluorescence.NEP-related genes SOX1 and HOXA3,MNP-related genes OLIG2 and PAX6,and pluripotency genes SOX2 and OCT4 were detected with real-time quantitative polymerase chain reaction. Results iPSCs expressed pluripotency markers,while NEP and MNP expressed high levels of neural related markers and low levels of pluripotency markers in two systems. The expression of the genes SOX1, HOXA3, OLIG2 and OCT4 was higher in the feeder system,and there was no significant difference in the expression of genes SOX2 and PA X 6. Conclusion iPSCs can differentiate into MNP in culture systems of feeder and feeder-free,and the induction efficiency is higher in the feeder system.

Cholinergic signal regulates neural stem cell differentiation in perilesional zone after ischemic stroke / 实用医学杂志

Objective To investigate the effects of cholinergic signal on neural stem cell(NSC)differenti-ation in peri-infarction region after ischemic stroke. Methods Mice were randomly assigned into sham + vehicle group,middle cerebral artery occlusion(MCAO)+ vehicle group,MCAO + donepezil group and MCAO + atro-pine group(n = 25). MCAO was induced by thread-occlusion method. Modified neurological severity score (mNSS)was used to evaluate neurological function recovery,and the brain water content was measured by dry-wet weight method. NeuN/5-bromodeoxyuridine(BrdU),CNPase/BrdU,GFAP/BrdU double-labeled cells were tested by immunofluorescence. Results Brain water content of MCAO + vehicle group was significantly higher than that of sham operation group(P < 0.05). Donepezil-treated MCAO mice had lower neurologic deficit scores and brain water content than of MCAO + vehicle group(P < 0.05). On day 14 and day 28 after MCAO,the NeuN/BrdU, CNPase/BrdU and GFAP/BrdU immune-positive cells of MCAO + vehicle group were markedly increased as com-pared with that of sham+vehicle group(P<0.05).Compared with that of MCAO+vehicle group,the number of NeuN/BrdU-positive cells,CNPase/BrdU-positive cells and GFAP/BrdU-positive cells was higher in MCAO+done-pezil group,and the number of NeuN/BrdU-positive cells and CNPase/BrdU-positive cells of MCAO + atropine group was lower(P < 0.05). Conclusions Cholinergic signal could promote NSCs differentiation in peri-infarc-tion region,a lleviate cerebral edema,and improve the brain function restoration after stroke.

Generation of induced pluripotent stem cells and neural cells from a patient with critical illness polyneuropathy / 中华急诊医学杂志

Objective To establish disease-associated or cell type relevant neuron model generated from patients with Critical Illness Polyneuropathy (CIP) by making CIP patient-derived induced pluripotent stem (iPS) cell lines and neurons to provide a cell-based disease model of CIP.Methods Skin tissue of CIP patient was obtained clinically,and specific skin fibroblasts were isolated and cultured.The iPS cells were derived from CIP patient by introducing 4 transcription factors,namely Oct4,Klf4,Sox2,c-Myc,into patient-specific fibroblast cells by Millipore's Human STEMCCA Constitutive Polycistronic (OKSM) Lentivirus Kit.Colony morphology,alkaline phosphatase (AP) activity,immunofluorescence staining,quantitative reverse transcription polymerase chain reaction (RT-PCR),and differentiation ability were used to identify the pluripotency of these iPS cell lines.In addition,neurons were derived from these iPS cells by inhibiting SMAD pathway.Results The CIP-iPS cells presenting morphological and growth characteristics of human embryonic stem cell (hES) showed the presence of alkaline phosphatase detected by histochemical staining,and the expression of ESC-marker genes.The relative expressions of endogenous pluripotency genes,namely Sox2,REX1,NANOG and OCT4,in iPS cell lines were significantly increased compared with their primary fibroblasts (t values were-9.020,-10.753,-13.295,-12.677,P＜0.01).Subcutaneous injection of iPS cells into NOD-SCID mice resulted in teratomas containing tissues from all the 3 germ layers.Furthermore,cholinergic neurons were successfully induced from CIP-iPS cells.Conclusion The CIP patient-specific iPS cell line and cholinergic neurons were successfully established.Furthermore,the CIP-iPS cell line can be used as models for further elucidating the cellular pathology and developing therapeutic strategies for Critical Illness Polyneuropathy.

Metodologías utilizadas en la diferenciación de células madre mesenquimales a linaje neuronal / Methodologies used in the differentiation of mesenchymal stem cells to neuronal lineage

RESUMEN INTRODUCCIÓN: Las células madre mesenquimales son células con la capacidad de autorrenovarse y diferenciarse a linajes mesenquimales e inclusive a células de origen no mesenquimal, como las del tejido nervioso. Teniendo en cuenta la idoneidad de estas células para diferenciarse en neuronas, han sido utilizadas con propósitos terapéuticos debido a que son capaces de restaurar neuronas que se deterioran en diversas enfermedades neurodegenerativas. OBJETIVO: Informar y comparar los métodos más utilizados para inducir a las células madre mesenquimales a diferenciarse en neuronas, además de mencionar las ventajas y desventajas de cada una de estas. METODOLOGÍA: La primera metodología para inducir a la diferenciación neural fue utilizada en 1999 y a partir de ese momento, se han empleado compuestos químicos, factores de crecimiento, compuestos sintéticos, entre otros métodos para diferenciar este tipo de células a linaje neuronal. El problema radica en que algunas de estas metodologías son tóxicas para las células, costosos o presentan otro tipo de efecto colateral. CONCLUSIÓN: La elección de uno de estos métodos depende de los intereses y las condiciones con las que cuente cada investigador. Además, es indispensable conocer las falencias que tenemos en este campo de la investigación con el propósito de continuar con la búsqueda de alternativas que no tengan desventajas, si no por el contario, reúna todas las ventajas de los métodos aquí mencionados.

SUMMARY INTRODUCTION: Mesenchymal stem cells are cells with the ability to self-renew and differentiate into mesen-chymal lineages and even cells of non-mesenchymal origin, such as those of nerve tissue. Taking into account the suitability of these cells to differentiate into neurons, they have been used for therapeutic purposes since they are able to restore neurons that deteriorate in various neurodegenerative diseases. OBJECTIVE: To inform and to compare the methods most used to induce mesenchymal stem cells to differentiate into neurons and to mention the advantages and disadvantages of each of these. DEVELOPMENT: The first methodology to induce neural differentiation was used in 1999 and since then, chemical compounds, growth factors, synthetic compounds have been used, among other methods to differentiate this type of cells into neuronal lineage. The problem is that some of these methodologies are toxic to cells, expensive or have other side effects. CONCLUSION: The choice of one of these methods depends on the interests and the conditions that each researcher has. In addition, it is indispensable to know the shortcomings that we have in this field of research with the purpose of continuing with the search for alternatives that do not have disadvantages, if not by the contrary, gather all the advantages of the methods mentioned here.

Differentiation of Human Umbilical Cord Mesenchymal Stem Cells into Hair-cell Like Cells in Vitro / 听力学及言语疾病杂志

Objective To induce human umbilical cord mesenchymal stem cells (hUC-MSCs) to hair-cell like cells in the inner ear, using a two-step neural differentiation method.Methods The hUC-MSCs were obtained from human umbilical cords by tissue adherence culture,whose surface antigen CD29, CD34, CD44, CD45, CD90, HLA-ABC, and HLA-DR could be identified by flow cytometry.In the neural stem cells induced phase, the NSE positive cells were analyzed by microscope and immunohistochemistry.In the second stage, the expression of hair-cell like cells markers (Math1, MyosinⅦa, Brn3c) were tested by qRT-PCR and immunofluorescence method.Results The control group and the protocol group had little NSE after differentiation while the protocol B group presented a neurobiological structure and demonstrated a higher NSE positive ratio after 5 days' neural stem cells induction (P<0.05).Compared to the control group, the mRNA and protein level of Math1, MyosinⅦa, and Brn3c exhibited a significant increase in the differential group,which induced for 4 weeks in the hair-cell like cells in the inner ear's induced phase(P<0.05).Conclusion The two-stage induction (hUC-MSCs-neural stem cells-hair-cell like cells) could produce more MyosinⅦa,Brn3c and Math1,which may provide an appropriate way to treat sensorineural deafness.

Protective Effects of Hydroxysafflor Yellow A against Oxidative Damage of β-Mercaptoethanol During Neural Differentiation of Mesenchymal Stem Cells / 中草药·英文版

Objective To study the protective effects of hydroxysafflor yellow A (HSYA) against the oxidative damage caused by β-mercaptoethanol (BME) during neural differentiation of mesenchymal stem cells (MSCs) in vitro. Methods When the confluence reached 50%-60%, 4th passage MSCs were divided into three groups to culture. G1: normal group which was cultured using basic medium (DMEM containing 10% FBS all the time); G2: unprotected group which was continuously cultured using basic medium for 24 h, and then cultured using pre-induction medium (DMEM containing 10% FBS and 1 mmol/L BME); G3: protected group which was firstly cultured using protective medium (DMEM containing 10% FBS and 160 mg/L HSYA) for 24 h, and then cultured using pre-induction medium for 24 h. After these treatments as above, cell viability, relative levels of SOD/GSH and apoptosis rate were respectively detected. The expression of Bcl and Bax was examined by Western blotting. After HSYA protection and BME pre-induction, neural induction was performed. The expression of NSE and MAP-2 was respectively analyzed on cellular and molecular levels. Results Compared with unprotected group, 160 mg/L HSYA could obviously improve cells viability, maintain high level of SOD and GSH in MSCs, reduce apoptosis rate and improve the ratio of Bcl/Bax. After protection with 160 mg/L HSYA, the survival time of neuron-like cells could be extended. Immunocytochemical staining showed that after 10 h of neural induction, the differentiated neuron-like cells in protected group were still in a good state, and the mRNA levels of NSE and MAP-2 were increased during the induction course checked. Conclusion HSYA could improve the resistance of cells to the oxidative damage caused by BME.

Isolation and Characterization of Human Chorionic Membranes Mesenchymal Stem Cells and Their Neural Differentiation

Mesenchymal stem cells (MSCs) can be obtained from a variety of human tissues. Placenta has become an attractive stem cell source for potential applications in regenerative medicine and tissue engineering. The aim of this study was to localize and characterize MSCs within human chorionic membranes (hCMSCs). For this purpose, immunofluorescence labeling with CD105 and CD90 were used to determine the distribution of MSCs in chorionic membranes tissue. A medium supplemented with a synthetic serum and various concentrations of neurotrophic factors and cytokines was used to induce hCMSCs to neural cells. The results showed that the CD90 positive cells were scattered in the chorionic membranes tissue, and the CD105 positive cells were mostly located around the small blood vessels. hCMSCs expressed typical mesenchymal markers (CD73, CD90, CD105, CD44 and CD166) but not hematopoietic markers (CD45, CD34) and HLA-DR. hCMSCs differentiated into adipocytes, osteocytes, chondrocytes, and neuronal cells, as revealed by morphological changes, cell staining, immunofluorescence analyses, and RT-PCR showing the tissue-specific gene presence for differentiated cell lineages after the treatment with induce medium. Human chorionic membranes may be the source of MSCs for treatment of nervous system injury.

Fetal Cerebrospinal Fluid Promotes Proliferation and Neural Differentiation of Stromal Mesenchymal Stem Cells Derived from Bone Marrow

ABSTRACT Embryonic cerebrospinal fluid (E-CSF) contains many neurotrophic and growth factors, acts as a growth medium for cortical progenitors, and can modulate proliferation and differentiation of neural stem cells. Mesenchymal stem cells (MSCs) are multipotential stem cells that can differentiate into several types of mesenchymal cells as well as nonmesenchymal cells, such as neural cells. In the present study, the effect of E-CSF on proliferation and neural differentiation of bone marrow mesenchymal stem cells (BM-MSCs) was investigated to test whether E-CSF is capable of driving these cells down the neuronal line. To verify the multipotential characteristics of BM-MSCs, the cells were analyzed for their osteogenic and adipogenic potential. Expression of the neural markers, MAP-2 and β-III tubulin, was determined by Immunocytochemistry. BM-MSCs differentiate into neuronal cell types when exposed to b-FGF. BMMSCs cells cultured in medium supplemented with CSF showed significantly elevated proliferation relative to control cells in media alone. E-CSF (E17-E19) supports viability and stimulates proliferation and, significantly, neurogenic differentiation of BM-MSCs. The data presented support an important role for CSF components, specifically neurotrophic factors, in stem cell survival, proliferation and neuronal differentiation. It is crucial to understand this control by CSF to ensure success in neural stem cell therapies.

Zinc finger protein 521 overexpression increased transcript levels of Fndc5 in mouse embryonic stem cells.

Zinc finger protein 521 is highly expressed in brain, neural stem cells and early progenitors of the human hematopoietic cells. Zfp521 triggers the cascade of neurogenesis inmouse embryonic stemcells through inducing expression of the early neuroectodermal genes Sox1, Sox3 and Pax6. Fndc5, a precursor of Irisin has inducing effects on the expression level of brain derived neurotrophic factor in hippocampus. Therefore, it is most likely that Fndc5 may play an important role in neural differentiation. To exhibit whether the expression of this protein is under regulation with Zfp521, we overexpressed Zfp521 in a stable transformants of mESCs expressing EGFP under control of Fndc5 promoter. Increased expression of Zfp521 enhanced transcription levels of both EGFP and endogenous Fndc5. This result was confirmed by overexpression the aforementioned vectors in HEK cells and indicated that Zfp521 functions upstream of Fndc5 expression. It is most likely that Zfp521 may act through the binding to its response element on Fndc5 core promoter. Therefore it is concluding that an enhanced expression of Fndc5 in neural progenitor cells is stimulated by Zfp521 overexpression in these cells.

Cell Cycle Synchronization Methods of Bone Marrow Mesenchymal Stem Cells and Its Effect on Neural Differentiation / 中国康复理论与实践

@#Objective To analyze the effect of different treatment conditions on cells synchronization in G0/G1 phase to get the best con-dition, and to explore its effect on neural differentiation of bone marrow mesenchymal stem cells (BMSCs) induced by basic fibroblast growth factor (bFGF). Methods BMSCs were isolated and cultured in 5%, 1%, 0.5%, 0.1%, 0 fetal bovine serum (FBS) respectively, for 24 hours and 48 hours. After PI staining, cell cycle proportions of each phase were detected by flow cytometry, and were compared with the normal group (10%FBS). After the optimal treatment condition was got, 20 ng/ml bFGF was added into synchronization group and unsyn-chronization group 3 days and 7 days, respectively. The expression of Nestin and Tuj-1 were detected with immunofluorescence. Results Adult rat BMSCs were isolated from bone marrow and cultured, after passage, the cells were with long spindle shape. Compared with the normal group, the cell proportion of G1/G0 phase increased under different treatments, peaked with (94.274 ± 0.468)%under 1%FBS, 48 hours (F=39.91, P<0.001). After bFGF induction for 3 days, the Nestin+cell number was higher in the synchronization group than in the un-synchronization group [(80.3 ± 2.4)%vs. (12.1 ± 1.5)%] (F=28.25, P<0.001). After bFGF induction for 7 days, the Tuj-1+cell number was higher in the synchronization group than in the unsynchronization group [(74.8±3.2%)%vs. (19.3±2.5)%] (F=17.95, P<0.001). Conclusion 1%FBS, 48 hours is the optimal condition to BMSCs synchronization in G0/G1 phase, which can promote the neural differentiation of BM-SCs.

Effects of Transfection of Mash-1 Gene on Neural Differentiation of Embryonic Stem Cell in Spinal Cord Injury Mice / 中国康复理论与实践

Objective To investigate the effects of overexpression of Mash-1 gene on functional recovery and neural differentiation of embryonic stem cells in spinal cord injury mice. Methods CE3 cell line with overexpression of Mash-1 gene was generated with murine stem cell virus. Spinal cord injury model was established with forceps compression in 4-week-old KM mice. Normal saline (model group, n=12), CE3 cells with or without overexpression Mash-1 gene (CE3-Mash-1 and CE3 groups, n=12 respectively) were transplanted into the ar-eas of injury 3 days after injury. They were assessed with the Basso Mouse Scale (BMS) 1, 7, 14, 21, and 28 days after injury. 6 mice from each group were sacrificed 14 and 28 days after injury respectively. The spinal cord area remained were observed with HE stained, and the expression of Oct3/4, nestin,β-tubulin III and glial fibrillary acidic protein (GFAP) were detected with immunofluorescence in the injured spinal cord in the CE3 and CE3-Mash-1 groups. Results The score of BMS significantly improved in CE3 and CE3-Mash-1 groups com-pared with that of the model group (F>84.471, P49.990, P0.05). Conclusion Overexpression of Mash-1 gene promotes CE3 cells to differentiate into neurons in spinal cord injury mice, and improve the motor function recovery.

Generation of induced pluripotent stem cells and neural cells from urine-derived cells of Alzheimer disease patients / 中国病理生理杂志

[ ABSTRACT] AIM:In this study, we aim to obtain the induced pluripotent stem cells ( iPSCs) from the patients with sporadic Alzheimer disease ( AD) .METHODS:Three typical Alzheimer’ s patients were chosen, and the epithelial cells were isolated from their urine.We reprogrammed these cells into induced pluripotent stem cells by transfection of 4 factors (Oct4, Sox2, Klf4 and SV40LT) with the technique of electro-transfection.After getting these iPSCs, we continue to differentiate them into neural cells by a specific method—dual inhibition of Smad signaling.RESULTS: The primary cells from 3 AD patients were successfully reprogrammed to iPSCs, and these patients-derived iPSCs were differentiated into neural cells.There was no significant difference, during iPSCs reprogramming and neural differentiation, between cells from AD patients and normal people.CONCLUSION: The urine cells from AD patients were able to transfer to iPSCs, functional neurons and neurogliocytes.

Meta-analysis of the relationship of NeuroD1 Ala45Thr polymorphism with type 2 diabetes / 中华内分泌代谢杂志

Objective To investigate the association of neural differentiation factor 1 (NeuroD1) Ala45Thr polymorphism with type 2 diabetes by meta-analysis.Methods The electronic databases were retrieved from PubMed,EMBASE,CNKI,VIP fulltext database,and Wanfang fulltext database.Literatures about the association of NeuroD1 Ala45Thr polymorphism with type 2 diabetes from 2004 to 2012 were searched.The selected studies from 2004 to 2012 were included to analyze data based on the published meta-analysis of the NeuroD1 (Ala45Thr) gene polymorphism and type 2 diabetes written by Kavvoura,et al.The odds ratio of NeuroD1 (Ala45Thr) genotype distributions in type 2 diabetic patients compared with healthy control were analyzed.Rev-Man 5.0 software was applied for investigating hereogeneity among individual studies and summarizing effects with proper statistical methods.Thirteen case control studies were enrolled.Results A total of 3 896 patients and 3 186 control subjects were enrolled for the study.The pooled ORs of Thr45Thr45/(Ala45Thr+ Ala45Ala45) genotype in type 2 diabetes mellitus and control groups were 3.16 (95％ CI0.99-10.11) in the subgroup of yellow race and 1.09(95％ CI0.90-1.32) in the subgroup of white race,with no statistically significant difference(P＞0.05).The pooled OR of G/A allele in type 2 diabetes mellitus and control groups was 0.85 (95 ％ CI 0.72-0.99)/1.18 (95％ CI 1.07-1.38),with statistically significant difference (Z =2.02,P =0.04).Conclusions The A allele of NeuroD1 (Ala45Thr) locus may be a risk factor for type 2 diabetes mellitus.

Basic Fibroblast Growth Factor-Chitosan Carriers Induce Neural Stem Cells to Differentiate into Neurons and Form Synapses / 中国康复理论与实践

Objective To explore the effect of basic fibroblast growth factor (bFGF)-chitosan carriers on neural differentiation of neural stem cells (NSCs). Methods NSCs were isolated from spinal cord of a neonatal Wistar rat and cultured. Purity of cultured NSCs was identi-fied with Nestin immunofluorescent staining. The 10 mg/ml chitosan carriers, 20 ng/ml bFGF or 10 mg/ml bFGF-chitosan carriers were add-ed into medium of P3~P4 NSCs respectively. NSCs were observed with immunofluorescent staining: 3 days after incubation with Nestin andβ-tubulin III;7 days after incubation with microtubule-associated protein-2 (MAP2), glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP);and 14 days after incubation with synapsin-1 and MAP2. The electrophysiological activity of cells was detected with MED64. Results 3 days after incubation, all the NSCs differentiated into Nestin+/β-tubulin III+, and the length of neurofilament was the high-est in those co-cultured with bFGF-chitosan carriers. 7 days after incubation, NSCs differentiated into MAP2+, GFAP+and MBP+, and more NSCs differentiated into MAP2+with bFGF-chitosan carriers. 14 days after incubation, NSCs differentiated with bFGF-chitosan carriers ex-press synapsin-1+/MAP2+and showed electrophysiological activity. Conclusion bFGF-chitosan carriers can induce NSCs to differentiate in-to neuron with high percentage and the differentiated neurons can form synapses with electrophysiology activity.

Experimental study on repairing spinal cord injury by human umbilical cord blood CD34+cell transplantation at different time points / 天津医药

Objective To investigate the effect of transplantation of human umbilical cord blood CD34+cells on spinal cord injury. Methods CD34+cells were separated from fresh human umbilical cord blood by magnetic cell sorting. Ninety-six female Wistar rats were injured at T10 by IMPACTOR MODEL-Ⅱ, and then randomly assigned to three groups:Cyclo?sporin A (CsA)+Dexamethasone (Dex) treated group (Ⅰ, n=32), local transplantation of cells+CsA+Dex treated group (Ⅱ) at the first day after operation (DAO 1, n=32), local transplantation of cells+CsA+Dex treated group (Ⅲ) at DAO 6 (n=32). BBB locomotor scoring system was used to assess the recovery of the lower limbs. The survival and neural differentiation of transplanted cells at the injury site were observed by double immunofluorescence. The tissue vitality at the injury site was ob?served by 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) staining, and the blood vessel density was observed by infusing mixture of Chinese ink and glutin followed by HE staining. Results BBB score at DAO 8-56 was significantly higher inⅡgroup than that of other two groups (P<0.05). TTC staining showed that the proportion of decreased vitality area was signifi?cantly smaller inⅡgroup than that of other two groups (P<0.01). The result of gelatin ink perfusion showed that the blood vessel density at the injury site was significantly bigger inⅡgroup than that of other two groups (P<0.01). There were more survival transplanted cells inⅡgroup than those of III group (per visual field, 7.51 ± 1.00 vs 5.51 ± 0.89,t=6.051, P<0.01). All the transplanted cells didn’t differentiate into neural cells. Conclusion Human umbilical cord blood CD34+cells can promote the recovery of the lower limbs after spinal cord injury by repairing blood vessels to increase tissue vitality at the in?jury site in rats.

Basic Fibroblast Growth Factor-Chitosan Carriers Induce Neural Stem Cells to Differentiate into Neurons and Form Synapses / 中国康复理论与实践

@#Objective To explore the effect of basic fibroblast growth factor (bFGF)-chitosan carriers on neural differentiation of neural stem cells (NSCs). Methods NSCs were isolated from spinal cord of a neonatal Wistar rat and cultured. Purity of cultured NSCs was identified with Nestin immunofluorescent staining. The 10 mg/ml chitosan carriers, 20 ng/ml bFGF or 10 mg/ml bFGF-chitosan carriers were added into medium of P3~P4 NSCs respectively. NSCs were observed with immunofluorescent staining: 3 days after incubation with Nestin and β-tubulin III; 7 days after incubation with microtubule-associated protein-2 (MAP2), glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP); and 14 days after incubation with synapsin-1 and MAP2. The electrophysiological activity of cells was detected with MED64. Results 3 days after incubation, all the NSCs differentiated into Nestin+/β-tubulin III+, and the length of neurofilament was the highest in those co-cultured with bFGF-chitosan carriers. 7 days after incubation, NSCs differentiated into MAP2+, GFAP+ and MBP+, and more NSCs differentiated into MAP2+ with bFGF-chitosan carriers. 14 days after incubation, NSCs differentiated with bFGF-chitosan carriers express synapsin-1+/MAP2+ and showed electrophysiological activity. Conclusion bFGF-chitosan carriers can induce NSCs to differentiate into neuron with high percentage and the differentiated neurons can form synapses with electrophysiology activity.