ObjectiveTo explore the effect of bFGF-chitosan carriers on inducing bone marrow-derived mesenchymal stem cells (MSCs) to differentiate into nerve cells.MethodsMSCs were detected by immunohistochemistry and Western blot after they were induced by bFGF-chitosan carriers to differentiate into neurons. The MTT chromometry assay was carried out to determine cell viability.ResultsThe proportion of express neural stem cells marker Nestin, and neuronal markers class Ⅲ β-tubulin and MAP-2 was 83.54% after MSCs induced by bFGF-chitosan carriers.ConclusionbFGF-chitosan carriers can induce MSCs to differentiate into nerve cells with a high percentage.

Objective Objective The present study was to increase the awareness of nonalcoholic Wernicke's encephalopathy ( WE) to reduce its misdiagnosis.Methods The clinical features and MR imaging findings in 6 patients with nonalcoholic WE were retrospectively analyzed.Results All patients exhibited different degrees of unconsciousness.Only two patients presented with the typical triad of neuro-ophthalmologic manifestations, ataxia, and global confusion.All patients presented with typical MR features characterized by bilaterally altered signal of the medial thalamus, periventricular region of the third ventricle and periaqueductal area. In addition, two patients developed symmetric cortical and facial nerve nucleus involvements with deep coma, which was clinically rare.The average clinical recovery and MRI imaging recovery times were 7.5 months and 2.8 months, respectively,.Two patients with deep coma showed a poor prognosis:1 patient died, and the other had a sever spastic paralysis of her extremities and mental retardation during a follow -up of 2 years.Two patients with deep coma showed symmetric hyperintensities on diffusion -weighted imaging ( DWI) .Conclusions MRI images are useful in the early diagnosis of nonalcoholic WE.Cortical and cranial nerve nucleus involve-ment in nonalcoholic WE patients may be an indication of irreversible damage and a poor prognosis.In addition, hyperintensities on DWI may also indicate an unfavorable prognosis.

Objective To investigate the effect of sodium citrate on the efficacy of oral midazolam premedication in children with congenital heart disease. Methods Forty ASA Ⅱ or Ⅲ children, aged 2-6 yr, weighing 12-20 kg, undergoing cardiac surgery, were randomly divided into 2 groups (n = 20 each): control group (gronp C) and sodium citrate group (group S). Group S received oral mixture of midazolam 0.12 ml/kg (0.6 my/kg), ketamine 0.12 ml/kg (6 my/kg), glucose 0.12 ml/kg (60 mg/kg) and sodium citrate 0.12 ml/kg (3 mg/kg), total volume 0.48 ml/kg. Group C received oral mixture of midazolam 0.12 ml/kg, ketamine 0.12 ml/kg and glucose 0.24 ml/kg, total volume 0.48 ml/kg. Hydrochloric acid (pH value 1.75) was mixed with the mixtures in the two groups and pH values were measured. Preoperative anxiety scale and the onset time,sedation score and parental separation score after receiving oral drugs were recorded in preparation room for anesthesia. After entering the operating room, HR, MAP and SpO2 were monitored, and the response to venepuncture in children and the adverse effects associated with oral drugs were also observed and recorded. Results The pH value was 1.97 in group C and 4.52 in group S. The parental separation score, sedation score and response score were significantly lower and the onset time was significantly shorter in group S than in group C. HR, MAP and SpO2 were in the normal range after entering the operating room. There was no obvious adverse effect after administration of oral drugs in the two groups. Conclusion Application of sodium citrate in the oral premedication in children with congenital heart disease can raise the pH value, shorten the onset time of midazolam, and enhance the sedative efficacy.

BACKGROUND:Neural stem cells (NSCs) hold self-renewal and multi-directional differentiation potential. NSCs differentiation into neurons in high proportion under induction conditions exhibits broad application prospect. OBJECTIVE:To explore the effect of basic fibroblast growth factor (bFGF)-chitosan carrier on the NSCs differentiation into neurons in vitro, and whether the differentiated neurons could form synaptic-like connection with myocytes. METHODS:After purification, the NSCs were co-cultured with chitosan, soluble bFGF or bFGF-chitosan carrier. After 7-day induction, the NSCs differentiation into neurons was observed by immunofluorescence staining of beta tubulin Ⅲ. The NSCs differentiation into cholinergic neurons was observed through double immunofluorescence staining of ChaT and beta tubulin Ⅲ. The synaptic-like connection between the neurons and myocytes was observed by triple staining of beta tubulin Ⅲ and MHC. RESULTS AND CONCLUSION:The percentage of differentiated neurons in the bFGF-chitosan carrier group was 74%, which was significantly higher than that in the other two groups. Additionally, the synaptic-like connection formed between the differentiated neurons and myocytes. To conclude, the bFGF-chitosan carrier promotes the NSCs differentiation into neurons to form synaptic-like connection with the co-cultured myocytes.

Objective:To investigate the regulation of proliferation and differentiation of bone marrow mesenchymal stem cells(BM-SCs)by CKIP-1 in vitro.Methods:BMSCs from CKIP-1 nock out(KO)and wild type(WT)C57 mice were isolated and cultured u-sing adherence method in vitro.BMSCs of the 3rd passage were induced to osteogenic and adipgenic differentiation.Cell proliferation was examined by MTT assay.Cell surface markers were tested by FCM.The osteogenic and adipogenic differentiation was studied by alkaline phosphatase (ALP)staining,alizarin red staining and oil red O staining.Results:The proliferation and cell marke expression of the 2 groups were similar.ALP staining of KO group was strong than that of WT group after osteogenic induction.Alizarin red stai-ning showed that there were more mineralized nodules in BMSCs of KO group than in those of WT group.Oil red O staining of KO mice BMSCs was stronger than that of WT.Conclusion:CKIP-1 deficiency can enhance the osteogenic and adipogenic differentiation without influence on the proliferation of BMSCs.

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.

Objective To explore the induction of basic fibroblast growth factor (bFGF)-chitosan carrier transforming the adult rat bone marrow mesenchymal stem cells (rMSCs) into nerve cells. Methods rMSCs were detected qualitatively and counted quantitatively by immunohistochemistry after they were induced into nerve cells, such as neural stem cells neurons and astrocytes. The methyl thiazolyl tetrazolium (MTT) chromometry assay was carried out to determine the cell viability. Results rMSCs induced by bFGF-chitosan carrier expressed neural stem cell marker nestin, neuron marker β-tubulin Ⅲ and astrocytes marker glial fibrillary acidic protein (GFAP). Nestin expressed more in the bFGF-chitosan group, and reached its maximum (49.40%) at the 9th day. Conclusion bFGF-chitosan carrier can induce the adult rMSCs differentiate into neural stem cells in a high proportion.

Objective To observe the effects of neurotrophin 3 (NT3)-chitosan on motor function, and proliferation and differentiation of the neural stem cells (NSCs) in the injury area and subventricular zone (SVZ) in rats with motor cortex injury. Methods Sixty-five Wistar rats were divided into control group (n=7), injury group (n=29) and NT3-chitosan group (n=29). The motor cortex was aspirated and re-moved as cerebral injury model. NT3-chitosan was immediately implanted into the injured area after operation, and the control group re-ceived no intervention. Pellet reaching test was performed to detect the recovery of the forelimb function, HE staining was used to observe the lesion cavity size, and immunofluorescence staining was used to observe the proliferation and differentiation of NSCs 3 days, 7 days, 14 days, 1 month, 2 months and 3 months after operation. Results The grasp success rate was higher (F>6.00, P≤0.05), and the lesion cavity size was significantly smaller (F>629.5, P<0.001) in the NT3-chitosan group than in the injury group. In the NSCs differentiation experi-ment, the number of BrdU cells at all the time points was significantly higher in the NT3-chitosan group than in the injury group (F>171.43, P<0.001). In the NSCs proliferation experiment, the number of BrdU positive cells was still significantly higher in the NT3-chitosan group than in the control group and in the injury group (F>155.06, P<0.001), the number of Dcx positive cells was significantly higher in the NT3-chitosan group than in the injury group (F=62.367, P<0.001), and the number of BrdU/Dcx positive cells was significantly higher in the NT3-chitosan group than in the control group (F=33.527, P<0.001). Conclusion NT3-chitosan could activate NSCs in the SVZ, and pro-mote endogenous neurogenesis and forelimb function recovery in rats after motor cortex injury.

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.

Objective To compare the application of CUBIC and iDISCO clearing methods in observing 3D imaging of spinal cord with immunofluorescent staining. Methods 1 mm thick spinal cord coronal sections were processed with CUBIC and iDISCO, respectively. The neurofilament (NF) protein was detected by immunofluorescent staining and then was observed by a laser confocal microscope. Results Compared with CUBIC, iDISCO had the advantages of shorter time, higher transparency (F=6.64, P<0.01), and deeper penetration (F=5117.55, P<0.01). Conclusion Immunofluorescent staining combined with iDISCO could completely observe the spinal axons with shorter time and better stain effect.