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BACKGROUND: Dopaminergic neurons differentiated from neural stem cells have been successfully used in the treatment of Parkinson's disease rats; however, the survival rate of transplanted cells has been low. Most cells die of apoptosis as a result of the formation of oxygen free radical and lipid peroxidation.OBJECTIVE: To observe resveratrol (Res) effects on survival of transplanted cells, transplanted efficacy and dopaminergic differentiation from neural stem cells in a rat model of Parkinson's disease.DESIGN, TIME AND SETTING: Randomized controlled animal experiments were performed at the Animal Experiment Center,Sun Yat-sen University from October 2007 to June 2008.MATERIALS: Thirty-two adult, healthy, male Sprague Dawley rats were equally and randomly assigned to model control, dopaminergic neuron, Res and combination groups. Four healthy Sprague Dawley rat embryos at gastational days 14-15 were selected and fetal rats were used for isolation and culture of neural stem cells. Res (Jingmal Biotech, Shenzhen, China) was used for this study.METHODS: Neural stem cells derived from the mesencephalon of embryonic rats were isolated and cultured in vitro, and passaged in serum-free culture medium containing epidermal growth factor and basic fibroblast growth factor, and then differentiated into dopaminergic neurons in differentia.ion medium. Parkinson's disease rat models were established by the injection of 6-hydroxydopamine in each group. Rats in the dopaminergic neuron group was injected with 3 pL cell suspension (1×10 cells/μL) containing dopaminergic neurons in the corpus striatum. Rats in the Res group received 3 μL of Ras (40 mg/L).Rats in the combination group were subjected to 3 μL of Res (40 mg/L) + 3 μL cell suspension (1×105 calls/μL) containing dopaminergic neurons. Rats in the model control group received 3 ×L of DMEM/F12 culture medium.MAIN OUTCOME MEASURES: The percentage of tyrosine hydroxylase-positive neurons in differentiated cells. The alteration of rotational asymmetry and the survival of tyrosine hydroxylase-positive neurons in graft areas of Parkinson's disease rats after transplantation.RESULTS: Flow cytometry demonstrated that survival rate of tyrosine hydroxytase-positive neurons was (17.8 ±4.2)% at 6 days following differentiation. Compared to the model control group, the rotational asymmetry was significantly improved at 10 days (P < 0.01), was significantly decreased at 20 days following transplantation in the combination group (P < 0.01). At 10-60 days following transplantation, the number of rotational asymmetry was significantly lower in the combination group than in the dopaminergic neuron group (P < 0.01). Tyrosine hydroxylase-positive neurons were not determined in the Res and model control groups. The number of tyrosine hydroxylase-positive neurons was significantly more in the combination group than in the dopaminergic neuron group (P < 0.01).CONCLUSION: Res can increase survival rate of transplanted cells in the corpus striatum, and improve rotational asymmetry in rat models of Parkinson's disease following transplantation of dopaminergic neurons differentiated from neural stem cells.Ke CL, Chen BL, Yu ZH, Huang ZS.Effects of resveratrol on the transplantation of neural stem call-derived dopaminergic neurons in a rat model of Parkinson's disease.Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu. 2009;13(27): 5281-5285.[http://www.crter.on http://en.zglckf.com]
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BACKGROUND: Differentiation inducing factors and gestational age influence the differentiation potential of embryonic neuralstem cells.OBJECTIVE: This study was designed to observe the differentiation potential of rat mesencephalic neural stem cells at differentgestational ages towards dopaminergic neurons.DESIGN: A randomized controlled observation.SETTING: Department of Neurosurgery, First Affiliated Hospital of Sun Yat-sen University, Guangzhou City, GuangdongProvince, China.MATERIALS: This study was performed at the Laboratory of the First Affiliated Hospital of Sun Yat-sen University between Marchand September 2007. Thirty adult gestational SD rats, weighing 350 400 g, were provided by the Laboratory Animal Center of SunYat-sen University (Permission No. 2007-0034). The protocol was performed in accordance with ethical guidelines stated in Guide forthe use and care of laboratory animals, approved by the Committee on the Care and Use of Laboratory Animals of the Institute ofLaboratory Animal Resources Commission on Life Sciences, National Research Council, China (1985). DMEM/F12 serum-free medium,B27 additives, epidermal growth factor, basic fibmblast growth factor, and fetal bovine serum (volume fraction:0, 1) were purchased fromGibco Company, British; Interleukin lα, interleukin 11, and glial cell-derived neurotrophic factors were purchased from R&D Company,USA; In addition, leukaemia inhibitory factor (Perpotech, British), tyrosine hydroxylase(Santa Cruz, USA), nidogen antibody,microtubule-associated protein 2 antibody, and glial fibrillary acidic protein antibody(Chemicon, USA) were also used.METHODS: Six rats were randomly selected at each time point (on days 10,12,14,16, and 18 after gestation). After anesthesia, therats were sacrificed. Under the aseptic condition, fetal rat was harvested. Rat mesencephalic ventral brain tissue was isolated forculture of neural stem cells. Different gestational ages of rat brain-derived neural stem cells were separately cultured in theserum-free medium containing epidermal growth factors and basic fibroblast growth factors. After passage and amplification, theneural stem cells were induced to differentiate towards dopaminergic neurons in the medium containing interleukin lu, interleukin11, leukaemia inhibitory factors, glial cell-derived leukaemia inhibitory factors. On day 6 after induction and differentiation, thedopaminergic neurons were observed and identified by immunocytochemistry. After labeled by tyrosine hydroxylase, thedifferentiated dopaminergic neuron proportion was detected by a flow cytometer.MAIN OUTCOME MEASURES: The growth state of differentiated rat neural stem cells at different gestational ages and theimmunocytochemistry results. The tyrosine hydroxylase staining-positive neural stem cell proportion after induction anddifferentiation.RESULTS: Rat mesencephalic neural stem cell spheres on days 10,12, 14, 16, and 18 after gestation adhesively grew in thedifferentiation-inducing medium. The neural stem cells in the spheres gradually grew in radial tendency. On day 6 afterdifferentiation, most of the neural stem cells exhibited 1-2 long processes or several short processes. After nidogenimmunocytochemical staining, most of neural stem cells exhibited cytoplasm-positive. After culture for 6 days in the differentiationinducing medium, rat mesencephalic neural stem cells at gestational 10,12, 14, 16, and 18 days were detected by a flow cytometer.Results demonstrated that the proportion of tyrosine hydroxylase-positive cells was (10.3±2.5)%, (21.6±3.4)%, (16.7±2.8)%,(14.2±3.2)%, and (8.9±1.8)%, respectively. There was a significant difference in the proportion of tyrosine hydroxylase-positivecells among the cells at different gestational days (P < 0.05). Rat neural stem cells at gestational 12 days could be induced todifferentiate into dopaminergic neurons at the highest proportion.CONCLUSION: Mesencephalic neural stem cells of rats at different gestational days have different capabilities to differentiatetowards dopaminergic neurons. The proportion of dopaminergic neurons is the highest when mesencephalic neural stem cells ofrats at gestational 12 days.
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BACKGROUND: Recently, several scientists have found that differentiation of neural stem cells (NSCs) towards dopaminergic neurons may be increased in vitro by combination of some special cytokines. They have also found that dopaminergic neurons differentiated from NSCs can be used for the treatment of Parkinsn's disease. To improve the therapeutic effects of/n vitro transplantation, we should further study the biologic characteristics of NSCs at the induction and differentiation.OBJECTIVE: To explore the differentiation of NSCs which were incubated in differentiation solution for different time towards dopaminergic neurons in vitro.DESIGN: Single sample observation.SETTING: Department of Neurosurgery, First Affiliated Hospital of Sun Yat-sen University.MATERIALS: This study was performed at the Department of Ncurosurgery, First Affiliated Hospital of Sun Yat-sen University from May to October 2007. Six healthy Sprague Dawley (SD) rats, gestational age 14 days, of clean grade, weighing 350-400 g,were provided by the Laboratory Animal Center, Sun Yat-sen University[permission No. SCXK (yue)2007-0034]. The protocol was performed in accordance with ethical guidelines for the use and care of animals.METHODS: NSCs derived from rat embryonic mesencephalon were cultured in serum-free culture medium containing epidermal growth factors and basic fibroblast growth factors. After passage, the NSCs were induced to differentiate towards dopaminergic neurons in the differentiation medium supplemented with interleukin 1o, interleukinl 1, human leukaemia inhibitory factors, and glial cell line-derived neurotrophic factors. The percentage of tyrosine hydroxylase positive neurons in differentiated ceils was detected with flow cytometer when NSCs were cultured in differentiation solution for 2, 4, 6, 8 and 10 days, respectively.MAIN OUTCOME MEASURES: Cellular morphological alteration of rat NSCs after differentiation. The percentage of tyrosine hydroxylase positive neurons in differentiated cells derived from NSCs.RESULTS: In differentiation medium, NSC spheres attached the bottom of plates and began to collapse. Cells inside the spheres grew out gradually and became irregular in shape. Six days later, most of the cells had I or 2 long processes and a few short processes. The percentage of tyrosine hydroxylase positive neurons in differentiated cells was respectively (3.2_+_0.9)%, (6.8 +1.6)%, (16.7-+2.6)%, (14.8_+1.8)% and (12.2_+2.5)% after culture for 2, 4, 6, 8 and 10 days, with significant differences (F =26.449, P ＜ 0.05).CONCLUSION: Induction time influences the differentiation of NSCs towards dopaminergic neurons in vitro. The percentage of dopaminergic neurons is the highest in differentiated cells derived from NSCs which are cultured in differentiation solution for 6 days.
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Objective To evaluate the role of 3 D spiral computerized tomographic angiography (SCTA) in microsurgery of separating craniopagus twins Methods A case of craniopagus twins underwent spiral CTA Image reconstruction was made by shaded surface display Result SCTA demonstrated osseous fusion in twins skull Parts of bone and dura mater were found to be absent in the fused area Two parts of brain tissues were linked together, and some area were fused Twins' one third of posterior superior sagittal sinuses were fused and converged into a common sinus One side of their transverse sinuses were fused Conclusion SCTA can clearlly reveal the abnormalities of skull, cerebrum, intracranial blood vessels, especially the stereostruction between intracranial blood vessels and skull SCTA is useful for intraoperative bone design and decision of surgical route and has important value in formulating surgical plans