摘要
BACKGROUND： Dopaminergic neurons differentiated from neural stem cells have been successfully used in the treatment of rat models of Parkinson＇s disease; however, the survival rate of transplanted cells has been low. Most cells die by apoptosis as a result of overloaded intracellular calcium and the formation of oxygen free radicals. OBJECTIVE： To observe whether survival of transplanted cells, transplantation efficacy, and dopaminergic differentiation from neural stem cells is altered by Panax notoginseng saponins （PNS） in a rat model of Parkinson＇s disease. DESIGN, TIME AND SETTING： Cellular and molecular biology experiments with randomized group design. The experiment was performed at the Animal Experimental Center, First Hospital of Sun Yat-sen University from April to October 2007. MATERIALS： Thirty-two adult, healthy, male Sprague Dawley rats, and four healthy Sprague Dawley rat embryos at gestational days 14-15 were selected. The right ventral mesencephalon was injected with 6-hydroxydopamine to establish a model of Parkinson＇s disease. 6-hydroxydopamine and apomorphine were purchased from Sigma, USA. METHODS： Neural stem cells derived from the mesencephalon of embryonic rats were cultivated and passaged in serum-free culture medium. Lesioned animals were randomly divided into four groups （n = 8）： dopaminergic neuron, dopaminergic neuron ＋ PNS, PNS, and control. The dopaminergic neuron group was injected with 3 μL cell suspension containing dopaminergic neurons differentiated from neural stem cells. The dopaminergic neurons ＋ PNS group received 3 μ L dopaminergic cell suspension combined with PNS （250 mg/L）. The PNS group received 3 μL PNS （250 mg/L）, and the control group received 3 μL DMEM/F12 culture medium. MAIN OUTCOME MEASURES： The rats were transcardially perfused with 4% paraformaldehyde at 60 days post-grafting for immunohistochemistry. The rats were intraperitoneally injected with apomorphine （0.5 mg/kg） to induce rotational behavior. RESULTS： Cell counts of tyrosine hydroxylase-positive neurons in the dopaminergic neuron ＋ PNS group were （732±82.6） cells/400-fold field. This was significantly greater than the dopaminergic neuron group [（326 ± 34.8） cells/400-fold field, P 〈 0.01]. Compared to the control group, the rotational asymmetry of rats that received dopaminergic neuron transplants was significantly decreased, beginning at 20 days after operation （P 〈 0.01）. Rotational asymmetry was further reduced between 10-60 days post-surgery in the dopaminergic neuron ＋ PNS group, compared to the dopaminergic neuron group （P 〈 0.01）. CONCLUSION： Panax notoginseng saponins can increase survival and effectiveness of dopaminergic neurons differentiated from neural stem cells for transplantation in a rat model of Parkinson＇s disease.
BACKGROUND： Dopaminergic neurons differentiated from neural stem cells have been successfully used in the treatment of rat models of Parkinson＇s disease; however, the survival rate of transplanted cells has been low. Most cells die by apoptosis as a result of overloaded intracellular calcium and the formation of oxygen free radicals. OBJECTIVE： To observe whether survival of transplanted cells, transplantation efficacy, and dopaminergic differentiation from neural stem cells is altered by Panax notoginseng saponins （PNS） in a rat model of Parkinson＇s disease. DESIGN, TIME AND SETTING： Cellular and molecular biology experiments with randomized group design. The experiment was performed at the Animal Experimental Center, First Hospital of Sun Yat-sen University from April to October 2007. MATERIALS： Thirty-two adult, healthy, male Sprague Dawley rats, and four healthy Sprague Dawley rat embryos at gestational days 14-15 were selected. The right ventral mesencephalon was injected with 6-hydroxydopamine to establish a model of Parkinson＇s disease. 6-hydroxydopamine and apomorphine were purchased from Sigma, USA. METHODS： Neural stem cells derived from the mesencephalon of embryonic rats were cultivated and passaged in serum-free culture medium. Lesioned animals were randomly divided into four groups （n = 8）： dopaminergic neuron, dopaminergic neuron ＋ PNS, PNS, and control. The dopaminergic neuron group was injected with 3 μL cell suspension containing dopaminergic neurons differentiated from neural stem cells. The dopaminergic neurons ＋ PNS group received 3 μ L dopaminergic cell suspension combined with PNS （250 mg/L）. The PNS group received 3 μL PNS （250 mg/L）, and the control group received 3 μL DMEM/F12 culture medium. MAIN OUTCOME MEASURES： The rats were transcardially perfused with 4% paraformaldehyde at 60 days post-grafting for immunohistochemistry. The rats were intraperitoneally injected with apomorphine （0.5 mg/kg） to induce rotational behavior. RESULTS： Cell counts of tyrosine hydroxylase-positive neurons in the dopaminergic neuron ＋ PNS group were （732±82.6） cells/400-fold field. This was significantly greater than the dopaminergic neuron group [（326 ± 34.8） cells/400-fold field, P 〈 0.01]. Compared to the control group, the rotational asymmetry of rats that received dopaminergic neuron transplants was significantly decreased, beginning at 20 days after operation （P 〈 0.01）. Rotational asymmetry was further reduced between 10-60 days post-surgery in the dopaminergic neuron ＋ PNS group, compared to the dopaminergic neuron group （P 〈 0.01）. CONCLUSION： Panax notoginseng saponins can increase survival and effectiveness of dopaminergic neurons differentiated from neural stem cells for transplantation in a rat model of Parkinson＇s disease.
基金
the National Natural Science Foundation of China, No.30300115
