BACKGROUND:It has been suggested that melatonin(MT)can protect secondary neuronal injury.However,the protective effect of MT on neuronal injury in ischemia/reperfusion models in vitro still has not been proved.OBJECTI...BACKGROUND:It has been suggested that melatonin(MT)can protect secondary neuronal injury.However,the protective effect of MT on neuronal injury in ischemia/reperfusion models in vitro still has not been proved.OBJECTIVE:To investigate the protective effect of MT on central ischemic injury of nerve cells and analyze its possible mechanism.DESIGN:Contrast observational study.SETTING:Department of Biochemistry and Molecular Biology,Tongji Medical College,Huazhong University of Science and Technology.MATERIALS:Rats aged 7-8 days and weighing 10-12 g were provided by Medical Experimental Animal Center,Tongji Medical College,Huazhong University of Science and Technology.MT was provided by Sigma Company,USA.METHODS:The experiment was carried out in the Laboratory of Biochemistry and Molecular Biology,Tongji Hospital,Huazhong University of Science and Technology from October 2002 to March 2004.The effects of MT on the neurodegeneration induced by oxygen-glucose-deprivation(OGD)were tested in cultured rat cerebellar granule cells.Neuron damage was quantitatively assessed by Typan Blue exclusion and MTT assay at different time points after oxygen-glucose-deprivation(90 minutes).DNA gel electrophoresis and acridine orange stain were performed to determine the nature of cell damage.And fluorescence spectrophotometer was used for quantification of intracellular malondialdehyde(MDA)at various time intervals.MAIN OUTCOME MEASURES:Correlation between degrees of neuronal injury and reperfusion times,apoptosis,and production of MDA in cells.RESULTS:①The neuron injury was aggravated with reperfusion time.②The protective effect of MT was time-and dose-dependent when its concentration was not higher than 10μmol/L.③When neurons were exposed to OGD for 90 minutes,part of the cells exhibited typical features of apoptosis:internucleosomal DNA condensation and DNA ladder on agarose gel electrophoresis.MT added to cells recovering from OGD exerted neuroprotective action against OGD-induced apoptosis.④In OGD exposed cultures,the production of MDA burst 12 hours after OGD,while MT significantly decreased the generation of MDA(P<0.05)in a time-dependent manner.CONCLUSION:MT may have therapeutic potential in the prevention and treatment of ischemic/hypoxic neuronal damage,and this neuroprotective action may contribute to the antioxidant nature of MT.展开更多
BACKGROUND: Sodium valproate (VPA) is used to be an effective anti-epileptic drug. VPA possesses the characteristics of penetrating rapidly through the blood-brain barrier (BBB) and increasing levels of Bcl-2 and grow...BACKGROUND: Sodium valproate (VPA) is used to be an effective anti-epileptic drug. VPA possesses the characteristics of penetrating rapidly through the blood-brain barrier (BBB) and increasing levels of Bcl-2 and growth cone-associated protein (GAP) 43 in spinal cord. OBJECTIVE: To observe the effect of VPA on Bcl-2 expression and motor neuronal apoptosis in spinal cord of rats following sciatic nerve transection. DESIGN: Randomized controlled experiment. SETTING: Department of Hand Surgery and Microsurgery, Wuhan Puai Hospital. MATERIALS: A total of 30 male healthy SD rats of clean grade and with the body mass of 180-220 g were provided by Experimental Animal Center of Medical College of Wuhan University. Sodium Valproate Tablets were purchases from Hengrui Pharmaceutical Factory, Jiangsu. METHODS: The experiment was performed in the Central Laboratory of Wuhan Puai Hospital and Medical College of Wuhan University from February to May 2006. Totally 30 rats were randomly divided into two groups: treatment group (n =15) and model group (n =15). Longitudinal incision along backside of right hind limbs of rats was made to expose sciatic nerves, which were sharply transected 1 cm distal to the inferior margin of piriform muscle after nerve liberation under operation microscope to establish sciatic nerve injury rat models. Sodium Valproate Tablets were pulverized and diluted into 50 g/L suspension with saline. On the day of operation, the rats in the treatment group received 6 mL/kg VPA suspension by gastric perfusion, once a day, whereas model group received 10 mL/kg saline by gastric perfusion, once a day. L4-6 spinal cords were obtained at days 1, 4, 7, 14 and 28 after operation, respectively. Terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) technique and immunohistochemical method (SP method) were used to detect absorbance (A) of neurons with positive Bcl-2 expression. Apoptotic rate of cells (number of apoptotic cells/total number of cells×100%) was calculated. MAIN OUTCOME MEASURES: A value of neurons with positive Bcl-2 expression and apoptotic rate in spinal cord of rats in the two groups. RESULTS: A total of 30 SD rats were involved in the result analysis. ①expression of positive Bcl-2 neurons: A value of positive Bcl-2 neurons were 0.71±0.02, 0.86±0.04, 1.02±0.06 at days 4, 7 and 14, respectively after operation in the treatment group, which were obviously higher than those in the model group (0.62±0.03, 0.71±0.05, 0.89±0.04, t = 3.10-4.50, P < 0.05). ②apoptotic result of motor neurons: Apoptotic rate of motor neurons in spinal cord was (6.91±0.89)% and (15.12±2.34)% at days 7 and 14 in the treatment group, which was significantly lower than those in the model group [(9.45±1.61)%, (19.35±0.92)%, t = 2.39, 3.03. P < 0.05]. CONCLUSION: VPA can increase expression of Bcl-2 in spinal cord and reduce neuronal apoptosis in rats following sciatic nerve injury, and has protective effect on motor neuron in spinal cord of rats.展开更多
基金the Natural Science Foundation of Hygienic Committee of Hubei Province,No:WJ01510
文摘BACKGROUND:It has been suggested that melatonin(MT)can protect secondary neuronal injury.However,the protective effect of MT on neuronal injury in ischemia/reperfusion models in vitro still has not been proved.OBJECTIVE:To investigate the protective effect of MT on central ischemic injury of nerve cells and analyze its possible mechanism.DESIGN:Contrast observational study.SETTING:Department of Biochemistry and Molecular Biology,Tongji Medical College,Huazhong University of Science and Technology.MATERIALS:Rats aged 7-8 days and weighing 10-12 g were provided by Medical Experimental Animal Center,Tongji Medical College,Huazhong University of Science and Technology.MT was provided by Sigma Company,USA.METHODS:The experiment was carried out in the Laboratory of Biochemistry and Molecular Biology,Tongji Hospital,Huazhong University of Science and Technology from October 2002 to March 2004.The effects of MT on the neurodegeneration induced by oxygen-glucose-deprivation(OGD)were tested in cultured rat cerebellar granule cells.Neuron damage was quantitatively assessed by Typan Blue exclusion and MTT assay at different time points after oxygen-glucose-deprivation(90 minutes).DNA gel electrophoresis and acridine orange stain were performed to determine the nature of cell damage.And fluorescence spectrophotometer was used for quantification of intracellular malondialdehyde(MDA)at various time intervals.MAIN OUTCOME MEASURES:Correlation between degrees of neuronal injury and reperfusion times,apoptosis,and production of MDA in cells.RESULTS:①The neuron injury was aggravated with reperfusion time.②The protective effect of MT was time-and dose-dependent when its concentration was not higher than 10μmol/L.③When neurons were exposed to OGD for 90 minutes,part of the cells exhibited typical features of apoptosis:internucleosomal DNA condensation and DNA ladder on agarose gel electrophoresis.MT added to cells recovering from OGD exerted neuroprotective action against OGD-induced apoptosis.④In OGD exposed cultures,the production of MDA burst 12 hours after OGD,while MT significantly decreased the generation of MDA(P<0.05)in a time-dependent manner.CONCLUSION:MT may have therapeutic potential in the prevention and treatment of ischemic/hypoxic neuronal damage,and this neuroprotective action may contribute to the antioxidant nature of MT.
文摘BACKGROUND: Sodium valproate (VPA) is used to be an effective anti-epileptic drug. VPA possesses the characteristics of penetrating rapidly through the blood-brain barrier (BBB) and increasing levels of Bcl-2 and growth cone-associated protein (GAP) 43 in spinal cord. OBJECTIVE: To observe the effect of VPA on Bcl-2 expression and motor neuronal apoptosis in spinal cord of rats following sciatic nerve transection. DESIGN: Randomized controlled experiment. SETTING: Department of Hand Surgery and Microsurgery, Wuhan Puai Hospital. MATERIALS: A total of 30 male healthy SD rats of clean grade and with the body mass of 180-220 g were provided by Experimental Animal Center of Medical College of Wuhan University. Sodium Valproate Tablets were purchases from Hengrui Pharmaceutical Factory, Jiangsu. METHODS: The experiment was performed in the Central Laboratory of Wuhan Puai Hospital and Medical College of Wuhan University from February to May 2006. Totally 30 rats were randomly divided into two groups: treatment group (n =15) and model group (n =15). Longitudinal incision along backside of right hind limbs of rats was made to expose sciatic nerves, which were sharply transected 1 cm distal to the inferior margin of piriform muscle after nerve liberation under operation microscope to establish sciatic nerve injury rat models. Sodium Valproate Tablets were pulverized and diluted into 50 g/L suspension with saline. On the day of operation, the rats in the treatment group received 6 mL/kg VPA suspension by gastric perfusion, once a day, whereas model group received 10 mL/kg saline by gastric perfusion, once a day. L4-6 spinal cords were obtained at days 1, 4, 7, 14 and 28 after operation, respectively. Terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) technique and immunohistochemical method (SP method) were used to detect absorbance (A) of neurons with positive Bcl-2 expression. Apoptotic rate of cells (number of apoptotic cells/total number of cells×100%) was calculated. MAIN OUTCOME MEASURES: A value of neurons with positive Bcl-2 expression and apoptotic rate in spinal cord of rats in the two groups. RESULTS: A total of 30 SD rats were involved in the result analysis. ①expression of positive Bcl-2 neurons: A value of positive Bcl-2 neurons were 0.71±0.02, 0.86±0.04, 1.02±0.06 at days 4, 7 and 14, respectively after operation in the treatment group, which were obviously higher than those in the model group (0.62±0.03, 0.71±0.05, 0.89±0.04, t = 3.10-4.50, P < 0.05). ②apoptotic result of motor neurons: Apoptotic rate of motor neurons in spinal cord was (6.91±0.89)% and (15.12±2.34)% at days 7 and 14 in the treatment group, which was significantly lower than those in the model group [(9.45±1.61)%, (19.35±0.92)%, t = 2.39, 3.03. P < 0.05]. CONCLUSION: VPA can increase expression of Bcl-2 in spinal cord and reduce neuronal apoptosis in rats following sciatic nerve injury, and has protective effect on motor neuron in spinal cord of rats.
