ObjectiveTo investigate the protective effect of LY367385 on impairment of cultured mouse cerebral cortical neurons induced by sodium glutamate (Glu) or oxygen-glucose deprivation (OGD).MethodsNeuron damage induced by Glu or OGD, as well as the action of (S)-(+)-a-amino-4-carboxy-2-methylbenzeneacetic acid (LY367385) were measured by determining the leakage of lactate dehydrogenase (LDH) from neurons. Immunocytochemistry and immunofluorescent methods were used to detect the expression of anti-mGluR1α. Morphological observation of primary cortical neurons was performed by phase contrast microscope.ResultsFollowing the exposure to 0.1 mmol/L Glu for 1 h or OGD for 1 h, LDH leakage from neurons obviously increased (P< 0.01 ). 50 mmol/L LY367385, when co-incubated with Glu or OGD, markedly reduced the LDH leakage (P<0.01). The 24-h leakage of LDH was increased from cells exposed to 0.1 mmol/L Glu for 15 min. Pre-and post-treatment with LY367385 (50 mmol/L ) decreased the leakage of LDH. The cultured neurons expressed mGluR1α.ConclusionLY367385 has protective effect on neurons damaged by Glu or OGD. It may be related to antagonizing mGluR1α.

ObjectiveTo observe the activities of cultured neurons incubated with Earle's solution, new minimal essential medium (MEM) or former MEM.MethodsActivity changes of cultured neurons were measured by determining the leakage of lactate dehydrogenase (LDH) and metabolic rate of 3-(4,5-dimethylthiazol)2,5 diphenyltetrazolium bromide (MTT).ResultsActivities of neurons incubated with different culture medium for 12 h or 24 h were significantly different (P<0.01).ConclusionDifferent culture medium can influence neuronal activities.

BACKGROUND: Total flavone of ginkgo biloba(TFG) can affect on free radical, but the effect on apoptosis induced by cerebral ischemia is unclear.OBJECTIVE: To study the inhibitory effect of TFG on apoptosis induced by cerebral ischemia.DESIGN: Completely randomized controlled experimental study based on experimental animals.SETTING: Department of pharmacology in a university.MATERIALS: Totally 24 SD rats in half genders with clean grade and body mass of(250 ± 50) g, were divided into 4 groups at random: sham-operation group, model group, TFG 40 rmg/kg group and TFG 80 mg/kg group (Certificate No. 01).METHODS: This study was completed in the Department of Pharmacology,Anhui Medical University during October 2001 and January 2002. Incomplete cerebral ischemia was made by ligating bilateral common carotid arteries(CCA) in rats. The cerebral injury was evaluated by brain edema. The apoptosis was determined by terminal deoxy-nucleotidyl transforase-mediated dUTP-digoxigenin nick end-labeling(TUNEL) and transmission electron microscopy (TEM) method. The DNA fragmentation analysis was measured with the diphenylamine reagent method.MAIN OUTCOME MEASURES: Major factor: Effect of TFG on ultrastructral alteration of apoptotic cerebral cortex cells; Secondary factor: Effect of TFG on DNA fragmentation induced by cerebral ischemia.RESULTS: Ligating of bilateral CCA markedly induced apoptotic cell in cerebral cortex. TFG 80 mg/kg significantly inhibited brain edema( P ＜ 0.05 )and decreased the numbers of apoptotic cells in cortex( P ＜ 0.01 ) and improved ultrastructral alteration of apoptotic cells; TFG 40, 80 rmg/kg also inhibited the increase of DNA fragmentation induced by cerebral ischemia (P ＜0.05, P ＜0.01).CONCLUSION: TFG has inhibitory effect on ischemia-induced apoptosis of cerebral cortex and improve the ultrastructual changes of apoptosis. Moreover,TFG can relieve the occurrence of edema of ischemic brain tissue and inhibit the increase of DNA section induced by cerebral ischemia.

OBJECTIVE:To study the effects of rosiglitazone(RH)on body weight,FPG,FFA and other indicators in diabe-tes model rabbit. METHODS:18 rabbits were evenly randomized into control group,model group and dextran group. The latter 2 groups were given alloxan intravenously to induce diabetes model. 3 groups were given RH(0.5 mg/kg)intragastrically,and dex-tran group was additionally given dextran 40 glucose injection(5 ml/kg)intravenously,once a day,for consecutive 3 weeks. Body weight,serum level of FPG,FFA,AngⅡ and NO were determined before and after medication. RESULTS:Compared with be-fore medication,body weight of rabbits in control group were increased after medication,while the levels of FFA and AngⅡ were decreased;the levels of FPG and FFA were decreased in model group;body weight of rabbits were decreased in dextran group af-ter medication,with statistical significance (P<0.01 or P<0.05);other indicators had no statistically significant difference (P>0.05). The level of FFA in dextran group was higher than in model group after medication,with statistical significance(P<0.01). CONCLUSIONS:Rosiglitazone can lead to weight gain by a mechanism which reduce the level of FFA.

Objective To provide the experimental basis for the further research of the interacting proteins with Stathmin ,the Stathmin gene Pichia pastoris expression system was constructed ,the expressed Stathmin product was purified and identified .Meth‐ods Stathmin gene was amplified from tumor cell line of SKBR3 by PCR method and cloned into the yeast expression vector pPIC3 .5K .The recombinant vector pPIC3 .5K‐Stathmin was constructed and transformed into Pichia pastoris GS115 .The positive clones were screened by YPD medium containing Geneticin 600 μg/mL .Expression was induced with 0 .5% methanol and expres‐sion products were identified by SDS‐PAGE and Western Blotting .Results DNA sequencing result showed that the gene fragment was consistent with Stathmin gene sequence .pPIC3 .5K‐Stathmin was selected from YPD culture medium containing Geneticin ,and the positive clones were identified by PCR .SDS‐PAGE showed that a 37 × 103 protein band could be seen on the PAGE gel after Coomassie Blue staining ,which was further confirmed and identified as Stathmin protein by Western Blotting .Conclusion Stathmin yeast expression vector is successfully constructed and expressed in Pichia pastoris ,which laid the foundation for the study of inter‐acting proteins with Stathmin ,and for the preparation of the biological treatment drugs of Stahtmin target .

Objective To observe the protective effect of total flavone of Camellia (TFC)against cerebral ischemic injury.Methods Decapitation method and close hypoxia method were used to observe the effect of TFC on anoxic tolerance of mice and step down test was used to observe the effect of TFC on learning and memory; after ischemia, the content of malondialdehyde (MDA)and nitric oxide (NO)and the activity of lactate dehydrogenase (LDH)were detected. Rat models with acute incompletely cerebral ischemia were established by means of ligating right common carotid arteries and effect of TFC on cerebral water volume, permeability of cerebral vessels and cerebral histopathological changes were also observed.Results TFC prolonged the grasping time after decapitation and the survival time after anoxia in mice, and improve the learning and memory during the step down test. TFC decreased MDA and NO contents, counteracted the de creases of LDH activities in the mice cerebral cortex, reduced the water volume and permeability of cerebral vessels in ischemic rats and improve the cerebral hitstopathological changes. Conclusion TFC has protective effects against cerebral ischemic injury and the mechanism may be related to the inhibition of free radicals and NO production.

Aim To study protective effects of Total of flacone C(TFC) against cerebral ischemia-reperfusion injury.Methods Four-vessel occlusion method was used to make acute cerebral ischemia-reperfusion model. Rats were initiated by ischemia for 30 min followed by 40 min of reperfusion.The electroencephalography(EEG) during cerebral ischemia and reperfusion was recorded.The level of intracellular calcium ion concentration([Ca~(2+)]i) in cerebral cells after ischemia was measured by using a Ca~(2+) indicator Fura-2/AM.Superoxide dismutase(SOD),Glutathione peroxidase(GSH-Px),Lactate dehydrogenase(LDH),nitric oxide Synthase(NOS) activeties and Malondialdehyde(MDA),Nitric Oxide(NO)contents in the ischemia cerebral cortex were measured.Results TFC can improved the EEG change,significantly attenuated the decrease of the intracellular calcium ion concentration([Ca~(2+)]_i), remarkly increased GSH-Px,SOD and NOS activities in the cerebrum,inhibit the decrease of LDH activity and NO,MDA contents.Conclusion TFC has protective effects on cerebral ischemia-reperfusion injury,the mechanism may be related to attenuating free radical,[Ca~(2+)]i overload and NO.

AIM: To investigate the protective effects of total of flavone C (TFC) on acute cerebral ischemia in mice and focal cerebral ischemia in rats. METHODS: The occlusion of bilateral common carotid arteries with vagus nerves in mice was used for make the acute cerebral ischemia models. The survival time and the death rate were observed. The permanent occlusion of the proximal of the right middle cerebral artery (MCA) was used for make the focal cerebral ischemia models. The extent of neurological deficits was observed, and the infarct area was measured by NBT staining technique. The activity of LDH and the content of MDA and NO in the ischemic cerebral cortex were determined. RESULTS: TFC of 80 and 40 mg?kg -1 prolonged the survival time and decreased the death rate of mice with acute cerebral ischemia injury. TFC of 60, 30, and 15 mg?kg -1 ameliorated neurologic deficits score and the infarct size of rats with MCAO. CONCLUSION: TFC provides significant protective effects against cerebral ischemia injury.

Objective To reproduce a scratch-wound model in cultured rat astrocytes (AST).Methods The secondary cultured AST prepared from newborn Wistar rat cerebral cortex were scratched with plastic pipette tips. The morphologic change of AST was observed through microscope at 10 min before and 1, 3, 6, 12, and 24 h after injury, meanwhile the lactate dehydrogenase (LDH) leakages in the cultured medium were determined.Results Immediately after injury the edge of the scratch was lined with irregularly shaped cell. 6 h after injury the AST processes began extending to cell-free area, and elongated further at 12 and 24 h after injury, with presented of new generated cells in the denuded area. At different times after injury, the LDH leakages of the experiment group were higher than that before injury ( P<0.05), and were higher than that of the control group ( P<0.05).Conclusion According to observed AST morphologic changes and determined LDH leakages in culture medium, the scratch-wound model in cultured rat AST is successfully reproduced.

Objective:To screen for the pathogenesis-related genes of osteosarcoma and to assess their roles for the de- velopment of osteosareoma.Methods:Total RNA was extracted from 3 ATCC osteosarcoma cell lines and an osteoblastic cell line and was used to synthesize biotinylated cRNAs;the latter were hybridized to Affymetrix~(?)GeneChip~(?)U133A ar- rays and a gene with more than 2 folds of change was selected.Ten of the differentially expressed genes were chosen and the primers were designed and the synthesized.Then SYBR~(?)Green real-time PCR(RT-PCR)method was used to detect the expression of the 10 genes in 9 fresh osteosarcoma specimens.ABI Prism 7 000 system was used to analyze the differ- ent expression between osteosarcoma cell line and osteoblastic cell line.Results:We identified 58 up-regulated and 142 down-regulated genes in the 3 osteosareoma cell lines.Many of the genes were firstly reported to be related to the patho- genesis of osteosarcoma.These differentially expressed genes were mainly involved in energy and material metabolism,on- cogene,signal transduction gene,transcription- related genes,cell cycle-related genes,cell apoptosis-related gene,im- mune response gene,tumor suppressor genes,etc.The array results of 10 randomly selected genes were further verified by the RT-PCR in 9 fresh osteosarcoma specimens.Conclusion:Many genes are involved in the pathogenesis of osteosarcoma. Gene microarray can help to discover the genes related to the pathogenesis of osteosarcoma,which may lay a foundation for studying the molecular mechanism of osteosarcom.