Cadmium Activates Reactive Oxygen Species-dependent AKT/mTOR and Mitochondrial Apoptotic Pathways in Neuronal Cells / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To examine the role of Cd-induced reactive oxygen species (ROS) generation in the apoptosis of neuronal cells.</p><p><b>METHODS</b>Neuronal cells (primary rat cerebral cortical neurons and PC12 cells) were incubated with or without Cd post-pretreatment with rapamycin (Rap) or N-acetyl-L-cysteine (NAC). Cell viability was determined by MTT assay, apoptosis was examined using flow cytometry and fluorescence microscopy, and the activation of phosphoinositide 3'-kinase/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and mitochondrial apoptotic pathways were measured by western blotting or immunofluorescence assays.</p><p><b>RESULTS</b>Cd-induced activation of Akt/mTOR signaling, including Akt, mTOR, p70 S6 kinase (p70 S6K), and eukaryotic initiation factor 4E binding protein 1 (4E-BP1). Rap, an mTOR inhibitor and NAC, a ROS scavenger, blocked Cd-induced activation of Akt/mTOR signaling and apoptosis of neuronal cells. Furthermore, NAC blocked the decrease of B-cell lymphoma 2/Bcl-2 associated X protein (Bcl-2/Bax) ratio, release of cytochrome c, cleavage of caspase-3 and poly(ADP-ribose) polymerase (PARP), and nuclear translocation of apoptosis-inducing factor (AIF) and endonuclease G (Endo G).</p><p><b>CONCLUSION</b>Cd-induced ROS generation activates Akt/mTOR and mitochondrial pathways, leading to apoptosis of neuronal cells. Our findings suggest that mTOR inhibitors or antioxidants have potential for preventing Cd-induced neurodegenerative diseases.</p>

Activation of α7 nicotinic acetylcholine receptor alleviates cerebral cortical neuron injury induced by oxygen- glucose deprivation / 第二军医大学学报

Objective To investigate the effect of activatinga7 nicotinic acetylcholine receptor (α7nAchR) on cerebral cortical neurons injury induced by oxygen-glucose deprivation (OGD) and the possible mechanism. Methods Cerebral cortical neurons cultured for 7 d were randomly divided into three groups; control group, OGD group (Cells experienced a 12 h oxygen- glucose deprivation) and OGD group treated with PNU-282987 (Cells experienced a 12 h oxygen-glucose deprivation with PNU- 282987 pretreatment for 24 h). Cell viability was determined by CCK-8 assay, lactate dehydrogenase (LDH) was examined to reflect cell injury, apoptosis and reactive oxygen species (ROS) production were analyzed by flow cytometry, and expression of hemeoxygenase-1 (HO-1) and hypoxia inducible factor-1a (HIF-1a) were detected by Western blotting analysis. Results OGD resulted in cell death, LDH increase, and cell apoptosis. Compared with the OGD group, PNU-282987 pretreated group had significantly increased cell survival (P α protein expression was significantly reduced in PNU-282987 pretreated group compared with the OGD group (Pα7nAchR can protect cerebral cortical neurons against OGD-induced injury, which may be related to the anti-oxidative stress.

Antioxidant and protective effects of LWMH-SOD on neurocytes / 中国药理学通报

Aim To investigate the protective effects of LMWH-SOD(Low molecular weight heparin- Superoxide dismutase Conjugate)on the injuries induced by oxygen and glucose deprivation in cultured neurons. Methods The cortical neurons of fetal rat were cultured in vitro. The antioxidant and protective effects of LMWH-SOD were observed by treating neurons with oxygen and glucose deprivation. Results LMWH-SOD reduced the number of cell death and the efflux of LDH and the content of NO,MDA and increased the membrane fluidity after the injuries of cells. Conclusion LMWH-SOD has protective effects on cerebral cortical neurons through its action of scavenging free radicals.

Protective effects of amitriptyline on glutamate-induced neurotoxicity in cerebral neuronal cultures / 中国临床药理学与治疗学

AIM: To study the protective effects of amitriptyline (Ami) on glutamate induced neurotoxicity in cultured rat cerebral cortical neurons. METHODS: Cortical neurons of fetal rat were cultured in vitro. The protective effects of Ami on glutamate induced neurotoxicity were observed. RESULTS: Exposure of rat fetus cerebral cells to Glu medium developed a neurotoxicity, expressed in the increase of LDH, NO and MDA, and the decrease of activity of SOD,as well as the development of morphological injury. Ami (10 -6 ,10 -7 , 10 -8 mol?L -1 ) significantly protected neurons against above demage. CONCLUSION: Ami can prevent rat cerebral neurons injury from the toxicity of Glu.

The Effects of Glutamate Receptor Antagonists on Cultured Cerebral Cortical Neurons of Neonatal Mouse Damaged by Oxidative Stress

PURPOSE: To evaluate neurotoxic effects induced by oxygen-radicals, which were generated by adding xanthine oxidase(XO) and hypoxanthine(HX), and protective effects of glutamate receptor antagonist such as MK-801 and 6-cyano-7-nitroquinoxaline(CNQX). METHODS: Dissociated cell cultures were prepared from cerebrum of neonatal mouse. Tissues were dissected and diced into small pieces in phosphate buffered saline and were incubated at 37degrees C. Isolated cells were resuspended in Eagle's minimum essential medium and plated poly-L-lysine coated plastic coverslips in 96 well multichambers at a cell density of 3x105 cells/well. Cells were grown in a 5% CO2/95% air atmosphere at 37degrees C. Cytotoxic effects were examined in cerebral cortical neurons cultured for 3 hours in media containing various concentration of XO and HX. The protective effects of glutamate receptor antagonist were also examined by MTT assay and neurofilament enzymeimmunoassay(EIA). Microscopic examinations were also done. RESULTS: Oxygen radicals markedly induced decrement of the cell viability of cultured mouse cerebral cortical neurons in a dose-dependent manner. Midpoint cytotoxicity value was 30mU/ml XO/0.1mM HX, when mouse cerebral cortical neurons were incubated for 3 hours with various concentrations of XO and HX. The number of cells and neurites was decreased when cerebral cortical neurons were cultured for 3 hours in a medium containing 30mU/ml XO/0.1mM HX. MK- 801 was very effective in blocking oxidant-induced neurotoxicity, while CNQX falied to show any protective effect in these cultures. CONCLUSION: It is suggested that oxygen radicals are neurotoxic, and selective N-methyl-D-aspartate antagonists such as MK-801 are very effective in protecting neurotoxicity induced by oxygen radicals in cultured cerebral cortical neurons of neonatal mouse.

Protective effects of total salvianolic acids on cerebral hypoxia in mice / 中国临床药理学与治疗学

Aim The protective effect of total salvianolic acids (Sal) on cerebral hypoxia inmice was studied. Methods Acute cerebral hypoxia was induced by sodium nitrite scand decapitation.The effect of Sal on acute cerebral hypoxia in mice and neuronalhypoxia injury induced by sodium dithionite in primary cultures were ob-served. Results Sal in the doses of 10, 20 mg?kg-1 iv protected mice against theacute cerebral hypoxia and inhibited the production of lipid peroxidation in brain tis-sue of mice caused by cerebral hypoxia. Sal in the doses of 1~10 ?g? L-1 reduced therate of cell death and the content of MDA and lowered LDH content in extra-cellularbathing media in oxygen deprived cortical cultures.Conclusions Sal protects miceagainst cerebral hypoxia by suppressing the generation of lipid peroxide.

NEUROTROPHIC ACTIVITY IN BRAIN WOUND TISSUE EXTRACT AND THE ORIGIN OF THE ACTIVE FACTORS / 解剖学报

Bilateral ablation of the cerebral parietal cortex in adult rats was performed. After appropriate days, the tissue surrounding the wound was removed and the brain wound tissue extract (EWTE) was prepared. Newborn rat cerebral cortical neurons were used as a culture model to test the neuronotrophic factors (NTFs) and neuritepromoting factors (NPFs) in EWTE. In order to investigate the origin of above mentioned factors whether related to the macrophages which appeared in the brain wound region at early stage, we designed to culture macrophages and collected the macrophage conditioned medium (M?CM) to measure their NTF and NPF activities for cultured cerebral cortical neurons. On the other hand, we also observed the effect of EWTE and M?CM on PC 12 (phehrmytema) cells and further studied the action of NPFs. Cur experimental results show that EWTE and M?CM contained NTFs and NPFs for cultured cerebral cortical neurons. These factors appeared in EWTE at 4 days post-lesion, with maximal level of their activities reached between 5 and 6 days post-lesion and there was another peak of NPF activity at 9 days post-lesion, until 13 days post-lesion also detected their activities. The NTF activity in M?CM was lower than that in BWTE, in contrast, the NPF activity in M?CM was higher than that in BWTE. There was NPF activity to PC 12 cells in BWTE and M?CM. According to the experimental assays, we suppose that the neurotrophic factors in BWTE mainly come from the macrophages which appear in the lesion site at early stage of injured brain, subsequently, it may relate to the astrocytes. The components of these factors may be complexity and multiplicity that remain to be solved.