Effect of lipopolysaccharide on astrocyte proliferation and release of its calcium and nitric oxide and reactive oxygen species
- VernacularTitle:脂多糖对星形胶质细胞增殖和细胞内钙离子水平及活性氧与一氧化氮释放的影响
- Author:
Yuan ZHOU
;
Jie LI
;
Chunfeng LIU
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2006;10(33):171-173,封三
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND: Astrocyte distributes the most widely in the central nerve system,but the role of astrocyte in the pathogenisis of neurodegenerative diseases is still unclear.OBJECTIVE :To investigate the effects of lipopolysaccharide (LPS) on the level of the free intracelluar calcium, the production of reactive oxygen species (ROS) and nitric oxide (NO), and the cell viability of astrocytes.DESIGN: Randomized grouping controlled experiment.SETTING: The Brain Circulation Research Laboratory, Second Affiliated Hospital of Suzhou University.MATERIALS: The experiment was carried out in the Brain Circulation Research Laboratory, Second Affiliated Hospital of Suzhou University from May 2005 to August 2005. The astrocytes were derived from the cerebral cortex of 8 Newborn SD rats aged 1-3 days provided by the Animal Experimental Center of Suzhou University. LPS was purchased from Sigma, MTT assay kit and NO assay kit were obtained from Beyotime Biotechnology.METHODS: The astrocytes were isolated and cultured using the method described by Kevin St. P. McNaught. The cells were purified by proliferation repeatedly. Astrocytes were divided into control group, LPS5, 10, 20,40 mg/L group according to different dosage of LPS added. The cell viability at 30 min and 60 min respectively was measured using MTT method.The NO accumulation in cultured cell supernatant at 30 min was assayed with Griess reagent after the treatment of LPS. Calcium and ROS accumulation in cultured astrocyte of rats stimulated by different dosage of LPS was measured at 30 min and 60 min respectively by confocal laser scan microscope (CLSM).MAIN OUTCOME MEASURES: Cell viability of astrocyte, the change of level of NO, calcium and reactive oxygen species of astrocyte.RESULTS: ① The cell viability 60 min after administration of LPS 5 mg/L,LPS 10 mg/L and LPS 20 mg/L was higher than that of control group(P ＜ 0.05). The cell viability of LPS 10 mg/L group was higher than that of LPS 5, 20 mg/L group.② NO production 30 min after LPS treatment of LPS 40 mg/L group was higher than control and the other 3 experimental groups (P ＜ 0.05 ). ③ The calcium of the LPS 5, 10 and 20 mg/L group was 200-400 times higher than that of the base state. The calcium of the LPS 40 mg/L group was 1 000 times than its baseline state; Absolute fluorescence intensity of ROS was superior than the detect range of CLSM, we could speculate that the relative fluorescence intensity of the group LPS 40 mg/L was at least 300-400 times higher than that of its base state.CONCLUSION: LPS could increase the level of the free intracelluar calcium and induce the production of ROS and NO in astrocytes.
