首页|锰对人源性脉络丛乳头状瘤细胞系合成GDF-15的影响及机制研究

锰对人源性脉络丛乳头状瘤细胞系合成GDF-15的影响及机制研究

扫码查看
原文链接
  • NETL
  • NSTL
  • 万方数据
目的 应用人源性脉络丛乳头状瘤细胞系(hCPP)探究锰对脑脉络丛上皮细胞中生长分化因子-15(growth differentiation factor-15,GDF-15)合成的影响及其调控机制.方法 应用CCK-8试剂盒检测氯化锰(以下简称锰)处理后hCPP的细胞活力;不同浓度的锰(25、50、75、100、150和200 µmol/L)处理hCPP细胞24、48和72 h,通过Western blot、Real-Time PCR检测GDF-15蛋白表达和Gdf-15 mRNA水平在锰作用下的剂量和时间-效应关系.给予hCPP细胞100 μmol/L的锰处理24 h,通过Western blot检测p38 MAPK、NF-κB、AMPK、PI3K/AKT通路的激活情况,随后,给予相应的通路抑制剂后检测GDF-15的蛋白表达.100 µmol/L的锰处理hCPP细胞0、6、12、24、36、48、72 h,检测GDF-15、p38 MAPK蛋白表达的时间-效应关系.使用荧光探针试剂盒检测hCPP锰处理(100 μmol/L,24h)后细胞活性氧(reactive oxygen species,ROS)的含量,给予 ROS 抑制剂 N-acetylcysteine(NAC)后检测 GDF-15、p38 MAPK 蛋白表达.结果 用200 µmol/L以下的不同浓度的锰处理hCPP细胞24、48和72 h,可引起细胞活力降低、GDF-15蛋白合成增加、Gdf-15 mRNA水平增加,且随锰浓度的增高而变化.100 μmol/L的锰处理hCPP细胞24 h,通过增加p38 MAPK、NF-κB p65和AMPKα的磷酸化水平激活p38 MAPK、NF-κB和AMPK信号通路,给予SB 202190(p38 MAPK通路抑制剂)可逆转锰处理hCPP引起的GDF-15合成增加效应.100 µmol/L的锰处理hCPP细胞0、6、12、24、36、48和72 h,GDF-15蛋白水平和p38 MAPK磷酸化蛋白水平随着锰处理时间的增加而增高.100 μmol/L的锰处理hCPP细胞15、30、60 min以及24h,细胞ROS水平与对照组相比差异无统计学意义,给予NAC处理不能逆转锰引起的GDF-15合成增加及p38 MAPK通路的激活状态.结论 本研究表明,锰处理可增加hCPP细胞中GDF-15的合成,这可能是p38 MAPK信号通路激活后介导的.这些发现为今后阐明GDF-15在锰所致中枢神经系统的毒性效应及分子机制提供了新的线索和科学依据.
Exploring the influence and mechanism of manganese on GDF-15 in human-derived choroid plexus papilloma cell line
Objective This study aimed to investigate the effect and mechanism of manganese on Growth differentiation factor 15(GDF-15)in a newly established human-derived Choroid Plexus Papilloma Cell Line(hCPP).Methods The dose-dependent and time-related toxic effects of manganese on hCPP cell viability were assessed using the CCK-8 assay.The expression of GDF-15 protein and the level of Gdf-15 mRNA were examined through Western blot and Real-Time PCR,respectively.After treatment with 100 μmol/L MnCl2 for 24 hours,the phosphorylation levels of p38 MAPK,NF-κB p65,and AMPKα were evaluated through Western blot,followed by the detection of GDF-15 protein expression after treatment with corresponding pathway inhibitors.The content of reactive oxygen species(ROS)following manganese exposure(100 μmol/L,24 h)was measured using the fluorescent probe Dihydroethidium kit.Additionally,GDF-15 and p38 MAPK protein levels were assessed through Western blotting after treatment with N-acetylcysteine(NAC),a ROS inhibitor.Results Treatment of hCPP with different concentrations of manganese for 24 h,48 h,and 72 h led to time-and concentration-dependent decreases in cell viability,accompanied by an increase in GDF-15 protein synthesis and the level of Gdf-15 mRNA.Treatment with 100 µmol/L manganese for 24 h activated several signaling pathways,evidenced by increased phosphorylation levels of p38 MAPK,NF-κB p65,and AMPKα.Furthermore,treatment with SB 202190(a p38 MAPK pathway inhibitor)reversed the effect of manganese stimulation on hCPP-induced increase in GDF-15 synthesis.Stimulation of hCPP with 100 μmol/L manganese for various durations revealed time-dependent increases in GDF-15 protein and p38 MAPK protein phosphorylation levels.However,no significant difference in ROS levels was observed between the control group and cells treated with 100 μmol/L manganese for 15 min,30 min,60 min,and 24 h.The lack of ROS involvement was further supported by the failure of acetylcysteine treatment to reverse the manganese-induced increase in GDF-15 synthesis.Conclusion This study demonstrates that manganese treatment enhances the expression of GDF-15 in hCPP cells,likely mediated through the activation of the p38 MAPK pathway.These findings provide insight for future studies to elucidate the role of GDF-15 in manganese-mediated brain toxicity.

Growth differentiation factor-15(GDF-15)Manganese chlorideChoroid plexus(CP)Human-derived Choroid Plexus Papilloma Cell Line(hCPP)p38 MAPK pathway

张雨、张国艳、李子南、田永吉、宁钧宇、洪昭雄、高珊、谭壮生、敬海明、李国君

展开 >

首都医科大学公共卫生学院,北京 100069

北京市疾病预防控制中心,食物中毒诊断溯源技术北京市重点实验室,北京 100013

首都医科大学附属北京天坛医院小儿神经外科,北京 100070

美国国立卫生环境健康研究所神经生物学实验室,德汉姆北卡州27709

展开 >

生长分化因子-15(GDF-15) 氯化锰 脉络丛 人源性脉络丛乳头状瘤细胞系(hCPP) p38 MAPK信号通路

首都高层次公共卫生技术人才建设卫生毒理学科"领军人才"项目

02-03

2024

毒理学杂志
北京市预防医学研究中心 北京大学医学部公共卫生学院

毒理学杂志

CSTPCD
影响因子：0.504
ISSN：1002-3127
年,卷(期)：2024.38(3)