紫花牡荆素对H446细胞系肺癌干样细胞球形成能力和p-Akt蛋白表达的影响(R)医学论坛网-网聚医学的力量
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紫花牡荆素对H446细胞系肺癌干样细胞球形成能力和p-Akt蛋白表达的影响
&&&&&&&&&　　近期，湖南省胸科医院研究人员发表论文，旨在研究紫花牡荆素（canstin，CAS）抑制人小细胞肺癌H446细胞系肺癌干细胞样细胞球形成作用，探讨其作用机制是否涉及降低Akt磷酸化蛋白表达水平。研究指出，CAS具有抑制H446细胞系干细胞样细胞球形成作用；Akt特异性抑制剂有效增强CAS抑制肺癌干细胞样细胞球形成与Akt蛋白磷酸化。该文发表在2015年第03期《湖南师范大学学报(医学版)》杂志
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　　近期，湖南省胸科医院研究人员发表论文，旨在研究（canstin，CAS）抑制人H446细胞系球形成作用，探讨其作用机制是否涉及降低Akt磷酸化蛋白表达水平。研究指出，CAS具有抑制H446细胞系干细胞样细胞球形成作用；Akt特异性抑制剂有效增强CAS抑制肺癌干细胞样细胞球形成与Akt蛋白磷酸化。该文发表在2015年第03期《湖南师范大学学报(医学版)》杂志上。
　　肿瘤球形成法获得H446细胞系第2代球形成细胞，并检定CAS、及与LY294002合用对肿瘤球形成率的影响。Western&blot分析、Akt蛋白表达。
　　CAS以浓度依赖方式降低H446细胞系第2代球形成细胞肿瘤球形成率和Akt蛋白磷酸化水平。LY.0&mol/L）增强CAS抑制球形成以及下调p-Akt蛋白表达作用。
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邮&&&&箱：上传用户：vpnnzobcxa资料价格：5财富值&&『』文档下载 ：『』&&『』学位专业：&关 键 词 ：&&&&&权力声明：若本站收录的文献无意侵犯了您的著作版权，请点击。摘要:（摘要内容经过系统自动伪原创处理以避免复制，下载原文正常，内容请直接查看目录。）目标：采取年夜鼠局灶性脑缺血再灌注模子商量长途创伤预处置(remote preconditioning of trauma, RPCT)对脑缺血再灌注毁伤的影响,研讨p-Akt和caspase-3在长途创伤预处置脑掩护机制中的感化。办法：(一)70只安康雄性SD年夜鼠,体重250-300g,随机分红七组(n=10)：假手术组(Sham组)、缺血再灌注组(I/R组)、RPCT0.5h组、RPCT1h组、RPCT2h组、RPCT4h组和RPCT24h组。用以测定脑梗逝世容积及神经功效缺损评分。(二)40只安康雄性SD年夜鼠随机分红四组(n=10)Sham、I/R组、RPCT1h组和RPCT24h组。用以组织形状学、免疫组化及Western Blot法检测。①Sham组：仅分别年夜鼠颈部血管,不拔出拴线壅塞年夜脑中动脉。②I/R组：采取Longa线栓法制造MCAO模子办法树立局灶性脑缺血再灌注模子。③RPCT组：在MCAO开端前分离0.5h、1h、2h、4h和24h做2cm腹部正中瘦语并切开腹壁全层,随即缝合伤口。各组年夜鼠均于缺血再灌注后不雅察24h,在24h对年夜鼠行神经功效缺损评分；行TTC染色法测定脑梗逝世容积；HE染色法不雅察脑组织形状；Nissl染色法不雅察神经细胞存活情形；免疫组化及Western Blot法不雅察并测定缺血半影区Akt、p-Akt及caspase-3卵白的表达。成果：①TTC染色：Sham组未见脑梗逝世；与I/R组比拟,RPCT1h组、RPCT2h组、RPCT4h组及RPCT24h组脑梗逝世容积明显减小(P0.05)。②神经功效缺损评分：RPCT1h组、RPCT2h组、RPCT4h组和RPCT24h组Garcia评分较I/R组明显进步(P0.05)。③HE染色：Sham组年夜鼠年夜脑皮质神经元形状正常；与I/R组比拟,RPCT1h组及RPCT24h组缺血半影区神经元数量较多,条理较清晰,细胞形状较完全,细胞间质水肿加重；I/R组、RPCT1h组及RPCT24h组缺血中间区均涌现细胞坏逝世,神经元数量削减,核固缩深染,组织松散。④Nissl染色：Sham组年夜脑皮质神经元形状正常；与I/R组比拟,RPCT1h组及RPCT24h组缺血半影区存活神经元数目明显增长(P0.05)；I/R组、RPCT1h组及RPCT24h组p-Akt卵白和caspase-3卵白在缺血半影区的表达均明显高于Sham组(P<0.05)；与I/R组比拟,RPCT1h组及RPCT24h组缺血半影区的p-Akt表达明显增长(P<0.05),而caspase-3表达明显削减(P<0.05)。结论：长途创伤预处置可以或许加重年夜鼠局灶性脑缺血再灌注毁伤,其机制能够与激活Akt卵白、克制caspase-3表达有关。Abstract:Objective: take the eve of the rats with focal cerebral ischemia re perfusion model to discuss long trauma pre disposal (remote preconditioning of trauma, RPCT) on cerebral ischemia and reperfusion damage effect and research of p-Akt and Caspase-3 in long-distance trauma preconditioning cerebral protective mechanism in action. Approach: (a) 70 healthy male SD big rat, weight 250-300g, stochastic dividend seven groups (n = 10): sham operation group (sham group), ischemia and reperfusion group (I / R group), group RPCT0.5h, RPCT1h, RPCT2h, RPCT4h group and RPCT24h group. In order to measure the cerebral infarction volume and neurological deficit score died. (two) 40 Ankang male SD rats were randomly divided into four groups (n=10) Sham, I/R group, RPCT1h group and RPCT24h group. Tissue shape, immunohistochemistry and Blot Western assay. Group Sham: the rats were only neck vessels, not pull out the line congestion in the brain artery. I/R group: take the Longa line bolt method to make the MCAO model method to establish the focal cerebral ischemia reperfusion model. RPCT group: before the start of MCAO 0.5h, 1H, 2h, 4h, 24h and to do 2cm abdominal median thin language and open the whole layer of abdominal wall, then suture the wound. The eve of the rats in each group were in ischemia again after perfusion not Yacha 24h, in 24 hours on the eve of the rat nerve fu TTC staining method determination of the volume of cere he staining observations o Nissl staining not Yacha neural cel observations immunohistochemical and Western blot method and determination of ischemic penumbra of Akt, p-Akt and caspase-3 protein expression. Results: 1. TTC staining: sham group ha with I / R group match, RPCT1h, RPCT2h, RPCT4h group and RPCT24h cerebral infarction group died volume decreased significantly (P & 0.05). RPCT0.5h group and I / R group, the cerebral infarction died volume without significant difference (P & 0.05). 2. Nerve function deficit score: group RPCT1h, RPCT2h, RPCT4h group and group RPCT24h Garcia score than those in I / R group significantly increased (P & 0.05). RPCT0.5h group and I / R group Garcia score no si longa five scores in each group no significant difference (P & 0.05). 3 HE staining: normal sham group rat brain of the eve of the co compared with I / R group, RPCT1h group and RPCT24h group ischemia penumbra neuron number, consecution is clear, cell shape is complete, intercellu intermediate zone of I / R group, RPCT1h group and group RPCT24h ischemia were emerging bad cell death, reduction in the number of neurons, karyopyknosis hyperchromatic, loosely organized. Nissl (4): sham group cerebral cortical neurons of normal shape. Compared with I / R group, RPCT1h group and group RPCT24h ischemia penumbra zone survival number of neurons increased significantly (P & 0.05). Fifthly, immunohistochemistry and Western blot results: Akt in ischemic penumbra expression stability, each group no statistical difference (P & 0.05); I / R group, RPCT1h group and RPCT24h group of p-Akt protein and caspase-3 protein expression in the ischemic penumbra were significantly higher than those in sham group (P & 0.05); with I / R group than intended, the penumbra area group and RPCT24h ischemia group RPCT1h p-Akt expression increased significantly (P & 0.05), and the expression of Caspase-3 significantly reduced (P & 0.05). Conclusion: pretreatment of the long-distance trauma may have heightened the eve of the rats with focal cerebral ischemia reperfusion injury, the mechanism can and activation of Akt protein and restraint on expression of Caspase-3.目录:摘要4-6ABSTRACT6-7英文縮略词表10-11第一章 前言11-13第二章 材料与方法13-22&&&&2.1 实验材料13-14&&&&&&&&2.1.1 实验动物13&&&&&&&&2.1.2 主要实验仪器与器材13-14&&&&&&&&2.1.3 主要实验试剂与耗材14&&&&2.2 实验方法14-21&&&&&&&&2.2.1 实验分组14-16&&&&&&&&2.2.2 MCAO模型建立16&&&&&&&&2.2.3 远程创伤预处理方法16&&&&&&&&2.2.4 神经功能缺损评分16-17&&&&&&&&2.2.5 脑梗死体积的测量17&&&&&&&&2.2.6 组织切片制备17-18&&&&&&&&2.2.7 HE染色18&&&&&&&&2.2.8 Nissl染色18&&&&&&&&2.2.9 免疫组化法检测缺血半影区Akt、p-Akt及caspase-3的表达18-19&&&&&&&&2.2.10 Western blot法检测缺血半影区Akt、p-Akt及caspase-3的表达19-21&&&&2.3 统计学分析21-22第三章 结果22-33&&&&3.1 七组大鼠TTC染色结果及梗死容积测定22-23&&&&3.2 脑组织HE染色结果23-24&&&&3.3 脑组织Nissl染色结果24-25&&&&3.4 脑组织Akt、p-Akt和caspase-3免疫组化及Western blot检测结果25-31&&&&&&&&3.4.1 脑组织缺血半影区Akt的表达25-27&&&&&&&&3.4.2 脑组织缺血半影区p-Akt的表达27-29&&&&&&&&3.4.3 脑组织缺血半影区caspase-3的表达29-31&&&&3.5 大鼠神经功能缺损评分(NDS评分)31-33第四章 讨论33-39&&&&4.1 远程创伤预处理的实施与脑保护作用33-35&&&&&&&&4.1.1 远程创伤预处理的的实施33-34&&&&&&&&4.1.2 远程创伤预处理的脑保护作用34-35&&&&4.2 脑缺血再灌注损伤与细胞凋亡35-36&&&&4.3 PI3K/Akt信号通路与脑保护36-39第五章 结论39-40参考文献40-47综述47-61&&&&参考文献55-61致谢61分享到：相关文献|扫二维码下载作业帮
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为什么要磷酸化,像AKT,跑出来的western 图AKT就没什么变化,而p-AKT就会减少?真不懂!
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Akt的Ser473可以被PDK1磷酸化而活化,介导下游信号传导,引起细胞的抗凋亡、抑自噬等生物学效应；经western blot 检测后,总的Akt变化可能不大,PI3K-Akt通路被激活时,p-Akt会增加；如果量减少,可能是信号通路过程中蛋白分子的调节作用,用于信号传导的中断.仅供参考.
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