目的:研究第10号染色体缺失的磷酸酶张力蛋白同源物基因(phosphatase and tensin homologue deleted on chromosome ten,PTEN)/磷脂酰肌醇3-激酶(phosphoinositide 3-kinase,PI3K)在肝缺血再灌注损伤中的调控作用及可能的机制.方法:夹闭...目的:研究第10号染色体缺失的磷酸酶张力蛋白同源物基因(phosphatase and tensin homologue deleted on chromosome ten,PTEN)/磷脂酰肌醇3-激酶(phosphoinositide 3-kinase,PI3K)在肝缺血再灌注损伤中的调控作用及可能的机制.方法:夹闭C57BL/6J小鼠的肝动脉和门静脉以阻断肝脏向头侧肝叶血供90 min,随后取下血管夹恢复血供,建立肝缺血再灌注损伤模型.实验分为四组:正常对照组(sham)、缺血再灌注模型组(IR)、PTEN抑制剂bpv(HOpic)干预组(BPV+IR)、PI3K抑制剂wortmannin干预组(WM+IR).检测小鼠血清ALT水平,行肝组织病理检查,以评估抑制PTEN/PI3K对肝脏IRI的影响.采用Western blot法检测p-AKT、p-GSK3β的表达,分析抑制PTEN/PI3K对其下游关键效应分子AKT/GSK3β磷酸化的调控.采用定量PCR法检测炎性因子IL-12p40、IL-10及TNF-α的基因表达,分析抑制PTEN/PI3K对TLR4介导的炎症反应的影响.结果:血清ALT及肝组织学改变均提示,抑制PTEN使IR诱导的肝损害明显减轻,相反,抑制PI3K则使IRI加剧.随着再灌注的发生,缺血期去磷酸化的AKT/GSK3β逐渐恢复其磷酸化活性.抑制PTEN增强了再灌注触发的AKT/GSK3β磷酸化,而阻断PI3K则使其明显削弱.阻断PTEN抑制了促炎基因IL-12p40、TNF-α的表达,并使IL-12/IL-10比值下调,该结果与抑制PI3K产生的效应完全相反.结论:PTEN/PI3K在肝缺血再灌注损伤中发挥重要的调控作用,PTEN抑制和/或PI3K活化可能通过上调p-AKT/p-GSK3β以及抑制炎症反应明显减轻肝缺血再灌注损伤.展开更多
目的通过癌症基因组图谱(The Cancer Genome Atlas,TCGA)数据库分析弥漫大B细胞淋巴瘤(diffuse large B-cell lymphoma,DLBCL)中铁死亡相关基因的表达及其与程序性死亡受体配体-1(programmed death ligand-1,PD-L1)和免疫细胞的关系,为D...目的通过癌症基因组图谱(The Cancer Genome Atlas,TCGA)数据库分析弥漫大B细胞淋巴瘤(diffuse large B-cell lymphoma,DLBCL)中铁死亡相关基因的表达及其与程序性死亡受体配体-1(programmed death ligand-1,PD-L1)和免疫细胞的关系,为DLBCL的治疗提供新的靶标。方法通过TCGA数据库查找获得22个铁死亡相关基因。从TCGA数据库获取48例DLBCL(DLBCL组)及54例反应性淋巴结增生患者(对照组)淋巴结标本的铁死亡相关基因以及PD-L1的表达数据。使用Wilcoxon秩和检验进行组间差异性表达分析。基因表达相关性分析采用Spearman相关性分析。采用R软件包pheatmap分析DLBCL中铁死亡相关基因表达与免疫细胞的相关性。采用R软件GSVA包分析铁死亡相关基因表达与磷脂酰肌醇-3-激酶-蛋白激酶B-哺乳动物雷帕霉素靶蛋白(phosphatidylinositol 3 kinase-protein kinase B-mammalian target of rapamycin,PI3K-Akt-mTOR)信号通路的相关性。结果DLBCL中周期素依赖性激酶抑制因子1A(cyclin dependent kinase inhibitor 1A,CDKN1A)、70 kDa热休克蛋白5(heat shock 70 kDa protein 5,HSPA5)、内质膜蛋白复合体亚基2(endoplasmic membrane protein complex subunit 2,EMC2)、溶质载体家族7成员11(solute carrier family 7,member 11,SLC7A11)、金属硫蛋白1G(metallothionein 1G,MT1G)、热休克蛋白B1(heat shock protein B1,HSPB1)、谷胱甘肽过氧化酶4(glutathione peroxidase4,GPX4)、范可尼贫血互补群D2(Fanconi anemia complementary group D2,FANCD2)、柠檬酸合成酶(citrate synthase,CS)、CDGSH铁硫结构域1(CDGSH iron sulfur domain 1,CISD1)、法尼基二磷酸法尼基转移酶1(farnesyl diphosphate farnesyltransferase 1,FDFT1)、SLC1A5、转铁蛋白受体(transferrin receptor,TFRC)、核糖体蛋白L8(ribosomal protein L8,RPL8)、核受体共激活因子4(nuclear receptor coativator 4,NCOA4)、二肽基肽酶Ⅳ(dipeptidyl peptidaseⅣ,DPP4)和花生四烯酸15脂氧合酶(arachidonate-15-lipoxygenase,ALOX15)基因表达均上调(均P<0.05)。免疫细胞相关分析显示,铁死亡相关基因可激活体内巨噬细胞M1(P<0.05)。DLBCL中长链脂酰辅酶A合成酶4(acyl-CoA synthetase long chain family member 4,ACSL4)、CDKN1A、DPP4、EMC2、谷氨酰胺酶2(glutaminase 2,GLS2)、HSPA5、溶血卵磷脂酰基转移酶3(lysophosphatidylcholine acyltransferase 3,LPCAT3)、MT1G、NCOA4、红细胞衍生核因子2样蛋白2(nuclear factor erythroid 2-like-2,NFE2L2)、精脒/精胺N1-乙酰基转移酶1(spermidine/spermine N1-acetyltransferase 1,SAT1)、SLC7A11和TFRC这些铁死亡相关基因的表达均与PD-L1表达呈正相关(均r>0.4,均P<0.05)。铁死亡相关基因LPCAT3、NCOA4和TFRC的表达均与PI3K-AktmTOR通路呈正相关(均r>0.4,均P<0.05)。结论多数铁死亡相关基因在DLBCL组织中高表达,且与PD-L1、免疫浸润及PI3K-Akt-mTOR通路有关。展开更多
Approaches for controlling inflammatory responses and reducing the mortality rate of septic patients remain clinically ineffective; new drugs need to be identified that can induce anti-inflammatory responses. Ephedrin...Approaches for controlling inflammatory responses and reducing the mortality rate of septic patients remain clinically ineffective; new drugs need to be identified that can induce anti-inflammatory responses. Ephedrine hydrochloride （EH） is a compound that is widely used in cardiovascular diseases, especially to treat hypotension caused by either anesthesia or overdose of antihypertensive drugs. In this study, we reported that EH also plays an important role in the control of the inflammatory response. EH increased IL-IO and decreased proinflammatory cytokine （IL-6, tumor-necrosis factor （TNF）-a, IL-12 and IL-11~） expression in primary peritoneal macrophages and Raw264.7 cells treated with peptidoglycan （PGN）, a Gram-positive cell wall component. The anti-inflammatory role of EH was also demonstrated in an experimental mouse model of peritonitis induced by intraperitoneal PGN injection. The phosphatidylinositol 3-kinase （PI3K）/Akt pathway was found to be responsible for the EH-mediated increase in IL-IO production and decrease in IL-6 expression. Therefore, our results illustrated that EH can help maintain immune equilibrium and diminish host damage by balancing the production of pro- and anti-inflammatory cytokines after PGN challenge. EH may be a new potential anti-inflammatory drug that can be useful for treating severe invasive Gram-positive bacterial infection.展开更多
[目的]探究shRNA靶向敲低Cx43基因表达对人多发性骨髓瘤细胞株MM.1S生存的影响及其机制。[方法]针对Cx43 mRNA不同位点构建sh/Cx43-1、sh/Cx43-2、sh/Con重组质粒,感染体外培养的MM.1S细胞,RT-PCR或Western Blotting检测Cx43 mRNA或蛋...[目的]探究shRNA靶向敲低Cx43基因表达对人多发性骨髓瘤细胞株MM.1S生存的影响及其机制。[方法]针对Cx43 mRNA不同位点构建sh/Cx43-1、sh/Cx43-2、sh/Con重组质粒,感染体外培养的MM.1S细胞,RT-PCR或Western Blotting检测Cx43 mRNA或蛋白表达,CCK-8、流式细胞术检测细胞增殖、凋亡情况,Western Blotting检测磷脂酸肌醇-3-激酶(PI3K)、丝苏氨酸蛋白激酶(AKT)通路蛋白表达。[结果]sh/Cx43-1组、sh/Cx43-2组Cx43 mRNA或蛋白均低于sh/Con组,且sh/Cx43-1组低于sh/Cx43-2组(0.18±0.05 vs 0.75±0.07,0.21±0.03 vs 0.81±0.06,P<0.05);培养24h时各组细胞增殖水平无明显差异(P>0.05),培养48 h、72 h时sh/Cx43-1组、sh/Cx43-2组细胞增殖水平均低于sh/Con组(0.42±0.05 vs 0.58±0.09,1.02±0.10 vs 1.08±0.13,P<0.05);sh/Cx43-1组、sh/Cx43-2组细胞总凋亡率高于sh/Con组,PI3K、AKT蛋白表达均低于sh/Con组,且sh/Cx43-1组变化较sh/Cx43-2组更明显(P<0.05)。[结论]通过shRNA靶向敲低Cx43基因表达能显著抑制MM.1S细胞增殖,促进其凋亡,这可能与敲低Cx43后PI3K/AKT通路受到抑制有关。展开更多
目的探究m iR 451对结肠癌移植瘤发生和发展的抑制作用。方法裸鼠随机分为M ock组和miRW51组,记录裸鼠存活率和肿瘤体积变化;检测肿瘤重量;qRT-PCR检测m iR 451表达水平;免疫组化检测Ki67、Caspase-3、MMP-2、VEGF、P13K、p-A K T和p-m...目的探究m iR 451对结肠癌移植瘤发生和发展的抑制作用。方法裸鼠随机分为M ock组和miRW51组,记录裸鼠存活率和肿瘤体积变化;检测肿瘤重量;qRT-PCR检测m iR 451表达水平;免疫组化检测Ki67、Caspase-3、MMP-2、VEGF、P13K、p-A K T和p-mTOR表达水平。结果结肠癌癌组织miR-451水平显著低于癌旁组织;结肠癌细胞中m iR 451水平显著低于NCM460细胞。与MOCK组相比,miK-451组裸鼠存活率显著上升,肿瘤体积显著减小,肿瘤重量显著降低,肿瘤组织中m iR 451水平显著升高,Ki67、MMP-2、VEGF、P13K、p-A K T和P-mTOR表达水平显著降低,Caspase-3表达水平显著升高。结论m iR 451抑制结肠癌移植瘤的生长和发展,其作用机制可能与P13K/AKT通路的抑制相关。展开更多
基金This work was supported by grants from the National Key Basic Research Program of China （2010CB529901 and 2010CB530600）, the National Natural Science Foundation of China （31100619 and 30972705）, the China Postdoctoral Science Foundation （20110490186）, the Chen-guang Plan Project of Shanghai Educational Municipal Education Commission （11CG48 and szyl0004）, the Specialized Research Fund for the Doctoral Program of Higher Education （20113107120014） and the Leading Academic Discipline Project of Shanghai Municipal Education Commission （J50301） （Yuejuan Zheng, Shanghai University of T.C.M）.
文摘Approaches for controlling inflammatory responses and reducing the mortality rate of septic patients remain clinically ineffective; new drugs need to be identified that can induce anti-inflammatory responses. Ephedrine hydrochloride （EH） is a compound that is widely used in cardiovascular diseases, especially to treat hypotension caused by either anesthesia or overdose of antihypertensive drugs. In this study, we reported that EH also plays an important role in the control of the inflammatory response. EH increased IL-IO and decreased proinflammatory cytokine （IL-6, tumor-necrosis factor （TNF）-a, IL-12 and IL-11~） expression in primary peritoneal macrophages and Raw264.7 cells treated with peptidoglycan （PGN）, a Gram-positive cell wall component. The anti-inflammatory role of EH was also demonstrated in an experimental mouse model of peritonitis induced by intraperitoneal PGN injection. The phosphatidylinositol 3-kinase （PI3K）/Akt pathway was found to be responsible for the EH-mediated increase in IL-IO production and decrease in IL-6 expression. Therefore, our results illustrated that EH can help maintain immune equilibrium and diminish host damage by balancing the production of pro- and anti-inflammatory cytokines after PGN challenge. EH may be a new potential anti-inflammatory drug that can be useful for treating severe invasive Gram-positive bacterial infection.
文摘[目的]探究shRNA靶向敲低Cx43基因表达对人多发性骨髓瘤细胞株MM.1S生存的影响及其机制。[方法]针对Cx43 mRNA不同位点构建sh/Cx43-1、sh/Cx43-2、sh/Con重组质粒,感染体外培养的MM.1S细胞,RT-PCR或Western Blotting检测Cx43 mRNA或蛋白表达,CCK-8、流式细胞术检测细胞增殖、凋亡情况,Western Blotting检测磷脂酸肌醇-3-激酶(PI3K)、丝苏氨酸蛋白激酶(AKT)通路蛋白表达。[结果]sh/Cx43-1组、sh/Cx43-2组Cx43 mRNA或蛋白均低于sh/Con组,且sh/Cx43-1组低于sh/Cx43-2组(0.18±0.05 vs 0.75±0.07,0.21±0.03 vs 0.81±0.06,P<0.05);培养24h时各组细胞增殖水平无明显差异(P>0.05),培养48 h、72 h时sh/Cx43-1组、sh/Cx43-2组细胞增殖水平均低于sh/Con组(0.42±0.05 vs 0.58±0.09,1.02±0.10 vs 1.08±0.13,P<0.05);sh/Cx43-1组、sh/Cx43-2组细胞总凋亡率高于sh/Con组,PI3K、AKT蛋白表达均低于sh/Con组,且sh/Cx43-1组变化较sh/Cx43-2组更明显(P<0.05)。[结论]通过shRNA靶向敲低Cx43基因表达能显著抑制MM.1S细胞增殖,促进其凋亡,这可能与敲低Cx43后PI3K/AKT通路受到抑制有关。
文摘目的探究m iR 451对结肠癌移植瘤发生和发展的抑制作用。方法裸鼠随机分为M ock组和miRW51组,记录裸鼠存活率和肿瘤体积变化;检测肿瘤重量;qRT-PCR检测m iR 451表达水平;免疫组化检测Ki67、Caspase-3、MMP-2、VEGF、P13K、p-A K T和p-mTOR表达水平。结果结肠癌癌组织miR-451水平显著低于癌旁组织;结肠癌细胞中m iR 451水平显著低于NCM460细胞。与MOCK组相比,miK-451组裸鼠存活率显著上升,肿瘤体积显著减小,肿瘤重量显著降低,肿瘤组织中m iR 451水平显著升高,Ki67、MMP-2、VEGF、P13K、p-A K T和P-mTOR表达水平显著降低,Caspase-3表达水平显著升高。结论m iR 451抑制结肠癌移植瘤的生长和发展,其作用机制可能与P13K/AKT通路的抑制相关。
