miR-217靶向MAPK1抑制胃癌细胞侵袭转移作用研究

目的：研究miR-217靶向MAPK1与胃癌侵袭转移的关系。方法在3个不同分化的胃癌细胞株MKN-28、SGC-7901、BGC-823中通过qRT-PCR技术检测miR-217情况,Western blot检测MAPK1表达,Transwell实验分析miR-217下调MAPK1表达后胃癌细胞侵袭转移能力改变。结果低分化人胃癌细胞株中MAPK1低表达,而miR-217高表达,中、高分化人胃癌细胞株中MAPK1高表达,而miR-217低表达（P＜0.05）;SGC-7901-miR-217mimics细胞株中miR-217表达增高,而MAPK1表达下调（P＜0.05）;而SGC-7901、SGC-7901-miR-Scramble、SGC-7901-miR-217-PM 3个细胞株中miR-217表达水平较低,而MAPK1表达水平较高（P＞0.05）。 SGC-790-miR-217 mimics侵袭细胞计数明显低于其他3组（P＜0.05）,SGC-7901、SGC-7901-miR-Scramble、SGC-7901-miR-217-PM 3组之间侵袭细胞计数无显著性差异（P＞0.05）。结论 miR-217能靶向结合MAPK1,降低其表达水平并抑制胃癌细胞的侵袭和转移。     

结肠癌患者Wnt信号通路的变化和作用

目的探讨结肠癌患者Wnt信号通路的变化情况及其作用意义。方法利用Human Ge-nome U133Plus2.0基因芯片对结肠正常黏膜组织和癌组织基因表达谱进行检测,分析Wnt信号通路基因表达变化情况。结果Wnt信号通路中FRP、Frizzled、CK2、p53、PP2A、CKIα、JNK、CaMKII、CaN、c-jun、cyc-D等关键基因均呈下调,调控着细胞周期、基因转录等。结论Wnt信号通路可能是调控结肠癌演变过程的重要信号通路之一。     

JNK信号通路通过介导细胞凋亡对肺纤维化的调控作用

目的观察肺纤维化中肺组织caspase-3和磷酸化c-Jun氨基末端激酶(p-JNK)蛋白表达的变化,并探讨JNK信号转导通路对肺纤维化的调控作用。方法 72只健康雄性Wistar大鼠随机分为正常对照组(N组)、肺纤维化模型组(M组),肺纤维化+JNK抑制剂干预组(I组),每组24只,分别于第3、7、14、28天处死,每组6只。留取肺组织,对其行苏木素-伊红(HE)染色,观察其病理学改变,碱水解法测定其中羟脯氨酸的含量,免疫组织化学法测肺组织中caspase-3和p-JNK的变化。结果模型组早期肺泡炎症明显,后期肺组织实变,caspase-3和p-JNK蛋白表达含量均显著增加(P<0.05),14d时达峰值。干预组能明显阻断JNK的激活(P<0.05),干预组caspase-3表达较模型组显著减少(P<0.05),14d时减少最明显。结论细胞凋亡是肺纤维化的一个重要病理组织学特点。JNK信号转导通路在肺纤维化中被激活,发挥促细胞凋亡效应。     

Ⅱ°烧烫伤模型大鼠体内TGF-β、p38MAPK、IL-1β的表达分析

目的:探讨Ⅱ°烧烫伤模型大鼠体内转化生长因子β(TGF-β)、p38丝裂原活化蛋白激酶(MAPK)和白细胞介素1β(IL-1β)的表达情况。方法:取大鼠分为4组,每组14只,分别为正常对照组、NaOH烧伤组、水烫伤组和乙醇烧伤组,分别建立相应的Ⅱ°烧烫伤模型,24 h后观察各组大鼠病理变化,酶联免疫吸附剂测定法检测各组大鼠建模后第1、4、7天血清中TGF-β、p38 MAPK和IL-1β的水平。结果:与正常对照组比较,其余3组大鼠均可见表皮及真皮有不同程度的缺损或坏死,炎性细胞向肌层组织浸润,其中水烫伤组和乙醇烧伤组大鼠皮下水肿明显;NaOH烧伤组大鼠血清中TGF-β和p38 MAPK水平在第4、7天均明显升高(P<0.05或P<0.01),IL-1β表达无明显变化;水烫伤组大鼠血清中TGF-β水平在第1、4、7天均明显升高(P<0.05),p38 MAPK和IL-1β水平在第4、7天明显升高(P<0.05或P<0.01);乙醇烧伤组大鼠血清中TGF-β水平在第4天明显升高(P<0.05),p38 MAPK和IL-1β水平在第1、4、7天均明显升高(P<0.05或P<0.01)。结论:水烫伤和乙醇烧伤可致大鼠Ⅱ°烧烫伤模型中TGF-β、p38 MAPK和IL-1β水平均有不同程度的升高,并可能激发TGF-β、p38 MAPK和IL-1β参与的炎症信号通路。     

17β雌二醇对人精子功能的非基因组调节效应及其机制的研究

目的：探讨17β雌二醇对人精子功能非基因组效应的可能机制。方法：运用计算机辅助精子分析系统与人精子穿透去透明带金黄地鼠卵异种体外受精实验,评价17β雌二醇对精子功能的调节作用；精子经信号转导途径抑制剂处理后,再采用流式细胞术检测精子胞内Ca~(2+)浓度的变化。结果：1μmol/L和5μmol/L E_2-BSA能提高人精子的A级精子百分比、平均曲线运动速度、平均直线运动速度、平均路径速度(P＜0．05)；自身受精率偏低的继发不育患者精子经1μmol/L和5μmol/L E_2-BSA作用后受精率提高(P＜0．05),但对本身受精率较高的精子影响不大(P＞0．05)；应用腺苷酸环化酶、磷脂酶C和酪氨酸蛋白激酶特异性抑制剂分别作用后均明显抑制E_2-BSA引起的胞内Ca~(2+)浓度的上升(P＜0．05)。结论：17β雌二醇能显著提高精子的运动能力和受精率,17β雌二醇可能通过腺苷酸环化酶、磷脂酶C或酪氨酸蛋白激酶信号转导途径实现对人精子功能调节的非基因组效应,为改善人精子功能提供了可能的新途径。     

mTOR信号通路与支气管肺发育不良

mTOR信号通路是在进化上高度保守的细胞内信号通路,参与多条信号通路的传导,主要包括 PI3K/AKT/mTOR 通路、AKT/TSC1-TSC2/Rheb/mTOR 通路、LKB1-AMPK-TSC-mTOR 通路和 FGF-10-Spry2-mTORC1-STAT3/HIF-1α-VEGF-A通路。该信号通路从多个水平多个方面参与肺发育及肺部多种疾病的调控过程,可能与支气管肺发育不良( bronchopulmonary dysplasia,BPD)有关。 BPD是早产儿十分常见的一种慢性肺疾病( chronic lung disease,CLD),是各种理化因素对发育不成熟肺造成急性肺损伤及损伤后异常修复、肺纤维化的过程。该文总结了mTOR信号通路与肺发育、急性肺损伤及肺纤维化可能存在的关系,探索mTOR信号通路在BPD形成过程中的作用,以期为BPD的防治提供新的切入点。     

Functional and molecular characterization of multiple K-Cl cotransporter isoforms in corneal epithelial cells

The dependence of regulatory volume decrease (RVD) activity on potassium-chloride cotransporter (KCC) isoform expression was characterized in corneal epithelial cells (CEC). During exposure to a 50% hypotonic challenge, the RVD response was larger in SV40-immortalized human CEC (HCEC) than in SV40-immortalized rabbit CEC (RCEC). A KCC inhibitor-[(dihydroindenyl)oxy] alkanoic acid (DIOA)-blocked RVD more in HCEC than RCEC. Under isotonic conditions, N-ethylmaleimide (NEM) produced KCC activation and transient cell shrinkage. Both of these changes were greater in HCEC than in RCEC. Immunoblot analysis of HCEC, RCEC, primary human CEC (pHCEC), and primary bovine CEC (BCEC) plasma membrane enriched fractions revealed KCC1, KCC3, and KCC4 isoform expression, whereas KCC2 was undetectable. During a hypotonic challenge, KCC1 membrane content increased more rapidly in HCEC than in RCEC. Such a challenge induced a larger increase and more transient p44/42MAPK activation in HCEC than RCEC. On the other hand, HCEC and RCEC p38MAPK phosphorylation reached peak activations at 2.5 and 15 min, respectively. Only in HCEC, pharmacological manipulation of KCC activity modified the hypotonicity-induced activation of p44/42MAPK, whereas p38MAPK phosphorylation was insensitive to such procedures in both cell lines. Larger increases in HCEC KCC1 membrane protein content correlated with their ability to undergo faster and more complete RVD. Furthermore, pharmacological activation of KCC increased p44/42MAPK phosphorylation in HCEC but not in RCEC, presumably a reflection of low KCC1 membrane expression in RCEC. These findings suggest that KCC1 plays a role in (i) maintaining isotonic steady-state cell volume homeostasis, (ii) recovery of isotonic cell volume after a hypotonic challenge through RVD, and (iii) regulating hypotonicity-induced activation of the p44/42MAPK signaling pathway required for cell proliferation.     

Rho/Rock信号转导通路中的关键因子RhoA、Rock Ⅰ、p-LMC在后发性白内障动物模型中的表达

目的 建立SD大鼠后囊膜混浊(posterior capsular opacification,PCO)动物模型,检测Rho/Rock通路中的关键因子RhoA、Rho激酶Ⅰ(Rho associated coiledcoil forming protein kinase-Ⅰ,Rock Ⅰ)和肌球蛋白轻链(phosphorylated myosin light chain,p-MLC)在PCO中的表达.探讨Rho/Rock通路在PCO形成过程中的作用.方法 选用健康SD大鼠30只,随机分为对照组(0d)和实验组(3d、7d、14 d、28 d),每个时间点各6只.所有大鼠(右眼)均行晶状体囊外摘出术,显微镜下观察不同时间点术眼前节的炎症反应及PCO发生情况.ELISA法检测不同时间点房水中TGF-β的表达.处死大鼠后摘除眼球行组织病理学检查,免疫组织化学和Western Blot法检测RhoA、Rock Ⅰ、p-MLC在晶状体后囊上的表达.结果 随着术后观察时间的推移,PCO混浊程度逐渐增加,HE染色显示晶状体上皮细胞不同程度的增生、移行.对照组术后0d、3d、7d、14 d、28 d房水中TGF-β的表达量分别为(37.28±1.31)μg·L-1、(40.29±3.17) μg·L-1、(42.12±3.45) μg·L-1、(60.42±4.75) μg·L-1、(68.69±8.06)μg·L-1,实验组各时间点与对照组相比,差异有统计学意义(F=137.158,P＜0.05);不同时间点两两相比,差异均有统计学意义(均为P＜0.05).术后0d、3d、7d、14 d和28 d晶状体后囊膜上RhoA蛋白相对表达量(RhoA/GAPDH)分别为1.00±0.00、1.49±0.01、1.57±0.03、1.77±0.02和1.97±0.02,实验组各时间点与对照组相比,差异有统计学意义(F=94.546,P＜0.05);不同时间点两两相比,差异均有统计学意义(均为P＜0.05).Rock Ⅰ蛋白的表达量(Rock Ⅰ/GAPDH)分别为1.00±0.00、0.35±0.12、1.42 ±0.14、1.78±0.09和1.95±0.14,实验组各时间点与对照组相比,差异有统计学意义(F=30.942,P＜0.05);不同时间点两两相比,差异均有统计学意义(均为P＜0.05).p-MLC蛋白的相对表达量(p-MLC/GAPDH)分别为1.00±0.00、1.44±0.08、1.47±0.13、1.63±0.10和2.12±0.20,实验组各时间点与对照组相比,差异有统计学意义(F =34.313,P＜0.05);不同时间点两两相比,差异均有统计学意义(均为P＜0.05).结论 RhoA、Rock Ⅰ、p-MLC在PCO形成过程中表达逐渐增多,Rho/Rock信号转导通路可能与PCO的发生、发展有关.     

Cancer chemoprevention by tea polyphenols through modulating signal transduction pathways

The action mechanisms of several chemopreventive agents derived from herbal medicine and edible plants have become attractive issues in cancer research. Tea is the most widely consumed beverage worldwide. Recently, the cancer chemopreventive actions of tea have been intensively investigated. It have been demonstrated that the active principles of tea were attributed to their tea polyphenols. Recently, tremendous progress has been made in elucidating the molecular mechanisms of cancer chemoprevention by tea and tea polyphenols. The suppression of various tumor biomarkers including growth factor receptor tyrosine kinases, cytokine receptor kinases, PI3K, phosphatases, ras, raf, MAPK cascades, N x FB, I x B kinase, PKA, PKB, PKC, c-jun, c-fos, c-myc, cdks, cyclins, and related transducing proteins by tea polyphenols has been studied in our laboratory and others. The I x B kinase (IKK) activity in LPS-activated murine macrophages (RAW 264.7 cells) was found to be inhibited by various tea polyphenols including (-) epigallocatechin-3-gallate (EGCG), theaflavin (TF-1), theaflavin-3-gallate (TF-2) and theaflavin-3,3'-digallate (TF-3). TF-3 inhibited IKK activity in activated macrophages more strongly than did the other tea polyphenols. TF-3 inhibited both IKK1 and IKK2 activity and prevented the degradation of I x B x and I x B x in activated macrophage cells. The results suggested that the inhibition of IKK activity by TF-3 and other tea polyphenols could occur by a direct effect on IKKs or on upstream events in the signal transduction pathway. TF-3 and other tea polyphenols blocked phosphorylation of IB from the cytosolic fraction, inhibited NFB activity and inhibited increases in inducible nitric oxide synthase levels in activated macrophage. TF-3 and other tea polyphenols also inhibited strongly the activities of xanthine oxidase, cyclooxygenase, EGF-receptor tyrosine kinase and protein kinase C. These results suggest that TF-3 and other tea polyphenols may exert their cancer chemoprevention through suppressing tumor promotion and inflammation by blocking signal transduction. The mechanisms of this inhibition may be due to the blockade of the mitogenic and differentiating signals through modulating EGFR function, MAPK cascades, NFkappaB activation as well as c-myc, c-jun and c-fos expression.     

MAPK信号转导通路在糖皮质激素耐药白血病中的研究进展

丝裂原活化蛋白激酶(MAPK)信号通路作为信号传递网络中的重要途径之一,在细胞凋亡及生存中发挥重要作用,其中最主要的是ERK、JNK及p38 MAPK途径。近来研究发现,MAPK信号转导通路与白血病的发生发展及耐药的产生有密切关系,将有可能成为白血病治疗的新靶点。本文主要综述了MAPK信号转导通路与白血病发病、治疗作用机制及与糖皮质激素耐药的关系。