Nobiletin Inhibits Hypoxia-Induced Placental Damage via Modulating P53 Signaling Pathway

In this study, we aimed to evaluate the effect of Nobiletin (NOB) on the placenta of Sprague–Dawley (SD) rats that had undergone reduced uterine perfusion pressure (RUPP) surgery and to evaluate the safety of NOB intervention during pregnancy. The results showed that NOB alleviated placental hypoxia, attenuated placental cell apoptosis, and inhibited placental damage in RUPP rats. No side effect of NOB intervention during pregnancy was observed. BeWo cell lines with P53 knockdown were then constructed using lentiviral transfection, and the P53 signaling pathway was found to be essential for NOB to reduce hypoxia-induced apoptosis of the BeWo cell lines. In summary, NOB attenuated hypoxia-induced placental damage by regulating the P53 signaling pathway, and those findings may contribute some insights into the role of NOB in placental development and the prevention of placental-related diseases.     

HIF-1α与VEGF在子宫内膜癌组织血管生成中的作用

目的：研究HIF-1α与VEGF在正常子宫内膜、子宫内膜不典型增生和子宫内膜癌组织中的表达情况及在肿瘤血管生成中的作用。方法：应用免疫组织化学SP法分别检测各类子宫内膜组织中的HIF-1α和VEGF的表达,并计数MVD。结果：子宫内膜癌HIF-1α阳性表达率63.6%（28/44）,正常子宫内膜为21.4%（3/14）,两者差异有统计学意义,P〈0.01。VEGF在子宫内膜癌、子宫内膜非典型增生和正常子宫内膜组织中的阳性率分别是56.8%、25.0%和14.3%,差异有统计学意义,P〈0.01。HIF-1α与VEGF均与MVD呈正相关（rs=0.579,P〈0.01;rs=0.607,P〈0.01）。结论：HIF-1α及其靶基因VEGF可能在子宫内膜病变由良性到恶性转变过程中,以及恶性肿瘤的进展中起了重要作用,可能作为肿瘤生物学行为的标志,并成为抗肿瘤治疗的新靶向。     

人脑胶质瘤组织HIF-1α和COX-2及CD105蛋白表达与临床病理学关系的研究

目的:探讨缺氧诱导因子-1α(HIF-1α)、环氧化酶-2(COX-2)及CD105在人脑胶质瘤组织中的表达及与临床病理特征的关系。方法:应用免疫组化SP法检测70例胶质瘤和10位正常人脑组织中HIF-1α、COX-2和CD105的表达,并测定CD105标记的MVD。结果:1)胶质瘤组织中HIF-1α、COX-2和CD105蛋白的阳性表达率分别为64.3%、61.4%和74.3%,明显高于正常人脑组织。2)HIF-1α、COX-2表达和MVD与胶质瘤病理分级密切相关。3)胶质瘤组织中HIF-1α和COX-2蛋白表达的一致符合率为70.35%,且两者表达呈正相关,r=0.5837,P=0.004。4)HIF-1α和COX-2阳性表达组的MVD(35.87±4.55和36.00±4.57)均高于阴性表达组(29.19±4.69和29.48±4.71),P值均为0.0000;两者均为阳性表达时的MVD(36.22±4.62)最大,明显高于其他各组(33.75±3.59和28.29±4.33),P=0.0001;两者表达均与MVD呈正相关,r值分别为0.5405和0.4652,P值分别为0.005和0.014。结论:HIF-1α、COX-2和CD105过表达可能在胶质瘤的恶性进展中发挥重要作用,并与肿瘤血管形成密切相关,三者的联合检测可更准确地揭示胶质瘤的生物学特性。     

Exosomes Derived from Glioma Cells under Hypoxia Promote Angiogenesis through Up-regulated Exosomal Connexin 43

Glioblastoma multiform (GBM) is a highly aggressive primary brain tumor. Exosomes derived from glioma cells under a hypoxic microenvironment play an important role in tumor biology including metastasis, angiogenesis and chemoresistance. However, the underlying mechanisms remain to be elucidated. In this study, we aimed to explore the role of connexin 43 on exosomal uptake and angiogenesis in glioma under hypoxia. U251 cells were exposed to 3% oxygen to achieve hypoxia, and the expression levels of HIF-1α and Cx43, involved in the colony formation and proliferation of cells were assessed. Exosomes were isolated by differential velocity centrifugation from U251 cells under normoxia and hypoxia (Nor-Exos and Hypo-Exos), respectively. Immunofluorescence staining, along with assays for CCK-8, tube formation and wound healing along with a transwell assay were conducted to profile exosomal uptake, proliferation, tube formation, migration and invasion of HUVECs, respectively. Our results revealed that Hypoxia significantly up-regulated the expression of HIF-1α in U251 cells as well as promoting proliferation and colony number. Hypoxia also increased the level of Cx43 in U251 cells and in the exosomes secreted. The uptake of Dio-stained Hypo-Exos by HUVECs was greater than that of Nor-Exos, and inhibition of Cx43 by ^37,43gap27 or lenti-Cx43-shRNA efficiently prevented the uptake of Hypo-Exos by recipient endothelial cells. In addition, the proliferation and total loops of HUVECs were remarkably increased at 24 h, 48 h, and 10 h after Hypo-Exos, respectively. Notably, ^37,43gap27, a specific Cx-mimetic peptide blocker of Cx37 and Cx43, efficiently alleviated Hypo-Exos-induced proliferation and tube formation by HUVECs. Finally, ^37,43gap27 also significantly attenuated Hypo-Exos-induced migration and invasion of HUVECs. These findings demonstrate that exosomal Cx43 contributes to glioma angiogenesis mediated by Hypo-Exos, and suggests that exosomal Cx43 might serve as a potential therapeutic target for glioblastoma.     

PC12细胞化学缺氧复氧损伤与缺氧诱导因子1表达的关系

目的 研究缺氧诱导因子1(HIF1)在PC12细胞化学缺氧复氧损伤中的作用。方法 在培养液中加入和去除氯化钴模拟化学缺氧和复氧，以乳酸脱氢酶(LDH)漏出和细胞超微结构改变作为细胞损伤指标，观察化学缺氧和复氧后不同时间细胞损伤和HIF1 α蛋白变化。结果 在氯化钴模拟化学缺氧实验中，LDH漏出明显增加，8h达高峰，随后逐渐下降，电镜结果与LDH改变相一致，HIF1 α蛋白表达在化学缺氧后2，4，8和12h均明显增加，8h达高峰，提示化学缺氧8h后细胞损伤逐渐减轻可能与HIF1 α蛋白水平升高有关。在模拟复氧实验中，LDH和细胞形态学改变都显示化学复氧8h细胞损伤最为严重，而HIF1 α蛋白表达在化学复氧4和8h均明显下降，提示细胞化学复氧损伤可能与HIF1 α蛋白水平下降有关。结论 HIF1对神经细胞化学缺氧复氧损伤具有保护作用。     

新生大鼠脑缺氧缺血后脑组织硝基酪氨酸测定

目的：探讨缺氧缺血(HI)后脑组织硝基酪氨酸形成的时相与分布特征及其与细胞死亡的关系。方法：采用7d龄Wistar大鼠42只制成HI脑损伤模型,分别于HI后30min、1h、3h、8h、14h、24h、72h处死6只,脑组织制作切片,取海马平面进行硝基酪氨酸和微管相关蛋白-2(MAP-2)免疫组化染色及硝基酪氨酸与发夹寡核苷酸探针(HPP)双染色,观察和计数皮层及僵核硝基酪氨酸阳性细胞数。以初生第7d(P7)和第10d(P10)的大鼠为对照。结果：硝基酪氨酸阳性细胞主要分布在Hl侧MAP-2阴性区域,包括皮层、丘脑、纹状体、海马、皮层下白质、脑室下区。在HI后早期,对照及对侧皮层血管内皮细胞及脑室下区也可偶见阳性细胞。在HI后30min皮层及僵核硝基酪氨酸阳性细胞计数明显升高并在HI后3h达到峰值,此后阳性细胞数逐渐下降;随时间的延长,阳性细胞逐渐缩小。荧光双染色结果显示,HI后早期硝基酪氨酸阳性细胞数明显高于HPP阳性细胞数,晚期则相反。结论：硝基酪氨酸形成与脑组织损伤密切相关,是反映细胞损伤的早期指标。     

黄连溶液对大鼠早期压疮缺血再灌注损伤中HIF-1α与VEGF的影响

目的:探讨黄连溶液对早期(Ⅰ、Ⅱ期)压疮缺血再灌注损伤中缺氧诱导因子-1α(HIF-1α)、血管内皮生长因子(VEGF)含量变化的影响。方法:SD大鼠40只随机分为4组,正常组、模型组、酒精组、黄连溶液组,每组10只。正常组大鼠备皮后不做任何处理,余各组采用埋置铁片外加磁铁施压的方法复制压疮模型;酒精组、黄连溶液组给予相应药物外涂。5 d后处死大鼠,取压疮创面皮肤肌肉组织,观察病理组织学变化,检测组织中HIF-1α、VEGF因子的表达。结果:病理组织学观察发现,模型组肌纤维萎缩,肌间隙间水肿明显,伴有大量炎性细胞浸润;酒精组镜下变化与模型组相似,而黄连溶液组细胞及组织水肿较模型组减轻、炎症细胞明显减少。组织中HIF-1α、VEGF表达显示,模型组较正常组增多,黄连溶液组干预后HIF-1α、VEGF表达较模型组进一步升高(P0.05),而酒精组与模型组无明显差异(P0.05)。结论:黄连溶液具有减轻早期压疮缺血再灌注损伤的作用,其机制可能与通过促进创伤组织HIF-1α及其下游因子VEGF的表达有关。     

Adv-HIF-1α^mu转染内皮祖细胞对CXCL12表达影响的研究

目的研究转染低氧诱导因子-1α突变型（hypoxiainduciblefactor-1α^mu,HIF-1α^mu）后的外源性内皮祖细胞（endothelialprogenitorcells,EPCs）中CXCL12的表达情况及对EPCs的影响。方法密度梯度离心结合差速贴壁法分离、培养EPCs并进行鉴定;以Adv—HIF-1α^mu的最佳转染指数转染EPCs,获得含有目的基因处理的EPCs细胞,设为A组。B组为单纯转染Adv的EPCs。设未转染Adv—HIF-1α^mu的EPCs为C组。WesternBlot检测转染后的EPCs中HIF-1的表达,检测转染后的EPCs培养液上清液中的CXCL12浓度及其对外源性EPCs迁移的影响。结果密度梯度离心结合差速贴壁法获得的EPCs能够结合UEA-1和摄取ac—LDL,并具有成血管作用;转染Adv—HIF-1α^mu的EPCs表达的HIF-1及CXCL12高于单纯转染Adv的EPCs及未转染Adv—HIF-1α^mu的EPCs（P〈0．05）,且能诱导外源性EPCs的迁移。结论密度梯度离心结合差速贴壁法能够分离、培养符合特征的EPCs,转染Adv—HIF-1α^mu的EPCs能够在常氧条件下高表达HIF-1及CXCL12,并诱导外源性EPCs的迁移。     

三仙信力胶囊对小鼠耐缺氧能力的实验研究

目的:探讨三仙信力胶囊对小鼠耐缺氧能力的影响.方法:对实验小鼠进行耐缺氧等实验观察.结果:三仙信力胶囊对小鼠缺氧等应激耐受性的影响显著,可明显延长实验动物在缺氧情况下的生存时间.结论:三仙信力胶囊具有明显的增强机体耐缺氧能力的作用.