microRNA-192在晚期糖基化终产物诱导人腹膜间皮细胞上皮-间叶转化中的作用

目的探讨microRNA-192(miR-192)对晚期糖基化终产物(AGEs)诱导人腹膜间皮细胞上皮-间叶转化(EMT)的调控作用。方法人腹膜间皮细胞、转染miR-192抑制物后人腹膜间皮细胞和转染miR-192抑制物阴性对照的人腹膜间皮细胞在含AGEs的培养基培养72 h,以M199培养基和含80 m M BSA的M199培养基为对照,然后运用实时荧光定量PCR法检测miR-192和mRNA的表达情况,运用蛋白质印迹法检测蛋白的表达情况。结果在AGEs刺激后,人腹膜间皮细胞miR-192、胶原蛋白I(Collagen I)mRNA、α-平滑肌肌动蛋白(α-SMA)mRNA和蛋白表达水平显著上升(P0.05),而E-钙黏蛋白(E-cadherin)mRNA和蛋白表达水平显著下降(P0.05)。与转染miR-192抑制物阴性对照的人腹膜间皮细胞相比,转染miR-192抑制物的人腹膜间皮细胞miR-192、Collagen I mRNA、α-SMAmRNA和蛋白表达水平显著下降(P0.05),而E-cadherin mRNA和蛋白表达水平显著上升(P0.05)。结论 AGEs可能通过上调miR-192表达诱导人腹膜间皮细胞EMT。miR-192抑物可能通过下调miR-192表达阻止AGEs诱导人腹膜间皮细胞EMT。miR-192在AGEs诱导人腹膜间皮细胞EMT中起重要调控作用。     

骨形成蛋白-7对晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化的影响

目的探讨骨形成蛋白-7(BMP-7)对晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化(EMT)的影响。方法晚期糖基化终产物诱导发生上皮-间叶转化的体外培养大鼠腹膜间皮细胞,分别经含5 ng/mL及80 mmol/L晚期糖基化终产物的M199培养基和含10 ng/mL BMP-7及80 mmol/L晚期糖基化终产物的M199培养基培养48 h,以含80 mmol/L晚期糖基化终产物的M199培养基为对照,应用实时定量PCR法检测间皮细胞E-cadherin、α平滑肌肌动蛋白(α-SMA)、Ⅰ型胶原(Collagen I)、转化生长因子β1(TGF-β1)、血管内皮生长因子(VEGF)mRNA的表达;应用Western印迹法检测E-cadherin、α-SMA蛋白的表达;应用ELISA法检测间皮细胞TGF-β1、VEGF蛋白表达水平。结果 BMP-7作用后,上皮-间叶转化的大鼠腹膜间皮细胞E-cadherin mRNA和E-cadherin蛋白表达水平显著增加(P<0.05)。BMP-7作用后,上皮-间叶转化的大鼠腹膜间皮α-SMA、Collagen I、TGF-β1、VEGFmRNA和α-SMA、TGF-β、VEGF蛋白表达水平显著下降(P<0.05)。结论 BMP-7能上调上皮-间叶转化的大鼠腹膜间皮细胞E-cadherin表达和下调α-SMA、Collagen I、TGF-β、VEGF表达,BMP-7能逆转晚期糖基化终产物诱导大鼠腹膜间皮细胞EMT。     

高浓度葡萄糖诱导大鼠腹膜间皮细胞上皮-间叶转化

目的探讨高浓度葡萄糖对大鼠腹膜间皮细胞上皮-间叶转化(EMT)的影响,建立大鼠腹膜间皮细胞EMT模型。方法体外培养的大鼠腹膜间皮细胞经含60、120 mmol/L葡萄糖的M199培养基分别培养48、72 h;以正常M199培养基和含120 mmol/L甘露醇的M199培养基为对照。采用相差显微镜观察细胞形态学改变,应用实时定量PCR法检测间皮细胞E-cad-herin、α-SMA、Collagen I mRNA的表达;应用Western印迹法检测E-cadherin、α-SMA蛋白的表达。结果高浓度葡萄糖(60、120 mmol/L)刺激后,大鼠腹膜间皮细胞由典型的上皮细胞形态逐渐变为梭形及不规则形,类似肌成纤维细胞;大鼠腹膜间皮细胞α-SMA、Collagen I mRNA和α-SMA蛋白表达水平显著增加(P0.05);大鼠腹膜间皮细胞E-cadherin mRNA和E-cad-herin蛋白表达水平显著下降(P0.05)。结论高浓度葡萄糖能上调α-SMA、Collagen I表达和下调E-cadherin表达,高浓度葡萄糖诱导大鼠腹膜间皮细胞EMT。     

高浓度葡萄糖诱导大鼠腹膜间皮细胞上皮-间叶转化

目的探讨高浓度葡萄糖对大鼠腹膜间皮细胞上皮-间叶转化（EMT）的影响,建立大鼠腹膜间皮细胞EMT模型。方法体外培养的大鼠腹膜间皮细胞经含60、120 mmol/L葡萄糖的M199培养基分别培养48、72 h;以正常M199培养基和含120 mmol/L甘露醇的M199培养基为对照。采用相差显微镜观察细胞形态学改变,应用实时定量PCR法检测间皮细胞E-cad-herin、α-SMA、Collagen I mRNA的表达;应用Western印迹法检测E-cadherin、α-SMA蛋白的表达。结果高浓度葡萄糖（60、120 mmol/L）刺激后,大鼠腹膜间皮细胞由典型的上皮细胞形态逐渐变为梭形及不规则形,类似肌成纤维细胞;大鼠腹膜间皮细胞α-SMA、Collagen I mRNA和α-SMA蛋白表达水平显著增加（P〈0.05）;大鼠腹膜间皮细胞E-cadherin mRNA和E-cad-herin蛋白表达水平显著下降（P〈0.05）。结论高浓度葡萄糖能上调α-SMA、Collagen I表达和下调E-cadherin表达,高浓度葡萄糖诱导大鼠腹膜间皮细胞EMT。     

晚期糖基化终产物在动脉粥样硬化中的作用

2型糖尿病患者动脉粥样硬化的发病率明显增高。糖尿病通过增加常规的危险因素(如血脂异常、高血压)或糖尿病特有的危险因子(如晚期糖基化终产物AGEs),促进动脉粥样硬化的发展。AGEs作为蛋白质或脂质与还原糖糖基化的终产物,在血管细胞中通过与RAGE结合促进炎症因子的表达,参与动脉粥样硬化的发展。近年来,关于AGEs及其受体(RAGE)对糖尿病动脉粥样硬化影响的研究较为深入。本文旨在对近年来有关AGEs/RAGE在糖尿病加速的动脉粥样硬化中作用的研究进展作一综述。     

吡哆胺对体外晚期糖基化及其终产物形成的抑制作用

目的体外观察吡哆胺对晚期糖基化和晚期糖基化终产物(AGEs)形成的抑制作用。方法应用牛血清白蛋白-葡萄糖试验和N-乙酰-甘氨酰-赖氨酸甲基酯-核糖两种体外试验,观察吡哆胺对糖基化反应及AGEs形成的抑制效果。结果牛血清白蛋白-葡萄糖试验显示,吡哆胺对糖基化反应具有明显抑制效果。在50 mmol/L和200 mmol/L葡萄糖浓度下,当吡哆胺浓度为50 mmol/L时,相对抑制率均在50%以上。200 mmol/L吡哆胺对糖基化的抑制作用最强,其相对抑制率分别达到92.12%和86.73%。N-乙酰-甘氨酰-赖氨酸甲基酯-核糖试验证明,吡哆胺能明显抑制晚期糖基化产物AGEs的形成。随着吡哆胺剂量的增大,吡哆胺对AGEs形成的抑制作用明显增强。结论吡哆胺对体外晚期糖基化反应和AGEs形成有明显抑制作用。     

晚期糖基化终产物与冠心病发病机制的研究进展

晚期糖基化终产物(AGEs)密切参与了血管平滑肌细胞(VSMCs)分化与增殖以及冠心病等心血管疾病的病理生理过程。AGEs能通过单核细胞趋化蛋白(ERK)、丝氨酸/苏氨酸蛋白激酶(Akt)信号通路来诱导VSMCs的自噬作用,可依赖骨髓基质细胞衍生因子-1(SDF-1)/趋化因子受体CXCR4轴信号通路促进心肌微血管内皮细胞(CMECs)的增生。AGEs-2和AGEs-3上调了单核细胞AGEs受体(RAGE)的表达。AGEs能抑制内皮祖细胞(EPCs)的增殖、迁移和黏附功能并诱导EPCs凋亡;能增加平滑肌细胞结缔组织因子(CTGF)mRNA和蛋白质的表达,刺激心肌成纤维细胞增殖并分泌转化生长因子-β1(TGF-β1),同时诱导Smad2及Smad4的表达。羧甲基赖氨酸(CML)/RAGE轴通过主动脉平滑肌成骨细胞的分化,诱导巨噬细胞凋亡,从而使AGEs在糖尿病动脉粥样硬化中发挥重要作用。可溶性RAGE(sRAGE)可作为RAGE配体的诱饵来防止动脉粥样硬化,其灵敏度和阴性预测值在判定冠状动脉介入治疗(PCI)术后再狭窄方面均高于AGEs/sRAGE比值。ALT-711是一种AGEs的裂解剂,能明显抑制AGEs介导的活性氧(ROS)产生、细胞外信号调节激酶磷酸化及环氧合酶-2的表达;色素上皮衍生因子(PEDF)能抑制AGEs诱导的血小板CD40配体(CD40L)表达,从而有可能成为预防冠心病的一个治疗靶点。他汀类药物亦能抑制AGEs诱导主动脉平滑肌细胞的增殖及ROS的产生。通过以上诸多因素的研究,可揭示冠心病的某些发病机制,为相应干预药物的研究及调整临床治疗策略提供依据和方向。     

肥胖儿童血清晚期糖基化终产物的变化及其临床意义

目的 评价肥胖儿童血清晚期糖基化终产物(AGEs)在肥胖儿童心血管病中的临床意义.方法 采用标准方法 进行体重分组,检测血压,以RF-540荧光分光光度计测定20例肥胖儿童及20例肥胖伴高血压和20名正常体重儿童(对照组)的血清AGEs含量.结果 肥胖组、肥胖伴高血压组及正常对照组AGEs分别为(7.373±1.816)、(10.971±0.861)、(4.830±1.056)mg/L,肥胖儿童组及肥胖伴高血压组儿童血清AGEs明显高于对照组,差异均有统计学意义(t值分别为5.41,20.16,P均<0.05).肥胖组与肥胖伴高血压组比较血清AGEs亦升高且差异有统计学意义(t=8.01,P<0.05).结论 肥胖及肥胖伴高血压儿童血清AGEs明显升高,AGEs可能参与了儿童心血管疾病的发生.     

有氧运动对2型糖尿病大鼠血浆晚期糖基化终产物的影响

目的 探讨有氧运动对2型糖尿病大鼠血浆晚期糖基化终产物(AGEs)的影响.方法 将52只8周龄健康雄性SD大鼠分为正常对照组10只和2型糖尿病造模组42只.造模组大鼠用高糖、高脂和高能量饲料喂养4周后,予以一次性腹腔注射链脲佐菌素(30 ms/kg体重),以建立2型糖尿病大鼠模型.取30只成功造模的2型糖尿病大鼠,分为糖尿病对照组、小强度运动组和中强度运动组,每组10只.小、中强度运动组实施运动方案.结果 与正常对照组比较,糖尿病对照组、中强度运动组AGEs水平显著升高(P<0.01);与糖尿病对照组比较,小强度运动组AGEs水平显著降低(P<0.05).结论 有氧运动可降低2型糖尿病大鼠血浆AGEs水平,其作用与血糖降低和运动强度有关.

Abstract：

Objective To determine the influence of aerobic exercise on advanced glycation end products (AGEs)in plasma using a rat model of type 2 diabetes,and to provide an experimental basis for explaining the rehabilitative mechanism of aerobic exercise in type 2 diabetes.Methods Fifty-two healthy 8-week-old male SpragueDawley rats were allocated at random into a normal control group(n=10)and a type 2 diabetes model group(n=42).The latter were overfed with a high-sugar,high-fat and high-energy diet for 4 weeks,then 30ms/kg of streptozotocin was injected intraperitoneally to create a model of type 2 diabetes.Thirty diabetic rats were then allocated at random into a diabetes control group,a low intensity exercise group and a moderate intensity exercise group with 10 in each group.The treadmill exercise was administered to the animals in the low and moderate exercise groups accordingly.Results Compared with the normal control group,plasma AGEs increased significantly in the diabetes control and moderate intensity exercise groups.Compared with the diabetes control group,AGEs in the lower intensity exercise group were significantly lower.Compared with the moderate intensity exercise group,plasma AGEs in the low intensity exercise group were somewhat lower,but not significantly.Conclusion Aerobic exercise can reduce plasma AGE levels in rats with a model of type 2 diabetes.The effect is probably related to decreasing excessive blood glncose and the exercise intensity.     

晚期糖基化终产物在β样淀粉蛋白25-35诱导PC12细胞氧化损伤中的作用

目的探讨晚期糖基化终产物(AGEs)在β样淀粉蛋白25-35(Aβ25-35)诱导PC12细胞氧化损伤中的作用。方法将实验对象分为五组:10、20、304、0μmol/L四种浓度的Aβ25-35干预组和正常对照组。采用MTT法测定细胞生存率,采用ELISA法测定AGEs的含量。结果Aβ25-35诱导PC12细胞,细胞生存率为50%时,Aβ25-35浓度约为30μmol/L;与正常对照组相比,30μmol/L Aβ25-35干预组细胞生存率明显降低(P<0.001)、细胞内AGEs含量明显升高(P<0.05)。结论AGEs参与了β样淀粉蛋白25-35诱导PC12细胞氧化损伤的过程。     

Influence of bicarbonate/low-GDP peritoneal dialysis fluid (BicaVera) on in vitro and ex vivo epithelial-to-mesenchymal transition of mesothelial cells

♦ Background: Peritoneal membrane damage induced by peritoneal dialysis (PD) is largely associated with epithelial-to-mesenchymal transition (EMT) of mesothelial cells (MCs), which is believed to be a result mainly of the glucose degradation products (GDPs) present in PD solutions.♦ Objectives: This study investigated the impact of bicarbonate-buffered, low-GDP PD solution (BicaVera: Fresenius Medical Care, Bad Homburg, Germany) on EMT of MCs in vitro and ex vivo.♦ Methods: In vitro studies: Omentum-derived MCs were incubated with lactate-buffered standard PD fluid or BicaVera fluid diluted 1:1 with culture medium.Ex vivo studies: From 31 patients randomly distributed to either standard or BicaVera solution and followed for 24 months, effluents were collected every 6 months for determination of EMT markers in effluent MCs.♦ Results: Culturing of MCs with standard fluid in vitro resulted in morphology change to a non-epithelioid shape, with downregulation of E-cadherin (indicative of EMT) and strong induction of vascular endothelial growth factor (VEGF) expression. By contrast, in vitro exposure of MCs to bicarbonate/low-GDP solution had less impact on both EMT parameters.Ex vivo studies partially confirmed the foregoing results. The BicaVera group, with a higher prevalence of the non-epithelioid MC phenotype at baseline (for unknown reasons), showed a clear and significant trend to gain and maintain an epithelioid phenotype at medium- and longer-term and to show fewer fibrogenic characteristics. By contrast, the standard solution group demonstrated a progressive and significantly higher presence of the non-epithelioid phenotype. Compared with effluent MCs having an epithelioid phenotype, MCs with non-epithelioid morphology showed significantly lower levels of E-cadherin and greater levels of fibronectin and VEGF. In comparing the BicaVera and standard solution groups, MCs from the standard solution group showed significantly higher secretion of interleukin 8 and lower secretion of collagen I, but no differences in the levels of other EMT-associated molecules, including fibronectin, VEGF, E-cadherin, and transforming growth factor β1.Peritonitis incidence was similar in both groups. Functionally, the use of BicaVera fluid was associated with higher transport of small molecules and lower ultrafiltration capacity.♦ Conclusions: Effluent MCs grown ex vivo from patients treated with bicarbonate/low-GDP BicaVera fluid showed a trend to acquire an epithelial phenotype, with lower production of proinflammatory cytokines and chemokines (such as interleukin 8) than was seen with MCs from patients treated with a lactate-buffered standard PD solution.     

Targeting the receptor for advanced glycation end products (RAGE) in type 1 diabetes

Type 1 diabetes (T1D) is one of the most common chronic diseases manifesting in early life, with the prevalence increasing worldwide at a rate of approximately 3% per annum. The prolonged hyperglycaemia characteristic of T1D upregulates the receptor for advanced glycation end products (RAGE) and accelerates the formation of RAGE ligands, including advanced glycation end products, high-mobility group protein B1, S100 calcium-binding proteins, and amyloid-beta. Interestingly, changes in the expression of RAGE and these ligands are evident in patients before the onset of T1D. RAGE signals via various proinflammatory cascades, resulting in the production of reactive oxygen species and cytokines. A large number of proinflammatory ligands that can signal via RAGE have been implicated in several chronic diseases, including T1D. Therefore, it is unsurprising that RAGE has become a potential therapeutic target for the treatment and prevention of disease. In this review, we will explore how RAGE might be targeted to prevent the development of T1D.     

晚期氧化蛋白产物通过活性氧诱导人腹膜间皮细胞TGF-β1的表达

目的探讨晚期氧化蛋白产物（AOPP）诱导体外培养的人腹膜间皮细胞（HPMCs）对转化生长因子（TGF-β1）表达的影响及内源性活性氧（ROS）在此过程中的调节机制。方法体外制备AOPP-HSA模型，原代培养人腹膜间皮细胞。采用ELISA法检测TGF-β1蛋白水平，采用RT-PCR半定量分析HPYCs中TGF-β1 mRNA的基因表达水平，同时应用流式细胞仪检测细胞内活性氧的水平。结果AOPP-HSA能显著诱导人腹膜间皮细胞TGF-β1的分泌与基因表达，同时增加细胞内ROS的生成，呈时间与剂量依赖关系（P〈0．01），不同浓度的抗氧化剂维生素E和N-乙酰半胱胺酸预处理细胞，可明显地降低细胞内ROS的生成量，同时显著地抑制AOPP-HSA诱导人腹膜间皮细胞TGF-β1的分泌与基因表达，呈剂量依赖关系（P〈0．01），其中维生素E（50μmol／L）组和NAC（10mmol／L）组抑制更加显著。结论体外制备的AOPP可显著诱导人腹膜间皮细胞TGF-β1的分泌与基因表达，部分可能通过内源性ROS介导细胞内信号转导途径调节此过程，抗氧化剂维生素E和N-乙酰半胱胺酸可显著降低细胞ROS的生成量和显著抑制TGF-β1的分泌与基因表达。此研究揭示抗氧化剂在防治腹膜纤维化发生过程也许是一种可行的治疗策略。