炎性环境下内皮微粒介导单核细胞促内皮细胞增殖的作用

目的 探讨炎性环境中内皮微粒(EMPs)在单核细胞参与的促内皮细胞增殖中的作用,并探讨EMPs对单核细胞表达内皮细胞标志物的影响.方法 体外培养人脐静脉内皮细胞(HUVECs),炎性因子肿瘤坏死因子α(TNF-α)刺激内皮细胞,超速离心法获取EMPs;用透射电镜观察其形态和结构;运用酶联免疫吸附法(ELISA)检测经EMPs激活的单核细胞株THP-1对血管内皮生长因子(VEGF)中VEGF-A、VEGF-C的表达情况;将HUVECs和THP-1细胞或者经EMPs诱导的THP-1细胞共培养,检测HUVECs的增殖情况;运用RT-PCR和免疫荧光细胞化学技术,观察经EMPs激活的THP-1细胞对内皮细胞标志物vWF和血管内皮生长因子受体2(VEGFR-2)的表达情况.结果 内皮细胞经炎性因子刺激后释放EMPs,EMPs刺激的THP-1细胞上清中VEGF-A和VEGF-C蛋白水平明显升高;HUVECs细胞增殖实验中,与THP-1细胞或者经EMPs诱导的THP-1共培养的两种情况下,HUVECs均显著增多;与EMPs共培养3d后,THP-1中vWF和VEGFR-2蛋白表达为阳性,基因表达为阴性,共培养8d后,vWF和VEGFR-2的蛋白和基因表达均为阴性.结论 炎性环境中内皮细胞释放EMPs可以促使单核细胞分泌VEGF-A和VEGF-C,后两者可使HUVECs增殖,EMPs仅能使单核细胞一过性呈现内皮表型,而不能将其转分化为内皮细胞.     

白藜芦醇联合间充质干细胞改善损伤内皮细胞的代谢记忆效应

背景：白藜芦醇联合间充质干细胞是否能够进一步改善高糖诱导内皮细胞产生的不良记忆目前鲜有报道。 目的：验证不同浓度的白藜芦醇联合间充质干细胞对高糖诱导人脐静脉内皮细胞损伤代谢记忆效应的保护作用。 方法：将人脐静脉内皮细胞分为10组：正常对照组、甘露醇对照组、高糖组、白藜芦醇低剂量组(0.1 μmol/L)、白藜芦醇中剂量组(1 μmol/L)、白藜芦醇高剂量组(10 μmol/L)、干细胞组、白藜芦醇低剂量+干细胞组、白藜芦醇中剂量+干细胞组及白藜芦醇高剂量+干细胞组。分别于5.5 mmol/L葡萄糖培养第1,4,6天收集细胞培养上清液、提取细胞核蛋白,应用ELISA法检测细胞培养上清中血管细胞黏附分子1、单核细胞趋化蛋白1、纤溶酶原激活剂抑制剂1的表达水平,以Western blot法检测细胞内核因子κB的蛋白水平。 结果与结论：与正常对照组相比,高糖可诱导内皮细胞核因子κB表达上调,同时血管细胞黏附分子1 、单核细胞趋化蛋白1和纤溶酶原激活剂抑制剂1水平升高,且在糖浓度恢复正常后仍持续上升。与高糖组相比,白藜芦醇及其联合干细胞干预后可以使内皮细胞核因子κB表达及血管细胞黏附分子1、单核细胞趋化蛋白1和纤溶酶原激活剂抑制剂1水平呈白藜芦醇剂量依赖性降低。干细胞组上述指标与高糖组比差异无显著性意义,甘露醇组与正常对照组相比差异无显著性意义。结果提示白藜芦醇可能通过核因子κB通路降低血管细胞黏附分子1、单核细胞趋化蛋白1和纤溶酶原激活剂抑制剂1水平,改善高糖代谢记忆效应介导的内皮细胞损伤。而间充质干细胞发挥内皮细胞保护作用可能不仅有赖于核因子κB通路。     

Notch信号通路对烧伤大鼠血清诱导的肺血管内皮细胞细胞间黏附分子-1的影响

目的探讨Notch信号通路对烧伤大鼠血清诱导的肺血管内皮细胞(PMVEC)细胞间黏附分子(ICAM)-1的影响。 方法42只SD大鼠(6～8周龄)中,随机选取30只,按随机数字表法分为假伤组(n＝15)和烧伤组(n＝15),烧伤组大鼠于95 ℃热水浸浴背部18 s造成30%总体表面积Ⅲ度烧伤,假伤组大鼠于37 ℃水浴中浸浴背部18 s模拟致伤。伤后6、12、24、48、72 h,烧伤组随机分别取3只大鼠,腹主动脉采血,酶联免疫吸附试验(ELISA)法检测血清中ICAM-1含量,假伤组大鼠行相同检测。取剩下的12只大鼠(6～8周龄)中的6只,按前述方法造成30%总体表面积Ⅲ度烧伤,伤后24 h制备烧伤大鼠血清;剩下的6只大鼠不作处理,制备健康大鼠血清。切取10只出生3 d的SD大鼠的肺外边缘组织,组织块法培养大鼠PMVEC,倒置相差显微镜下观察原代细胞培养2、6 d后的形态特征,流式细胞术对原代培养7 d的细胞进行细胞鉴定。取处于对数生长期的第4代PMVEC进行实验,将细胞接种于6孔板中,待细胞生长至80%融合时按随机数字表法分为3组(每组设3个复孔)：对照组(培养液中分别加入体积分数10%健康大鼠血清),二甲基亚砜(DMSO)+烧伤血清组、γ-分泌酶抑制剂(GSI)+烧伤血清组(分别加入3 μL/mL DMSO、75 μmol/L GSI,培养24 h后,加入体积分数10%烧伤大鼠血清刺激培养24 h),ELISA法检测PMVEC上清中ICAM-1的含量;流式细胞仪检测各组PMVEC中ICAM-1的含量;采用蛋白质印迹法检测ICAM-1蛋白表达水平,计算相对蛋白表达量;检测PMVEC的细胞黏附能力。数据比较采用单因素方差分析和LSD-t检验。 结果(1)烧伤组大鼠伤后6、12、24、48、72 h血清中的ICAM-1含量分别为(19.77±3.03)、(22.09±3.65)、(22.44±4.04)、(25.40±2.51)、(26.37±3.07) pg/mL,显著高于假伤组的[(10.60±1.51)、(11.03±1.95)、(10.87±0.89)、(9.30±0.89)、(10.93±1.22) pg/mL],2组比较差异均有统计学意义(t＝4.699、4.466、3.181、10.490、8.097,P<0.05)。(2)原代细胞培养2 d后可见细胞从组织块边缘移出,生长较慢;培养6 d后可见细胞融合,细胞呈铺路石样生长。对原代培养7 d的细胞进行流式细胞术鉴定,结果显示CD31阳性细胞占98.6%,确定为PMVEC。(3)血清刺激培养24 h后,对照组,DMSO+烧伤血清组、GSI+烧伤血清组细胞上清中ICAM-1含量分别为1.21±0.25、2.16±0.12、3.07 ±0.30,3组比较差异有统计学意义(F＝46.72,P<0.05);与对照组比较,DMSO+烧伤血清组和GSI+烧伤血清组细胞上清中ICAM-1含量均明显增加,差异均有统计学意义(t＝5.977、8.274,P<0.05);GSI+烧伤血清组细胞上清中ICAM-1含量明显高于DMSO+烧伤血清组,2组比较差异有统计学意义(t＝4.847,P<0.05)。(4)血清刺激培养24 h后,对照组,DMSO+烧伤血清组、GSI+烧伤血清组PMVEC中ICAM-1平均荧光强度分别为2 582±143、3 453±204、4 559±414,3组比较差异有统计学意义(F＝37.84,P<0.05);与对照组比较,DMSO+烧伤血清组、GSI+烧伤血清组细胞中ICAM-1含量均明显增加,差异均有统计学意义(t＝6.056、7.817,P<0.05);GSI+烧伤血清组ICAM-1含量明显高于DMSO+烧伤血清组,2组比较差异有统计学意义(t＝4.149,P<0.05)。(5)血清刺激培养24 h后,对照组、DMSO+烧伤血清组、GSI+烧伤血清组的ICAM-1/GAPDH比值分别为0.74±0.08、1.07±0.08、1.38±0.28,3组总体比较差异有统计学意义(F＝30.76,P<0.05); DMSO+烧伤血清组和GSI+烧伤血清组的ICAM-1/GAPDH比值均明显高于对照组,差异均有统计学意义(t＝5.182、6.990,P<0.05);GSI+烧伤血清组ICAM-1/GAPDH比值明显高于DMSO+烧伤血清组,差异有统计学意义(t＝3.770,P<0.05)。(6)PMVEC与人单核细胞系THP-1细胞共培养2 h后,倒置荧光显微镜下观察对照组、DMSO+烧伤血清组、GSI+烧伤血清组细胞黏附的细胞数呈递增趋势,3组的细胞数分别为(152.00±21.07)、(265.33±36.83)、(345.67±30.66)个,3组比较差异有统计学意义(F＝31.09,P<0.05);DMSO+烧伤血清组和GSI+烧伤血清组黏附的细胞数均明显高于对照组,比较差异均有统计学意义(t＝4.626、9.016,P<0.05);GSI+烧伤血清组黏附的细胞数明显高于DMSO+烧伤血清组,差异有统计学意义(t＝2.903,P<0.05)。 结论大鼠烧伤后血清中的ICAM-1含量明显上升,烧伤大鼠血清刺激PMVEC可以导致细胞内ICAM-1含量以及分泌到上清中的ICAM-1含量增加,应用GSI阻断Notch信号通路可以增加ICAM-1的产生及表达,可增加PMVEC对人单核细胞系THP-1细胞的黏附作用。     

去甲肾上腺素/烧伤血清诱导大鼠神经胶质细胞VEGF基因表达

目的： 观察血管内皮细胞生长因子（VEGF）在去甲肾上腺素（NE）/烧伤血清诱导神经胶质细胞时基因表达变化。 方法： 用免疫荧光法观察NE/烧伤血清刺激神经胶质细胞后24 h VEGF在细胞内的分布。用免疫蛋白印迹法检测细胞VEGF蛋白表达。用荧光定量聚合酶联反应（PCR）法检测细胞VEGF mRNA表达。 结果： ①高剂量NE（50 μmol/L）刺激时，神经胶质细胞胞浆内绿色荧光增强，烧伤血清刺激后绿色荧光明显增强，当烧伤血清加高剂量NE刺激时绿色荧光显著增强。②烧伤血清组细胞VEGF蛋白表达增加，烧伤血清加中剂量（20 μmol/L）、高剂量NE组VEGF的蛋白表达明显增加。③烧伤血清刺激后，细胞VEGF mRNA表达增加，烧伤血清与NE同时刺激时，随着NE剂量的增加，细胞中VEGF mRNA表达明显增加。 结论： NE/烧伤血清能够诱导神经胶质细胞VEGF的基因表达增强，提示烧伤应激因素诱导神经胶质细胞分泌VEGF可能是烧伤后脑水肿形成的一个重要因素。     

血小板微粒通过激活单核细胞HIF-1α促进血管生成的作用

目的:探讨血小板微粒(PMP)通过单核细胞促进血管生成的作用及机制。方法:制备人PMP,采用基质胶栓实验检测PMP对体内新生血管形成的影响。在体外常氧和缺氧条件下,使PMP与人单核细胞系THP-1结合,ELISA法检测血管内皮生长因子(VEGF)的分泌水平,RT-qPCR检测THP-1细胞VEGF和低氧诱导因子1α(HIF-1α)mRNA的表达,转录活性实验检测THP-1细胞HIF-1α的转录活性;利用共培养系统检测PMP与THP-1细胞相互作用对人脐静脉内皮细胞(HUVECs)体外管腔形成的影响。结果:PMP诱导在体基质胶栓表面形成新生血管,HIF-1α选择性抑制剂chetomin可明显减弱该效应(P0.01)。在常氧和缺氧条件下,PMP均呈剂量依赖性地促进THP-1细胞表达和释放VEGF,上调HIF-1αmRNA的表达并增强其转录活性;阻断PMP与THP-1细胞的相互作用可以抑制THP-1细胞HIF-1α的转录活性以及VEGF的生成。PMP激活的单核细胞可促进HUVECs在体外基质胶上形成管腔,chetomin干预导致管腔的数量明显减少。结论:PMP通过激活单核细胞的HIF-1α诱导其释放VEGF,导致新生血管的形成,这可能是PMP促进动脉粥样硬化血管生成的一种新机制。     

牙龈卟啉单胞菌在单核细胞对内皮细胞黏附作用中的影响

目的 观察牙龈卟啉单胞菌(P.gingivalis)W83和ATCC33277株侵入在单核细胞对内皮细胞黏附作用中的影响,及在内皮细胞细胞间黏附分子l(ICAM-1)转录和翻译中的作用. 方法 建立体外P.gingivalis侵入内皮细胞模型,孟加拉玫瑰红活细胞染色法测定P.gingivalis侵入前后单核细胞对内皮细胞黏附的变化;RT-PCR和mRNA比色定量法检测内皮细胞ICAM-1基因表达;West-ern blot检测ICAM-1蛋白水平的变化. 结果 P.gingivalis W83和ATCC33277株侵入可增加单核细胞对内皮细胞的黏附,抗ICAM-1抗体部分抑制P.gingivalis侵入介导的单核细胞对内皮细胞黏附增加;P.gingivalis侵入上调内皮细胞ICAM-l基因和蛋白的表达,W83诱导单核细胞对内皮细胞黏附增强及内皮细胞ICAM-1表达的能力强于ATCC33277. 结论 ICAM-1在P.gingivalis介导的单核细胞对内皮细胞黏附增强过程中起部分作用,P.gingivalis侵入内皮细胞诱导ICAM-1表达可能是其诱发动脉粥样硬化疾病的机制之一.     

严重烧伤大鼠血清对脂肪间质干细胞生物学特性的影响

目的 探讨严重烧伤大鼠血清对体外培养的大鼠脂肪间充质干细胞(ADSCs)生物学特性的影响.方法 取雄性SD大鼠双侧腹股沟脂肪组织,采用胶原酶消化法、分离纯化大鼠ADSCs,取第3代细胞,采用流式细胞仪检测细胞表面标志物,行成脂成骨诱导分化鉴定;ADSCs传代后按随机数字表法分为10％胎牛血清组(正常培养组)、10％正常大鼠血清组,10％烧伤大鼠血清组,采用噻唑蓝(MTT)法测定各组细胞的生长曲线.通过流式细胞仪检测各组细胞的生长周期、Real-time PCR法检测各组细胞血管内皮生长因子(VEGF)、凋亡相关因子Caspase-3的表达情况.结果 分离培养的ADSCs传至第3代时形态规则,经流式细胞仪检测,所培养的ADSCs均表达CD29、CD90、CD105,阳性率分别为88.6％、99.7％、92.8％,而CD31、CD34、CD45阳性率分别为4.4％、4.7％、3.3％;经诱导培养后可向成骨成脂分化.MTT法检测结果显示:10％烧伤大鼠血清组细胞增殖较快,各时间点吸光度值(A值)明显高于10％胎牛血清组及10％正常大鼠血清组.流式细胞仪检测结果显示:培养1周后10％烧伤大鼠血清组、10％正常大鼠血清组、10％胎牛血清组ADSCs G1期细胞比例分别为:(72.20±5.13)％、(83.50±4.74)％、(90.20±6.37)％;S期细胞比例分别为:(21.40±2.84)％、(12.50±1.96)％和(8.60±1.31)％,10％烧伤大鼠血清组ADSCs G1期细胞比例显著低于10％正常大鼠血清组、10％胎牛血清组(t=2.39、4.86;P ＜0.05);10％烧伤大鼠血清组ADSCs S期细胞比例显著高于10％正常大鼠血清组、10％胎牛血清组(=5.54,7.93;P＜0.01).Real-Time PCR检测结果显示:10％烧伤大鼠血清组VEGF的表达明显上调,Caspase-3表达显著下调与10％正常大鼠血清组、10％胎牛血清组相比差异均有统计学意义(P＜0.05).结论 10％烧伤大鼠血清可明显促进ADSCs的增殖、抑制细胞凋亡发生、促进ADSCs的分泌功能,严重烧伤后可望移植ADSCs用?     

PMA诱导的THP-1细胞分化过程中MMP-9及MMP-2的表达与细胞侵蚀性的关系

目的：应用单核细胞THP-1株建立单核细胞向巨噬细胞分化的体外模型，并探讨THP-1细胞分化过程中基质金属蛋白酶-9(MMP-9)及基质金属蛋白酶-2(MMP-2)的表达与细胞侵蚀性的关系以及巨噬细胞住类风湿关节炎(RA)发病机制中的作用。方法：用乙酸肉豆蔻佛波酯(PMA)诱导THP-1细胞向成熟的单核-巨噬细胞分化后，采用倒置显微镜观察分化细胞的形态学变化。用流式细胞术测定细胞表面特异性抗原CD14的表达。采用明胶酶谱(gelatin—zymogram)分析法测定分化前后THP-1细胞中MMP-9和MMP-2的表达。采用侵蚀性小室(Boyden Chamber-Matrigel人工重组基底膜)法，观察分化前后的THP-1细胞侵蚀性的变化。结果：THP-1细胞向单核-巨噬细胞分化过程中，细胞由分散、悬浮转变为黏附、贴壁，且出现CD14的高表达分化后的THP-1细胞中MMP-9和MMP-2的表达明显增多，侵蚀性明显增强结论：建立了THP-1单核细胞的体外分化模型。该细胞在分化过程中，MMP-9和MMP-2的表达增多、侵蚀性增强，为巨噬细胞在RA发病机制的作用提供了一定的实验依据。     

去甲肾上腺素诱导严重烧伤大鼠脑组织VEGF表达的实验研究

目的：观察去甲肾上腺素（NE）诱导严重烧 伤后24h大鼠脑组织血管内皮细胞生长因子(VEGF)的表达变化。方法:(1 )采用40%全身总体表面积（TBSA）Ⅲ°烧伤大鼠模型，用干、湿重法检测NE刺激烧伤24h大 鼠脑组织含水量；(2)用高效液相色谱法，检测烧伤大鼠脑组织去甲肾上腺素活性水平的变 化;(3) 用免疫蛋白印迹法（Western blotting）检测NE诱导脑组织VEGF蛋白表达变化。 结果：(1) 烧伤大鼠和NE刺激后的烧伤大鼠脑组织水肿明显；(2)烧伤后2 4 h大鼠脑组织NE活性增加，而NE刺激后的烧伤大鼠，脑组织NE的活性增加显著;(3) NE刺 激剂量增大时，脑组织VEGF的蛋白表达逐渐增加，烧伤后VEGF的蛋白表达增加明显，随NE刺 激剂量增大，烧伤后VEGF的蛋白表达明显增强。结论:NE可诱导严重烧 伤大鼠脑组织VEGF的蛋白表达，提示烧伤后NE水平增高是脑水肿形成的一个重要因素。     

脉冲电磁场对人单核细胞激活的影响

研究不同参数的脉冲电磁场(PEMF)干预对人单核(THP-1)细胞激活的影响。将体外培养的THP-1细胞分别以频率为32Hz或64Hz、强度为1mT的PEMF进行干预,2次/d,30min/次,间隔8h,共3d,以频率为0Hz作为对照组。采用计数法观察PEMF处理后THP-1细胞与人脐静脉内皮细胞(HUVECs)之间的黏附变化;ELISA法检测THP-1细胞培养基中单核细胞趋化因子1(MCP-1)的分泌情况;荧光定量PCR法观察THP-1细胞MCP-1基因表达改变。结果观察到,PEMF干预能明显抑制THP-1细胞与HUVECs间黏附,降低THP-1细胞MCP-1基因表达及其蛋白分泌水平。本实验提示,频率为32Hz或64Hz、场强为1mT的PEMF干预3d后,能明显抑制THP-1细胞激活。     

Estradiol reduces basal and cytokine induced monocyte adhesion to endothelial cells

OBJECTIVE: To investigate the effect of 17beta-estradiol (E2) on binding of monocytes to human aortic endothelial cells (HAECs) with or without cytokine induction. METHODS: Confluent monolayers of HAECs were incubated with or without E2 for 48 h prior to the monocyte adhesion assay. In studies with cytokines, 1 ng/ml tumor necrosis factor-alpha (TNF-alpha), 20 U/ml interleukin-1beta (IL-1beta) or both were added to the culture medium for the final 24 or 4 h. For the measurement of monocyte adhesion, 3H-thymidine labeled human THP-1 monocytes (4 x 10(5) cells per well) were added to the confluent monolayer of HAECs and incubated at 37 degrees C for 90 min. The unbound THP-1 cells were removed by gentle washing, and bound cells were digested with NaOH and quantified by measuring radioactivity. RESULTS: When HAECs were pretreated for 48 h with E2 the basal adhesion of THP-1 cells was reduced by an average of 28%. Estrogen significantly reduced cytokine-induced adhesion by 30-35% when the cytokines were added for 4 h. When the cytokine treatment was prolonged to 24 h, pretreatment of HAECs with E2 had no effect on THP-1 cell adhesion. CONCLUSIONS: E2 reduces basal and short-term cytokine induced monocyte binding to HAECs. Since monocyte adhesion to vascular endothelial cells is one of the initial steps in the pathogenesis of atherosclerosis, E2 may mediate vascular protection by reducing monocyte-endothelial cell binding in the early stages of atherogenesis.     

Associations between insulin-like growth factor I,vascular endothelial growth factor and its soluble receptor 1 in umbilical serum and endothelial cells obtained from normotensive and preeclamptic pregnancies

Abstract

The aim of this study was to investigate the associations between insulin-like growth factor I (IGF-I) with vascular endothelial growth factor (VEGF) and its soluble receptor 1 (sFlt-1) in umbilical serum and to study the effects of IGF-I upon sFlt-1 synthesis in human umbilical vein endothelial cells (HUVEC) in normotensive (NT) and preeclamptic (PE) pregnancies. As compared with the NT group, umbilical serum IGF-I and VEGF levels were lower in the PE group, while sFlt-1 concentrations were higher. Levels of sFlt-1 correlated with IGF-I in the NT group and with VEGF in the PE group. Basal concentration of sFlt-1 in HUVEC culture media was higher in the PE group. IGF-I stimulated sFlt-1 synthesis only in the NT group. In summary, umbilical serum sFlt-1 is associated with IGF-I in normotensive pregnancy and with VEGF in preeclampsia. IGF-I stimulates sFlt-1 synthesis in endothelial cells in normotensive but not in PE pregnancies.     

Inhibition of plasma-dependent monocyte chemokinesis and cytokine-triggered endothelial activation for neutrophil transmigration by administration of clopidogrel in man

Mediators released by spontaneously activated platelets may contribute to alterations in endothelial and leukocyte dysfunctions. We investigated the roles of clopidogrel and aspirin in ex vivo endothelial activation for interactions with leukocytes. Eight healthy volunteers received clopidogrel or aspirin for 8 days. Blood samples were taken before, during, and after treatment. Levels of adhesion molecules and platelet-derived mediators in these samples were measured using commercially available test kits, and effects of plasma on endothelial cells and leukocytes were investigated in neutrophil transendothelial migration, monocyte-endothelial adhesion and leukocyte migration assays. Plasma samples from clopidogrel-treated persons induced diminished chemokinesis of monocytes. Tumour necrosis factor-induced priming of endothelial cells for enhanced neutrophil transmigration was also diminished by pretreatment of endothelial cells, but not of neutrophils, with plasma derived from subjects during clopidogrel treatment. Plasma from the aspirin group had no such effects. Administration of clopidogrel but not aspirin significantly decreased serum levels of soluble intercellular adhesion molecule-1, whereas no changes in levels of soluble vascular cell adhesion molecule-1, P-selectin, L-selectin, von Willebrand factor, platelet-derived growth factor, vascular-endothelial growth factor, and transforming growth factor-beta were observed. Inhibition of plasma-promoted endothelial activation by clopidogrel may indicate a novel role in the prevention of atherosclerosis.     

Vascular endothelial growth factor and its receptor, Flt-1, in smokers and non-smokers

Raised levels of plasma vascular endothelial growth factor (VEGF) are found in some cancers, diabetes, and certain other conditions, but levels of its receptor, soluble Flt-1 (sFlt-1), in these diseases have yet to be reported. We hypothesised that smoking would influence levels of these molecules. Consequently, we measured VEGF and sFlt-1 by enzyme-linked immunosorbent assay (ELISA) in plasma from 92 non-smokers and 35 smokers. No difference in VEGF was seen between the groups but, despite considerable overlap, sFlt-1 was significantly lower in smokers (P = 0.027). VEGF and sFlt-1 correlated strongly with each other (P < 0.001). Although VEGF may arise from a number of cell types, including endothelial cells, the primary source of sFlt-1 is thought to be the endothelium; however, neither VEGF nor sFlt-1 correlated with levels of the endothelial cell activation/damage marker soluble thrombomodulin. Our data point to changes in levels of the VEGF receptor, sFlt-1--but not VEGF itself--in smokers, which appears to be unrelated to endothelial cell function.     

血管内皮生长因子及其可溶性血管内皮生长因子受体-1水平与胎儿出生体重

目的本研究通过对比血管内皮生长因子(VEGF)、可溶性血管内皮生长因子受体-1(sFlt-1)水平差异与新生儿出生体重的关系,以探讨其在胎儿出生体重发生中的作用。方法采用免疫组织化学法检测40例分娩正常出生体重儿组(AGA组)、30例高出生体重儿组(LGA组)及30例低出生体重儿组(SGA组)胎盘组织中VEGF、sFlt-1的表达水平。结果①LGA组胎盘组织中VEGF的表达高于AGA组,sFlt-1的表达水平低于AGA组,差异有统计学意义(χ2=21.17,P<0.01)。SGA组胎盘组织中VEGF的表达低于AGA组,sFlt-1的表达水平高于AGA组,差异有统计学意义(χ2=8.44,P=0.04)。②胎盘组织中VEGF的表达水平与胎儿出生体重呈正相关(r=0.427,P<0.01),胎盘组织中sFlt-1的表达水平与胎儿出生体重呈负相关(r=-0.569,P<0.01)。结论孕妇胎盘组织中VEGF及sFlt-1表达水平的变化可能与胎儿出生体重有关。     

血管内皮生长因子与血管细胞粘附分子在妊高征发病中的作用

目的探讨血管内皮生长因子及血管细胞粘附分子-1在妊高征发病中的作用。方法选取49例妊高征妇女(其中轻度15例、中度16例及重度18例),35例正常晚期妊娠妇女为研究对象,采用ELISA方法测定其血精VEGF、VCAM-1的水平。结果妊高征组血清VEGF浓度明显低于正常妊娠组(P<0.01).而VCAM-1浓度则明显高于正常妊娠组(P<0.01),轻度妊高征组血清VEGF、VCAM-1浓度与正常妊娠组无明显差别(P>0.05),中、重度妊高征组血清VEGF浓度明显低于正常妊娠组(P<0.01).而血清VCAM-1浓度明显高于正常妊娠组(P<0.01)。结论血清VEGF浓度的降低可能与胎盘浅着床有关,血清VCAM-1的升高参与了妊高征血管内皮损伤过程,VEGF、VCAM-1均可能与妊高征的发病机制有关。     

Endothelial cell-derived foam cells fail to express adhesion molecules (ICAM-1 and VCAM-1) for monocytes

The purpose of this study was to assess the expression of cell adhesion molecules ICAM-1 (intercellular adhesion molecule-1) and VCAM-1 (vascular cell adhesion molecule-1) in endothelial cell-derived foam cells. Hamster aortic endothelial cells (HAEC) in culture were exposed to hypercholesterolemic or normal homologous serum for 24 h. At the end of the incubation period, HAEC exposed to hypercholesterolemic serum exhibited numerous lipid droplets and had a general aspect of foam cells. When examined for the expression of ICAM-1 and VCAM-1 (by indirect immunofluorescence) normal HAEC expressed constitutively (to low level) on their surface these adhesion molecules; however HAEC-derived foam cells failed to display any labeling. To further assess these results, HAEC were first incubated with normal or hypercholesterolemic sera (as above) and then exposed to freshly isolated normal hamster blood monocytes. These experiments showed that monocytes adhered in small number to normal cells and failed to adhere to the surface of HAEC-derived foam cells. Together these data indicate that endothelial cell-derived foam cells: a) do not express ICAM-1 and VCAM-1 on their surface; b) have low or no adhesion properties for monocytes and c) may represent an appropriate experimental model to study the cellular alterations that take place in the advanced stages of atherosclerosis.     

Down-modulation of monocyte transendothelial migration and endothelial adhesion molecule expression by fibroblast growth factor: reversal by the anti-angiogenic agent SU6668

Basic and acidic fibroblast growth factor (bFGF and aFGF, respectively) and vascular endothelial growth factor (VEGF) exert angiogenic actions and have a role in wound healing, inflammation, and tumor growth. Monocytes and endothelial cells are involved in these processes, but the effect of FGF and VEGF on monocyte-endothelial cell interactions has not been defined. We observed that monocyte adhesion to resting or cytokine (tumor necrosis factor-alpha or interleukin-1 alpha)-stimulated human umbilical vein endothelial cells (HUVECs) was markedly inhibited (40 to 65%) by culture (1 to 6 days) of HUVECs with aFGF or bFGF. Monocyte transendothelial migration induced by C5a and chemokines (MCP-1, SDF-1 alpha, RANTES, MIP-1 alpha) was also suppressed (by 50 to 75%) on bFGF-stimulated HUVECs. VEGF did not have these effects at the concentrations used (10 to 20 ng/ml), although like bFGF, it promoted HUVEC proliferation. Culture of HUVECs with bFGF and aFGF significantly down-regulated intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and E-selectin expression on resting or tumor necrosis factor-alpha-stimulated HUVECs, but had no influence on platelet endothelial cell adhesion molecule (PECAM)-1 and VE-cadherin expression. bFGF also inhibited MCP-1 production by HUVECs. The inhibitory effects of bFGF on monocyte transendothelial migration and adhesion molecule expression were reversed by SU6668, an anti-angiogenic agent and bFGF receptor tyrosine kinase inhibitor. Our results suggest that bFGF and aFGF may suppress endothelial-dependent monocyte recruitment and thus have an anti-inflammatory action during angiogenesis in chronic inflammation but inhibit the immunoinflammatory tumor defense mechanism. However, SU6668 is an effective agent to prevent this down-regulatory action of bFGF on monocyte-endothelial cell interactions.     

The levels of circulating vascular endothelial growth factor and soluble Flt-1 in pregnancies complicated by preeclampsia

To evaluate the role of vascular endothelial growth factor (VEGF) in the pathogenesis of preeclampsia, we measured total VEGF, free VEGF and soluble Flt-1 (sFlt-1) concentrations and determined their relationships. Maternal serum samples were collected from 20 patients with preeclampsia and 20 normotensive women with uncomplicated pregnancies matched with the patients with preeclampsia for gestational age and parity. The serum concentrations of total VEGF (2.39+/-0.75 vs. 0.28+/-0.14) and sFlt-1 (934.5+/-235.5 vs. 298.0+/-161.2) were significantly increased in the patients with preeclampsia compared to the women with uncomplicated pregnancies. However the serum concentration of free VEGF (21.5+/-6.3 vs. 134.0+/-16.3) was lower in patients with preeclampsia. There was a positive correlation between the serum concentrations of total VEGF and sFlt-1 with systolic and diastolic blood pressure, respectively. There was a negative correlation between the serum concentration of free VEGF and systolic and diastolic blood pressure. There was a strong negative correlation between free VEGF and sFlt-1 concentrations. In conclusion, we found VEGF and sFlt-1 were related to the pathogenesis of preeclampsia. Although reduced concentrations of free VEGF might interfere with endothelial cell function and survival, further studies are required to clarify its specific role in the pathogenesis of preeclampsia.     

Soluble Flt-1 regulates Flk-1 activation to control hematopoietic and endothelial development in an oxygen-responsive manner

Vascular endothelial growth factor (VEGF) and the vascular endothelial growth factor receptors (VEGFRs) regulate the development of hemogenic mesoderm. Oxygen concentration-mediated activation of hypoxia-inducible factor targets such as VEGF may serve as the molecular link between the microenvironment and mesoderm-derived blood and endothelial cell specification. We used controlled-oxygen microenvironments to manipulate the generation of hemogenic mesoderm and its derivatives from embryonic stem cells. Our studies revealed a novel role for soluble VEGFR1 (sFlt-1) in modulating hemogenic mesoderm fate between hematopoietic and endothelial cells. Parallel measurements of VEGF and VEGFRs demonstrated that sFlt-1 regulates VEGFR2 (Flk-1) activation in both a developmental-stage-dependent and oxygen-dependent manner. Early transient Flk-1 signaling occurred in hypoxia because of low levels of sFlt-1 and high levels of VEGF, yielding VEGF-dependent generation of hemogenic mesoderm. Sustained (or delayed) Flk-1 activation preferentially yielded hemogenic mesoderm-derived endothelial cells. In contrast, delayed (sFlt-1-mediated) inhibition of Flk-1 signaling resulted in hemogenic mesoderm-derived blood progenitor cells. Ex vivo analyses of primary mouse embryo-derived cells and analysis of transgenic mice secreting a Flt-1-Fc fusion protein (Fc, the region of an antibody which is constant and binds to receptors) support a hypothesis whereby microenvironmentally regulated blood and endothelial tissue specification is enabled by the temporally variant control of the levels of Flk-1 activation. Disclosure of potential conflicts of interest is found at the end of this article.