Circulating Endothelial Cells and Endothelial Progenitor Cells in Obstructive Sleep Apnea

Increased circulating endothelial cells (CECs) have been observed in patients with vascular injury associated with acute myocardial infarction, pulmonary hypertension, and congestive heart failure. Decreased circulating endothelial progenitor cells (EPCs) have been observed in patients with risk factors for cardiovascular disease. Obstructive sleep apnea (OSA) is associated with increased risk of cardiovascular disease and endothelial dysfunction. Subjects were recruited from patients referred for overnight polysomnograms; 17 subjects had OSA and 10 control subjects did not have OSA. All subjects lacked vascular disease and risk factors for vascular disease. Peripheral blood was obtained from fasting subjects in the morning, following sleep studies. CECs and EPCs were quantified using magnetic bead separation with UV epifluorescence microscopy and flow cytometry immunophenotyping, respectively. Cell counts and demographic variables were compared using unpaired t tests. Regression analysis was performed comparing cell counts with the apnea-hypopnea index (AHI) and nadir SaO(2). Subjects with OSA and controls did not differ significantly in terms of age and body mass index. Subjects with OSA had higher AHI, lower nadir SaO(2), and greater sleepiness (Epworth Sleepiness Scale scores). There were no significant differences in CEC (7.0+/-1.5 vs. 4.9+/-0.9, p>0.05) or EPC (1077+/-318 vs. 853+/-176, p>0.05) between controls and OSA cases, respectively. In this small study, we found no differences in CECs or circulating EPCs between patients with OSA and controls. OSA may not be associated with these markers of vascular endothelial cell injury in patients with no concomitant vascular disease.     

冠心病患者循环内皮祖细胞与血管内皮功能的变化

目的:探讨冠心病患者循环内皮祖细胞(EPCs)数量及功能变化与血管内皮舒张功能的关系。方法:将58例患者分为对照组(20例)、稳定性心绞痛组(11例)、不稳定性心绞痛组(27例)。采用高分辨率二维超声检测肱动脉血流介导的内皮依赖性血管舒张功能(FMD)及硝酸甘油介导的非内皮依赖性血管舒张功能(NMD);用密度梯度离心法从外周血获取单个核细胞,将其接种在人纤维连接蛋白包被培养板,培养7天后贴壁细胞进行细胞化学分析,激光共聚焦显微镜鉴定异硫氰酸荧光素标记荆豆凝集素I和DiI标记的乙酰化低密度脂蛋白双染色阳性细胞为正在分化的EPCs,采用二苯基四氮唑嗅盐比色法、改良的Boyden小室和黏附能力测定实验观察内皮祖细胞的增殖能力、迁移能力和黏附能力。结果:①稳定性心绞痛组与不稳定性心绞痛组的FMD均明显低于对照组,有显著性差异(P<0．05～0．01),而稳定性心绞痛组的FMD较不稳定性心绞痛组也降低,有显著性差异(P<0．05);3组间NMD差异无统计学意义(P>0．05)。②稳定性心绞痛组、不稳定性心绞痛组较对照组循环EPCs数量明显减少,且黏附、迁移及增殖能力也明显下降,均有极显著性差异(P<0．01);不稳定性心绞痛组较稳定性心绞痛组EPCs数量及迁移能力无差异(P>0．05),但黏附和增殖能力降低(P<0．05)。③直线相关性分析发现不稳定性心绞痛组EPCs的数量及功能均与FMD呈正相关(P<0．05),而稳定性心绞痛组仅EPCs黏附功能与FMD呈正相关(P<0．05)。结论:冠状动脉EPCs数量及功能下降与血管内皮依赖性舒张功能障碍一致,提示当冠心病患者血管内皮功能受损而又缺乏足够有效的EPCs时,可能影响冠心病的病情程度及临床表现。     

小鼠骨髓源性内皮祖细胞的分离培养与鉴定

目的 建立一种高效、稳定的从小鼠骨髓中分离培养与定向诱导分化内皮祖细胞的方法.方法 通过密度梯度离心法从小鼠骨髓中分离单个核细胞,经差速贴壁结合特殊培养基扩增,诱导分化为内皮祖细胞.应用流式细胞技术鉴定内皮细胞系列标志:CD34、CD31、Flk-1和祖细胞标志CD133.结果 经密度梯度离心和差速贴壁法分离所得的细胞经EBM-2专用培养基培养后,第4天可见集落形成,培养第12天流式细胞仪检测其CD34、CD133、Flk-1、CD31的阳性率分别为65%±4%、48%±3%、37%±3%和51%±4%.结论 从小鼠骨髓中分离培养与定向诱导分化内皮祖细胞的方法效率高,稳定性和重复性好.     

Increased Endothelial Progenitor Cells and Nitric Oxide in Young Prehypertensive Women

This study investigated the effect of sex differences on circulating endothelial progenitor cells (EPCs) in prehypertension and its underlying mechanism. The authors found that premenopausal women show increased number and activity of circulating EPCs when compared with men, which was similar to enhanced nitric oxide (NO) level in plasma or culture medium. There was no difference in the number and activity of circulating EPCs and NO level between normotensive and prehypertensive premenopausal women. There was also no difference seen in levels of vascular endothelial growth factor and granulocyte macrophage colony‐stimulating factor. Both number and activity of circulating EPCs were correlated with the level of NO. The present study firstly demonstrated that the number and activity of circulating EPCs were preserved in prehypertensive premenopausal women, which was related to the restoration of NO production. The sex differences in EPCs in prehypertension may be involved in the mechanism underlying vascular protection in premenopausal women.     

内皮祖细胞的转化与扩增研究进展

内皮祖细胞移植有可能成为治疗缺血性脑血管病的新策略。内皮祖细胞数量有限成为限制其治疗应用的主要障碍。如何在一定条件下有效扩增内皮祖细胞或将其他组织来源的细胞转化为内皮祖细胞具有十分重要的意义。近年来已发现一些药物、其他组织来源的细胞以及某些大分子物质对增加内皮祖细胞数量有着积极的作用。     

阿托伐他汀增加高血压患者外周血内皮祖细胞数量

目的观察阿托伐他汀对无脂代谢紊乱的高血压患者外周血内皮祖细胞(EPCs)数量及血压的影响。方法原发性高血压患者38例随机分为单用常规降压药物组(常规组,n=18)和常规降压药物与阿托伐他汀(20mg 睡前)联合用药组(联合组,n=20)。8例健康志愿者口服阿托伐他汀8周作为对照组(n=8)。于治疗前和治疗后8周分别测血压并抽取外周血进行 EPCs 的分离培养,第10天对 EPCs 进行鉴定并于倒置相差显微镜下计数内皮祖细胞克隆形成单位(EPC-CFU)以评估外周血 EPCs 水平。结果 1)常规组和联合组治疗前后收缩压均有显著下降,分别为(165.8±10.3)vs(132.7±10.3)mmHg 和(163.7±10.2)vs(127.9±10.1)mmHg;加用阿托伐他汀血压下降幅度较单用降压药大[(35.7±3.4)vs(33.1±2.4)mmHg,P<0.05]。对照组服药前后收缩压的差异无统计学意义(114.2±18.4)vs(108.4±21.6)mmHg。2)常规降压药8周后 EPC-CFU 从(8.8±2.0)升为(12.1±2.2);联合用药后 EPC-CFU 从(9.2±1.9)升为(13...     

阿托伐他汀增加高血压患者外周血内皮祖细胞数量

目的 观察阿托伐他汀对无脂代谢紊乱的高血压患者外周血内皮祖细胞(EPCs)数量及血压的影响.方法 原发性高血压患者38例随机分为单用常规降压药物组(常规组,n=18)和常规降压药物与阿托伐他汀(20 mg睡前)联合用药组(联合组,n=20).8例健康志愿者口服阿托伐他汀8周作为对照组(n=8).于治疗前和治疗后8周分别测血压并抽取外周血进行EPCs的分离培养,第10天对EPCs进行鉴定并于倒置相差显微镜下计数内皮祖细胞克隆形成单位(EPC-CFU)以评估外周血EPCs水平.结果 1)常规组和联合组治疗前后收缩压均有显著下降,分别为(165 8±10 3)vs(132 7±10 3)mmHg和(163 7±10 2) vs(127 9±10 1)mmHg;加用阿托伐他汀血压下降幅度较单用降压药大[(35 7±3 4)vs(33 1±2 4)mmHg,P<0 05].对照组服药前后收缩压的差异无统计学意义(114 2±18 4)vs(108 4±21 6)mmHg.2)常规降压药8周后EPC-CFU从(8 8±2 0)升为(12 1±2 2);联合用药后EPC-CFU从(9 2±1 9)升为(13 6±2 2)个.两组EPC-CFU改变值[(3 3±0 6)vs(4 5±1 1)]的差异有统计学意义(P<0 05).对照组服药前后EPC-CFU为(13 2±2 8)vs(16 2±3 5).3)联合组治疗前后血压下降幅度与EPC-CFU改变值呈正相关(r=0 581,P=0 007).结论 高血压可减少外周血EPCs数量,在应用常规降压药物基础上加用阿托伐他汀可增加外周血EPCs数量并进一步降低血压.     

AMP-Activated Protein Kinase Inhibits Homocysteine-Induced Dysfunction and Apoptosis in Endothelial Progenitor Cells

Purpose  

Homocysteine (Hcy) has been shown to induce oxidative stress and apoptosis of endothelial progenitor cells (EPCs). AMP-activated protein kinase (AMPK) has been reported to have protective effects on endothelial function. However, effects of AMPK activation on Hcy-induced EPCs injury remain to be determined. In this study, we examined the effect of AMPK phosphorylation on Hcy-induced NO bioavailability impairment and NADPH oxidase 4 (Nox4) derived reactive oxygen species (ROS) accumulation in EPCs.     

缓激肽对内皮祖细胞存活、迁移及凋亡的影响

目的 观察不同浓度的缓激肽对人脐血来源的内皮祖细胞存活、迁移及凋亡的影响。方法 采用密度梯度离心法从人脐带血获取单个核细胞,将其接种在人纤维连接蛋白包被的培养板上,培养7天后,收集贴壁细胞。通过免疫荧光法鉴定,FITC标记的异凝集素和DiI标记的乙酰化低密度脂蛋白染色双染色阳性细胞为正在分化的内皮祖细胞,并经流式细胞仪检测技术进一步确定内皮祖细胞。以不同浓度缓激肽（1、10、100 nmol/L）或缓激肽B2受体阻断剂艾替班特+ 10 nmol/L 缓激肽干预内皮祖细胞 16 h。分别采用MTT比色法、Transwell小室和膜联蛋白V-异硫氰酸荧光素/碘化丙啶流式细胞术及Hoechst33342染色法观察缓激肽对人内皮祖细胞的存活、迁移以及凋亡影响。 结果 缓激肽在1、10 nmol/L浓度时可促进人内皮祖细胞存活、迁移,抑制其凋亡（P<0.05）。然而100 nmol/L缓激肽无促人内皮祖细胞存活、迁移和抑制其凋亡的作用（P>0.05）。缓激肽促人内皮祖细胞存活、迁移,抑制其凋亡的作用可被艾替班特阻断（P<0.05）。 结论 缓激肽在一定范围内可促进内皮祖细胞的存活、迁移,并抑制其凋亡,此作用主要由缓激肽B2受体介导。     

Systemic Inflammation,Vascular Function,and Endothelial Progenitor Cells after an Exercise Training Intervention in COPD

BackgroundExercise training is a cornerstone of the treatment of chronic obstructive pulmonary disease (COPD) in all disease stages. Data about the training effects with supplemental oxygen in nonhypoxemic patients remains inconclusive. In this study we set out to investigate the training and oxygen effects on inflammatory markers, vascular function, and endothelial progenitor cells in this population of increased cardiovascular risk.MethodsIn this prospective, randomized, double-blind, crossover study, 29 patients with nonhypoxemic COPD performed combined endurance and strength training 3 times a week while breathing medical air or supplemental oxygen for the first 6-week period, and were then reallocated to the opposite gas for the following 6 weeks. Exercise capacity, inflammatory biomarkers, endothelial function (peripheral arterial tone analysis), and endothelial progenitor cells were assessed. Data were also analyzed for a subgroup with endothelial dysfunction (reactive hyperemia index <1.67).ResultsFollowing 12 weeks of exercise training, patients demonstrated a significant improvement of peak work rate and an associated decrease of blood fibrinogen and leptin. Eosinophils were found significantly reduced after exercise training in patients with endothelial dysfunction. In this subgroup, peripheral arterial tone analysis revealed a significant improvement of reactive hyperemia index. Generally, late endothelial progenitor cells were found significantly reduced after the exercise training intervention. Supplemental oxygen during training positively influenced the effect on exercise capacity without impact on inflammation and endothelial function.ConclusionsThis is the first randomized controlled trial in patients with COPD to show beneficial effects of exercise training not only on exercise capacity, but also on systemic/eosinophilic inflammation and endothelial dysfunction.     

Inhibition of S1P Receptor 2 Attenuates Endothelial Dysfunction and Inhibits Atherogenesis in Apolipoprotein E-Deficient Mice

Aim: The bioactive lipid, sphingosine-1-phosphate (S1P), has various roles in the physiology and pathophysiology of many diseases. There are five S1P receptors; however, the role of each S1P receptor in atherogenesis is still obscure. Here we investigated the contribution of S1P receptor 2 (S1P2) to atherogenesis by using a specific S1P2 antagonist, ONO-5430514, in apolipoprotein E-deficient ( Apoe ^−/− ) mice. Methods: Apoe ^−/− mice fed with a western-type diet (WTD) received ONO-5430514 (30 mg/kg/day) or vehicle. To examine the effect on atherogenesis, Sudan IV staining, histological analysis, qPCR, and vascular reactivity assay was performed. Human umbilical vein endothelial cells (HUVEC) were used for in vitro experiments. Results: WTD-fed Apoe ^−/− mice had significantly higher S1P2 expression in the aorta compared with wild-type mice. S1P2 antagonist treatment for 20 weeks reduced atherosclerotic lesion development ( p ＜0.05). S1P2 antagonist treatment for 8 weeks ameliorated endothelial dysfunction ( p ＜0.05) accompanied with significant reduction of lipid deposition, macrophage accumulation, and inflammatory molecule expression in the aorta compared with vehicle. S1P2 antagonist attenuated the phosphorylation of JNK in the abdominal aorta compared with vehicle ( p ＜0.05). In HUVEC, S1P promoted inflammatory molecule expression such as MCP-1 and VCAM-1 ( p ＜0.001), which was attenuated by S1P2 antagonist or a JNK inhibitor ( p ＜0.01). S1P2 antagonist also inhibited S1P-induced JNK phosphorylation in HUVEC ( p ＜0.05). Conclusions: Our results suggested that an S1P2 antagonist attenuates endothelial dysfunction and prevents atherogenesis. S1P2, which promotes inflammatory activation of endothelial cells, might be a therapeutic target for atherosclerosis.     

Circulating Endothelial Progenitor Cells,Th1/Th2/Th17-Related Cytokines,and Endothelial Dysfunction in Resistant Hypertension

IntroductionA possible link between chronic vascular inflammation and arterial hypertension is now an object of intensive studies.ObjectiveTo compare Th1/Th2/Th17 cells-related cytokines, circulating endothelial progenitor cells (EPC), and endothelial function in subjects with resistant arterial hypertension (RAH) and controlled arterial hypertension (CAH).MethodsBlood pressure was measured by electronic sphygmomanometer. EPC were identified as CD34 +/ CD133 +/kinase insert domain receptor (KDR) + cells by flow cytometry. Th1/Th2/Th17 cells-related cytokines were identified using the Human Th1/Th2/Th17 Cytokines MultiAnalyte ELISArray Kit. Endothelium-dependent (FMD) vasodilatation of brachial artery was measured by Doppler ultrasound scanning.ResultsRAH group (n = 20) and CAH group (n = 20) and 17 healthy individuals (control group) were recruited. In the RAH group, lower blood levels of EPC number (42.4 ± 16.7 cells/mL) and EPC% (0.19 ± 0.08%) were observed than in the CAH group (93.1 ± 88.7 cells/mL; P = 0.017; 0.27 ± 0.17; P = 0.036) and control group (68.5 ± 63.6 cells/mL; P < 0.001; 0.28 ± 0.17%; P = 0.003), respectively. Plasma transforming growth factor-β1 levels were significantly higher in the RAH group (1767 ± 364 pg/mL) than in the CAH group (1292 ± 349; P < 0.001) and in control group (1203 ± 419 pg/mL; P < 0.001). In the RAH group, statistically significant negative correlation was observed between systolic blood pressure and EPC% (r = –0.72, P < 0.01). FMD in the RAH group was significantly lower (5.5 ± 0.8%) than in the CAH group (9.2 ± 1.4; P < 0.001) and in healthy controls (10.1 ± 1.1%; P < 0.001).ConclusionsRAH is characterized by reduced circulating EPC, substantial endothelial dysfunction, and increased plasma transforming growth factor-β1 levels.     

大鼠血管内皮祖细胞培养及鉴定

目的探索骨髓来源的血管内皮祖细胞的分离培养方法,为缺血性疾病治疗找到新的移植细胞来源。方法采集SD雄性大鼠骨髓细胞,用梯度密度离心法分离单核个细胞,种植于提前包埋了纤维连接蛋白的培养皿中培养,将血管内皮生长因子(VEGF)、重组纤维细胞生长因子(bFGF)和重组上皮生长因子(EGF)作为诱导剂,将72h后贴壁细胞(A组)和非贴壁细胞(B组)分别培养,观察细胞形态的变化以及数量变化,取A组在第14天,应用免疫组化和免疫荧光技术鉴定内皮祖细胞系列标志。结果A组细胞经过体外诱导后大量扩增,在28d时仍有强大扩增的活力,呈铺路石样,而B组细胞短期内可以大量增殖,呈长索样,之后逐渐变少;在细胞迁移实验中,A组细胞可以相互融合形成血管结构,而B组细胞之间则不能融合,随着时间推移,逐渐凋亡;A组经过CD34免疫荧光、CD133、FLK-1免疫组化鉴定呈阳性,并能特异性吸附FITC-UEA-和内吞DIL-Ac-LDL。结论自体骨髓细胞通过贴壁换液后可以分离培养出内皮祖细胞,取贴壁细胞经过体外诱导后大量扩增,并通过表面标记物鉴定可以确认为内皮祖细胞,而非贴壁细胞不能大量增殖,故将贴壁分离方法来筛选内皮祖细胞,此方法简易,并能满足细胞移植的需要,因而,在移植治疗脑缺血引起的内皮损伤应该有较好的临床应用前景。     

Potential Link Between C3a,C3b and Endothelial Progenitor Cells in Resistant Hypertension

IntroductionEndothelial progenitor cells (EPC) and complement C3 are involved in the pathophysiology of arterial hypertension. C3a is the negative regulator of progenitor cells egress during their mobilization from bone marrow. Previously, higher plasma concentration of C3 was observed in resistant arterial hypertension (RAH) than in controlled arterial hypertension (CAH). Thus, we hypothesized that RAH would be associated with complement C3 activation and reduced number of circulating EPCs.ObjectiveTo compare C3a, C3b and their correlation with circulating EPC in subjects with RAH and CAH.MethodsBlood pressure was measured by electronic sphygmomanometer. EPCs were identified as CD34+/CD133+/KDR+ cells by flow cytometry. C3a and C3b were determined using enzyme-linked immunosorbent assay (Quidel, CA).ResultsRAH group (n = 20) and CAH group (n = 20) and 17 healthy individuals (control group) were recruited. In the RAH group, C3a (858.1 ± 70.6 μg/dL) was higher than in the CAH group (816.1 ± 123.3 μg/dL; P < 0.001), and in the control group (751.3 ± 98.8; P < 0.001), C3b (564.1 ± 54.7 μg/dL) was higher than in the CAH group (490.2 ± 58.5 μg/dL; P < 0.001). In control group (456.3 ± 98.8; P < 0.001), statistically significant negative correlation was observed between C3a and blood levels of EPC (r = ?0.523, P = 0.018); statistically significant positive correlation was observed between systolic blood pressure and blood levels of C3a (r = 0.52, P = 0.02) and between systolic blood pressure and blood levels of C3b (r = 0.57, P = 0.009).ConclusionRAH is characterized by higher levels of C3 component fragments and a negative correlation between circulating C3a and EPCs.     

运动康复对心血管疾病内皮祖细胞的动员作用及机制

内皮祖细胞是一种新型生物标志物,其数量下降可预测心血管不良事件的高发生率。目前运动康复降低心血管疾病死亡率及提高患者生活质量已成为研究热点。大量研究证明运动康复可促进内皮祖细胞动员及改善内皮功能,但其具体机制尚不十分明确。目前认为其可能与缺血刺激和重要体液因子的调节密切相关。文章将运动康复对心血管疾病内皮祖细胞的动员作用及机制作一综述。     

培哚普利对冠心病患者循环血内皮祖细胞及血管内皮功能的影响

目的：探讨不同剂量培哚普利在冠心病治疗中对循环血内皮祖细胞（EPCs）水平及血管内皮功能的影响。
　　方法：选取我院经冠状动脉造影确诊为冠心病的患者84例,随机分为对照组（n=27,给予常规药物）,小剂量组（n=29,给予常规药物+4 mg培哚普利）和大剂量组（n=28,给予常规药物+8 mg培哚普利）。随访12周,治疗前后各组分别采用流式细胞术检测EPCs水平,采用超声测定肱动脉血管舒张功能（FMD）,同时检测高敏C反应蛋白（hs-CRP）、血管紧张素II（Ang II）水平。
　　结果：治疗12周后,对照组、小剂量组及大剂量组患者较治疗前循环血EPCs、肱动脉FMD均有不同程度的增高,hs-CRP水平均有不同程度的降低（ P<0.05）;小剂量组及大剂量组Ang II较治疗前均有不同程度的降低（P<0.05）;治疗后,小剂量组及大剂量组循环血EPCs、肱动脉FMD均显著高于对照组,hs-CRP及Ang II均显著低于对照组（ P<0.01）,同时,与小剂量组比较,大剂量组循环血EPCs及FMD均明显升高,hs-CRP及Ang II均明显降低（ P<0.05）。
　　结论：培哚普利对循环血EPCs有一定的动员作用,可显著改善血管内皮功能,且较大剂量效果更显著。