糖基化终末产物对人牙周膜干细胞生物学特性的影响

目的探讨糖基化终末产物(advanced glycation end products,AGEs)对人牙周膜干细胞(human periodontal ligament stem cells,HPDLSCs)生物学特性的影响研究。方法 HPDLSCs培养、提纯以及鉴定;检测不同质量浓度AGEs(50、100、200μg/ml)下HPDLSCs增殖能力、克隆能力、成骨分化能力。Real-time PCR检测不同质量浓度AGEs对HPDLSCs白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)和肿瘤坏死因子(TNF-α)m RNA表达的影响。结果 MTT结果显示,与正常组比较,不同质量浓度AGEs均抑制HPDLSCs的增殖,且200μg/ml AGEs抑制最明显。克隆结果显示,与正常组比较,不同质量浓度AGEs均抑制HPDLSCs的自我更新及克隆能力,且200μg/ml AGEs抑制最明显。成骨分化能力结果显示,与正常组比较,不同质量浓度AGEs均抑制HPDLSCs的成骨能力,且200μg/ml AGEs抑制最明显。RT-PCR结果显示,与正常组比较,不同质量浓度AGEs均促进IL-1β、IL-6和TNF-αm RNA的表达,且随着质量浓度的增加,炎性因子的表达增加。结论 AGEs成浓度依赖性抑制HPDLSCs的增殖能力、自我更新能力、克隆能力和成骨分化能力,同时刺激炎性因子的表达。因此,牙周膜干细胞治疗糖尿病伴牙周病导致的牙周组织损伤要考虑糖基化终末产物的影响。     

晚期糖基化终末产物诱导人卵巢颗粒细胞株COV434凋亡并上调促炎因子HMGB1自分泌

目的:探讨晚期糖基化终末产物(advanced glycation end products,AGEs)是否诱导人卵巢颗粒细胞株COV434凋亡,并探究其可能的机制。方法:采用不同浓度AGEs牛血清白蛋白与人卵巢颗粒细胞株COV434共同孵育,流式细胞术观察细胞凋亡率,Western blot观察caspase-3和cleaved caspase-3的蛋白水平,ELISA检测细胞培养液上清中高迁移率族蛋白B1(high mobility group box 1 protein,HMGB1)的含量。结果:与对照组比较,100 mg/L AGEs组和200 mg/L AGEs组早、晚期凋亡率显著增加,各组caspase-3蛋白水平的差异无统计学显著性,但与对照组比较,100 mg/L AGEs组和200 mg/L AGEs组cleaved caspase-3的蛋白水平显著增加(P0.05)。此外,与对照组比较,100 mg/L AGEs组和200 mg/L AGEs组细胞培养液上清中HMGB1促炎介质的水平显著上升(P0.05)。结论:晚期糖基化终末产物诱导人卵巢颗粒细胞株COV434凋亡可能与促炎反应有关。     

TRB3通过GSK-3β信号通路参与AGEs诱导胰岛细胞凋亡

<正>目的:探讨TRB3在晚期糖基化终末产物(AGEs)诱导β细胞凋亡中的作用及其与GSK-3β通路相关的分子机制。方法:首先检测糖尿病大鼠模型发病不同阶段体内AGEs水平、胰岛β细胞TRB3表达情况与β细胞凋亡的关系。细胞水平上,检测AGEs处理后大鼠胰岛β细胞系INS-1细胞TRB3表达水平与细胞凋亡的关系以及沉默或过表达TRB3对AGEs诱导INS-1     

硫化氢抑制糖基化终末产物引起上皮细胞钠通道异常激活的机制研究

<正>目的:探讨糖基化终末产物(AGEs)激活上皮细胞钠通道(ENa C)的作用,及H2S对抗AGEs引起ENa C异常激活的机制研究。方法:应用膜片钳记录爪蟾肾皮质集合管上皮细胞ENa C电流,研究H2S对抗AGEs引起ENa C激活的机制;应用共聚焦显微镜技术观察H2S对AGEs引起胞内活性氧水平升高的调节作用;应用分子生物学实验检测AGEs处理后过氧化氢酶的     

晚期糖基化终末产物对肠道L细胞凋亡及增殖活性的影响

目的探讨晚期糖基化终末产物(AGEs)对肠道L细胞株GLUTag细胞凋亡及细胞增殖活性的影响。方法肠道L细胞株GLUTag细胞由加拿大多伦多大学Druker实验室惠赠。将GLUTag细胞株分5组:空白对照组,牛血清白蛋白(BSA)对照组,AGEs 100μg/m L组,AGEs 200μg/m L组,AGEs 300μg/m L组。培养方法:空白对照组,将GLUTag细胞株培养于低糖DMEM中;BSA对照组,将GLUTag细胞株细胞培养于加入BSA的低糖DMEM中;AGEs 100μg/m L组,将GLUTag细胞株培养于终质量浓度100μg/m L AGEs的低糖DMEM中;AGEs 200μg/m L组,将GLUTag细胞株培养于终质量浓度200μg/m L AGEs的低糖DMEM中;AGEs 300μg/m L组,将GLUTag细胞株培养于终质量浓度300μg/m L AGEs的低糖DMEM中;每组细胞均培养24 h。将培养在终质量浓度200μg/m L AGEs中GLUTag细胞株分别培养0、12、24、48 h(即4组)。各组细胞干预结束后采用双染细胞凋亡检测方法检测细胞凋亡率;Hoechst33258荧光染色观察细胞形态;细胞计数试剂盒检测细胞增殖活性。结果各组细胞经药物干预24 h后,AGEs 100μg/m L组、AGEs 200μg/m L组、AGEs 300μg/m L组凋亡细胞百分率明显升高,均显著高于空白对照组和BSA对照组,且AGEs 300μg/m L组凋亡细胞百分率最高(P=0.000)。AGEs 200μg/m L作用12、24、48 h后凋亡细胞百分率显著高于阴性对照组(作用0 h),且作用48 h凋亡率最高(P=0.000)。100、200、300μg/m L AGEs作用GLUTag细胞24 h后,细胞活性光密度(OD)值显著低于空白对照组和BSA对照组,且AGEs 300μg/m L组OD值最低(P0.05)。AGEs 200μg/m L作用12、24、48 h后细胞OD值显著低于阴性对照组(作用0 h),其中48 h组OD值最低(P0.05)。结论 AGEs对肠道L细胞株GLUTag细胞的凋亡及增殖活性均产生影响,其细胞凋亡率在相同作用时间下随着AGEs的浓度增高而增多,且在相同剂量下随着AGEs的作用时间延长而增多。AGEs可诱导肠道L细胞凋亡,抑制细胞增殖活性,从而损伤肠道L细胞。     

高游离脂肪酸通过FOXO1途径抑制内皮祖细胞增殖并促进其凋亡

目的: 研究高游离脂肪酸(FFA)对糖尿病患者内皮祖细胞(EPCs)增殖及凋亡的影响,评价FFA在糖尿病血管并发症中的可能作用。方法: 分离培养2型糖尿病患者EPCs(DM组)和正常对照组EPCs(对照组),加入不同浓度(0、10和50 mmol/L)的FFA。MTT法检测EPCs增殖,Annexin-V/PI法检测其凋亡。RT-PCR法和Western blotting法检测核转录因子FOXO1的表达水平。结果: 随着FFA浓度的增加,DM组和对照组EPCs增殖率明显下降,而凋亡率明显增加,差异均显著(P＜0.05)。相同FFA浓度下,DM组增殖率低于对照组(P＜0.05)。随着FFA浓度的增加,DM组和对照组FOXO1的mRNA表达逐渐降低,差异显著(P＜0.05)。结论: 高FFA可能通过FOXO1途径抑制EPCs增殖并诱导其凋亡,可能在糖尿病血管并发症的发生中发挥作用。     

球状脂联素上调脂联素受体1抑制晚期糖基化终产物诱导人脐静脉内皮细胞凋亡的初步研究

目的:观察球状脂联素(gAd)对晚期糖基化终产物(AGEs)诱导人脐静脉内皮细胞(HUVECs)凋亡时脂联素受体1(AdipoR1)表达的影响。方法:体外培养HUVECs,用不同浓度AGEs作用HUVECs,以及预处理gAd后再联合AGEs作用HUVECs。MTT比色法测定细胞活性、Annexin V-FITC/PI双染标记流式细胞仪检测细胞凋亡、实时定量RT-PCR检测内皮细胞AdipoR1 mRNA的表达。结果:AGEs作用HUVECs,细胞凋亡具有AGEs浓度依赖性(P0.05)。相同浓度AGEs作用HUVECs,有和无gAd预处理的2组比较,用gAd预处理组明显拮抗HUVECs凋亡(P0.01)。实时定量RT-PCR检测发现,gAd具有上调AdipoR1 mRNA表达的作用,与相同浓度AGEs作用的无gAd预处理组比较,差异显著(P0.01)。结论:gAd可能通过上调AdipoR1表达拮抗AGEs诱导HUVECs的凋亡。     

黄芪甲苷对EPCs中SDF-1α/CXCR4的调控作用

目的:探讨黄芪甲苷(AS-IV)对内皮祖细胞(EPCs)中CXC趋化因子受体4(CXCR4)和基质细胞衍生因子1α(SDF-1α)的调控作用及其作用机制。方法:体外培养大鼠骨髓源性EPCs,观察应用AS-IV及CXCR4的特异性阻断剂AMD3100后EPCs的增殖、黏附、迁移、凋亡和管状结构形成能力的变化,并分析AS-IV对EPCs中SDF-1α及CXCR4的mRNA和蛋白,以及p-CXCR4蛋白水平变化的影响。结果:AS-IV可以显著提升EPCs的增殖、黏附、迁移和管状结构形成能力,减轻EPCs的凋亡,上调EPCs中SDF-1α和CXCR4的mRNA和蛋白及p-CXCR4蛋白的水平(P0.05);AMD3100可以阻断AS-IV对CXCR4的mRNA和蛋白及p-CXCR4蛋白水平的上调作用,但不影响AS-IV对SDF-1α的mRNA和蛋白水平的上调作用。结论:AS-IV可能通过调控EPCs中SDF-1α/CXCR4的表达而增强EPCs的生物学作用。     

转化生长因子α促进人内皮祖细胞增殖、迁移和黏附

目的:探讨转化生长因子α(transforming growth factorα,TGF-α)对人内皮祖细胞(endothelial progenitor cells,EPCs)单克隆形成、增殖、迁移和黏附细胞功能的影响及机制。方法:分离培养人内皮祖细胞,分别用浓度为1、5、10μg/L TGF-α诱导处理细胞,并设置PBS对照组和表皮生长因子受体酪氨酸激酶抑制剂EGFR-TKI组(同时加入10μg/L TGF-α和1∶1 000的EGFR-TKI)。利用单克隆形成实验、MTT法和Ed U法、Transwell法和细胞黏附实验检测不同浓度TGF-α对各组EPCs单克隆形成能力、细胞增殖、细胞迁移能力及黏附功能的影响;并通过Western blotting法检测不同浓度TGF-α对各组EPCs中表皮生长因子受体(epithelial growth factor receptor,EGFR)和血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的影响。结果:不同浓度TGF-α均能显著诱导EPCs单克隆形成能力、增殖、迁移和黏附细胞功能,差异有统计学意义(P0.05),并可以被EGFR-TKI抑制。Western blotting检测发现TGF-α显著诱导EPCs中EGFR和VEGF的表达(P0.05),并呈浓度依赖性。结论:TGF-α能够显著促进人EPCs细胞单克隆形成、增殖、迁移和黏附相关细胞功能,并通过与EGFR结合诱导VEGF表达来发挥作用。     

高糖对糖尿病患者内皮祖细胞增殖凋亡的影响及其机制探讨

目的：了解葡萄糖对糖尿病患者内皮祖细胞（EPCs）增殖及凋亡的影响，并探讨其可能机制。方法：以不同浓度葡萄糖分别作用于25例正常人(对照组)和25例2型糖尿病患者(糖尿病组)EPCs，MTT法检测其增殖，Annexin-V/PI法检测其凋亡。RT-PCR法检测bcl-2和bax的表达水平。结果：33 mmol/L葡萄糖使糖尿病组和对照组EPCs增殖率均减低，凋亡率增高；且能使糖尿病组和对照组EPCs表达bax增强，bcl-2表达无明显改变。而5 mmol/L葡萄糖作用下糖尿病组和对照组EPCs增殖率、凋亡率及bax和bcl-2表达无明显改变。结论：高糖使EPCs增殖功能减弱，凋亡增加，bax表达增加可能是其机制之一。     

AGEs诱发体外大鼠心脏微血管内皮细胞iNOS表达

目的观察体外原代培养心脏微血管内皮细胞内一氧化氮(NO)生成与诱导型一氧化氮合酶(iNOS)表达的变化及相关性。方法采用不同浓度牛血清白蛋白糖基化终末产物(BSA-AGEs)(50~200 mg/L)分别作用于心脏微血管内皮细胞(0~24 h),检测NO生成,免疫组化及Western blot检测iNOS表达。结果随着AGEs浓度增加及作用时间的延长,心脏微血管内皮细胞中NO生成增多,iNOS表达也增多(P<0.05)。结论BSA-AGEs可以诱发心脏微血管内皮细胞产生毒性NO,从而引发以微血管内皮功能障碍为特征的糖尿病性心肌病。     

AGEs Decreased SIRT3 Expression and SIRT3 Activation Protected AGEs-Induced EPCs’ Dysfunction and Strengthened Anti-oxidant Capacity

Advanced glycation end products (AGEs) have been confirmed to induce dysfunction in endothelial progenitor cells (EPCs) and play key roles in pathogenesis of diabetes-related vascular complications. The major function of sirtuin 3 (SIRT3) is to orchestrate oxidative metabolism and control reactive oxygen species (ROS) homeostasis, which are more closely related to EPCs’ dysfunction. Our study therefore was designed to explore the role of SIRT3 on AGEs-induced EPCs dysfunction of. EPCs isolated from healthy adults were stimulated with AGEs and the expression of SIRT3 was assessed. Then, EPCs transfected with ad-SIRT3 or siRNA-SIRT3 were cultured with or without AGEs. EPCs function, including proliferation, migration; expression of manganese superoxide dismutase (MnSOD), ROS production, and interleukin-8 (IL-8); and vascular endothelial growth factor (VEGF) production were measured. In some experiments, EPCs were pre-cultured with anti-receptor for advanced glycation end products (RAGE) antibody or anti-neutralizing antibody, and then proliferation, migration, expression of MnSOD, ROS production, and IL-8 and VEGF production were measured. Our results showed that SIRT3 expressed in EPCs and AGEs decreased SIRT3 expression. SIRT3 knockdown with siRNA-SIRT3 promoted dysfunction in EPCs whereas SIRT3 activation with ad-SIRT3 strengthened anti-oxidant capacity and protected AGE-impaired dysfunction. Moreover, RAGE may involve in AGEs-decreased SIRT3 expression in EPCs. These data suggested an important role of SIRT3 in regulating EPCs bioactivity.     

Anti-angiogenic effect of high doses of ascorbic acid

Pharmaceutical doses of ascorbic acid (AA, vitamin C, or its salts) have been reported to exert anticancer activity in vitro and in vivo. One proposed mechanism involves direct cytotoxicity mediated by accumulation of ascorbic acid radicals and hydrogen peroxide in the extracellular environment of tumor cells. However, therapeutic effects have been reported at concentrations insufficient to induce direct tumor cell death. We hypothesized that AA may exert anti-angiogenic effects. To test this, we expanded endothelial progenitor cells (EPCs) from peripheral blood and assessed, whether or not high dose AA would inhibit EPC ability to migrate, change energy metabolism, and tube formation ability. We also evaluated the effects of high dose AA on angiogenic activities of HUVECs (human umbilical vein endothelial cells) and HUAECs (human umbilical arterial endothelial cells). According to our data, concentrations of AA higher than 100 mg/dl suppressed capillary-like tube formation on Matrigel for all cells tested and the effect was more pronounced for progenitor cells in comparison with mature cells. Co-culture of differentiated endothelial cells with progenitor cells showed that there was incorporation of EPCs in vessels formed by HUVECs and HUAECs. Cell migration was assessed using an in vitro wound healing model. The results of these experiments showed an inverse correlation between AA concentrations relative to both cell migration and gap filling capacity. Suppression of NO (nitric oxide) generation appeared to be one of the mechanisms by which AA mediated angiostatic effects. This study supports further investigation into non-cytotoxic antitumor activities of AA.     

Biocompatibility of porcine small intestinal submucosa and rat endothelial progenitor cells in vitro

Objective: This study investigated the biocompatibility of the small intestinal submucosa (SIS) and endothelial progenitor cells (EPCs) by co-cultivating EPCs and SIS in vitro and observing EPC growth on the SIS. Methods: The porcine SIS was prepared and bone marrow mononuclear cells (BMMNCs) were isolated from 3 or 4-week old male SD rats. Cellular morphology was observed by light microscopy and scanning electron microscopy (SEM) and viabilities by the MTT assays. Endothelial progenitor cells (EPCs) were phenotyped by immunocytochemistry, immunofluorescence microscopy and flow cytometry. Vascular lumen formation was evaluated by the Matrigel tube formation assays. EPCs were seeded onto the SIS and production of angiogenin-1 and endothelial cell growth factor (VEGF) by EPCs was examined by ELISA and immunoblotting assays. Results: Light microscopy and SEM showed that the mechanically and chemically treated small intestinal submucosa was composed of cell-free extracellular matrix. Immunohistochemistry, and flow cytometry revealed that the EPCs expressed appropriate surface markers including CD34, CD133, and VEGFR-2. Furthermore, the EPCs formed lumen-like structures and the SIS significantly enhanced the growth of EPCs in vitro. Conclusion: SIS has good biocompatibility with EPCs. SIS pre-seeded with EPCs can be potentially applied as an alternative scaffold material in artificial blood vessel prosthesis.     

Hepatocyte growth factor induces a proangiogenic phenotype and mobilizes endothelial progenitor cells by activating Nox2

Hepatocyte growth factor (HGF) by stimulating the receptor tyrosine kinase c-Met induces angiogenesis and tissue regeneration. HGF has been shown to antagonize the angiotensin II-induced senescence of endothelial progenitor cells (EPCs), which is mediated by NADPH oxidase-dependent reactive oxygen species (ROS) formation. As growth factors, however, usually require ROS for their signaling, we hypothesized that the proangiogenic effects of HGF require NADPH oxidases and focused on the homolog Nox2, which is most abundantly expressed in EPCs and endothelial cells. Indeed, HGF increased the H(2)O(2) formation in EPCs and human umbilical vein endothelial cells (HUVECs), and this effect was not observed in Nox2-deficient cells. HGF induced the mobilization of EPCs and vascular outgrowth from aortic explants in wild-type (WT) but not Nox2(y/-) mice. HGF also stimulated migration and tube formation in HUVECs, and antisense oligonucleotides against Nox2 prevented this effect. To identify the signal transduction underlying these effects, we focused on the kinases Jak2 and Jnk. In HUVECs, HGF increased the phosphorylation of these in a Nox2-dependent manner as demonstrated by antisense oligonucleotides. Also, the HGF-induced Jak2-dependent activation of a STAT3 reporter construct was attenuated after downregulation of Nox2. Accordingly, the HGF-stimulated tube formation of HUVEC was blocked by inhibitors of Jak2 and Jnk. In vivo treatment with the Jnk inhibitor SP600125 blocked the HGF-induced mobilization of EPCs. Ex vivo, SP600125 blocked HGF-induced migration and tube formation. We conclude that HGF-induced mobilization of EPCs and the proangiogenic effects of the growth factor require a Nox2-dependent ROS-mediated activation of Jak2 and Jnk.     

人胚胎主动脉血管内皮祖细胞的分离、培养及鉴定

目的研究人胚胎血管内皮祖细胞(EPCs)分离、扩增及鉴定的方法,并评价其分化成血管内皮细胞的能力,为人胚胎血管来源EPCs作为干细胞技术治疗疾病的细胞材料提供依据。方法从14周龄流产人胚胎主动脉中应用胶原酶消化法分离获得EPCs,用含有碱性成纤维细胞生长因子、表皮生长因子和白血病抑制因子的低血清培养基体外扩增培养EPCs。分离培养的EPCs鉴定采用细胞免疫荧光染色、RT-PCR及流式细胞术,检测EPCs细胞的特异标志CD133、CD34和血管内皮细胞生长因子受体2(VEGFR2)。培养的EPCs应用VEGF进行诱导分化,并评价其分化成为血管内皮细胞的能力。结果分离的人胚胎主动脉EPCs细胞表达EPCs的标志分子CD133、CD34和VEGFR2。EPCs在体外特定低血清培养条件下表现很强的增殖能力。培养的EPCs细胞经过VEGF诱导后,细胞表达CD133明显降低,表达vWF、CD31和ELAM-1增强,并且体外成管能力和摄取Ac-LDL能力增强,表明细胞分化成为血管内皮细胞。结论人胚胎早期主动脉的EPCs具有很好的体外自我更新能力和分化成为血管内皮细胞的潜能,可作为EPCs治疗疾病的细胞材料。     

p53在层流剪切应力下对EPCs分化及功能的影响

目的:研究p53蛋白在层流剪切应力(LSS;12 dyn/cm²)作用下对内皮祖细胞(EPCs)分化及功能的影响,为心血管疾病的防治找到新的靶点。方法:采用密度梯度离心法获得大鼠骨髓单个核细胞,用EGM-2培养基诱导获得EPCs;通过Western blot方法检测p-p53及p53蛋白水平的变化;采用RT-qPCR及流式细胞术分别检测内皮标志物CD31和vWF的mRNA及蛋白表达;使用ELISA检测培养液上清中vWF含量;分别利用Matrigel、Boyden小室、细胞贴壁法、CCK-8法和EdU技术检测EPCs成血管、迁移、黏附和增殖能力。结果:LSS作用促进EPCs中pp53和p53蛋白水平升高;p53抑制剂pifithrin-α处理后,与单纯LSS组相比,EPCs的CD31和vWF表达、黏附、迁移及成血管能力均降低,但其增殖能力升高。结论:LSS是调节EPCs分化及其生物学功能的重要力学因素,而p53蛋白可能在该过程中起关键的调节作用。     

百里醌对血管生成及胰腺癌生长的影响

目的: 探讨百里醌对血管生成和胰腺癌生长的影响及其机制。方法: 采用贴壁选择法培养人脐血内皮祖细胞(EPCs),细胞免疫组化检验细胞中血管内皮生长因子受体-2(VEGFR-2)、VIII因子和 CD34表达,证实细胞属性;观察不同浓度(10 nmol/L、20 nmol/L和40 nmol/L)百里醌对EPCs小管形成的影响;不同浓度(20 μmol/L、40 μmol/L和80 μmol/L)百里醌作用人胰腺癌细胞株PANC-1后,Western blotting检测胰腺癌细胞中血管内皮生长因子(VEGF)表达的变化;建立裸鼠胰腺癌原位移植瘤模型,分成对照组和百里醌组,实验结束后观察百里醌对裸鼠胰腺癌生长的抑制作用;免疫组织化学法检测裸鼠胰腺肿瘤组织中Ki-67、CD34和VEGF的阳性表达。结果: 体外成功培养出人脐血EPCs,百里醌可显著抑制体外EPCs小管形成;并抑制体外人胰腺癌PANC-1细胞中VEGF的表达;与对照组相比较,百里醌可明显抑制荷瘤裸鼠中胰腺肿瘤生长,并下调Ki-67、CD34和VEGF在胰腺肿瘤组织中的阳性表达。结论: 百里醌可抑制体内外血管生长,有望作为治疗胰腺癌的血管抑制药物。     

过氧亚硝基阴离子对实验小鼠骨髓源内皮祖细胞的损伤作用

目的:观察过氧亚硝基阴离子(ONOO-)对实验小鼠骨髓源内皮祖细胞(EPCs)的体外损伤作用。方法:无菌抽取4周龄雄性C57小鼠骨髓,密度梯度离心法分离获取单个核细胞,EGM-2培养基诱导培养EPCs。用免疫荧光染色法进行EPCs表面标志物检测和功能试验鉴定EPCs,培养2～3代后用于实验。通过不同浓度的3-吗啉斯德酮亚胺(SIN-1,CONOO- 供体)作用于EPCs 24h,确定SIN-1的有效作用剂量。将EPCs分为空白对照组、SIN-1组和FeTMPyP(ONOO- 清除剂)干预组,分别用MTT法检测各组EPCs存活率,TUNEL法检测EPCs凋亡,免疫荧光染色法检测细胞内ONOO- 生成印记——3-硝基酪氨酸(NT)表达情况。结果:上述培养细胞表面标记物和功能试验均阳性,是EPCs。800μM SIN-1刺激24h后EPCs存活率较空白对照组明显下降(P<0.05),10μM FeTMPyP干预能抑制这种作用。SIN-1刺激后EPCs凋亡率较空白对照组明显增加(P<0.05),FeTMPyP干预能抑制SIN-1诱导的EPCs凋亡。SIN-1刺激EPCs后细胞内NT表达较空白对照组明显增加(P<0.05),FeTMPyP干预能减少NT的表达。结论:SIN-1能够介导硝化应激,诱导EPCs凋亡,造成EPCs损伤,FeTMPyP干预能抑制SIN-1的上述作用。     

黄芪多糖对2型糖尿病患者外周血内皮祖细胞PI3K/Akt/eNOS信号通路的影响

背景：前期研究证实黄芪通过P38MAPK通路促进内皮祖细胞增殖,其影响是否通过PI3K/Akt/eNOS途径实现？ 目的：观察黄芪多糖对2型糖尿病患者外周血内皮祖细胞蛋白激酶B、内皮型一氧化氮合酶表达的影响。 方法：采用密度梯度离心法获取糖尿病患者外周血单个核细胞,培养7 d后鉴定内皮祖细胞。观察0,50,200,800,3 200,6 400 mg/L黄芪多糖分别干预6,12,24,48 h对内皮祖细胞影响的量效和时效关系;用黄芪多糖及黄芪多糖与PI3K抑制剂LY294002联合干预糖尿病患者内皮祖细胞,Western blot检测磷酸化Akt及磷酸化内皮型一氧化氮合酶的表达水平。以未进行任何处理健康人内皮祖细胞作为对照组。 结果与结论：糖尿病患者内皮祖细胞的增殖能力较对照组明显下降(P  < 0.05)。黄芪多糖显著增加糖尿病患者内皮祖细胞的增殖能力,当黄芪多糖在200~800 mg/L质量浓度范围,干预6~24 h可呈时间及剂量依赖性增强内皮祖细胞的增殖能力(P < 0.01),并呈剂量依赖性升高内皮祖细胞磷酸化Akt及磷酸化内皮型一氧化氮合酶的表达(P < 0.05);PI3K抑制剂LY294002能阻断黄芪多糖诱导的Akt、内皮型一氧化氮合酶的磷酸化(P < 0.05)。说明黄芪多糖通过激活PI3K/Akt/eNOS信号通路促进内皮祖细胞增殖和向内皮细胞的分化。