沙利度胺通过诱导G1阻滞和凋亡抑制血管内皮细胞增殖

沙利度胺是一种抗血管生成药物,临床上用于治疗多种肿瘤,但其抗肿瘤血管生成机制尚不十分清楚. 本文采用MTT法观察沙利度胺对体外培养的血管内皮细胞增殖的影响. 结果发现,沙利度胺能够抑制血管内皮细胞的增殖,其IC50为16.47 μg/mL;然后采用Hoechst染色和流式细胞仪检测细胞凋亡和细胞周期,发现沙利度胺能够诱导内皮细胞凋亡,并干扰细胞的周期,出现G0/G1期阻滞. 最后,通过Western印迹方法分析沙利度胺对血管内皮细胞Bcl-2蛋白表达的影响,发现抗凋亡的Bcl-2蛋白表达水平随沙利度胺浓度增大而降低. 初步结果提示,沙利度胺可能通过阻遏抗凋亡分子Bcl-2表达,激活诱导G1期阻滞的信号通路而抑制内皮细胞新生,从而抑制肿瘤生长. 诱导内皮细胞凋亡及G1期阻滞的具体分子机制正在研究中.     

内皮细胞增殖抑制因子研究进展

内皮细胞过度增殖引起的病理性血管生成是肿瘤、类风湿性关节炎等发病的关键环节。内皮细胞增殖由血管内皮细胞生长因子等促血管生成因子提供促增殖信号,而新近发现的多种内皮增殖抑制因子,如血管内皮抑素、血管抑素、血小板反应蛋白-1、微囊蛋白1、某些microRNAs和某些抑癌基因等,则通过抑制促增殖信号、调节细胞周期、诱导细胞凋亡等途径下调内皮细胞的增殖及血管生成。内皮增殖抑制因子可望成为病理性血管生成防治的新靶点。     

血管能抑素,一种新的血管生成抑制因子

血管能抑素(canstatin)是新近发现的血管生成抑制因子,在体外能抑制血管内皮增殖与迁移,并诱导其凋亡,在体内可抑制肿瘤生长。血管能抑素的N端与C端均有抗血管生成和抑制内皮细胞增殖的作用．而N末端诱导内皮细胞的凋亡活性明显高于C末端。血管能抑素能抑制Akt、FAK的磷酸化。血管能抑素诱导的细胞凋亡与PI3K／Akt信号传导通路的抑制作用有关。     

人肿瘤抑素(Tumstatin)在E.coli中的克隆、表达及活性分析

从人胚肾2 93细胞中扩增肿瘤抑素(tumstatin)基因,进行原核表达,纯化和生物活性检测.利用原核表达载体pMAL c2在大肠杆菌BL2 1中表达肿瘤抑素,经AmyloseResin亲和层析柱和QSepharoseFastFlow柱纯化,通过体外内皮细胞增殖、内皮细胞凋亡和鸡尿囊绒膜新生血管生成试验检测其抑制活性.MBP tumstatin在BL2 1中表达率约2 0 % ,肿瘤抑素纯度可达95 % .肿瘤抑素可明显抑制内皮细胞增殖(IC50 约为15 μg ml)、诱导内皮细胞凋亡和抑制鸡尿囊绒膜新生血管生成.研究结果表明,肿瘤抑素对内皮细胞具有明显的抑制作用,提示其在肿瘤治疗中有潜在的应用前景.     

人血管生成抑制素抑瘤研究进展

肿瘤的发展和转移需要新生血管的形成。人血管生成抑制素是近年发现的能够专一性抑制血管内皮细胞的内皮抑制因子。大量研究表明,在体外用血管生成抑制素处理血管内皮细胞可以抑制新生血管的形成,在体内单独使用血管生成抑制素,或者将血管生成抑制素与其他物质如基质金属蛋白酶、尿激酶联合处理荷瘤小鼠,可以降低小鼠体内肿瘤组织新生血管密度,抑制肿瘤的生长和肿瘤细胞的迁移。简要综述了血管生成抑制素抑制肿瘤生长和转移及其作用机理。     

内皮抑制素的研究进展

肿瘤生长依赖新生血管的形成,通过抑制血管内皮细胞,切断肿瘤生长所需的营养途径,阻断新生血管形成,从而达到抑制肿瘤生长的目的,Endostatin（内皮抑制素）具有抑制肿瘤血管生成和肿瘤转移最强烈,最特异的生物学活性,且不产生耐药,是目前发现最为理想的血管生成抑制因子。     

原核表达Arresten蛋白抑制血管生成作用

目的:研究原核表达的Arresten蛋白纯化品对血管内皮细胞及血管生成的抑制作用。方法:MTT法检测Arresten蛋白对人脐静脉内皮细胞(HUVEC)增殖的影响;流式细胞仪分析Arresten蛋白作用下HUVEC凋亡的情况;细胞迁移实验观察Arresten蛋白对HUVEC迁移能力的影响;鸡胚绒毛尿囊膜(CAM)实验观察Arresten蛋白对新生血管的抑制情况。结果:原核表达的Arresten蛋白纯化品能特异性地抑制 HUVEC的增殖、迁移,诱导HUVEC的凋亡,并在一定范围内呈现出剂量—效应关系。Arresten蛋白能有效抑制鸡胚尿囊膜血管的生长(P<0.01)。结论:原核表达的Arresten蛋白纯化品对内皮细胞有特异的抑制作用,能有效抑制血管生成。     

一氧化氮和动脉粥样硬化

动脉粥样硬化是脂蛋白、单核细胞、巨噬细胞、T淋巴细胞与血管壁内皮细胞相互作用而导致的慢性炎症反应。这个炎症的过程由脂质浸润开始,涉及氧化应激反应,最终导致复杂的病理损伤和斑块的形成,斑块突出入血管,破裂形成血栓而导致急性的心肌梗塞或中风。激活内皮源性的一氧化氮合成酶而生成的一氧化氮（NO）能够预防动脉粥样硬化,并对不周发展阶段的动脉粥样硬化的病理形成均有改善和逆转作用。其生成的NO能抗氧化、清除自由基、抑制低密度脂蛋自在血管壁被氧化,防止氧化低密度脂蛋白（oxLDL）的产生,而影响脂质浸润;能抑制NFKB的激活和核内迁移,阻抑激活的内皮细胞表达黏附分子,减少嗜中性粒细胞和单核细胞的黏附和活化,减少血管壁的炎症反应;能抑制血小板黏附、聚集,抑制凝血酶诱导的血小板活性因子的表达以减少血栓形成;能阻止凋亡,保持内皮细胞的完整性;还能有效地抑制血管平滑肌细胞增殖、迁移和细胞外基质的合成,对动脉粥样硬化病理形成和发展具有阻抑作用。     

内皮抑素在新生血管形成相关疾病中的作用机制研究进展

新血管形成是许多生理、病理过程的关键步骤,受血管形成促进因子和抑制因子的调节。内皮抑素是最重要的血管形成抑制因子之一,可在体外抑制血管内皮细胞的增殖、迁移和血管化,在动物模型中抑制新血管形成,对新生血管形成相关疾病,特别是肿瘤有治疗作用。关于内皮抑素抑制新血管形成的分子机制尚无定论,已有线索表明,它可通过与VEGF、MMP-2、整合素以及VEGF受体KDR等相互作用,从而抑制内皮细胞增殖、迁移或通过多种途径促进内皮细胞凋亡。本就内皮抑素作用的分子机制,及其作用于新血管形成相关疾病的最新研究成果进行综述。     

缺氧条件下血管活性因子对血管内皮细胞中VEGF表达的影响

 探讨在缺氧条件下人脐静脉血管内皮细胞对血管内皮生长因子 (vascular endothelialgrowth factor,VEGF)表达及缩血管活性物质内皮素 (ET)、舒血管活性物质一氧化氮 (NO)和 NO抑制剂 LNNA对 VEGF基因表达的影响 .体外培养人脐静脉血管内皮细胞 ,经缺氧及血管活性物质处理 .Northern杂交、酶联免疫检测和计算机图象分析等观察 VEGF m RNA和蛋白表达水平 .发现缺氧 6h内皮细胞可见 VEGF表达 .ET可促进 VEGF m RNA的表达 ,NO可明显抑制 VEGFm RNA的表达 ,NO抑制剂 LNNA也影响 VEGF m RNA的表达 .ELISA检测 VEGF蛋白水平分别为 6h组 8.2± 1 .1 ng/ L,ET+6h组 9.37± 1 .0 2 ng/ L,NO+6h组 2 .86± 0 .91 ng/ L,L - NNA+6h组 1 4.75± 1 .87ng/ L.缺氧可诱导人脐静脉血管内皮细胞分泌 VEGF并受血管活性物质ET和 NO的调控 ,ET促进其表达 ,NO抑制其表达 .     

Antibiotic bedaquiline effectively targets growth,survival and tumor angiogenesis of lung cancer through suppressing energy metabolism

Tumor angiogenesis plays essential roles during lung cancer progression and metastasis. Therapeutic agent that targets both tumor cell and vascular endothelial cell may achieve additional anti-tumor efficacy. We demonstrate that bedaquiline, a FDA-approved antibiotic drug, effectively targets lung cancer cells and angiogenesis. Bedaquiline dose-dependently inhibits proliferation and induces apoptosis of a panel of lung cancer cell lines regardless of subtypes and molecular heterogeneity. Bedaquiline also inhibits capillary network formation of human lung tumor associated-endothelial cell (HLT-EC) on Matrigel and its multiple functions, such as spreading, proliferation and apoptosis, even in the presence of vascular endothelial growth factor (VEGF). We further demonstrate that bedaquiline acts on lung cancer cells and HLT-EC via inhibiting mitochondrial respiration and glycolysis, leading to ATP reduction and oxidative stress. Consistently, oxidative damage on DNA, protein and lipid were detected in cells exposed to bedaquiline. Importantly, the results obtained in in vitro cell culture are reproducible in in vivo xenograft lung cancer mouse model, confirming that bedaquiline suppresses lug tumor growth and angiogenesis, and increases oxidative stress. Our findings demonstrating that energy depletion is effectively against lung tumor cells and angiogenesis. Our work also provide pre-clinical evidence to repurpose antibiotic bedaquiline for lung cancer treatment.     

Vascular superoxide and hydrogen peroxide production and oxidative stress resistance in two closely related rodent species with disparate longevity

Vascular aging is characterized by increased oxidative stress, impaired nitric oxide (NO) bioavailability and enhanced apoptotic cell death. The oxidative stress hypothesis of aging predicts that vascular cells of long-lived species exhibit lower production of reactive oxygen species (ROS) and/or superior resistance to oxidative stress. We tested this hypothesis using two taxonomically related rodents, the white-footed mouse (Peromyscus leucopus) and the house mouse (Mus musculus), that show a more than twofold difference in maximum lifespan potential (MLSP = 8 and 3.5 years, respectively). We compared interspecies differences in endothelial superoxide (O2-) and hydrogen peroxide (H2O2) production, NAD(P)H oxidase activity, mitochondrial ROS generation, expression of pro- and antioxidant enzymes, NO production, and resistance to oxidative stress-induced apoptosis. In aortas of P. leucopus, NAD(P)H oxidase expression and activity, endothelial and H2O2 production, and ROS generation by mitochondria were less than in mouse vessels. In P. leucopus, there was a more abundant expression of catalase, glutathione peroxidase 1 and hemeoxygenase-1, whereas expression of Cu/Zn-SOD and Mn-SOD was similar in both species. NO production and endothelial nitric oxide synthase expression was greater in P. leucopus. In mouse aortas, treatment with oxidized low-density lipoprotein (oxLDL) elicited substantial oxidative stress, endothelial dysfunction and endothelial apoptosis (assessed by TUNEL assay, DNA fragmentation and caspase 3 activity assays). According to our prediction, vessels of P. leucopus were more resistant to the proapoptotic effects of oxidative stressors (oxLDL and H2O2). Primary fibroblasts from P. leucopus also exhibited less H2O2-induced DNA damage (comet assay) than mouse cells. Thus, increased lifespan potential in P. leucopus is associated with a decreased cellular ROS generation and increased oxidative stress resistance, which accords with the prediction of the oxidative stress hypothesis of aging.     

Involvement of Kv1.5 Protein in Oxidative Vascular Endothelial Cell Injury

Endothelial injury related to oxidative stress is a key event in cardiovascular diseases, such as hypertension and atherosclerosis. The activation of the redox-sensitive Kv1.5 potassium channel mediates mitochondrial reactive oxygen species (ROS)-induced apoptosis in vascular smooth muscle cells and some cancer cells. Kv1.5 channel is therefore taken as a new potential therapeutic target for pulmonary hypertension and cancers. Although Kv1.5 is abundantly expressed in vascular endothelium, there is little knowledge of its role in endothelial injury related to oxidative stress. We found that DPO-1, a specific inhibitor of Kv1.5, attenuated H₂O₂-evoked endothelial cell apoptosis in an in vivo rat carotid arterial model. In human umbilical vein endothelial cells (HUVECs) and human pulmonary arterial endothelial cells (HPAECs), angiotensin II and oxLDL time- or concentration-dependently enhanced Kv1.5 protein expression in parallel with the production of intracellular ROS and endothelial cell injury. Moreover, siRNA-mediated knockdown of Kv1.5 attenuated, whereas adenovirus-mediated Kv1.5 cDNA overexpression enhanced oxLDL–induced cellular damage, NADPH oxidase and mitochondria-derived ROS production and restored the decrease in protein expression of mitochondria uncoupling protein 2 (UCP2). Collectively, these data suggest that Kv1.5 may play an important role in oxidative vascular endothelial injury.     

Forkhead‐box R2 promotes metastasis and growth by stimulating angiogenesis and activating hedgehog signaling pathway in ovarian cancer

Overexpression of forkhead‐box R2 (FoxR2) is related to metastasis and progression of tumor. However, its biological functions in ovarian cancer (OC) progression remain unclear. Herein, we aimed to explore the changes in biological functions and molecular events related to FoxR2 overexpression. We found that FoxR2 is upregulated frequently in OC where these events are associated with worse histologic grade and poor survival. Enhanced expression of FoxR2 was related to cell growth, migration, and epithelial‐mesenchymal transition, whereas silencing of FoxR2 suppressed these malignant phenotypes. In addition, angiogenesis was stimulated by FoxR2 overexpression by enhancing vascular endothelial growth factor expression. Further mechanistic investigations revealed that the increase in cell surface FoxR2 promoted sonic hedgehog binding and signaling. Inhibiting hedgehog pathway with sonidegib decreased FoxR2‐induced migration and lung metastasis of OC cells, establishing the critical role of hedgehog signaling in mediating the effects of FoxR2 expression. Taken together, our results indicate that FoxR2 overexpression in OC contributes to malignant behavior in cancer cells, at least in part through stimulating angiogenesis and activation of the hedgehog signaling pathway. Hedgehog signaling pathway activation may be the key in tumor progression mediated by FoxR2.     

Suppression of tumorigenesis in mitochondrial NADP+-dependent isocitrate dehydrogenase knock-out mice

The tumor host microenvironment is increasingly viewed as an important contributor to tumor growth and suppression. Cellular oxidative stress resulting from high levels of reactive oxygen species (ROS) contributes to various processes involved in the development and progress of malignant tumors including carcinogenesis, aberrant growth, metastasis, and angiogenesis. In this regard, the stroma induces oxidative stress in adjacent tumor cells, and this in turn causes several changes in tumor cells including modulation of the redox status, inhibition of cell proliferation, and induction of apoptotic or necrotic cell death. Because the levels of ROS are determined by a balance between ROS generation and ROS detoxification, disruption of this system will result in increased or decreased ROS level. Recently, we demonstrated that the control of mitochondrial redox balance and cellular defense against oxidative damage is one of the primary functions of mitochondrial NADP⁺-dependent isocitrate dehydrogenase (IDH2) that supplies NADPH for antioxidant systems. To explore the interactions between tumor cells and the host, we evaluated tumorigenesis between IDH2-deficient (knock-out) and wild-type mice in which B16F10 melanoma cells had been implanted. Suppression of B16F10 cell tumorigenesis was reproducibly observed in the IDH2-deficient mice along with significant elevation of oxidative stress in both the tumor and the stroma. In addition, the expression of angiogenesis markers was significantly down-regulated in both the tumor and the stroma of the IDH2-deficient mice. These results support the hypothesis that redox status-associated changes in the host environment of tumor-bearing mice may contribute to cancer progression.     

IL-8 directly enhanced endothelial cell survival,proliferation, and matrix metalloproteinases production and regulated angiogenesis

IL-8, a member of the chemokine family, has been shown to play an important role in tumor growth, angiogenesis, and metastasis. The objective of this study was to determine the mechanism of IL-8-mediated angiogenesis. We examined the direct role of IL-8 in angiogenesis by examining IL-8 receptor expression on endothelial cells and their proliferation, survival, and matrix metalloproteinases (MMPs) production. We demonstrate that HUVEC and human dermal microvascular endothelial cells constitutively express CXCR1 and CXCR2 mRNA and protein. Recombinant human IL-8 induced endothelial cell proliferation and capillary tube organization while neutralization of IL-8 by anti-IL-8 Ab blocks IL-8-mediated capillary tube organization. Incubation of endothelial cells with IL-8 inhibited endothelial cell apoptosis and enhanced antiapoptotic gene expression. Endothelial cells incubated with IL-8 had higher levels of Bcl-x(L):Bcl-x(S) and Bcl-2:Bax ratios. Furthermore, incubation of endothelial cells with IL-8 up-regulated MMP-2 and MMP-9 production and mRNA expression. Our data suggest that IL-8 directly enhanced endothelial cell proliferation, survival, and MMP expression in CXCR1- and CXCR2-expressing endothelial cells and regulated angiogenesis.