选择性雌激素受体调节剂及其心血管保护作用

选择性雌激素受体调节剂 (SERMs)是一类在不同靶组织中可表现为雌激素激动剂或拮抗剂作用的化合物。雷洛昔芬是新一代SERMs代表药物。SERMs作用机制涉及雌激素受体、配体、应答元件、雌激素受体相关蛋白等方面。SERMs对于心血管疾病保护作用主要表现在对血脂的调节和对血管壁的直接作用。与激素替代疗法比较 ,SERMs对不同脂蛋白调节作用各异 ;SERMs具有调节血管壁增殖作用 ,其机制可能与eNOS(内皮一氧化氮合酶 )激活、Akt和ERK信号转导途径有关     

雌激素及雌激素受体对高血压心肌肥厚的影响

雌激素通过雌激素受体（estrogen receptor,ER）的介导对心血管系统发挥保护作用。雌激素被动扩散进入细胞后与雌激素受体结合,通过影响肾素血管紧张素系统（RAS）、NO合成、钙离子通道,改善脂质代谢等方面导致高血压心肌肥厚。雌激素受体调节剂（SERMs）如雷洛昔芬等也可以与ER结合,发挥抗心肌肥厚的作用。     

雌激素作用分子机制研究进展

雌激素在体内具有广泛的生物学活性。女性进入绝经期后 ,体内雌激素水平显著下降 ,出现与之相关的绝经期综合症 ,以及心血管疾病如冠心病、动脉粥样硬化和骨质疏松等。使用激素替代疗法能明显减少上述疾病的发生[1 ] 。本文重点在分子机制水平上对雌激素作用研究进展进行综述。     

雌激素对血管平滑肌细胞及内皮细胞功能的调节

在血管平滑肌细胞及内皮细胞上均存在雌激素受体 ,目前认为该受体有α和 β两种类型。雌二醇可直接与血管平滑肌细胞和内皮细胞上的雌激素受体结合 ,使血管内NO生成和释放量增加 ,NO可刺激cGMP增加 ,进而激活蛋白激酶G ,并导致血管平滑肌细胞上大电导Ca2 依赖性K通道开放 ,最后引起血管舒张。     

硫化氢在雌激素促血管内皮细胞增殖中的作用研究

目的研究硫化氢（H2S）在雌激素促进血管内皮细胞增殖中的作用。方法利用培养的脐静脉血管内皮细胞,以雌激素处理后,Western blot检测H2S产生关键酶CSE（胱硫醚-γ-裂解酶）的蛋白表达情况;亚甲基蓝法测定血管内皮细胞释放H2S含量;MTT法检测血管内皮细胞增殖情况。结果不同浓度（1nmo1／L～1μmol／L）的雌激素处理血管内皮细胞48h后．均可促进CSE蛋白表达和内皮H2S释放。不同浓度的雌激素均能明显促进血管内皮细胞增殖。以CSE特异性抑制剂PPG抑制H2S产生后,雌激素促进血管内皮增殖的能力受到显著抑制。结论雌激素可通过促进CSE蛋白表达促进血管内皮细胞释放H2S,进而促进血管内皮细胞增殖。     

大豆异黄酮的选择性雌激素受体调节剂作用

近年来 ,选择性雌激素受体调节剂和大豆异黄酮的研究不断深入 ,逐步证实大豆异黄酮在不同组织可以表现为雌激素激动剂和 (或 )拮抗剂的作用 ,本文将对大豆异黄酮的选择性雌激素受体调节剂的作用、作用机理以及应用和发展前景等进行概述。     

血管内皮细胞生长因子受体KDR胞外配基结合区单抗对内皮细胞增殖及血管形成的抑制

目的　研制能封闭血管内皮生长因子 (VEGF)受体KDR的单克隆抗体 (McAb) ,探讨其抑制VEGF诱导的体内外活性。方法　以原核表达的KDRⅠ～Ⅳ区融合蛋白免疫BALB/c小鼠 ,用杂交瘤技术制备抗KDRⅠ～Ⅳ的McAb ,并用免疫双扩、ELISA和Western免疫印迹鉴定其亚类和抗原结合特异性 ,用VEGF刺激内皮细胞增殖及鸡胚血管形成实验检测该抗体的中和活性。结果　通过筛选和鉴定获得了 2株KDRⅠ～Ⅳ区McAbs(3G9、1E4) ,其分泌抗体亚类分别为IgG1和IgM。 2株McAb均能明显抑制VEGF诱导的内皮细胞 (EC)增殖 ,经纯化的McAb 3G9能明显抑制经VEGF刺激的鸡胚血管形成。结论　KDRⅠ～Ⅳ区McAb可能通过封闭内源性KDR而抑制VEGF活性 ,McAb 3G9具有潜在治疗肿瘤的应用前景。     

选择性雌激素受体调节剂

雌激素替代疗法(estrogen replacement therapy,ERT)是治疗绝经后综合征的首选治疗方案,但是长期应用导致子宫内膜增生、乳腺癌等.选择性雌激素受体调节剂主要通过ER 亚型、共调节子、靶启动子、雌激素受体相关受体等机制实现其组织选择性,在发挥骨骼、心血管保护作用的同时,减少了对乳腺及生殖系统的副作用.目前,选择性雌激素受体调节剂的种类、作用的组织特异性及其临床应用在医学界引起广泛关注,具有广阔的发展前景.     

PES1对雌激素受体转录活性的调节作用

目的 探讨PES1与雌激素受体(ER)的相互作用及其对ER转录活性的影响.方法 将体外翻译的PES1与纯化的GST-ERα和GST-ERβ蛋白分别混合,用GST pull-down验证在体外PES1与ERα和ERβ是否存在相互作用.将HA-PES1与FLAG-ERα或FLAGC-ERβ共转染293T细胞后进行免疫共沉淀,以验证PES1与ER是否在体内有相互作用.用含雌激素受体作用元件的荧光素酶报告基因检测PES1对ERα和ERβ转录活性的影响.结果 PES1与ERα、ERβ在体内外均存在相互作用,而且PES1与ERα的结合比与ERβ的强.在体内,在雌激素(E2)存在下,E2可以增强PES1与ERα的结合,而对PES1与ERβ的结合没有明显影响.PES1对ER转录活性的影响是E2依赖性的,PES1能升高ERα的转录活性而降低ERβ的转录活性(P＜0.01).结论 PES1是一种新的ER共调节因子,能反向调节ERα和ERβ的转录活性,需要进一步研究的是其在ER信号通路以及在E2诱发的肿瘤发生发展中的作用.     

剪切应力对血管内皮细胞的影响

血管内表面覆盖有一层内皮细胞。这层内皮细胞曾被认为是血液成分与血管壁之间的屏障，仅在血管内外的养料、氧和一些大分子物质的交换时起中介作用。六十年代，病理解剖发现动脉粥样硬化常发生于血管分支处与血管弯曲部位，因而激发了人们对血管内皮细胞及其血流动力学环境的研究举。研究表明，血管内皮细胞在体内不断地受到脉动血流的剪切作用，这种血流动力学环境对内皮细胞生物学特性和内皮细胞正常功能至关重要。目前已研究发现     

Concentration-dependent effects of a selective estrogen receptor modulator raloxifene on proliferation and apoptosis in human uterine leiomyoma cells cultured in vitro

BACKGROUND: This study was conducted to elucidate the effects of raloxifeneon proliferation and apoptosis in cultured human uterine leiomyomacells. METHODS: The monolayer cultures were treated with graded concentrations(10^–9, 10^–8 and 10^–7 M) of raloxifeneand 10^–7 M 17-estradiol (E₂). Cell viability, percentageof proliferating cell nuclear antigen (PCNA)-positive cells,percentage of terminal deoxynucleotidyl transferase-mediated2'-deoxyuridine 5'-triphosphate nick-end labelling (TUNEL)-positivecells and the expression of PCNA and Bcl-2 proteins were assessedby 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxylphenyl)-2-(4-sulphophenyl)-2H-tetrazolium assay, immunocytochemistry, TUNEL assay and western blotanalysis, respectively. RESULTS: Compared with untreated cultures, the number of viable culturedcells, percentage of PCNA-positive cells and PCNA protein expressionwere significantly decreased by treatment with 10^–9 Mraloxifene, but increased by treatment with either 10^–8 Mor 10^–7 M raloxifene. In contrast, the percentageof TUNEL-positive cells was significantly increased and Bcl-2protein expression was significantly decreased by treatmentwith 10^–9 M raloxifene, whereas they were not affectedby treatment with either 10^–8 or 10^–7 M raloxifene. CONCLUSIONS: In cultured leiomyoma cells, low concentration (10^–9 M)of raloxifene may inhibit the growth of leiomyoma cells, whereashigh concentrations (10^–8 M, 10^–7 M) ofraloxifene may promote their growth.     

Rationale for using raloxifene to prevent both osteoporosis and breast cancer in postmenopausal women

Both osteoporosis with fracture and breast cancer are important health issues for postmenopausal women. It is well known that estrogen and estrogen receptors (ERs) play an important role in the pathogenesis of both diseases. In past decades, hormone therapy (HT), mainly estrogen plus progestin (EPT), has been frequently used for the purpose of preventing and treating postmenopausal osteoporosis because of its efficacy, but it also contributes to a significant increase in breast cancer. Currently, there is a dilemma regarding the use of estrogen for postmenopausal women. Fortunately, an increasing understanding of the action of estrogen has led ultimately to the design of new drugs that work by virtue of their interaction with the ER; these drugs have come to be known as selective estrogen receptor modulators (SERMs), and are not only effective in preventing osteoporosis and managing those with osteoporosis, but also in decreasing the incidence of breast cancer. Among these SERMs, raloxifene may be the most attractive agent based on the evidence from five recent large trials (Multiple Outcomes of Raloxifene Evaluation [MORE], Continuing Outcomes Relevant to Evista [CORE], Raloxifene Use for the Heart [RUTH], Study of Tamoxifen and Raloxifene [STAR], and Evista Versus Alendronate [EVA]). The former three trials showed that raloxifene not only decreases the incidence of osteoporosis-associated fractures, but also has efficacy in breast cancer prevention. The head-to-head comparison with the anti-fracture agent alendronate (EVA trial) and the chemoprevention agent tamoxifen (STAR trial) further confirmed that raloxifene is a better choice. We concluded that since there is an absence of a therapeutic effect on relieving climacteric symptoms and there is the presence of a potential risk of thromboembolism in the use of raloxifene, this drug can be prescribed for clear indications, such as the management of osteoporosis, the prevention of fracture, and decreasing the incidence of invasive breast cancer, with careful monitoring for thromboembolism. It is reasonable to use raloxifene as an appropriate medicine that targets climacteric symptom-free postmenopausal women because of its overall favorable risk-benefit safety profile using the global index proposed by the Women's Health Initiation (WHI).     

EMAS clinical guide: selective estrogen receptor modulators for postmenopausal osteoporosis

Osteoporosis and the resulting fractures are major public health issues as the world population is ageing. Various therapies such as bisphosphonates, strontium ranelate and more recently denosumab are available. This clinical guide provides the evidence for the clinical use of selective estrogen modulators (SERMs) in the management of osteoporosis in postmenopausal women.     

蜂胶对脂多糖诱导的血管内皮细胞中PC-PLC活性和TLR4表达的影响

目的: 探讨在脂多糖(LPS)诱导的条件下中国蜂胶对血管内皮细胞(VECs)磷脂酰胆碱特异性磷脂酶C (PC-PLC)活性和TLR4表达的影响。方法: 将100 μg/L LPS 加入到含0.5%血清的培养液中培养VECs,经12.5 mg/L的中国蜂胶分别处理12 h和24 h后,SRB法测定细胞存活率;化学法测定NO含量;以L-α-卵磷脂为底物测定PC-PLC的活性;Western blotting检测TLR4、核因子κB p65(NF-κB p65)和p53的表达;细胞内活性氧(ROS)和线粒体膜电位分别通过荧光探针DCHF和JC-1检测。结果: 中国蜂胶处理LPS诱导的血管内皮细胞24 h并不影响细胞存活率,但降低NO的含量和ROS的水平;处理12 h后降低PC-PLC活性和NF-κB p65表达;处理12 h和24 h后降低TLR4和p53的表达。此外,中国蜂胶不影响细胞内线粒体膜电位的水平。结论: 中国蜂胶通过降低PC-PLC活性和TLR4表达,抑制其下游信号分子NF-κB p65、p53的表达和ROS的水平,以及抑制NO的释放,从而发挥抗炎功效。     

阿托伐他汀钙对高血压大鼠血管内皮细胞的保护作用

目的:观察高血压状态下阿托伐他汀钙对SD大鼠胸主动脉血管内皮细胞的保护作用。方法:构建两肾一夹SD大鼠肾性高血压模型。研究动物随机分为高血压组、他汀治疗组和假手术组,每组8只,术后第4周他汀治疗组给予阿托伐他汀钙20mg·kg-1.d-1灌胃8周。术后第4、12周测血压、血脂,第12周通过扫描电镜方法观察胸主动脉内皮细胞层的完整性,检测血液中循环内皮细胞数量,并测定循环内皮祖细胞(CEPCs)的数量及增殖能力、黏附能力及凋亡情况。结果:他汀治疗组大鼠胸主动脉内皮细胞受损轻于高血压组,但重于假手术组,高血压组、他汀治疗组和假手术组大鼠循环内皮细胞数量(5.9×106、3.9×106、2.0×106)与CEPCs凋亡率(22.1%±2.1%、13.4%±1.6%、7.4%±1.3%)依次降低(均P0.01);而CEPCs数量(21.63±2.33、40.38±6.00、65.38±2.97)、CEPCs增殖能力(0.13±0.01、0.17±0.01、0.29±0.03)与CEPCs黏附能力(12.25±2.49、21.50±2.20、28.88±2.85)依次增高(均P0.01)。结论:(1)高血压状态可导致大鼠血管内皮细胞严重受损,CEPCs数量增加,而CEPCs数量与功能降低,凋亡率增高,继而引起CEPCs对血管内皮细胞的修复能力降低。(2)阿托伐他汀钙对血管内皮细胞具有直接保护作用,并可能通过提高CEPCs数量及功能、降低CEPCs凋亡、增强CEPCs对血管内皮细胞的修复能力而发挥作用。     

Estrogen receptor modulators and estrogen receptor beta immunolabelling in human umbilical vein endothelial cells

Human umbilical vein endothelial cells (HUVEC) exposed to the female sex hormone estradiol show different kinds of effects including increased elasticity, activation of plasma membrane Na⁺/H⁺ exchange, prostacyclin production, prevention of apoptosis and many others. The aim of this study was the systematic analysis of the immunolabelling of estrogen receptors (ERs), ERα and ERβ, in HUVEC after stimulation with different commercially available ER modulators and ER agonists or antagonists. HUVEC response to these substances was shown to be regulated via ERβ. ERα immunolabelling or up-regulation was abrogated after application of estrogen derivatives, selective estrogen receptor modulators (SERM) and ER agonists or antagonists. Immunolabelling of ERβ was significantly increased by estradiol, estrone, ethinylestradiol and tumour necrosis factor alpha (TNFα). SERM, such as Tamoxifen, and pure antagonists, such as ICI 182.780, stimulated ERβ in HUVEC at low concentrations, whereas higher concentrations inhibited ERβ immunolabelling. The pure estrogen receptor agonist 2,3-bis (4-hydroxyphenyl) proprionitrile (DPN) exhibited its activating potential at low concentrations. In contrast, higher concentrations resulted in a down-regulation of ERβ. Estrogenic effects in HUVEC, independent of stimulation or inhibition, are mediated via the ERβ. SERM such as Tamoxifen and ER antagonists such as ICI 182.780 act as ER activators in low concentrations, whereas higher concentrations lead to inhibitory effects.     

Toll样受体4介导的自噬减轻糖基化高密度脂蛋白诱导的血管内皮细胞凋亡

目的:探讨自噬对糖基化高密度脂蛋白(glycosylated high-density lipoprotein,gly-HDL)所致的血管内皮细胞凋亡的影响及其分子机制。方法:体外培养人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs),分别与100 mg/L HDL和不同浓度(25、50和100 mg/L)gly-HDL共同孵育24 h;另再培养HUVECs给予1μmol/L自噬诱导剂雷帕霉素或2 mmol/L自噬抑制剂3-甲基腺嘌呤(3-methyladenine,3-MA)预处理1 h,或5 mg/L抗Toll样受体4(Toll-like receptor 4,TLR4)单克隆中和抗体预处理30 min,再与gly-HDL(100 mg/L)共同孵育24 h。采用MTT法检测细胞活力,Annexin V-FITC/PI双染法检测细胞凋亡情况,试剂盒测定培养液中乳酸脱氢酶(lactate dehydrogenase,LDH)活性,采用Western blot技术检测自噬标志分子beclin-1和微管相关蛋白1轻链3-Ⅱ(microtubule-associated protein 1 light chain 3-Ⅱ,LC3-Ⅱ)、内质网应激凋亡途径关键分子caspase-12及TLR4的表达变化,采用激光共聚焦显微镜观测细胞内LC3的变化。结果:经gly-HDL处理的HUVECs活力下降,LDH漏出和细胞凋亡显著增加(P<0.01),且caspase-12被激活(P<0.05);雷帕霉素预处理HUVECs后,gly-HDL对细胞的损伤作用和对caspase-12的活化作用减弱(P<0.05);而3-MA预处理HUVECs后,gly-HDL对细胞的损伤作用和对caspase-12的活化作用则进一步加强(P<0.05)。gly-HDL显著上调TLR4的表达,并触发自噬反应,表现为beclin-1和LC3-Ⅱ表达上调及LC3显著颗粒化,且呈浓度依赖性(P<0.05);而抗TLR4单克隆中和抗体预处理可显著抑制gly-HDL所诱导的beclin-1上调和LC3颗粒化(P<0.01)。结论:TLR4介导gly-HDL对HUVECs自噬的诱导作用,而一定程度的自噬可通过抑制caspase-12活化减轻gly-HDL所诱导的HUVECs凋亡。     

Progesterone receptor modulator CDB-2914 down-regulates vascular endothelial growth factor, adrenomedullin and their receptors and modulates progesterone receptor content in cultured human uterine leiomyoma cells

BACKGROUND: This study was conducted to evaluate the effects of graded concentrations (10(-8), 10(-7) and 10(-6) M) of progesterone receptor (PR) modulator CDB-2914 on the protein contents of PR, of vascular endothelial growth factor (VEGF), adrenomedullin (ADM) and their receptors in cultured human uterine leiomyoma and matching myometrial cells. METHODS: PR-A, PR-B, VEGF-A, VEGF-B, VEGF receptor (VEGFR)-1, VEGFR-2, ADM and ADM receptor (ADMR) contents were assessed by Western blot analysis. RESULTS: Treatment with 100 ng/ml progesterone increased VEGF-A, VEGF-B and ADM contents in cultured leiomyoma cells and normal myometrial cells. The concomitant treatment with 10(-6) M CDB-2914 significantly decreased the progesterone-induced VEGF-A, VEGF-B and ADM contents in cultured leiomyoma cells but not in normal myometrial cells. CDB-2914 treatment alone decreased VEGFR-1, VEGFR-2 and ADMR contents in cultured leiomyoma cells but not in normal myometrial cells. CDB-2914 treatment increased PR-A and decreased PR-B contents in cultured leiomyoma cells in a dose-dependent manner compared with untreated cultures, whereas no significant changes in PR isoform contents were observed in normal myometrial cells. CONCLUSIONS: These results suggest that CDB-2914 down-regulates VEGF, ADM and their receptor contents and modulates PR isoform contents in cultured leiomyoma cells in a cell-type-specific manner.     

Effects of vascular endothelial growth factor released from cultured dermal substitute on proliferation of vascular endothelial cells in vitro

Allogeneic cultured dermal substitute (CDS) was prepared by culturing fibroblasts on a two-layered spongy matrix of hyaluronic acid and atelocollagen. CDS can be cryopreserved and transported to other hospitals in a frozen state. The present study was designed to analyze the amounts of vascular endothelial growth factor (VEGF) released from fibroblasts in fresh and cryopreserved CDS and to investigate the effects of this VEGF on proliferation of vascular endothelial cells in vitro. The culture medium used in preparing CDS (fresh CDS culture medium sample) was collected and stored at –30°C for the quantitative analysis of VEGF. After thawing cryopreserved CDS, it was recultured in a culture medium for 1 week. The culture medium used was collected and stored at –30°C for quantitative analysis of VEGF. The amounts of VEGF released from the fresh and cryopreserved CDS into the culture medium were about 610pg/ml and 640pg/ml, respectively. This finding suggests that the cryopreserved CDS retains its ability to release VEGF. Immunohistological analysis indicated that some of the VEGF adhered to the matrix. Human vascular endothelial cells were cultured in medium mixed with the fresh or cryopreserved CDS culture medium sample. Proliferation of vascular endothelial cells was enhanced by increasing the concentration of both CDS culture medium samples. When antihuman VEGF antibody was added to the culture medium, the proliferative activity of vascular endothelial cells was reduced. These findings confirm that VEGF released from CDS promotes proliferation of vascular endothelial cells.