Genistein对bFGF诱导的EJ细胞增殖的抑制作用

目的:观察碱性成纤维细胞生长因子(bFGF)对人膀胱癌细胞株EJ细胞的增殖作用;以及酪氨酸蛋白激酶(TPK)抑制剂Genistein对bFGF诱导的EJ细胞增殖的抑制作用。方法:以不同浓度的bFGF刺激EJ细胞,再对由bFGF引起的细胞增殖施加不同浓度的Genistein;以MTT法检测细胞的增殖程度,原位凋亡细胞检测(TUNEL)和流式细胞仪(FCM)检测Genistein对EJ细胞凋亡和细胞周期的影响。结果:bFGF可以使EJ细胞增殖比明显上升,Genistein可引起细胞增殖比明显下降,其程度均随浓度增高而增强;Genistein可以促进EJ细胞的凋亡,使细胞阻滞于G2/M期。结论:bFGF可以诱导EJ细胞的增殖,Genistein可以诱导EJ细胞凋亡,对bFGF诱导的细胞增殖有抑制作用,可能为膀胱肿瘤的治疗提供新的方法。     

大豆异黄酮苷元滴丸的制备和含量测定

目的:研究大豆异黄酮苷元滴丸的处方、制备工艺和含量测定方法.方法:用单因素实验法确定制备工艺中的各种条件;用正交设计法优选处方组成;用HPLC法测定异黄酮苷元的含量.结果:最佳工艺条件为:冷凝剂二甲基硅油温度10℃,药液温度80℃,滴距6 cm,滴速15滴/min.染料木素在3.6～36%、6.00%和0.89%.结论:本制备工艺稳定性好、成型率高,建立的含量测定方法可控性强、专属性高.     

Genistein-磁性纳米微粒对人肝癌细胞中FAK表达的影响

目的 观察不同浓度genistein-磁性纳米微粒对肝癌细胞株HepG2生长及FAK表达的影响.方法 四甲基偶氮唑盐(MTT)法测定genistein-磁性纳米微粒作用的肝癌细胞株HepG2活性,RT-PCR及免疫组化法分别检测FAK mRNA及FAK蛋白的表达.结果 HepG2细胞经genistein一磁性纳米微粒(浓度范围10～80mg/L)作用后细胞生长明显受到抑制,具有剂量依赖效应关系和时间依赖效应关系(P<0.05).RT-PCR检测不同浓度genistein-磁性纳米微粒作用48h后FAKmRNA表达的相对水平分别为1.242±0.031,1.213±0.443,0.834±0.044,0.652±0.039,0.446±0.041,对照组为1.443±0.053(F=21.97,P<0.05),FAK蛋白表达明显减弱,也有明显的剂量效应关系.结论 genistein-磁性纳米微粒能够降低肝癌细胞FAK-mRNA及蛋白的表达,这作用可能是其抑制肝癌细胞的生长的重要机制之一.     

Changes of growth hormone-releasing hormone and somatostatin neurons in the rat hypothalamus induced by genistein: a stereological study

Objectives: Genistein is a plant-derived estrogenic isoflavone commonly found in dietary and therapeutic supplements, due to its potential health benefits. Growth hormone-releasing hormone (GHRH) and somatostatin (SS) are neurosecretory peptides synthesized in neurons of the hypothalamus and regulate the growth hormone secretion. Early reports indicate that estrogens have highly involved in the regulation of GHRH and SS secretions. Since little is known about the potential effects of genistein on GHRH and SS neurons, we exposed rats to genistein.

Methods: Genistein were administered to adult rats in dose of 30 mg/kg, for 3 weeks. The estradiol-dipropionate treatment was used as the adequate controls to genistein. Using applied stereology on histological sections of hypothalamus, we obtained the quantitative information on arcuate (Arc) and periventricular (Pe) nucleus volume and volume density of GHRH neurons and SS neurons. Image analyses were used to obtain GHRH and SS contents in the median eminence (ME).

Results: Administration of estradiol-dipropionate caused the increase of Arc and Pe nucleus volume, SS neuron volume density, GHRH and SS staining intensity in the ME, when compared with control. Genistein treatment increased: Arc nucleus volume and the volume density of GHRH neurons (by 26%) and SS neurons (1.5 fold), accompanied by higher GHRH and SS staining intensity in the ME, when compared to the orhidectomized group.

Discussion: These results suggest that genistein has a significant effect on hypothalamic region, involved in the regulation of somatotropic system function, and could contribute to the understanding of genistein as substance that alter the hormonal balance.     

Genistein抗高血压机制的研究进展

近年来,Genistein(金雀异黄素/染料木素)的血管活性作用越来越受到的重视。目前研究表明,Genistein可通过内皮依赖性和内皮非依赖性血管舒缩系统调节血管平滑肌舒缩,这些影响涉及基因组和非基因组机制。Genistein还可与雌激素受体相互作用,通过多信号转导通路发挥类雌激素作用。另外,Genistein可作用于肾脏,增加肾血流量及尿钠排泄。Genistein也可针对体液调节系统(如肾素-血管紧张素系统)发挥作用。动物研究数据显示,以上作用相互协同可以达到降压的效果。     

高效液相色谱法测定食品中大豆异黄酮

目的：建立高效液相色谱法测定食品中大豆异黄酮（包括染料木黄酮和大豆苷元）的含量，探讨样品的提取方法及测定影响因素。方法：样品经0.50mol/L盐酸回液和乙醇提取大豆异黄酮后，采用甲醇－0.01mol/L乙酸铵溶液（pH4.5)(60 40)作为流动相分离待测物质，等度洗脱，采用紫外检测器于254nm波长下测定。结果：染料木黄酮的检出限为0．04μg/ml，大豆苷元为0．05μg/ml，样品的加标回收率为91．4％－113．1％，日内和日间相对标准偏差分别为1．1％－2．8％和2．9％－5．1％。结论：本法简便、快速，易于推广普及，为大豆及其制品和保健食品中大豆异黄酮的检测和膳食摄取植物性雌激素提供了分析方法。     

Mechanisms of serotonin-induced Ca2+ responses in mesangial cells

Smooth muscle cell-like mesangial cells play an important role in the regulation of glomerular blood flow and are involved in renal inflammatory reactions, thereby interacting with circulating cells. The platelet products serotonin (5-HT) and ATP induce similar, e.g. mitogenic, effects in mesangial cells, but differentially activate and induce inflammation-related genes. To get an insight into intracellular signaling steps, a very early step in the signaling cascade, the biphasic Ca²⁺ signal elicited by 5-HT and ATP in rat mesangial cells was investigated. Both phases of the Ca²⁺ signal, release from internal stores as well as influx of extracellular Ca²⁺, were dependent on phospholipase C activation as shown by the specific inhibitor U73122 (complete inhibition at 10μM U73122). There was no evidence for voltage-gated L-type channels in these cells, suggesting that Ca²⁺ influx was mediated by Ca²⁺ release-activated channels. The L-type channel blocker verapamil, however, dose-dependently (0.1–10μM) and specifically inhibited 5-HT-elicited Ca²⁺ signals by interference with binding of 5-HT to 5-HT_2A receptors. 5-HT-mediated Ca²⁺ release was reduced by 80% when protein kinase C was activated by the phorbolester TPA (0.1μM). Interaction of 5-HT_2A receptors with phospholipase C was also inhibited by genistein (30% at 5μM; 100% at 50μM), an inhibitor of tyrosine kinases. Binding of 5-HT to its receptor reduced subsequent ATP-mediated Ca²⁺ signaling. The cross talk between the receptors was sensitive to genistein. ATP-mediated Ca²⁺ signaling was attributed to different types of P_2y receptors and/or multiple G-proteins coupled, because the signal was partially inhibited by pertussis toxin (50%). In accordance, modulation of the ATP-mediated signaling by phosphorylation was less tightly controlled than 5-HT-mediated Ca²⁺ release. These data indicate that although the Ca²⁺ responses elicited by the two stimuli are comparable, interactions between receptors, G-proteins and target enzymes are regulated differentially. Received: 6 January 1997 / Accepted: 9 May 1997     

Potential role of Ca++ on the differentiation of erythroid progenitor cells

In order to gain insight into the mechanisms by which erythropoietin promotes erythropoiesis, effects of various inhibitors on the erythropoietin-promoted differentiation of erythroid progenitor cells and on the erythropoietin-promoted Ca⁺⁺ uptake in the progenitor cells were determined, and the relationship between the inhibitory activity of each inhibitor toward the differentiation and Ca⁺⁺ uptake were examined. The inhibitors used were a tyrosine kinase inhibitor (genistein), a Ca⁺⁺-channel blocker (verapamil), a Ca⁺⁺ chelator (EDTA) and a protein kinase C inhibitor (staurosporine). All of these agents inhibited both the erythropoietin-mediated differentiation of the erythroid progenitor cells, as determined by the incorporation of⁵⁹Fe into heme, and Ca⁺⁺ uptake in a concentration dependent manner. In the cases of verapamil and EDTA, the half-miximal inhibitory concentration (IC₅₀) values for differentiation of the progenitor cells may be the consequence of the inhibition of the Ca⁺⁺ uptake by the inhibitior. On the other hand, in the cases of genistein and staurosporine, the IC₅₀ values for inhibition of differentiation were significantly different from that for inhibition of Ca⁺⁺ uptake. These results suggest that the mechanism of inhibition of differentiation by these two inhibitors is complex. However, taken all together, the above results support the proposition that Ca⁺⁺ uptake may paly a role in the erythropoietin-mediated differentiation of erythoid progenitor cells.     

Taurine、EGCG和genistein联合对体外活化肝星状细胞自噬的影响

目的:探讨牛磺酸(taurine)、表没食子儿茶素没食子酸酯(EGCG)和三羟基异黄酮(genistein) 3种抗肝纤维化药物联合使用对体外活化大鼠肝星状细胞(HSCs)自噬的影响及可能机制。方法:体外培养大鼠肝星状细胞株HSC-T6,设立对照组、联合药物(taurine+EGCG+genistein,TEG)组、自噬抑制剂巴弗洛霉素A1(Baf-A1)组和联合药物+抑制剂(TEG+Baf-A1)组。采用CCK-8法分别检测各组HSCs的存活率,应用透射电镜观察HSCs内部自噬体超微结构变化,通过吖啶橙(AO)染色和荧光显微镜观察各组细胞内自噬溶酶体变化,Western blot分析自噬标志性蛋白LC3-Ⅱ和beclin-1的表达。结果:与对照组相比,TEG组、Baf-A1组和TEG+Baf-A1组细胞的存活率明显降低(P 0. 05);透射电镜观察到细胞内部出现双层或多层膜结构的自噬体,以及单层膜结构的自噬溶酶体。与对照组相比,AO染色结果显示TEG组和Baf-A1组红色荧光区域显著缩小,TEG组中LC3-Ⅱ和beclin-1的表达量显著降低(P 0. 05)。结论:活化的HSCs发生自噬现象,联合药物TEG可阻断HSCs的自噬过程,其作用机制可能与抑制HSCs中自噬体的生成有关。     

Enhanced migration of tissue inhibitor of metalloproteinase overexpressing hepatoma cells is attributed to gelatinases： Relevance to intracellular signaling pathways

AIM: To study the effect of gelatinases (especially MMP-9) on migration of tissue inhibitor of metalloproteinase (TIMP-1) overexpressing hepatoma cells. METHODS: Wild type HepG2 cells, cells stably transfected with TIMP-1 and TIMP-1 antagonist (MMP-9-H401A, a catalytically inactive matrix metalloproteinase (MMP) which still binds and neutralizes TIMP-1) were incubated in Boyden chambers either with or without Galardin (a synthetic inhibitor of MMP-1, -2, -3, -8, -9) or a specific inhibitor of gelatinases. RESULTS: Compared to wild type HepG2 cells, the cells overexpressing TIMP-1 showed 115% migration (P<0.05) and the cells overexpressing MMP-9-H401A showed 62% migration (P<0.01). Galardin reduced cell migration dose dependently in all cases. The gelatinase inhibitor reduced migration in TIMP-1 overexpressing cells predominantly. Furthermore, we examined intracellular signal transduction pathways of TIMP-1-dependent HepG2 cells. TIMP-1 deactivates cell signaling pathways of MMP-2 and MMP-9 involving p38 mitogen-activated protein kinase. Specific blockade of the ERK pathway suppresses gelatinase expression either in the presence or absence of TIMP-1. CONCLUSION: Overexpressing functional TIMP-1-enhanced migration of HepG2-TIMP-l cells depends on enhanced MMP-activity, especially MMP-9.