槲皮素诱导MCF-7细胞凋亡及其与Fas/FasL通路的相关性研究

目的:了解槲皮素诱导人乳腺癌MCF-7细胞凋亡的作用及其与Fas/Fas L通路的相关性。方法:建立槲皮素诱导的MCF-7细胞凋亡模型。采用透射电镜观察细胞核形态变化,Annexin V-FITC/PI和JC-1荧光标记流式细胞术检测细胞凋亡率、线粒体膜电位(Δψm)及Fas L中和抗体对细胞凋亡的阻断作用。采用免疫荧光法和流式细胞术检测细胞Fas/Fas L的表达。流式细胞术观察p38 MAPK抑制剂SB203580对细胞Fas/Fas L表达的影响。Western blot检测p38 MAPK和p-p38 MAPK蛋白水平的变化。结果:80.0μmol/L槲皮素处理MCF-7细胞48h,透射电镜可见染色质浓缩及边缘化现象;处理24 h、48 h和72 h,Δψm分别下降17.4%、44.3%和68.9%,细胞凋亡率分别为(10.2±3.3)%、(28.9±7.5)%和(39.2±8.9)%。Fas L中和抗体预处理细胞后,24 h、48 h和72 h的细胞凋亡率则分别为(8.2±2.8)%、(19.2±5.3)%和(22.5±6.9)%,细胞凋亡阻断率分别为19.6%、33.6%和42.6%。Fas/Fas L表达率随处理时间增加而升高,SB203580可显著抑制细胞Fas/Fas L的表达率。p38 MAPK的蛋白水平变化不大,而p-p38 MAPK的蛋白水平在48 h和72 h显著增高。结论:槲皮素可上调MCF-7细胞的Fas/Fas L表达,并经膜Fas/Fas L途径诱导MCF-7细胞凋亡,p-p38 MAPK可能是上调Fas/Fas L表达的重要信号分子。     

HPLC法同时测定楚雄臭菜中槲皮素和山奈酚的含量

目的建立HPLC法测定楚雄臭菜中的槲皮素和山奈酚含量的方法。方法采用色谱柱:InertSustain C18(250 mm×4.6 mm,5μm);以乙腈(A)-0.3%磷酸水溶液(B)为流动相进行梯度洗脱;流速:0.8 m L/min;检测波长:360 nm;柱温:30℃。结果槲皮素的进样量在0.034~0.272μg范围内(r=0.999 9)、山奈酚的进样量在0.039~0.317μg范围内(r=0.999 8)线性关系良好,槲皮素、山奈酚的平均回收率分别为99.33%(RSD=1.33%,n=6)、99.48%(RSD=1.85%,n=6)。结论本方法操作简便、快捷,稳定性高,重现性好,为楚雄臭菜药材的质量评价提供依据。     

槲皮素抑制血小板活化因子受体结合作用的研究

目的 :观察槲皮素对氚标记的血小板活化因子与血小板膜上受体结合的影响 ,试图证明该药为一新型血小板激活因子 (PAF)受体拮抗剂。方法 :以放射配基结合试验观察 [3H]PAF与兔血小板受体特异性结合的作用 ;以分光光度法测定PAF诱发血小板黏附的强度。结果 :槲皮素可浓度依赖地抑制2 5 ,5 0 ,10 0nmol·L- 1 [3H]PAF与兔血小板受体的特异性结合 ;该药可明显抑制PAF诱发的血小板黏附 ,具明显的量效关系 ,其IC50 为 39 8μmol·L- 1 。结论 :槲皮素具抗PAF作用 ,为一新的PAF受体拮抗剂。     

HPLC波长切换法同时测定杜仲双降袋泡剂中7种成分的含量

目的建立高效液相(HPLC)波长切换法同时测定杜仲双降袋泡剂中桃叶珊瑚苷、京尼平苷酸、绿原酸、京尼平苷、松脂醇二葡萄糖苷、芦丁及槲皮素7种成分含量的方法。方法以80%甲醇为溶剂,加热回流提取;色谱柱:Hydrosphere C18(4.6 mm×200 mm,5μm);柱温:30℃;流动相:乙腈-0.1%磷酸溶液,梯度洗脱;检测波长:208 nm(桃叶珊瑚苷)、235 nm(京尼平苷酸、绿原酸、京尼平苷、松脂醇二葡萄糖苷)、270 nm(芦丁、槲皮素);流速:0.9 ml/min。结果桃叶珊瑚苷、京尼平苷酸、绿原酸、京尼平苷、松脂醇二葡萄糖苷、芦丁、槲皮素的质量浓度分别在10.97~274.25μg/ml(r=0.999 7)、9.78~244.50μg/ml(r=0.999 8)、6.86~171.50μg/ml(r=0.999 6)、2.47~61.75μg/ml(r=0.999 7)、8.11~202.75μg/ml(r=0.999 1)、4.59~114.75μg/ml(r=0.999 9)、1.85~46.25μg/ml(r=0.999 5)范围内线性关系良好;平均加样回收率分别为99.85%、98.92%、97.52%、97.08%、98.51%、97.10%、96.91%,RSD分别为0.93%、0.74%、1.26%、1.37%、0.88%、1.05%、1.33%。结论所建立的HPLC波长切换法可以同时测定杜仲双降袋泡剂中桃叶珊瑚苷、京尼平苷酸、绿原酸、京尼平苷、松脂醇二葡萄糖苷、芦丁和槲皮素的含量,方法简便准确、灵敏度高,可用于杜仲双降袋泡剂的质量控制。     

Suppressive Effects of the Flavonoids Quercetin and Luteolin on the Accumulation of Lipid Rafts after Signal Transduction via Receptors

Quercetin (QUER) and luteolin (LUTE) are dietary flavonoids capable of regulating the production of cytokines, such as tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6). However, their mechanisms of action are not fully understood. In lipopolysaccharide-triggered (LPS)-triggered signaling via Toll-like receptor 4 (TLR4), QUER and LUTE suppresses not only the degradation of the inhibitor of κB (IκB), with resultant activation of nuclear factor-κB (NF-κB), but also the phosphorylation of p38 and Akt in bone marrow-derived macrophages that have been stimulated with LPS. We report here that, in TNF-α-induced signaling, QUER and LUTE significantly suppressed the production of IL-6 and activation of NF-κB. Accumulation of lipid rafts, the initial step in the signaling pathway, was significantly inhibited when macrophages were treated with QUER or with LUTE prior to exposure to LPS. Similarly, the accumulation of lipid rafts was inhibited by the flavonoids when B cells were activated via the membrane IgM and when T cells were activated via CD3. In contrast, QUER and LUTE did not inhibit the activation of phorbol myristate acetate-induced NF-κB in macrophages. Our observations suggest that QUER and LUTE interact with receptors on the cell surface and suppress the accumulation of lipid rafts that occurs downstream of the activation of the receptors.     

Quercetin regulates inflammation,oxidative stress,apoptosis, and mitochondrial structure and function in H9C2 cells by promoting PVT1 expression

ObjectiveTo investigate the effect and potential mechanism of quercetin on inflammation, oxidative stress, apoptosis, and mitochondrial structure and function in H9C2 cells.Materials and methodsH9C2 cells were obtained from the Shanghai Institutes for Biological Sciences, Chinese Academy of Science, and randomly divided into six groups: control, model, PVT1 overexpression (OV), quercetin, OV + quercetin, and NAC groups. The CCK-8 assay was performed to examine cell proliferation. Flow cytometry was used to examine cell apoptosis, cell membrane potential, and ROS levels. The expression of endothelial nitric oxide synthase (eNOS), malondialdehyde (MDA), and superoxide dismutase (SOD) was measured by ELISA and a Biochemical kit. Western blotting was used to determine the levels of p-DRP1 (s637), MFN2, NF-kB, p-NF-kB, IkB, and p-IkB. IL-6, IL-10, TNF-α, and IL-1β mRNA expression was examined by RT-PCR. Electron microscopy was used to observe the structure of mitochondria in H9C2 cells.ResultsMDA, p-NF-κB, p-IKB, IL-6, IL-1β, and TNF-α expression levels, and the cell apoptosis rate were significantly higher in the model group than in the control group (P < 0.05). In contrast, the cell proliferation rate and IL-10, SOD, eNOS, and ATP levels were significantly lower in the model group (P < 0.05). Moreover, MDA expression was significantly lower in the OV, quercetin, quercetin + OV, and NAC groups than in the model group (P < 0.05), while SOD, eNOS, and ATP levels were higher. The electron microscopy results showed that PVT1 overexpression or quercetin treatment could inhibit inflammation-induced mitochondrial fission and promote mitochondrial fusion.ConclusionQuercetin promotes the proliferation of H9C2 cells, while inhibiting inflammation, oxidative stress, and cell apoptosis, and alleviating the structural and functional dysfunction of mitochondria. These effects are achieved by promoting PVT1 expression.     

Quercetin reduces erosive dentin wear: Evidence from laboratory and clinical studies

ObjectiveThe aim of the present study was to evaluate the effect of quercetin on the acid resistance of human dentin through both laboratory and clinical studies.MethodsTwo hundred and twelve dentin blocks (2 mm × 2 mm × 2 mm) were prepared and used. For the laboratory study, dentin specimens were randomly divided into 8 groups (n = 12): deionized water, ethanol, 1.23 × 10⁴ μg/ml sodium fluoride (NaF), 120 μg/ml chlorhexidine, 183.2 μg/ml epigallocatechin gallate (EGCG), and 75 μg/ml, 150 μg/ml, and 300 μg/ml quercetin (Q75, Q150, and Q300). The specimens were treated with the respective solutions for 2 min and then subjected to in vitro erosion (4 cycles/d for 7 d). The surface microhardness loss (%SMH_l), erosive dentin wear, and surface morphology were evaluated and compared. For the impact on MMP inhibition, the release of crosslinked carboxyterminal telopeptide of type I collagen (ICTP) and the thickness of the demineralized organic matrix (DOM) were measured using additional dentin specimens. For the clinical study, the specimens were treated with NaF or Q300 for 2 min and then subjected to in vivo erosion (4 cycles/d for 7 d). The %SMH_l and erosive dentin wear of the specimens were measured to determine whether quercetin similarly inhibits erosion in situ.ResultsThe quercetin-treated group had a significantly lower %SMH_l and erosive dentin wear than any other group, and the effect was concentration-dependent in vitro (P < 0.05). Dentin treated with quercetin produced significantly less ICTP and had a thicker DOM than the control dentin (P < 0.05). After in vivo erosion, the %SMH_l and erosive dentin wear of the Q300 group were significantly lower than those of the control group (P < 0.05).SignificanceThe application of quercetin was shown, for the first time, to increase the acid resistance of human dentin, possibly through MMP inhibition and DOM preservation.     

槲皮素对人肝癌高转移细胞基质粘附能力的影响

目的 研究槲皮素对肝癌高转移细胞基质粘附能力的影响及其机制.方法 研究对象为人高转移肝细胞癌细胞HCCLM6,用细胞基质粘附实验检测槲皮素对细胞外基质粘附能力的影响,免疫荧光和Western blot 检测细胞内E-Cadherin蛋白表达.结果 槲皮素能抑制肝癌细胞对Marigel胶的粘附,免疫荧光和Western blot检测均发现槲皮素处理组肝癌细胞E-Cadherin蛋白表达较对照组增加(P＜0.05).结论 槲皮素能抑制肝癌高转移细胞的基质粘附能力,其机制之一可能在于槲皮素能上调肝癌细胞E-Cadherin蛋白表达.     

Protective effect of flavonoids against red blood cell hemolysis by free radicals

BACKGROUND:

Flavonoids are polyphenolic substances with antioxidant properties, and they are found in different vegetables and fruits. Epidemiological studies have shown that the consumption of flavonoids reduces the prevalence of cardiovascular diseases. The use of synthetic antioxidants, however, has been limited because of their toxicity. Therefore, medical researchers have intensified their quest to find natural antioxidants.

OBJECTIVES:

To investigate the effect of several pure flavonoids, such as kaempferol, quercetin, morin and rutin, on red blood cell hemolysis and evaluate their -SH capacity as an indicator of membrane protection.

METHODS:

The rate of hemolysis and cell membrane -SH capacity were determined by spectrophotometry. Red blood cell peroxidation was induced using 2,2′-azo-bis-(2-amidinopropane) dihydrochloride. The effect of each flavonoid on hemolysis was examined at three concentrations (0.5 μg/mL, 5 μg/mL and 10 μg/mL), however, only the greatest concentration (10 μg/mL) of each flavonoid was used to study the effect on -SH groups.

RESULTS:

In all cases, the antioxidant activity was dose-dependent. Rutin showed the highest inhibitory effect on hemolysis among flavonoids (42.5%). The protective effect of kaempferol, rutin and morin against -SH group oxidation measured 7.7%, 23.3% and 26.4%, respectively.

CONCLUSIONS:

Results showed that flavonoids and flavonoid-containing plants can be used as natural antioxidants for the treatment and prevention of disease conditions, the pathogenesis of which is mediated by lipid peroxidation.