绿茶表没食子儿茶素没食子酸酯对β淀粉样蛋白25-35诱导小鼠神经瘤母细胞N2a损伤的神经保护作用及其机制

目的探讨绿茶表没食子儿茶素没食子酸酯(green tea epigallocatechin-3-gallate,EGCG)对β淀粉样蛋白25-35(beta-amyloid 25-35,Aβ_(25-35))诱导的小鼠神经瘤母细胞N2a损伤的神经保护作用及其机制。方法体外培养N2a细胞,用含50μg/m L水溶性Aβ_(25-35)片段的DMEM培养液作用于N2a细胞后,加入不同浓度的EGCG(5、10、20、40μmol/L)作用不同时间(24、48、72 h),同时设模型组(未用EGCG处理)及DMSO组(仅用0.1%的DMSO处理)。MTT法检测EGCG对N2a细胞存活率的影响;JC-1法检测EGCG对线粒体膜电位的影响;半胱氨酸蛋白酶-3(caspase-3)活性检测试剂盒检测EGCG对N2a细胞中caspase-3活性的影响;Western blot法检测EGCG对N2a细胞内Bax、Bcl-2和caspase-3蛋白表达水平的影响。结果与模型组和DMSO组比较,20和40μmol/L的EGCG作用24 h时,N2a细胞存活率及caspase-3酶活性明显降低(P0.05),A_(590)/A_(530)值显著升高(P0.05);20μmol/L的EGCG作用24 h时,N2a细胞中Bax和caspase-3蛋白表达水平均明显降低(P0.05),Bcl-2蛋白水平明显升高(P0.05)。结论 EGCG对Aβ_(25-35)诱导的N2a细胞的损伤具有保护作用,其机制可能与EGCG调控细胞凋亡相关蛋白Bax、Bcl-2和caspase-3的表达水平有关。     

酯化型表没食子儿茶素没食子酸酯对乳腺癌细胞凋亡的影响

目的探讨酯化型表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)对人乳腺癌细胞株MCF-7(Myc依赖性)和MDA-MB-231(Myc非依赖性)凋亡的影响。方法采用脂肪酶催化法对EGCG进行酯化修饰,2次硅胶柱层析纯化后,通过高效液相色谱-质谱(HPLC-MS)进行鉴定,并分析纯化的酯化型EGCG的稳定性。流式细胞术检测纯化的酯化型EGCG对MCF-7和MDA-MB-231细胞凋亡的影响。结果经酶法修饰并纯化后获得了EGCG单棕榈酸酯(EGCG-C16),HPLC中不同保留时间的酯化产物的相对分子质量一致,均为696;EGCG在普通培养基里产生的H2O2量远高于其在酸性培养基里及EGCG-C16在普通培养基里产生的H2O2量(P0.000 1),表明EGCG-C16的稳定性较EGCG好;EGCG-C16对MCF-7细胞具有明确的促进凋亡效应,而对MDA-MB-231细胞则无此效应。结论酯化型EGCG对Myc依赖性乳腺癌细胞具有特异性的促进凋亡效应,为进一步深入研究EGCG的抗肿瘤效应奠定了基础。     

利用寡聚环糊精配基键合介质分离表没食子儿茶素没食子酸酯的新方法

研究了利用寡聚环糊精配基键合的高效凝胶介质分离纯化茶多酚中的表没食子儿茶素没食子酸酯的新方法。考察了不同种类的流动相对分离效果的影响。最终确定采用水／乙醇／乙腈为57／30／13（体积比）进行等梯度洗脱,一次色谱分离的收率和纯度分别为73％、98％,介质经过0．5mol／L的NaOH-水-30％乙酸简单再生后即可重复利用。     

没食子酸丙酯及其衍生物的合成研究进展

没食子酸丙酯(PG)是国内外广泛使用的食品及饲料抗氧化剂。对近期PG及其衍生物的催化合成研究进行了评述,提出了存在的问题及建议。     

绿茶提取物表没食子儿茶素-3-没食子酸酯对人卵巢癌HO-8910细胞增殖及Wnt/β-catenin信号通路相关基因表达的影响

目的研究绿茶提取物表没食子儿茶素-3-没食子酸酯(Epigallocatechin-3 gallste,EGCG)对人卵巢癌HO-8910细胞增殖及细胞内Wnt/β-catenin信号通路相关基因表达的影响,探讨EGCG抑制卵巢癌细胞生长的机制。方法用不同浓度的EGCG(10、20和40μg/ml)处理体外培养的人卵巢癌HO-8910细胞不同时间(24、48和72 h),采用MTT法检测细胞的增殖活力;流式细胞术检测细胞周期的变化;RT-PCR和Western blot分别检测细胞中β-catenin和下游靶基因CyclinD1 mRNA的转录水平和蛋白的表达水平。结果 EGCG可明显抑制HO-8910细胞的增殖活力,且抑制作用呈剂量-时间依赖性(P<0.05);40μg/ml EGCG干预后,HO-8910细胞主要阻滞于G0/G1期,且在48 h阻滞作用最为明显;EGCG可显著降低HO-8910细胞中β-catenin和CyclinD1基因mRNA的转录水平和蛋白的表达水平,且呈剂量-时间依赖性(P<0.01)。结论 EGCG可抑制HO-8910细胞的增殖,其机制可能与其抑制Wnt/β-catenin信号通路的活性有关,提示EGCG可能在卵巢癌的治疗中具有一定的应用前景。     

汉黄芩苷通过调节Nrf2/HO-1通路对大鼠心肌缺血再灌注(I/R)损伤的保护作用

目的:观察汉黄芩苷对大鼠心肌缺血再灌注(I/R)损伤的心肌保护作用并探讨其机制.方法:选用健康雄性SD大鼠制备心脏I/R模型大鼠,随机分为5组:假手术组、模型组、ZnPPIX组、汉黄芩苷组和汉黄芩苷+ZnPPIX组.再灌注后24 h,检测各组大鼠血清乳酸脱氢酶(LDH)、肌酸激酶同工酶(CK-MB)的表达水平.用ELI...     

基于没食子酸聚乙二醇酯的锈转化剂制备及其性能

以没食子酸丙酯和聚乙二醇为原料,通过酯交换反应合成了没食子酸聚乙二醇酯(GA-PEG),以此作为金属离子配体制备了新型锈转化剂。利用正交试验和单因素实验考察了各组分比例、转化方法与时间等因素对锈转化膜防腐蚀性能的影响。通过质谱仪确定了合成产物的结构,使用金相显微镜表征了转化膜的形态,通过电化学阻抗谱及Tafel曲线测量研究了转化膜的防腐蚀性能。结果表明：磷酸浓度、溶剂、转化方式和转化时间对锈转化膜的性能有显著影响。最佳的锈转化剂配方(按质量分数计)为GA-PEG 30%、磷酸5%和邻苯二酚1%,使用15%乙醇溶液作为溶剂。该配方制成的转化膜平整,缓蚀效率达83.9%,性能优于以单宁酸或没食子酸为配体的配方。     

过表达脑红蛋白对大鼠脊髓损伤的神经保护作用及其机制

目的 观察过表达脑红蛋白(neuroglobin,NGB)对大鼠脊髓损伤(spinal cord injury,SCI)后神经功能的影响,并探讨其可能的机制.方法 将90只SD大鼠随机分为5组:假手术组、SCI模型组(SCI)、阴性对照组(NC)、AAV-Ngb组(Ngb)和AAV-Ngb+LY249002组(Ngb+...     

Sox11基因对小鼠脑缺血再灌注损伤的保护作用及其机制

目的 探讨Sox11基因对小鼠脑缺血再灌注损伤(cerebral ischemia reperfusion injury,CIRI)的保护作用及其机制,为CIRI的治疗提供新的靶点。方法 建立小鼠大脑中动脉栓塞(middle cerebral artery occlusion,MCAO)模型和Neuro2A细胞氧糖剥离再灌注(oxygen glucose deprivation reperfusion,OGDR)模型,采用实时定量PCR、Western blot、免疫组织化学染色和免疫组织荧光染色技术检测Sox11在MCAO和OGDR模型中的时间和空间分布;Western blot法检测Sox11基因表达干扰后,OGDR模型中细胞凋亡和炎症反应等通路重要基因的表达改变。结果 Sox11 mRNA和蛋白表达水平在小鼠MCAO模型和Neuro2A OGDR模型中均显著上升(P=0.000 1～0.038 8);小鼠CIRI后,Sox11表达升高集中于海马体的海马齿状回(dentate gyrus,DG)区域;在OGDR模型中干扰Sox11表达后,凋亡相关蛋白Cleaved Caspase ...     

Protective effects of lemon flavonoids on oxidative stress in diabetic rats

The effects of lemon flavonoids, as crude flavonoids prepared from lemon juice, were investigated in diabetic rats. The oxidative stress of eriocitrin (eriodictyol 7-O-β-rutinoside) and hesperidin (hesperetin 7-O-β-rutinoside) on streptozotocin-induced diabetic rats was investigated. Diabetic rats were given a diet which contained 0.2% crude flavonoids, 0.2% eriocitrin, and 0.2% hesperidin. After the 28-d feeding period, the concentration of the thiobarbituric acid- reactive substance in the serum, liver, and kidney of diabetic rats administered crude flavonoids, eriocitrin, and hesperidin significantly decreased as compared with that of the diabetic group. The levels of 8-hydroxydeoxyguanosine, which is exchanged from deoxyguanosine owing to oxidative stress, in the urine of diabetic rats administered eriocitrin and hesperidin significantly decreased as compared with that of the diabetic rat group. Crude flavonoids, eriocitrin, and hesperidin suppressed the oxidative stress in the diabetic rats. These results demonstrated that dietary lemon flavonoids of eriocitrin and hesperidin play a role as antioxidant in vivo.     

Prior exercise and antioxidant supplementation: effect on oxidative stress and muscle injury

Background

Both acute bouts of prior exercise (preconditioning) and antioxidant nutrients have been used in an attempt to attenuate muscle injury or oxidative stress in response to resistance exercise. However, most studies have focused on untrained participants rather than on athletes. The purpose of this work was to determine the independent and combined effects of antioxidant supplementation (vitamin C + mixed tocopherols/tocotrienols) and prior eccentric exercise in attenuating markers of skeletal muscle injury and oxidative stress in resistance trained men.

Methods

Thirty-six men were randomly assigned to: no prior exercise + placebo; no prior exercise + antioxidant; prior exercise + placebo; prior exercise + antioxidant. Markers of muscle/cell injury (muscle performance, muscle soreness, C-reactive protein, and creatine kinase activity), as well as oxidative stress (blood protein carbonyls and peroxides), were measured before and through 48 hours of exercise recovery.

Results

No group by time interactions were noted for any variable (P > 0.05). Time main effects were noted for creatine kinase activity, muscle soreness, maximal isometric force and peak velocity (P < 0.0001). Protein carbonyls and peroxides were relatively unaffected by exercise.

Conclusion

There appears to be no independent or combined effect of a prior bout of eccentric exercise or antioxidant supplementation as used here on markers of muscle injury in resistance trained men. Moreover, eccentric exercise as used in the present study results in minimal blood oxidative stress in resistance trained men. Hence, antioxidant supplementation for the purpose of minimizing blood oxidative stress in relation to eccentric exercise appears unnecessary in this population.     

On-column decaffeination and HPLC analysis of epigallocatechin gallate in green tea nutraceuticals

Green tea (GT) derived epigallocatechin gallate (EGCG) is a commonly used nutraceutical for its antioxidant activity. Caffeine is the second major component and interferes with extraction of EGCG. On-column decaffeination was optimized to enable selective enrichment of EGCG, though less retained than caffeine. EGCG recovery and loss in caffeine were determined using RP-HPLC. Results indicated a significant improvement in caffeine removal and EGCG recovery. The SPE-HPLC protocol represents a model for an efficient and economic approach for improving dynamic capacity, selectivity, and sensitivity towards less retained analytes. This is particularly important for analysis of inherently complex samples of nutraceutical products.     

Quercetin and epigallocatechin gallate induce in vitro a dose-dependent stiffening and hyperpolarizing effect on the cell membrane of human mononuclear blood cells

The bioactivity of polyphenols is closely linked to their ability to interact with biological membranes. The study evaluates the in vitro effect of quercetin and epigallocatechin on the membrane anisotropy and transmembrane potential of peripheral blood mononuclear cells (PBMCs) isolated from 26 type 2 diabetes mellitus patients compared to 25 age matched controls. The in vitro assays were analyzed in correlation with the biochemical and inflammatory profile of the subjects and with insulin resistance parameters (HOMA-IR, plasma resistin) as well. For type 2 diabetes patients, the increase of HOMA-IR and resistin concentration was associated with a significant decrease of the PBMCs membrane anisotropy. The two tested polyphenols induced a dose-dependent hyperpolarizing effect and stiffening of the cell membranes for all tested subjects. Physiological levels of quercetin and epigallocatechin gallate had the tendency to normalize the PBMCs membrane anisotropy of the cells isolated from diabetes patients, bringing it to the level of cells isolated from normoglycemic ones. Epigallocatechin gallate induced higher effects compared to quercetin on the membranes isolated from subjects with higher cardiovascular risk. The decrease of membrane fluidity and the hyperpolarizing effect could explain the cardiovascular protective action of the tested compounds.     

Impaired mitochondrial respiratory functions and oxidative stress in streptozotocin-induced diabetic rats

We have previously shown a tissue-specific increase in oxidative stress in the early stages of streptozotocin (STZ)-induced diabetic rats. In this study, we investigated oxidative stress-related long-term complications and mitochondrial dysfunctions in the different tissues of STZ-induced diabetic rats (>15 mM blood glucose for 8 weeks). These animals showed a persistent increase in reactive oxygen and nitrogen species (ROS and RNS, respectively) production. Oxidative protein carbonylation was also increased with the maximum effect observed in the pancreas of diabetic rats. The activities of mitochondrial respiratory enzymes ubiquinol: cytochrome c oxidoreductase (Complex III) and cytochrome c oxidase (Complex IV) were significantly decreased while that of NADH:ubiquinone oxidoreductase (Complex I) and succinate:ubiquinone oxidoreductase (Complex II) were moderately increased in diabetic rats, which was confirmed by the increased expression of the 70 kDa Complex II sub-unit. Mitochondrial matrix aconitase, a ROS sensitive enzyme, was markedly inhibited in the diabetic rat tissues. Increased expression of oxidative stress marker proteins Hsp-70 and HO-1 was also observed along with increased expression of nitric oxide synthase. These results suggest that mitochondrial respiratory complexes may play a critical role in ROS/RNS homeostasis and oxidative stress related changes in type 1 diabetes and may have implications in the etiology of diabetes and its complications.     

白皮杉醇和乙酰化白藜芦醇对小鼠辐射损伤的防护作用

目的探讨白藜芦醇(resveratrol)衍生物白皮杉醇(piceatannol)和乙酰化白藜芦醇(acetylated resveratrol)对小鼠辐射损伤的防护作用。方法将昆明小鼠随机分为8组:正常对照组、照射对照组、白皮杉醇低、中、高剂量(50、100、200 mg/kg)组和乙酰化白藜芦醇低、中、高剂量(50、100、200 mg/kg)组,每组6只。小鼠经5.5 Gy 60Coγ射线一次性全身照射前2 h灌胃给药,对照组均灌服含2%DMSO的生理盐水溶液。照射后24 h,观察小鼠脾脏和胸腺系数、外周血淋巴细胞数量和百分率、血浆超氧化物歧化酶(superoxide dismutase,SOD)活力及骨髓嗜多染红细胞微核率的变化。结果与正常对照组相比,照射对照组小鼠脾脏和胸腺系数、外周血淋巴细胞数量和百分率及血浆SOD活力均显著降低(P<0.05或P<0.01),骨髓嗜多染红细胞微核率显著升高(P<0.01)。与照射对照组相比,白皮杉醇和乙酰化白藜芦醇各剂量组脾脏系数均显著增高(P<0.05或P<0.01),但胸腺系数差异无统计学意义(P>0.05);乙酰化白藜芦醇中、高剂量组外周血淋巴细胞数量和淋巴细胞百分率均显著升高(P<0.05或P<0.01);白皮杉醇中、高剂量组以及乙酰化白藜芦醇各剂量组血浆SOD活力均显著升高(P<0.05或P<0.01);白皮杉醇及乙酰化白藜芦醇各剂量组骨髓嗜多染红细胞微核率均显著降低(P<0.05或P<0.01)。结论白藜芦醇衍生物白皮杉醇和乙酰化白藜芦醇均对小鼠具有一定的辐射损伤防护作用,其作用机制与提高机体免疫能力、清除活性氧(reactive oxygen species,ROS)、保护造血组织免于辐射诱发的损伤有关。     

蛇床子素对鼻咽癌裸鼠移植瘤放疗作用的影响及作用机制

目的探讨蛇床子素(osthole)对鼻咽癌裸鼠移植瘤放疗作用的影响及其可能的机制。方法将鼻咽癌CNE2细胞株经NU/NU裸鼠右腋皮下接种,2×10~8个/只,待移植瘤长至150mm~3时,随机分为对照组(隔1 d经腹腔注射无菌生理盐水,0.2 m L/只)、单纯蛇床子素组[隔1 d经腹腔注射,1.5μg/(g·10μL)]、单纯放疗组(5 Gy/次,隔3 d X射线照射1次)、蛇床子素联合放疗组(经腹腔注射蛇床子素2 h后进行放射治疗,剂量与方法同单纯蛇床子素组及单纯放射组)。每3 d测量移植瘤体积,首次给药后第21天取出移植瘤,称瘤重;流式细胞术检测移植瘤细胞凋亡率;荧光定量PCR及Western blot法分别检测移植瘤细胞中血管内皮生长因子(vascular endothelial growth factor,VEGF)及乏氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)基因m RNA转录和蛋白表达水平。结果与对照组比较,单纯蛇床子素组、单纯放疗组及蛇床子素联合放射组裸鼠移植瘤体积及瘤重均明显下降(P0.05),细胞凋亡率明显升高(P0.05),其中蛇床子素联合放射组最显著,且该组裸鼠移植瘤中VEGF及HIF-1α基因m RNA转录及蛋白表达水平明显低于单纯蛇床子素组及单纯放疗组(P0.05)。结论蛇床子素可增强鼻咽癌裸鼠移植瘤对放疗的敏感性,其机制可能与下调VEGF及HIF-1α表达,抑制血管生成有关。     

可溶性鸟苷酸环化酶激活剂cinaciguat对糖尿病大鼠氧化应激的影响

目的分析可溶性鸟苷酸环化酶(soluble guanylate cyclase,s GC)激活剂cinaciguat对糖尿病大鼠氧化应激的影响。方法用高脂高糖饲料喂养及腹腔注射链脲佐菌素(streptozotocin,STZ)的方法建立2型糖尿病大鼠模型,随机分为模型组和cinaciguat低、中、高剂量[3.5、7、14μg/(kg·d)]组。各组大鼠饲养12周,cinaciguat干预14 d后,经下腔静脉采血,采用全自动生化分析仪检测大鼠血糖及血脂水平,比色法检测血清总氧化物歧化酶(superoxide dismu-tase,SOD)活性、丙二醛(malonaldehyde,MDA)含量,内皮素(endothelin-1,ET-1)ELISA试剂检测血浆ET-1含量。结果与模型组比较,cinaciguat各剂量组大鼠血糖值下降不明显(P均0.05);甘油三酯(triglycerides,TG)和总胆固醇(cholesterol,TC)含量均明显降低(P均0.05);cinaciguat各剂量组SOD活性明显升高,MDA及ET-1含量明显降低。结论 cinaciguat可能具有调节血脂,降低糖尿病大鼠氧化应激水平,改善血管内皮功能的作用。     

过表达脑红蛋白对SH-SY5Y细胞损伤的神经保护作用及其机制

目的探讨过表达脑红蛋白(neuroglobin,NGB)对水溶性β-淀粉样蛋白片段1-42(beta-amyloid1-42,Aβ1-42)诱导的神经母细胞瘤SH-SY5Y细胞损伤的神经保护作用及其机制。方法将质粒pEGFP-NGB转染经Aβ1-42诱导损伤的SH-SY5Y细胞,MTT法检测NGB对损伤细胞存活率的影响;JC-1染色法检测NGB对损伤细胞线粒体膜电位的影响;免疫细胞化学法及Western blot法分别检测损伤细胞中细胞色素C(cytochrome C,cytoC)和caspase-3、caspase-9的表达水平。结果过表达NGB可明显提高Aβ1-42诱导损伤的SH-SY5Y细胞的存活率(P<0.05),抑制损伤细胞线粒体膜电位的降低(P<0.01),使损伤细胞内cytoC和caspase-3、caspase-9蛋白的表达水平均明显降低(P<0.05)。结论 NGB可通过抑制与细胞凋亡密切相关的cytoC、caspase-3和caspase-9等蛋白的表达而发挥其神经保护作用。本实验为阿尔茨海默病(Alzheimer disease,AD)的治疗提供了实验依据。     

microRNA-143阻遏乳腺癌细胞免疫抑制的作用及其机制

目的探讨microRNA-143(miR-143)阻遏乳腺癌细胞免疫抑制的作用及其分子机制。方法采用MTT法检测乳腺细胞HBL-100和乳腺癌细胞MCF-7、MDA-MB-231、BT549补体依赖的细胞毒性(complement-dependent cytotoxicity,CDC);在脂质体介导下,将miR-143、miR-143对照物(scramble)和miR-143抑制剂(inhibitor)分别转染MCF-7细胞,将miR-143 scramble和miR-143 inhibitor分别转染HBL-100细胞,采用MTT法检测CDC,RT-PCR法检测细胞中补体调节蛋白CD46基因mRNA的转录水平,Western blot法检测CD46蛋白的表达水平。结果乳腺癌细胞MCF-7、MDA-MB-231、BT-549的A570值与正常乳腺细胞HBL-100相比,均明显升高(P<0.01);与miR-143 scramble转染的MCF-7细胞的A570值相比,miR-143转染组的A570值明显下降(P<0.001),与miR-143 scramble转染的HBL-100细胞的A570值相比,miR-143 inhibitor转染的HBL-100细胞的A570值明显上升(P<0.001);与HBL-100细胞相比,CD46蛋白在MCF-7、MDA-MB-231和BT-549细胞中表达均上调,且与抑制CDC的作用呈正相关;在HBL-100细胞中过表达miR-143 inhibitor能上调CD46蛋白的表达水平,而不影响CD46基因mRNA的转录水平;在MCF-7细胞中过表达miR-143能明显抑制CD46蛋白的表达水平。结论 CD46在乳腺癌细胞中的上调导致了癌细胞对CDC的抵抗作用,是产生免疫抑制的机制之一;miR-143通过抑制CD46蛋白的表达水平,使细胞对CDC的作用更为敏感,提高了补体系统对乳腺癌细胞的杀伤作用,降低了其免疫抑制能力。