复方总黄酮对ApoE基因敲除小鼠动脉粥样硬化的影响

目的探讨复方总黄酮对ApoE基因敲除小鼠动脉粥样硬化的保护作用。方法 15只7周龄的♂C57BL/6小鼠普通饮食为正常对照组;75只7周龄的♂ApoE基因敲除小鼠高脂饮食,随机分为5组:模型组、辛伐他汀组、低剂量复方总黄酮组、中剂量复方总黄酮组、大剂量复方总黄酮组。灌胃16周造模完成后,采集血清,分离胸主动脉。HE染色检查胸主动脉斑块,检测血脂4项(TC、TG、LDL-C、HDL-C)和血清SOD;ELISA检测IL-1β、NF-κB值。结果模型组胸主动脉斑块明显,复方总黄酮组斑块均不同程度变小。复方总黄酮组的TC、TG、LDL-C、IL-1β、NF-κB明显降低,HDL-C、SOD含量明显升高,与模型组比较差异有显著性(P<0.05)。结论复方总黄酮对小鼠早期动脉粥样硬化有良好的保护作用,可能与其降脂抗炎作用有关。     

基因敲除动脉粥样硬化小鼠模型研究进展

基因敲除小鼠动脉粥样硬化模型是目前较为认可的探讨动脉粥样硬化发病机制及寻找新药物靶点的关键工具,也应用于潜在抗动脉粥样硬化药物的药理和毒理研究。载脂蛋白E(ApoE)、低密度脂蛋白受体(LDLR)和B类Ⅰ型清道夫受体(SR-B1)等基因的表达促进机体脂质、胆固醇和低密度脂蛋白等转运和代谢,维持血管正常功能,敲除这些基因会引起脂质转运及代谢紊乱从而诱发动脉粥样硬化及并发症的发生发展。常见的基因敲除小鼠有ApoE基因敲除、LDLR基因敲除及其改良品系,这些模型小鼠能够客观反映敲除基因对动脉粥样硬化发生的影响,且广泛应用于动脉粥样硬化的非临床研究。本文阐述了当前基因敲除小鼠动脉粥样硬化模型的机制、应用和优缺点,以期为动脉粥样硬化发病机制研究及抗动脉粥样硬化药物筛选提供参考。     

银杏内酯B对ApoE基因敲除小鼠动脉粥样硬化的影响

目的探讨银杏内酯B(ginkgolide B,GB)对ApoE基因敲除(ApoE-/-)小鼠动脉粥样硬化的影响。方法 12只8周龄C57BL/6J小鼠为正常组,24只8周龄♂ApoE-/-小鼠为阳性模型组以及药物组;药物组每天给予GB 0.6 mg灌胃。8周后处死小鼠,用病理学方法观察小鼠动脉斑块、脂质沉积以及巨噬细胞表达。用ELISA方法测定血浆RAN-TES含量。结果电镜结果显示模型组动脉斑块较大,血管内膜损伤明显,而GB组小鼠动脉内表面损伤明显减轻,斑块较小。HE染色显示了同样的结果,模型组斑块较大,GB组斑块较小。血浆RANTES测定正常组为123.81 ng.L-1,模型组为359.16 ng.L-1,GB组为193.36 ng.L-1,各组之间差异存在统计学意义(P<0.01)。结论 GB能有效地减轻ApoE-/-小鼠的动脉粥样硬化损伤,并降低血浆RANTES含量,其药理学机制可能与抑制血小板功能相关。     

青蒿素对ApoE-/-基因敲除小鼠动脉粥样硬化的影响

目的探讨青蒿素对ApoE-/-基因敲除小鼠动脉粥样硬化以及炎症因子的影响。方法将20只ApoE-/-基因敲除小鼠分为青蒿素组和PBS处理组,每组10只,经高脂喂养8周,与对照组小鼠(C57BL/6J标准饮食小鼠,10只)比较,通过血管大体油红O染色评价斑块面积;HE染色观察病变形态及血浆中炎症因子的变化。结果与对照组比较,PBS处理组及青蒿素组均可见动脉硬化斑块,炎症因子水平升高。青蒿素组的动脉硬化程度及炎症因子水平均明显低于PBS处理组。结论青蒿素可以改善ApoE-/-基因敲除小鼠动脉粥样硬化进展。     

优降脂组方对ApoE基因敲除动脉粥样硬化模型小鼠的动脉粥样硬化相关细胞因子的影响

目的 探讨优降脂组方对ApoE基因敲除（ApoE-/-）动脉粥样硬化模型小鼠的动脉粥样硬化相关细胞因子的影响。方法 选择ApoE-/-小鼠45只,同时选择同遗传背景C57BL/6J小鼠雄性15只。造模成功后分为4组：模型组、正常对照组、优降脂组方实验组、阿托伐他汀对照组。实验结束后,测定4组小鼠血脂水平及动脉粥样硬化相关细胞因子水平。结果 模型组小鼠TC、TG、LDL-C均高于正常对照组,HDL-C低于正常对照组,差异有统计学意义（P<0.05）。优降脂组方实验组和阿托伐他汀对照组TC、TG、LDL-C均低于模型组,HDL-C均高于模型组,差异有统计学意义（P<0.05）。模型组小鼠hs-CRP、TNF-α、MDA均高于正常对照组,NO低于正常对照组,差异有统计学意义（P<0.05）。优降脂组方实验组和阿托伐他汀对照组hs-CRP、TNF-α、MDA均低于模型组,NO均高于模型组,差异有统计学意义（P<0.05）。结论 优降脂组方有助于降低血脂水平,对动脉粥样硬化发生和发展有一定抑制作用,为临床提供一定的实验理论依据。     

Caveolin-1在载脂蛋白E基因敲除小鼠动脉粥样硬化形成中的变化

目的：探讨caveolin-1在apoE基因敲除小鼠动脉粥样硬化形成过程中的变化。方法：取正常和apoE基因敲除小鼠(品系均为C57BL／6J)作为研究对象，分别观察不同周龄apoE基因敲除小鼠血清甘油三酯、总胆固醇、低密度脂蛋白、高密度脂蛋白的含量及主动脉横断面积、斑块面积的变化。免疫组织化学染色法对caveolin-1进行半定量及定位分析，Western-blotting检测caveolin-1在主动脉的表达。结果：apoE基因敲除小鼠的血清甘油三酯、总胆固醇、低密度脂蛋白水平与对照组比较显著升高，并随小鼠周龄增加而升高；斑块面积及斑块面积与主动脉面积的比值也随小鼠周龄增加而增大。免疫组织化学染色法显示，caveolin-1在实验组血管内膜表达呈阳性减弱，其程度随周龄增加有减少趋势。免疫印迹检测显示实验组caveolin-1的表达与对照组比较有所减弱，并与小鼠周龄呈负相关。结论：apoE基因敲除小鼠主动脉内皮细胞caveolin-1表达下调，可能与其动脉粥样硬化形成有关。     

抗炎抗凝融合蛋白TAP-SSL5对ApoE基因敲除小鼠动脉粥样硬化病变形成的影响

摘 要 目的：探讨重组融合蛋白蜱抗凝血肽（TAP） 金黄色葡萄球菌超抗原样蛋白5（SSL5）对ApoE基因敲除（ApoE-/-）小鼠动脉粥样硬化病变形成的影响。方法: 选取12周龄雄性ApoE-/-小鼠21只,随机分为3组：TAP-SSL5组（3 mg·kg^-1·d^-1）、SSL5组（2 mg·kg^-1·d^-1）、空白对照组（pH 7.4磷酸盐缓冲液）,ip,qd,连续给药12周,观察小鼠体质量变化。高脂饮食饲养12周后取材,检测血浆总胆固醇（TC）、三酰甘油（TG）、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)水平;对小鼠主动脉血管进行石蜡切片,常规HE染色,分析小鼠主动脉根部血管动脉粥样硬化斑块的形成情况,小鼠主动脉内膜面采用油红O染色,比较动脉粥样硬化斑块的分布情况。结果: 高脂饲养12周后,与空白对照组比较,TAP-SSL5组小鼠体质量增长和TC水平明显降低（P<0.001）,而TG、HDL-C、LDL-C水平无明显变化。HE染色结果表明,TAP-SSL5组主动脉根部切片斑块面积明显降低（P<0.05）;主动脉大体标本斑块油红O染色提示TAP-SSL5干预组主动脉动脉粥样硬化斑块形成明显小于空白对照组。结论:TAP-SSL5可显著抑制ApoE-/-小鼠动脉血管粥样硬化斑块的形成。     

新型他汀类药物—ZD4522对ApoE基因敲除小鼠血管内皮粘附功能的影响

目的观察他汀类新药ZD4522对ApoE基因敲除小鼠主动脉内皮粘附单核细胞的抑制效应.方法40只高胆固醇血症动物模型-ApoE基因敲除小鼠随机均分为A,B,C,D,E 5 组,各组ZD4522的皮下注射剂量分别为0,1,5,20和20mg·kg-1,qd,E组给药6周,其余4组均给药2周.2或6周后测定小鼠血浆胆固醇水平和主动脉内皮对单核细胞的粘附率.结果5组ApoE基因敲除小鼠血浆胆固醇水平均比正常小鼠显著升高(P<0.01),与A组比较,E组血浆胆固醇水平明显降低(P<0.01).5组ApoE基因敲除小鼠的主动脉内皮单核细胞粘附率均高于正常组,与A组比较,C,D,E组的单核细胞粘附率均显著降低(P<0.05,P<0.01,P<0.01).结论他汀类新药ZD4522对ApoE基因敲除小鼠血管内皮粘附单核细胞有明显的抑制作用,该抑制效应是独立于其降脂作用的.     

小鼠bcl10基因敲除载体的构建

根据已知的cDNA序列设计引物，通过RT-PCR获得小鼠bcl10基因cDNA片段作为探针，用噬斑原位杂交法克隆129品系小鼠的bcl10基因组DNA，在亚克隆完成序列结构分析的基础上，利用常规分子克隆技术，构建完成了针对bcl10基因的替代型基因敲除载体。两条同源臂分别为bcl10基因exon3上游2.4kb和exon4下游4.5kb的基因片段。构建完成替代型小鼠bcl10基因敲除载体，为后续获得bcl10基因缺陷型胚胎干细胞系奠定了实验基础。     

硫化氢对动脉粥样硬化胆固醇代谢的调节作用

目的观察H2S对动脉粥样硬化(AS)胆固醇代谢的影响,并探究其作用机制。方法用高脂饲料结合维生素D3的方法建立AS大鼠模型,并给与腹腔注射NaHS 56μmol·kg-1.d-1,持续12周。检测血脂水平及肝胆固醇含量。HepG2细胞给与NaHS 100μmol·L-16 h后,用RT-PCR方法检测羟甲基戊二酰辅酶A还原酶(HMGCoA reductase)和低密度脂蛋白受体(LDLR)mRNA表达。结果 NaHS组血清总胆固醇(TC)、甘油三酯(TG)、高密度脂蛋白胆固醇(HDL-Ch)和低密度脂蛋白胆固醇(LDL-Ch)均较模型组降低,肝胆固醇含量下降。NaHS使HepG2细胞HMGCoA Re-ductase mRNA表达下降,LDLR mRNA表达增加。结论H2S对AS胆固醇代谢具有调节作用,能下调肝HMGCoAReductase,使胆固醇合成下降;上调肝脏LDLR表达,降低血清总胆固醇水平。     

Taurine inhibits development of atherosclerotic lesions in apolipoprotein E-deficient mice

1. The effects of taurine on the development of atherosclerotic lesions were investigated in apolipoprotein (apo) E-deficient mice, an animal model with severe hypercholesterolaemia and extensive atherosclerosis. These mice were fed a normal laboratory chow containing 2% taurine for 12 weeks. 2. Serum total cholesterol was significantly elevated after 12 weeks treatment with taurine. This elevation was due to increases in very low-density lipoprotein- and low-density lipoprotein-cholesterol. 3. Despite such effects on serum lipoproteins, analysis using en face oil red O staining revealed that taurine reduced the area of arterial lipid accumulation by 28%, as measured quantitatively as an index of atherosclerosis. Histological examination also demonstrated a decrease in the size of aortic lesions in taurine-treated mice. 4. Serum levels of thiobarbituric acid reactive substances (TBARS) in apoE-deficient mice were higher than in normolipidaemic C57BL/6J mice. Serum TBARS levels were significantly decreased by 12 weeks treatment of apoE-deficient mice with taurine. 5. Thus, taurine prevents the formation of atherosclerotic lesions, independently of serum cholesterol levels, and the results suggest that the anti-oxidative effects of taurine are related to its anti-atherosclerotic actions.     

Digoxin increases hydrogen sulfide concentrations in brain, heart and kidney tissues in mice

The interest in digoxin has recently increased due to the expanding knowledge regarding endogenous cardiac glycosides and a potential oncological application of this drug. Hydrogen sulfide (H(2)S), a crucial co-modulator of various physiological processes, is involved in the pathophysiology of different disorders and may be useful in the treatment of some diseases. The interaction between cardiac glycosides and H(2)S is unknown. The aim of the study is to assess the influence of digoxin on H(2)S tissue concentrations in mouse brain, heart and kidney. Thirty male BALB/c mice were given intraperitoneal injections of digoxin at 0.5 mg/kg body weight (b.w.) per day (group D1, n = 10) or 1 mg/kg b.w. per day (group D2, n = 10). The control group (n = 10) received physiological saline. Free H(2)S tissue concentrations were measured via the Siegel spectrophotometric modified method. There was a significant, progressive increase in the H(2)S concentrations for both the low and high digoxin doses in the brain (7.7% and 8.5%, respectively), heart (by 6.0% and 22.1%, respectively) and kidney (by 7.6% and 13.0%, respectively). This report shows that digoxin administration is followed by an increase in the free H(2)S concentrations in mouse brain, heart and kidney tissues.     

Effects of hydrogen peroxide and apolipoprotein E isoforms on apolipoprotein E trafficking in HepG2 cells

1. The major source of apolipoprotein E (apoE) is the liver. In the present study, the effects of oxidative stress and apoE isoforms on apoE distribution and trafficking were established using the HepG2 liver tumour cell line. 2. Hydrogen peroxide (0, 25, 250 and 1000 μmol/L) was associated with rapid and concentration‐dependent redistribution of apoE into the early endosomal compartment. This redistribution was achieved with a much lower concentration (25 μmol/L) than that needed to induce changes in intracellular apoE mRNA expression, apoE protein levels and markers of oxidative stress (250–1000 μmol/L). 3. Live cell imaging of apoE3–green fluorescent protein revealed a significant decrease in traffic velocity in response to oxidative stress. 4. The E4 isoform was associated with reduced trafficking velocity compared with the E3 isoform under basal conditions. 5. The results indicate that oxidative stress and apoE isoforms influence apoE trafficking and distribution within HepG2 cells. Altered apoE hepatocyte trafficking may provide a mechanistic link between oxidative stress, ageing and some diseases in older people.     

Effects of taurine on serum cholesterol levels and development of atherosclerosis in spontaneously hyperlipidaemic mice

1. The effects of two naturally occurring substances, namely taurine and catechins, on serum cholesterol levels and on the progression of atherosclerotic lesions were evaluated using spontaneously hyperlipidaemic (SHL) mice as an animal model of atherogenesis. 2. Twelve weeks treatment of SHL mice with taurine (1% in drinking water) significantly elevated serum high-density lipoprotein-cholesterol (HDL-C) levels without affecting levels of low-density lipoprotein- and very low-density lipoprotein-cholesterol. In addition, taurine suppressed the development of atherosclerotic lesions by 29%, as determined by oil red O-stained areas in cross-sections of the aorta. 3. In contrast, 12 weeks treatment with a catechin mixture had no apparent effect on serum cholesterol levels and on the progression of atherosclerosis. 4. Serum levels of thiobarbituric acid-reactive substances, an index of oxidized substances, significantly decreased from 9.6 to 6.7 nmol/mL following taurine treatment. 5. We suggest that retardation of atherosclerosis by taurine in SHL mice may be related to decreases in oxidized substances and increases in serum HDL-C levels.     

A novel dendritic mesoporous silica based sustained hydrogen sulfide donor for the alleviation of adjuvant-induced inflammation in rats

PurposeS-propargyl-cysteine (SPRC), an excellent endogenous hydrogen sulfide (H₂S) donor, could elevate H₂S levels via the cystathionine γ-lyase (CSE)/H₂S pathway both in vitro and in vivo. However, the immediate release of H₂S in vivo and daily administration of SPRC potentially limited its clinical use.MethodsTo solve the fore-mentioned problem, in this study, the dendritic mesoporous silica nanoparticles (DMSN) was firstly prepared, and a sustained H₂S delivery system consisted of SPRC and DMSN (SPRC@DMSN) was then constructed. Their release profiles, both in vitro and in vivo, were investigated, and their therapeutical effect toward adjuvant-induced arthritis (AIA) rats was also studied.ResultsThe spherical morphology of DMSN could be observed under scanning Electron Microscope (SEM), and the transmission electron microscope (TEM) images showed a central-radiational pore channel structure of DMSN. DMSN showed excellent SPRC loading capacity and attaining a sustained releasing ability than SPRC both in vitro and in vivo, and the prolonged SPRC releasing could further promote the release of H₂S in a sustained manner through CSE/H₂S pathway both in vitro and in vivo. Importantly, the SPRC@DMSN showed promising anti-inflammation effect against AIA in rats was also observed.ConclusionsA sustained H₂S releasing donor consisting of SPRC and DMSN was constructed in this study, and this sustained H₂S releasing donor might be of good use for the treatment of AIA.     

硫化氢对自发性高血压大鼠胸主动脉舒张反应的影响

目的 探讨内源性及外源性硫化氢(H_2S)对自发性高血压大鼠(SHR)离体胸主动脉舒张反应的影响。方法 4wb WKY大鼠24只,随机分为WKY对照组(n=8)、WKY+H_2S组(n=8)及WKY+PPG组(n=8)。同样周龄SHR大鼠24只,随机分为高血压对照组(n=8)、高血压+H_2S组(n=8)及高血压+PPG组(n=8),高血压对照组及WKY对照组大鼠每日腹腔注射生理盐水,WKY+H_2S组及高血压+H_2S组每日腹腔注射硫氢化钠(NaHS),WKY+PPG组及高血压+PPG组每日腹腔注射PPG,5 wk后处死,取胸主动脉,观察其对乙酰胆碱(ACh)、硝普钠(SNP)及NaHS的舒张反应。结果 WKY+PPG组及高血压对照组、高血压+PPG组对ACh及SNP的舒张率较WKY组降低(P<0.05),而高血压+H_2S组较高血压对照组升高(P<0.05),与WKY对照组相似;各组大鼠血管环对SNP的舒 张反应均高于ACh(P<0.05);WKY对照组及高血压对照组对不同浓度的NaHS呈现剂量依赖性舒张反应,对于同样浓度的NaHS,高血压对照组较WKY对照组的舒张率高。结论 H_2S可以独立及与一氧化氮协同发挥舒张血管效应,在自发性高血压血管舒张功能异常的形成机制中占据重要地位。     

Metformin raises hydrogen sulfide tissue concentrations in various mouse organs

BackgroundThe epidemic of diabetes mellitus type 2 forces to intensive work on the disease medication. Metformin, the most widely prescribed insulin sensitizer, exerts pleiotropic actions on different tissues by not fully recognized mechanisms. Hydrogen sulfide (H₂S) is involved in physiology and pathophysiology of various systems in mammals and is perceived as a potential agent in the treatment of different disorders. The interaction between biguanides and H₂S is unknown. The aim of the study is to assess the influence of metformin on the H₂S tissue concentrations in different mouse organs.MethodsAdult SJL female mice were administered intraperitoneally 100 mg/kg b.w. per day of metformin (group D1, n = 6) or 200 mg/kg b.w. per day of metformin (group D2, n = 7). The control group (n = 6) received physiological saline. The measurements of the free and acid-labile H₂S tissue concentrations were performed with Siegel spectrophotometric modified method.ResultsThere was a significant progressive increase in the H₂S concentration along with the rising metformin doses as compared to the control group in the brain (D1 by 103.6%, D2 by 113.5%), in the heart (D1 by 11.7%, D2 by 27.5%) and in the kidney (D1 by 7.1%, D2 by 9.6%). In the liver, massive H₂S accumulation was observed in the group D1 (increase by 420.4%), while in the D2 group only slight H₂S level enhancement was noted (by 12.5%).ConclusionOur experiment has shown that metformin administration is followed by H₂S tissue concentrations increase in mouse brain, heart, kidney and liver.     

载脂蛋白E基因多态性对调血脂药临床疗效研究进展

载脂蛋白E是维持人类正常脂质代谢的重要物质,是一种遗传多态性蛋白,存在于多种脂蛋白中。不同基因表型调节着体内载脂蛋白E的代谢能力及其与受体的结合能力。本文对载脂蛋白E基因多态性与调血脂药临床疗效的相关性进行综述。