长程体外反搏对高胆固醇血症猪高级氧化蛋白产物和高敏C反应蛋白的影响

目的探讨长程体外反搏对高胆固醇血症猪血清中高级氧化蛋白产物(AOPP)和高敏C反应蛋白(hs-CRP)的影响.方法 18头雄性乳猪随机分为正常饲养组(n=6)、高脂饲养组(n=6)及高脂饲养+体外反搏组(n=6).后2组通过高脂饲养复制高胆固醇血症猪模型并对高脂饲养+体外反搏组进行36 d共36 h的长程增强型体外反搏.分别于分组饲养前、反搏前、反搏中期和反搏结束时留取3组动物静脉血,采用分光光度法检测血清AOPP浓度,采用乳胶凝集反应法检测血清hs-CRP浓度.结果高脂饲养组和高脂饲养+体外反搏组经高脂饲养后血清总胆固醇和低密度脂蛋白明显升高(P<0.05).血清AOPP和hs-CRP浓度在分组饲养前组差异无统计学意义(P>0.05).反搏前、反搏中期和反搏结束时,高脂饲养组与高脂饲养+体外反搏组血清AOPP和hs-CRP浓度较正常饲养组同时期均有显著增高(P<0.05);而反搏中期和反搏结束时高脂饲养+体外反搏组血清AOPP浓度较高脂饲养组显著降低[(95.38±12.66)μmol/L比(128.46±12.55)μmol/L;(85.78±10.33)μmol/L比(158.22±16.32)μmoL/L,P<0.05];且反搏中期和反搏结束时高脂饲养+体外反搏组血清hs-CRP浓度较高脂饲养组也有显著降低[(0.47±0.14)mg/L比(0.62±0.32)mg/L;(0.47±0.16)mg/L比(0.59±0.43)mg/L,P<0.05].结论 AOPP和hs-CRP参与了高胆固醇血症猪的发病过程.长程体外反搏可能通过减轻机体体内氧化应激和微炎性反应过程,从而阻止高胆固醇血症的病理生理进程.     

Differential effects of ceramide species on exocytosis in rat PC12 cells

Increases in several ceramide species have been shown by non-targeted lipid profiling (lipidomics) of the rat hippocampus after kainate lesions (Guan et al. FASEB J 20:1152–1161, 2006). This study was carried out to examine possible effects of ceramide species on exocytosis. Significant increase in membrane capacitance in voltage-clamped rat pheochromocytoma (PC12) cells, an indication of exocytosis, was detected immediately after external application of C2, C6, and C18 ceramide. In contrast, no increase in capacitance was found after addition of C16 and C20 ceramide, or DMSO vehicle. The effect of ceramide on exocytosis was dependent on the integrity of lipid rafts. Treatment of cells with the cholesterol binding agent/disruptor of lipid rafts, methyl β cyclodextrin, prior to addition of C18 ceramide suppressed the increase in capacitance induced by this lipid species. The ability of C2, C6 and C18 ceramide to trigger exocytosis was confirmed using total internal reflection fluorescence microscopy (TIRFM) experiments. External application of these species caused an exponential decrease in the number of subplasmalemmal neuropeptide Y (NPY)-enhanced green fluorescence protein (EGFP) labeled vesicles, indicating exocytosis. Interestingly, C18 is also the ceramide species that showed the greatest increase in the rat hippocampus after kainate excitotoxicity. It is postulated that C18 ceramide might facilitate exocytosis of glutamate from damaged neurons, thus propagating neuronal injury.     

中链甘油三酯对小鼠胆固醇代谢和胆固醇7α-羟化酶表达的影响

目的：研究中链甘油三酯对小鼠胆固醇代谢的影响及可能的机制。 方法： 分别用AIN-93G配方饲料（BC），含1％胆固醇的AIN-93G配方饲料（Chol），含1％胆固醇和14%富含豆蔻酸的长链甘油三酯（LCT）的AIN-93G配方饲料（Chol＋LCT），含1％胆固醇和14％的中链甘油三酯（MCT）的AIN-93G配方饲料(Chol+MCT)喂养C57小鼠6周，每组15只。观察血清总胆固醇（TC）、肝脏胆固醇含量、机体总胆汁酸含量以及肝脏胆固醇7α-羟化酶（CYP7A1）的表达。 结果： Chol＋MCT组小鼠血清TC低于（P＜0.01）、肝脏胆固醇高于（P＜0.01）、机体总胆汁酸含量低于（P＜0.01）、CYP7A1表达低于（P＜0.01）Chol组小鼠；Chol＋MCT组小鼠血清TC低于（P＜0.01）、机体总胆汁酸含量以及CYP7A1 表达高于（P＜0.01）Chol+LCT组小鼠、肝脏胆固醇含量不变。 结论： MCT对机体胆固醇水平的影响与CYP7A1表达有关。     

针对晚期氧化蛋白产物的抗体制备与应用

目的:制备特异性识别晚期氧化蛋白产物(Advanced oxidation protein products,AOPP)的多克隆抗体,为研究AOPP的致病机理及与临床病情相关性提供有效工具。方法:用次氯酸氧化家兔血清白蛋白以获得AOPP-RSA,以此为免疫原免疫家兔,亲和层析纯化免疫血清,ELISA鉴定多克隆抗体的效价及特异性,Western blot检测慢性肾脏病(chronic kidney disease,CKD)患者血浆中AOPP,免疫组织化学方法检测肾组织中AOPP分布与定位。结果:获得效价达10-6的抗AOPP免疫血清,纯化抗体可特异地与不同种属来源的氧化修饰白蛋白结合,而与正常白蛋白及糖基化蛋白终产物无交叉反应。该抗体可识别人血浆中的AOPP,可特异地与大鼠CKD模型及各种炎症性CKD患者肾组织成份反应。结论:成功制备了可与不同种属AOPP结合的特异性多克隆抗体,并首次用于检测存在于CKD患者血浆、肾组织及模型动物肾组织中的AOPP,为深入研究AOPP的病理作用及其在组织定位提供了有效工具。     

Anti-inflammatory properties of lipid oxidation products

Oxidative modification of lipids occurs during inflammatory processes and leads to the formation and accumulation of biologically active lipid oxidation products that induce specific cellular reactions. These reactions lead to a modulation of the inflammatory process and may determine the fate and outcome of the body's reaction in acute inflammation during host defense. The processes by which oxidized lipids may play an important role include resolution of inflammation involving apoptosis, chronic inflammatory processes, and innate and adaptive immune responses. The classical view of lipid oxidation products is that they can induce and propagate chronic inflammatory reactions. However, evidence is accumulating that cells and tissues respond towards these oxidatively formed stress signals also by activation of anti-inflammatory processes. These include defense strategies such as (a) induction of signaling pathways leading to the upregulation of anti-inflammatory genes, (b) inhibition of signaling pathways coupled to the expression of proinflammatory genes, and (c) preventing the interaction of proinflammatory bacterial products with host cells. This contribution summarizes recent findings on the anti-inflammatory action of oxidized lipoproteins and lipid oxidation products. We discuss confirmed and suggested mechanisms as well as the (patho)physiological significance of these findings.     

Effects of 2-deoxyglucose on plasma catecholamines in adult and aged rats

To examine the effects of aging on the responsiveness of the sympathetic-adrenal medullary system, I have measured plasma levels of norepinephrine (NE) and epinephrine (EPI) in adult (6 months old) and aged (24 months old) Fischer 344 male rats. Two days prior to testing, rats were surgically prepared with chronic tail artery catheters to permit remote sampling of blood in conscious, unrestrained animals. Following collection of basal blood samples, each rat received a single injection of 2-deoxyglucose (2-DG, 250 or 500 mg/kg, IP) and additional blood samples were collected 1, 2 and 4 hours later. 2-DG, a glucose analogue, stimulates a centrally mediated activation of the adrenal medulla and to a lesser extent the postganglionic sympathetic neurons. For purposes of analysis, data were excluded from animals which died within 4 hours after injection. Basal plasma levels of both catecholamines were similar in adult and aged rats. Administration of 2-DG was attended by significant and sustained increases in plasma NE and EPI in rats of both ages. A greater proportion of aged rats died following administration of 2-DG compared to adult rats. At the higher dose of 2-DG, plasma levels of NE were significantly higher in 6 month old rats at 1 and 2 hours post-injection. In contrast, plasma levels of EPI were significantly higher in 24 month old rats at 1 and 2 hours after administration of 250 mg/kg 2-DG and at 1 hour after administration of 500 mg/kg 2-DG. These findings indicated that the adrenal medullary response of aged rats to an intense and prolonged stressor is greater than that of adult rats. The attenuated plasma NE response of aged rats to 2-DG may be due to a lessor release of NE from the adrenal medulla and/or a diminished activation of postganglionic sympathetic nerves.     

Effects of 2-deoxyglucose on plasma catecholamines in adult and aged rats

To examine the effects of aging on the responsiveness of the sympathetic-adrenal medullary system, I have measured plasma levels of norepinephrine (NE) and epinephrine (EPI) in adult (6 months old) and aged (24 months old) Fischer 344 male rats. Two days prior to testing, rats were surgically prepared with chronic tail artery catheters to permit remote sampling of blood in conscious, unrestrained animals. Following collection of basal blood samples, each rat received a single injection of 2-deoxyglucose (2-DG, 250 or 500 mg/kg, IP) and additional blood samples were collected 1, 2 and 4 hours later. 2-DG, a glucose analogue, stimulates a centrally mediated activation of the adrenal medulla and to a lesser extent the postganglionic sympathetic neurons. For purposes of analysis, data were excluded from animals which died within 4 hours after injection. Basal plasma levels of both catecholamines were similar in adult and aged rats. Administration of 2-DG was attended by significant and sustained increases in plasma NE and EPI in rats of both ages. A greater proportion of aged rats died following administration of 2-DG compared to adult rats. At the higher dose of 2-DG, plasma levels of NE were significantly higher in 6 month old rats at 1 and 2 hours post-injection. In contrast, plasma levels of EPI were significantly higher in 24 month old rats at 1 and 2 hours after administration of 250 mg/kg 2-DG and at 1 hour after administration of 500 mg/kg 2-DG. These findings indicated that the adrenal medullary response of aged rats to an intense and prolonged stressor is greater than that of adult rats. The attenuated plasma NE response of aged rats to 2-DG may be due to a lessor release of NE from the adrenal medulla and/or a diminished activation of postganglionic sympathetic nerves.     

Effect of hypertension on the in vitro incorporation of [1-14C]-oleate into phospholipid and cholesterol ester in the aortae of normal-fed and cholesterol-fed rabbits

Summary The effect of prior hypertension on the in vitro incorporation of [1-¹⁴C]-oleate into phospholipid and cholesterol ester in aortae from cholesterolfed and normal fed rabbits was studied.Incorporation of [1-¹⁴C]-oleate into phospholipid was not increased in aortae from either hypertensive normal-fed or hypertensive cholesterol-fed rabbits when compared to the appropriate normotensive controls.In the normal-fed rabbits, incorporation of [1-¹⁴C]-oleate into cholesterol ester was increased by hypertension in all aortic regions. In cholesterol-fed rabbits cholesterol esterification was found to be proportional to the intimal cholesterol concentration, irrespective of the prior blood pressure or the particular aortic region studied.It is concluded that the increased lipid synthesis in atherosclerotic vessels from hypertensive rabbits is a consequence of the increased lipid accumulation produced by hypertension and not the result of hypertension directly stimulating arterial wall metabolism.     

阿魏酸钠对高脂血症致动脉粥样硬化过程中胆固醇和甘油三酯代谢的影响

目的：观察阿魏酸钠(SF)对高脂血症引起的动脉粥样硬化(AS)过程中胆固醇和甘油三酯代谢的影响，进一步探讨其抗动脉粥样硬化的作用机制。方法: 高胆固醇喂养复制AS动物模型；以氧化型低密度脂蛋白（ox-LDL）与小鼠巨噬细胞株RAW264.7细胞体外共培养，诱导建立泡沫细胞模型，与人肝母细胞瘤细胞株HepG2细胞体外共培养，建立损伤细胞模型；分别给予SF进行干预。检测实验动物AS斑块面积和血脂水平，油红O染色观察RAW264.7细胞内脂质堆积情况，高效液相色谱定量分析RAW264.7细胞内胆固醇的含量，逆转录聚合酶链反应（RT-PCR）测定HepG2细胞肝脂酶（HL）mRNA的表达水平。结果: (1)与高脂组相比，高脂+SF组动物主动脉粥样斑块面积明显减小（P<0.01），血浆甘油三酯水平显著降低（P<0.01），但血浆胆固醇水平无明显改变（P>0.05）；(2) 与ox-LDL组相比，ox-LDL+SF处理组中HepG2细胞HL mRNA表达水平显著增加（P<0.01），但RAW264.7细胞内胆固醇含量无明显改变（P>0.05）。结论: SF能降低高胆固醇导致的AS斑块面积，具有抗动脉粥样硬化的作用，其作用机制可能与上调HL mRNA表达水平从而降低血浆甘油三酯水平有关，而与血浆胆固醇水平和ox-LDL诱导的巨噬泡沫细胞形成无关。提示血浆甘油三酯升高是AS的独立危险因素，HL介导的甘油三酯代谢途径可能是AS治疗的潜在靶点。     

阳极氧化产品表面元素定性分析探讨

目的对表面经阳极氧化的钛及钛合金产品进行表面元素定性分析,并对其数据进行探讨。方法采用扫描电镜及能谱仪对表面元素进行测定。结果阳极氧化产品表面经扫描电镜及能谱仪分析,所包含元素除基体元素外,还可包含少量Na、Si、Ca、P等元素。结论纯钛、钛合金类产品植入物表面形成的氧化膜具有良好的生物相容性,通过对钛及钛合金表面阳极氧化工艺和成分控制,可以有效降低产品生物安全风险。     

胆固醇自稳失调对SP2/0骨髓瘤细胞免疫原性的影响

目的　探讨胆固醇自稳失调对SP2 0骨髓瘤细胞免疫原性的影响。方法　以去脂血清 (LDS)和 或洛伐他汀 (LOV)处理SP2 0细胞 ,导致胆固醇自稳失调 ,以密度梯度离心法分离细胞膜 ;以rmGM CSF和rmIL 4诱导获得小鼠骨髓细胞来源的DC ,混合淋巴细胞反应 (MLR)检测其功能 ;DC负载细胞膜抗原活化T细胞获得肿瘤特异性CTL ,MTT法检测其对SP2 0细胞的杀伤效果。结果　MLR结果表明DC具有较强的刺激同种T细胞增殖的能力 ;与对照组膜抗原诱导的CTL相比 ,经胆固醇自稳失调处理的SP2 0细胞膜负载DC诱导的CTL的杀伤活性增强 ,尤以高效靶比 (2 0∶1)时细胞杀伤作用增强更为明显 ,CTL细胞毒性具有肿瘤特异性。结论　胆固醇自稳失调的SP2 0细胞膜负载DC致敏的CTL的细胞毒作用增强 ,并与胆固醇失调严重程度有一致性。     

氧化低密度脂蛋白及溶血卵磷脂对巨噬泡沫细胞胆固醇外流的影响

目的:观察氧化低密度脂蛋白(OxLDL)及其成分溶血卵磷脂(LPC)对小鼠腹腔巨噬泡沫细胞胆固醇外流的影响及其机制的初步探讨。方法:(1)以载脂蛋白AI(apoAI)分别诱导OxLDL和乙酰化低密度脂蛋白(AcLDL)负载小鼠腹腔巨噬细胞所形成的巨噬泡沫细胞, 观察其胆固醇外流情况。(2)分离正常及apoE基因缺陷(E⁰)小鼠腹腔巨噬细胞, 以AcLDL负载形成巨噬泡沫细胞, 分别以LPC及apoAI作为诱导物, 观察其胆固醇外流情况。结果:(1)apoAI能引起AcLDL组巨噬泡沫细胞胆固醇大量外流, 而OxLDL组外流明显受阻。(2)LPC能引起正常组小鼠腹腔巨噬泡沫细胞胆固醇外流, 且呈剂量效应关系, 但E⁰组未见明显外流;apoAI能引起两组的胆固醇外流, 且外流量显著高于LPC。结论:(1)OxLDL能造成胆固醇外流途径受阻。(2)LPC能促进巨噬泡沫细胞胆固醇外流, 可能主要通过apoE途径来进行。     

Effects of dietary cholesterol and voluntary exercise on histopathology,plasma and hepatic lipids of the male rat

Summary To avoid the stress of imposed activity, male weanling rats were allowed to exercise voluntarily in individual activity wheels. The exercising animals, which were compared to sedentary controls, ran over 26 km/wk (over 2 miles/day). Half of the animals in each group were fed a 10% coconut oil diet; the other half were fed the same diet with 1% added cholesterol.Plasma cholesterol was monitored throughout the 23-week regime. Consistently lower plasma cholesterol values were shown by the exercising animals during the first weeks of the study, the differences being statistically significant at the end of the 8th week. Dietary cholesterol sharply elevated plasma cholesterol, which reached a peak at the 5th week, then declined to basal levels by the 10th week.Both neutral glycerides and cholesterol levels of the livers were elevated considerably by the addition of cholesterol to the diet. Exercising, however, had a lowering effect on both liver cholesterol and neutral glycerides. The weights of the hearts of the exercising rats were increased, while those of the other organs selected were unchanged.Histologic examination of sections of livers showed fat infiltration of hepatic cells varying in severity, depending on the diet. Greater damage to liver cells was noted when cholesterol was added to the basal diet. Fat infiltration was lessened considerably in exercising rats on the basal diet; exercising partially overcame the effects of added cholesterol.This work was supported in part by Grant-In-Aid from The Nutrition Foundation, Inc.; State of Washington Initiative 171 Fund and USPHS Research Grant HD-03475 from the National Institutes of Health.     

Effects of endurance training on lactate removal by oxidation and gluconeogenesis during exercise

This report describes the effects of 9 weeks of endurance-training on the relative rates of lactate removal via oxidation and gluconeogenesis in humans. Before and after training, eight subjects performed incremental (60 W plus 40 W every 6 min) exercise tests, while¹⁴C-lactate was infused into one forearm vein and arterialized venous blood was sampled from the other forearm. During the trial, the volume of expired¹⁴CO₂ and circulating¹⁴C-lactate and¹⁴C-glucose specific radioactivities were measured. Such measurements revealed that training increased the estimated oxidation of equivalent venous blood lactate concentrations [VLa] of greater than 1.6 mmol/l. These increases in lactate oxidation were more than would be predicted from the approximately 40% higher O₂ uptake values at any [VLa] after training. At a [VLa] of 6 mmol/l, rates of lactate oxidation were increased by some 100% following training, from 105±12 to 208±33 mol/min/kg (P<0.01). Improvements in lactate oxidation after training reduced the estimated rates of lactate-to-glucose conversion from 40±3 to 9±2 mol/min/kg at a [VLa] of 2.5 mmol/l (P<0.01). Thus, unlike in rats, human endurance-training does not increase gluconeogenesis. In the final stages of progressive exercise after training, more than 80% of lactate was oxidised and accounted for approximately 45% of overall carbohydrate oxidation.     

阿魏酸钠对糖尿病大鼠肾脏非酶糖基化和氧化的影响

目的:探讨阿魏酸钠(SF)对糖尿病(DM)大鼠肾脏非酶糖基化和氧化的影响。方法:对链脲佐菌素(STZ)诱导的DM大鼠灌胃给予SF 110 mg·kg^-1·d^-1,治疗8周,测定各组大鼠肾重／体重、肌酐清除率(Ccr)、24 h尿蛋白定量、血清和肾皮质果糖胺(FMN)、血清和肾皮质丙二醛(MDA)含量及抗氧化酶活性,并分别测定肾皮质糖基化终产物(AGEs)含量,观察肾脏病理改变。结果:糖尿病对照组(DM组)大鼠肾重／体重、Ccr、24 h尿蛋白定量、血清FMN、肾皮质FMN和AGEs显著高于正常对照组(N组);SF治疗组(SF组)肾重／体重、Ccr、24 h尿蛋白定量、血清FMN和肾皮质AGEs显著低于DM组;DM组大鼠肾皮质和血清超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性显著低于N组,MDA含量显著高于N组,SF治疗组大鼠肾皮质和血清SOD、CAT活性显著高于DM组,MDA含量显著低于DM组:DM组大鼠肾脏病理改变异常显著,SF组的肾脏病理学改变轻于DM组大鼠。结论: SF通过保护肾脏抗氧化酶,减轻氧化应激,抑制AGEs在肾脏的沉积对DM大鼠肾脏产生保护作用。     

高脂高胆固醇饮食对apoE-/-/LDLR-/-/Leprdb/db小鼠肝组织基因表达差异的影响

目的：研究高脂高胆固醇饮食对三基因突变(apoE-/-/LDLR-/-/Leprdb/db)小鼠肝脏基因表达的影响, 分析这些差异表达基因与血脂代谢紊乱和动脉粥样硬化（AS）的关系。 方法： 利用cDNA表达谱芯片检测普通饮食和高脂高胆固醇饮食喂养的三基因突变小鼠肝脏基因表达差异;分别用COD-PAP,GPO-PAP法测定血浆总胆固醇（TC）和甘油三酯（TG）浓度;并观察了主动脉形态变化。 结果： 在被测的4 000条基因中，高脂高胆固醇饮食组较普通饮食组小鼠上调基因为78条，下调114条，包括脂代谢、糖代谢、细胞骨架蛋白和免疫与炎症等相关基因。高脂高胆固醇饮食组血浆TC和TG水平均高出普通饮食组。5周龄时，两组小鼠均有主动脉内膜损伤，而高脂高胆固醇饮食组重于普通饮食组，并随年龄增长而加重。 结论： 高脂高胆固醇饮食对三基因突变小鼠血脂代谢紊乱乃至AS的发生发展有明显促进作用。     

Adiponectin 基因转染对肌细胞糖原合成和葡萄糖氧化的影响

目的：研究adiponectin对C2C12肌细胞糖原合成和葡萄糖氧化的影响。 方法： 用阳离子脂质体介导转染和随后G418筛选建立稳定转染小鼠adiponectin cDNA真核表达质粒（pcDNA3.0-mad）及空载pcDNA3.0的C2C12细胞株并鉴定。C2C12肌细胞糖代谢实验分对照组、空载体组和pcDNA3.0-mad(mad)组共3组进行，每组又分 0、0.5、5、100 nmol/L胰岛素刺激4个亚组。通过液闪测定细胞合成的糖原中［14C］的放射活性和氧化产生的［14CO2］，分别检测肌细胞的糖原合成和葡萄糖氧化情况。 结果： Western blotting和免疫组化检测证实mad组细胞表达adiponectin蛋白。Mad组葡萄糖氧化量随胰岛素浓度增加的速率较其它两组快，对照、空载体和mad组线性回归系数分别为23.34、23.23和26.06。Mad组C2C12肌细胞基础状态下和胰岛素刺激下的葡萄糖氧化和糖原合成与其它两组无显著差异（P>0.05）。 结论： 转染adiponectin基因对C2C12肌细胞葡萄糖氧化和糖原合成无显著影响。