热毒清拮抗内毒素损伤线粒体的实验研究

本实验制作内毒素所致家兔DIC模型。观察到肝细胞线粒体在超微结构和功能方面均受到明显损害。而应用热毒清注射液之模型动物肝细胞超微结构基本正常;线粒体呼吸功能、ATP酶,肝匀浆和血清过氧化脂质(LPO)接近正常,与模型组比较差异有显著性意义(P<0.01)或<0.05)。体外实验与体内实验结果一致,从而提示热毒清有拮抗内毒素损伤线粒体的功能。     

热毒清的拆方研究——对家兔抗脂质过氧化、清除氧自由基作用的比较

为进一步探索热毒清拆方后各单味中药是否也具有抗脂质过氧化、清除自由基的作用,采用原弥散性血管内凝身(DIC)家兔模型,分别观察热毒清复方与各单味中药对DIC家兔血液超氧化物歧化酶(SOD)、过氧化脂质(LPO)影响.结果显示:热毒清组可提高SOD活性、降低LPO水平,与模型组比较具有显著性差异(P<0.01),各单味药组虽也有该二种作用,但结果不及热毒清组,与之比较具有显著性差异(P<0.05,P<0.01).结果表明,在清除氧自由基方面热毒清之效果明显优于组方内各单味药.     

褪黑素对免疫性肝纤维化大鼠抗氧化酶活性的影响

目的 研究褪黑素对免疫性肝纤维化大鼠血清及肝匀浆中SOD、GSHpx活性的影响。方法 采用人血清白蛋白诱导大鼠免疫性肝纤维化模型,实验大鼠分为正常组、模型组、褪黑素预防组及阳性药对照组,检测各组动物血清及肝匀浆中SOD、GSHpx活性。结果 模型大鼠血清及肝匀浆中SOD、GSHpx活性明显降低;而自造模的尾静脉攻击阶段起,每ig褪黑素(0.3、0.6、0.9mg·kg-1),均可恢复模型大鼠血清及肝匀浆中SOD、GSHpx的活性。结论 褪黑素对免疫性肝纤维化大鼠模型具有一定程度的保护作用,其机制可能与增强其抗氧化酶活性有关。     

热毒清的拆方研究──对DIC家兔肝线粒体呼吸功能和体外稳定溶酶体膜的影响

对组成复方热毒清的４味中药进行拆方研究，分别观察其对播散性血管内凝血（ＤＩＣ）模型家兔肝细胞线粒体呼吸功能和体外稳定溶酶体膜的影响。结果表明：线粒体呼吸控制率（ＲＣＲ），模型组明显低于热毒清组、地塞米松组和正常组（Ｐ＜０．０１）；但组成热毒清的４味中药单方各项指标与模型组比较无明显统计学意义（Ｐ＞０．０５）。在体外稳定溶酶体膜实验中，除热毒清组溶酶体标志酶酸性磷酸酶（ＡＣＰ）和β－葡萄糖苷酸酶在溶酶体上清液中之酶比活性较低（Ｐ＜０．０５）外，余４味中草药均无统计学意义。由此可见：复方热毒清在保护ＤＩＣ家兔肝线粒体损害和体外稳定溶酶体膜方面优于组方中各单味药，从而从一个侧面验证了热毒清组方的合理性。     

珍珠梅提取物中活性成分的分离及对自由基的清除作用

[目的] 探讨珍珠梅提取物中活性成分的分离方法及对自由基的清除作用。[方法] 用硅胶柱色谱法对珍珠梅提取物进行分离纯化，通过物理化学常数和光谱分析法鉴定分离出的化合物结构，用四氯化碳制作急性肝损伤模型，观察珍珠梅活性成分对大鼠血清、肝匀浆、肝线粒体超氧化物歧化酶及谷胱甘肽过氧化物酶活性及丙二醛含量的影响。[结果] 从珍珠梅提取物中分离出4种化合物，分别为5，2?A′，4′-三羟基-6，7，5′-三甲氧基黄酮(Ⅰ)、5，7-二羟基-8-甲氧基黄酮(Ⅱ)、丁二酸(Ⅲ)及对羟基苯甲酸(Ⅳ)；珍珠梅提取物中的活性成分对急性肝损伤大鼠血清、肝匀浆及线粒体超氧化物歧化酶及谷胱甘肤过氧化物酶的活性具有明显的升高作用，并具有降低丙二醛含量的作用。[结论] 从珍珠梅提取物中分离出的4种化合物中化合物Ⅰ，Ⅲ是首次得到的，具有清除自由基的作用。     

肠毒清颗粒对肝硬变内毒素血症大鼠模型内毒素及炎症因子的干预作用

目的：探讨肠毒清颗粒对大鼠肝硬变内毒素血症的干预作用。方法：70只大鼠分为正常对照组10只,余60只制作大鼠肝硬变模型组。模型制作成功后,分为模型组、肠毒清高剂量组、肠毒清低剂量组、乳果糖治疗组。观察各组大鼠的内毒素、TNF-α、IL-6及NO水平。结果：模型组与正常组比较,血浆内毒素、肝匀浆TNF-α、IL-6及NO水平明显升高（ P<0.05）;与模型组比较,肠毒清高剂量组、肠毒清低剂量组、乳果糖组大鼠血浆内毒素、肝匀浆TNF-α、IL-6及NO水平明显降低（ P<0.05）。肠毒清高剂量组与乳果糖组比较,内毒素、TNF-α、IL-6差异显著（ P<0.05）;与肠毒清低剂量组比较,血浆内毒素、IL-6差异有统计学意义。乳果糖组TNF-α、NO与肠毒清低剂量组比较,有显著性差异（ P<0.05）。结论：肠毒清颗粒能够防治肝硬变内毒素血症。     

四氯化碳致大鼠肝损伤机理的初步探讨

本文初步探讨了CCl_4致大鼠急性肝损伤的机理。一次经口给予大鼠CCl_42.5ml/kg 后12、24和48小时,血清ALT及AST活性显著增高。染毒后12小时,肝微粒体细胞色素P-450含量明显减少。肝匀浆GSH含量以及15000g 沉淀的谷胱甘肽—过氧物酶和过氧化氢酶活性下降。肝匀浆、线粒体和微粒体膜MDA含量增高,且呈时间依赖关系。血清MDA含量也明显增高。与脂质过氧化同时,线粒体和微粒体膜流动性增高;肝匀浆脂肪酸组成发生变化。这些结果提示,CCl_4在肝细胞色素P-450催化下产生自由基,进而导致氧化应激,可能是引起大鼠肝损伤的机理。     

橙皮苷对刀豆蛋白A所致小鼠免疫性肝损伤的保护作用

目的探讨橙皮苷对刀豆蛋白A(ConA)所致小鼠免疫性肝损伤的保护作用。方法小鼠随机分为正常组、模型组、联苯双酯组、橙皮苷低、中、高剂量组,各组分别灌胃给药7 d,采用尾静脉注射ConA建立急性肝损伤模型,检测各组小鼠肝脾指数,测定血清谷丙转氨酶(ALT)和谷草转氨酶(AST)水平,检测肝匀浆中谷胱甘肽(GSH)、丙二醛(MDA)含量和caspase-3活性和TNF-α水平,观察肝组织病理学变化。结果与正常组比较,模型组肝脾指数显著增高,肝组织炎性坏死明显,血清ALT、AST升高,肝匀浆中GSH含量显著降低,MDA含量显著升高,同时肝组织caspase-3活性和TNF-α水平亦显著增高(均P0.01)。与模型组比较,橙皮苷能显著降低肝脾指数,改善肝组织损伤程度,降低血清ALT、AST水平,提高肝匀浆中GSH水平,降低MDA含量,降低caspase-3活性和TNF-α水平(P0.05,P0.01)。结论橙皮苷对ConA诱导的小鼠免疫性肝损伤具有一定的保护作用,其机制可能与抑制氧化应激反应,清除自由基,抑制TNF-α释放及caspase-3活化有关。     

长波紫外线大鼠损伤模型的构建

目的探讨长波紫外线（UVA）照射对大鼠机体氧化损伤的影响。方法采用长波紫外线为照射源,对大鼠进行照射,检测各组血清及肝脏匀浆中氧化物歧化酶（SOD）和谷胱甘肽（GSH）活性,丙二醛（MDA）含量及抑制羟自由基的能力。结果经981.71 J/cm2 UVA照射后,模型组血清中SOD、GSH活性明显低于正常组（P〈0.05）;羟自由基、MDA含量显著高于正常组（P〈0.05）。肝组织匀浆中,模型组SOD活性、抑制羟自由基能力明显低于正常组（P〈0.05）;但MDA与GSH与正常组无统计学意义（P〉0.05）。结论 UVA可对大鼠造成氧化损伤。     

热毒清对红细胞膜Na~+,K~+-ATP酶保护作用的实验研究

热毒清(原名抗炎6号)系我校附属同济医院用四味清热解毒中药制成的静脉注射液,具有抑菌、抗炎、解毒、退热、增强免疫功能及稳定溶酶体膜、保护线粒体H~+-ATP合酶等多方面的作用,其多相药理作用可能与保护生物膜活性有关。本实验研究热毒清对内毒素所致家兔红细胞膜Na~+,K~+-ATP酶活性损害作用的影响,以探讨热毒清治疗疾病的机理。 一、材料和方法 1.动物、健康日本大耳白兔24只,体重2～2.5 kg, 雌雄不拘,随机分为4组,每组6只。①正     

Preventive effect of re du qing on hepatocytes and mitochondria damaged by lipid peroxidation in experimental rabbits with endotoxin-induced disseminated intravascular coagulation

In this study, the general Shwartzman reaction of rabbits induced by Escherichia Coli endotoxin was made as DIC models. The experiments showed that the levels of lipid peroxide (LPO) in hepatic tissue and mitochondria in the model group were increased significantly compared with the control group (P less than 0.01), while superoxide dismutase (SOD) activity in hepatic tissue and glutathione peroxidase (GSH-Px) activity in hepatic tissue and mitochondria were decreased significantly (P less than 0.01). The levels of LPO in hepatic tissue and mitochondria in Re Du Qing (RDQ) group and vitamin E (VE) group were decreased significantly (P less than 0.01 and P less than 0.05 respectively) compared with the model group. The levels of LPO in the RDQ group did not differ from the control group (P greater than 0.05), but the levels of LPO in the VE group were still higher than those in the control group significantly (P less than 0.05). The SOD activity in hepatic tissue and GSH-Px activity in hepatic tissue and mitochondria in both RDQ group and VE group were also significantly higher than those in the model group (P less than 0.01). These data suggest that the levels of oxygen free radicals were increased in hepatocytes and mitochondria. This is related to the decreased activities of SOD and GSH-Px in the course of pathogenesis of endotoxin-induced DIC. This study indicates that lipid peroxidation might be one of the important mechanisms resulting in hepatocellular and mitochondria from oxidative damage.     

Experimental studies on the antagonistic effect of re du qing on endotoxin-induced damage of lysosomes and mitochondria

A rabbit endotoxic DIC model was preliminarily performed. The structures of lysosomes and mitochondria in liver cells were evidently destroyed as observed under electronic microscope, whereas those rabbits pretreated with Re Du Qing (RDQ, formerly named anti-inflammatory No. 6)--a mixture of Chinese traditional herbs providing antipyretic and detoxifying action, showed principally normal ultrastructure in liver cells. In lysosomal functional studies, the activity of the lysosomal marker enzyme--acid phosphate (ACP) was 79.0 +/- 4.7% (M +/- SD) in model group, higher than pretreated group (54.01 +/- 4.0%, P less than 0.01). Studies on the mitochondrial function showed that the significant criteria of the respiratory activity of mitochondria--respiratory control ratios (RCR) was 2.83 +/- 1.08 in model group and markedly lower than pretreated group (5.46 +/- 1.25, P less than 0.01). Mitochondrial ATPase activity (mu mol pi/min/mg pr.) was lower in model group (0.280 +/- 0.015) than in pretreated group (0.341 +/- 0.018, P less than 0.05). Lipid peroxide (LPO) in liver homogenates and serum were 1.86 +/- 0.43 n mol MDA/mg pr. and 12.26 +/- 0.84 n mol MDA/ml respectively in model group, whereas in pretreated group they gave a much lower value (1.19 +/- 0.12 and 6.55 +/- 2.97) respectively. Those data showed very significant difference between two groups (P less than 0.01). All of the above indices of pretreated group yielded values close to those of normal control group. The results of experimental study in vitro were identical to those of experimental study in vivo. These experimental studies suggested that RDQ provide antagonistic effect on endotoxin induced damage of lysosomes and mitochondria.     

Effect of Reduqing on TNF-α, IL-1, IL-6, IL-8 and PAF levels in endotoxin-lnduced DIC model of rabbits) on TNF-α, IL-1, IL-6, IL-8 and PAF levels in endotoxin-lnduced DIC model of rabbits

Objective: To study the mechanism of anti-endotoxemic effect of Reduqing Injection (RDQ) and to explore the essence of traditional Chinese “heat-clearing and detoxifying therapy”.Methods: A disseminated intravascular coagulation (DIC) model was made in rabbits by intravenous injection with E. coli endotoxin. Increased plasma levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8) and platelet activating factor (PAF), as well as the ex-vivo LPS-induced interleukin-1 (IL-1) production in peripheral blood monocytes were observed in model (M) group.Results: Same parameters were significantly lower in RDQ and dexamethasone (DXM) groups than those in the M group (P<0.01,P<0.05). Histopathological examination showed severe damage of the liver, lungs and kidneys in the rabbits of M group, in contrast, only mild affects were seen in the RDQ and DXM groups.Conclusions: RDQ exhibits protective effect on rabbits against endotoxin-induced DIC. The suppression of cytokines and inflammatory mediator PAF by RDQ may play a central role in the inhibition of endotoxin-induced DIC cascade.     

热毒清的拆方研究──体内及体外对内毒素的影响

从体内、外观察复方热毒清及其各单味药对内毒素的影响。结果显示：热毒清具有①体外能裂解大肠杆茵Ｏ１１１Ｂ４内毒素的网状结构；②体内能减少家兔内毒素性播散性血管内凝血肾小球微血栓的形成；③减轻内毒素引起的家兔发热反应，与模型组比较，均为Ｐ＜０．０１。而单味药中大青叶和鱼腥草只具有①②两项作用，金银花具有②③两项作用，蒲公英只具有第③项作用。表明复方的整体作用要优于各单味药。     

大承气汤抗自由基损伤的实验研究

采用内毒素所致家兔全身性Ｓｈｗａｒｔｚｍａｎ反应作为ＤＩＣ模型，检测肝线粒体及血清过氧化脂质（ＬＰＯ）含量，结果表明，模型组ＬＰＯ明显升高（Ｐ〈０．０５）；地塞米松组与正常对照组相似，大承气汤组中然较正常对照组稍高，但无统计学差异（Ｐ〉０．０５），提示：大承气汤能够拮抗内毒素诱导的脂质过氧化损害，保护细胞器。     

Scavenging effect of re-du-qing on free radicals

The general Shwartzman reactions of rabbits were induced by intravenously injecting endotoxin twice with 24 hours interval. The lipid peroxides (LPO) in sera and liver homogenates of rabbits in normal saline group (12.26 +/- 0.84 n mol MDA/ml, 1.86 +/- 0.43 n mol MDA/mg protein respectively) were significantly higher than those of rabbits in normal control group (7.93 +/- 2.90, 1.31 +/- 0.22, both P less than 0.01), and LPO in sera and liver homogenates of rabbits in Re-Du-Qing group (6.55 +/- 2.97, 1.19 +/- 0.12) were evidently lower than those of rabbits in normal saline group (P less than 0.01), accessed to the LPO level of rabbits in normal control group. In in vitro experiment, LPO of mitochondria in Re-Du-Qing group (1.50 +/- 0.43 n mol MDA/mg protein) dramatically decreased, compared with that of endotoxin group (2.39 +/- 0.69, P less than 0.05) and of control group (2.23 +/- 0.75, P less than 0.05). The findings of both in vivo and in vitro experiments showed that Re-Du-Qing possesses scavenging effect on free radicals. In view of the detoxification mechanism of Re-Du-Qing, in addition to inhibiting bacteria and degrading. endotoxin, the scavenging effect of Re-Du-Qing on free radicals are also included.     

The Prevenlive－The rapeutic Effects of Reduqing（热毒清）on Endotoxin－Induced DIC in Rabbits（Continued）Protec

ＴｈｅＰｒｅｖｅｎｌｉｖｅ－ＴｈｅｒａｐｅｕｔｉｃＥｆｆｅｃｔｓｏｆＲｅｄｕｑｉｎｇ（热毒清）ｏｎＥｎｄｏｔｏｘｉｎ－ＩｎｄｕｃｅｄＤＩＣｉｎＲａｂｂｉｔｓ（Ｃｏｎｔｉｎｕｅｄ）ＰｒｏｔｅｃｔｉｏｎｏｆＨｅｐａｔｏｃｙｔｉｃＭｉｃｒｏｓｏｍｅｓ，Ｍａ...     

The preventive-therapeutic effects of reduqing (热毒清) on endotoxin-induced DIC in rabbits (Continued) protection of hepatocytic microsomes, maintenance of calcium homeostasis and reduction of oxygen free radicals

Disseminated intravascular coagulation (DIC) model of rabbits was made by the injection of endotoxin. Cytochrome P-450 content and aniline hydroxylase activity in hepatocytic microsomes in DIC rabbits were significantly reduced. The microviscosity of mitochondrial membrane was elevated, while the fluidity of the membrane, and the activities of calcium-magnesium-adenosine triphosphatase and calcium-adenosine triphosphatase were reduced. Lipid peroxide and xanthine oxidase activities were increased, while superoxide dismutase and glutathione peroxidase activities were significantly decreased (P<0.05 or <0.01). The changes in the Reduqing test group were not prominent, and the parameters of the test group were close to those of the control group. This study suggests that in endotoxin induced DIC rabbits the hepatocytic microsomes are injured, calcium homeostasis is disturbed and oxygen free radicals are notably accumulated. Whereas Reduqing may have a protective effect on hepatocytic microsomes from the injury, maintain calcium homeostasis and reduce oxygen free radicals in DIC rabbits.