大鼠再生肝对二乙基亚硝胺启动作用的敏感性

目的比较再生肝和正常肝对二乙基亚硝胺（ＤＥＮ）启动作用的敏感性。方法以２／３肝叶切除后８周末的大鼠为实验组,正常大鼠为对照组,作如下比较：肝重、常规组织学检查及３Ｈ－ＴｄＲ掺入试验;用修改的Ｓｏｌｔ－Ｆａｒｂｅｒ模型,通过对ＧＧＴ阳性癌前病灶的体视学测量,观察肝脏对ＤＥＮ的启动效应;在体内和体外（无血清原代培养肝细胞）经ＤＥＮ攻击后,以核酸原位缺口标记方法观察肝细胞ＤＮＡ的损伤程度。结果２／３肝叶切除后８周末的实验组肝脏的修复过程已完成,未见肝细胞继续增生的表现;经ＤＥＮ攻击后,实验组肝癌前病灶在数密度和体积密度上都显著高于对照组;无论在体内或体外接受ＤＥＮ攻击后,实验组肝细胞ＤＮＡ的损伤程度都显著大于对照组。结论即使再生过程已经完成,再生肝仍比正常肝具有较高的致癌敏感性,这与再生肝肝细胞在ＤＥＮ攻击后其ＤＮＡ损伤较重相关。     

Thymidine transport in hepatocytes

Thymidine transport and phosphorylation were investigated in isolated rat hepatocytes and AS 30 D hepatoma cells. In contrast to hepatoma cells, hepatocytes exhibited a minimum of thymidine phosphorylation due to a 100-fold smaller thymidine kinase activity. In hepatocytes thymidine is transported by two transport systems: a specific concentrative high affinity system and an unspecific non-concentrative low affinity system. In hepatoma cells only the low affinity system could be detected. A single dose of 20 or 50 mg diethylnitrosamine/kg body weight induced in hepatocytes a remarkable increase of thymidine kinase activity and a decrease of the transport by the high affinity system. Thymidine transport and phosphorylation by hepatocytes are considered to be sensitive markers for early recognition of toxin-induced liver regeneration.

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Zusammenfassung Thymidintransport und -phosphorylierung wurden in isolierten Rattenhepatozyten und AS 30 D-Hepatomzellen untersucht. Hepatozyten wiesen im Gegensatz zu Hepatomzellen aufgrund einer 100fach niedrigeren Thymidinkinaseaktivität eine äußerst niedrige Thymidinphosphorylierungsrate auf. In Hepatozyten wurde Thymidin durch zwei Transportsysteme aufgenommen: ein spezifisches, konzentratives high affinity System und ein unspezifisches, nichtkonzentratives low affinity System. In ATP-freien Hepatomzellen konnte nur das low affinity System nachgewiesen werden. Eine einmalige Dosis von 20 bzw. 50 mg Diäthylnitrosamin/kg Körpergewicht bewirkte in Hepatozyten einen Anstieg der Thymidinkinaseaktivität und eine Verminderung des Thymidintransports über das high affinity System. Thymidintransport und -phosphorylierung in Hepatozyten erwiesen sich als sensitives System zur frühen Erkennung beginnender Leberregeneration nach toxischer Vorschädigung.

     

"调肝颗粒剂"阻断肝癌前病变过程中的抗氧化和清除自由基作用

目的探讨氧自由基与肝细胞癌变的关系,以及调肝颗粒剂的抗氧化作用.方法:用二乙基亚硝胺(DEN)建立SoltFarber二步肝癌前病变短期动物模型,用调肝颗粒剂进行干预,电子自旋共振(ESR)技术测定羟自由基清除率,改良的硫代巴比土酸(TBA)法测定过氧化脂质(LPO),用双硫代硝基苯(DTNB)直接显色法测定谷胱甘肽过氧化物酶(GSH-Px),用黄嘌呤-黄嘌呤氧化酶法测定超氧化物歧化酶(SOD).结果:调肝颗粒剂能明显抑制肝癌前病变过程中产生的丙二醛(MDA),并对羟自由基具有良好的清除作用.与此同时提高肝组织及血浆中的SOD、GSH-Px活力.结论:调肝颗粒剂可以通过抗氧化来干预、阻断肝癌前病变.     

Carcinogenicity of diethylnitrosamine in newborn,infant, and adult mice

Summary Modifying effects of age, sex, and mouse strain on diethylnitrosamine (DEN) carcinogenesis have been investigated in C57BL/6Jx C3HeB/FeJ F₁ (B6C3F₁) and C3HeB/FeJxA/J F₁ (C3AF₁) hybrid mice. Animals each received four IP injections of 1.5 or 3.0 g DEN/g body weight. The first injections were administered on days 1, 15, or 42 of life. Subsequent treatments were delivered at 3-, 6-, and 6-day intervals, respectively. Mice were kept under observation for the remaining life-span. DEN treatment induced tumors in liver, lungs, and forestomach in descending order of frequency. The majority of the induced liver tumors were hepatocellular carcinomas. Animals treated as newborns and infants developed significantly more liver tumors than animals that were treated as young adults. Newborn and infant females developed liver tumors at a later age (B6C3F₁) and with a lower incidence (C3AF₁) than similarly treated males. The B6C3F₁ mice developed more hepatocellular carcinomas and a higher rate of pulmonary metastases than the C3AF₁ mice. In contrast, C3AF₁ mice developed lung tumors with a higher incidence and multiplicity than B6C3F₁ hybrids. Forestomach tumors were observed also with a slightly but significantly higher incidence in C3AF₁ mice.Dedicated to Professor Hermann Druckrey on the occasion of his 80th birthdayThese investigations have been supported in part by Contracts NIH-NCI-69-2087 and NO1-CO-43317 from the National Cancer Institute     

Role of insulin receptor substrate‐1 for diethylnitrosamine plus high‐fat diet‐induced hepatic tumorigenesis in mice

We investigated the role of insulin receptor substrate (Irs)‐1 for diethylnitrosamine (DEN) plus high‐fat (HF) diet‐induced hepatic tumorigenesis in mice. We gave DEN by intraperitoneal injection at the dose of 80 mg/kg to 18‐week‐old wild‐type (WT) and Irs1‐knockout (Irs1^−/−) mice, which were fed a HF diet from 8 weeks‐of‐age until they were killed (52 weeks). The Irs1^−/− mice showed significantly lower plasma alanine aminotransferase levels, triglyceride contents in the liver and also lower expression levels of the genes encoding inflammatory cytokines than the WT mice. The incidence of DEN plus HF diet‐induced hepatic tumors was 71.4% in the WT mice, whereas it was just 14.3% in the Irs1^−/− mice. The present study showed that Irs1 played an important role in DEN plus HF diet‐induced hepatic tumorigenesis.     

斯钙素1基因在二乙基亚硝胺诱发大鼠肝癌发生中的表达

目的 检测哺乳动物斯钙素1(STC1) mRNA和STC1蛋白在二乙基亚硝胺(DENA)诱导大鼠肝癌发生不同阶段的表达水平. 方法 选取80只SD大鼠,随机分为两组,实验组按DENA 70 mg/kg体重灌胃,诱导大鼠肝癌模型,对照组等量蒸馏水灌胃.采用组织病理学、RT-PCR和Western blotting技术分析大鼠肝组织中的STC1基因和蛋白表达水平. 结果只有在DENA诱导的大鼠肝癌组织内才有STC1 mRNA表达和STC1分泌,并且与肝癌发生进程具有相关性. 结论 STC1基因表达和STC1分泌水平与肝细胞癌变具有相关性,可望成为用于肝癌诊断的分子标记物.     

Celecoxib enhances the detoxification of diethylnitrosamine in rat liver cancer

AIM： To study the effect of celecoxib （CXB） on diethylnitrosamine activation through the regulation of cytochrome P450 in a hepatocarcinogenesis model.METHODS： Six-week-old male Sprague-Dawley rats were randomly divided into five groups, a non-treated group （NT）, a diethylnitrosamine-treated group （DEN）, a DEN＋CXB-treated group （DEN＋CXB）,and CXB 8 d-treated and CXB 32 d-treated groups. The effects of celecoxib on the enzymatic activities of CYP1A1, 2A, 2B1/2, and 2E1 were assessed in hepatic microsomes 24 h after DEN administration.Changes in CYPIA1 and CYP2B1/2 protein expression were also evaluated. The rate of DEN metabolism was measured by the production of the deethylation metabolite acetaldehyde, and the denitrosation metabolite nitrite.RESULTS： DEN＋CXB administration produced a significant increase in the enzymatic activities ofCYP2B1/2 and 1A1, whereas it did not change the activities of CYP2A and 2E1, compared to that of the DEN group. CXB treatment for eight days did not produce a significant effect on enzymatic activity when compared to the NT group; however, when it was administered for prolonged times （CXB 32 d group）,the enzymatic activities were increased in a similar pattern to those in the DEN＋CXB group. The observed increase in the enzymatic activities in the DEN＋CXB group was accompanied by an increase in the CYP2B1/2 protein levels; no changes were observed in the levels of CYPIA1. In vitro, CXB increased the denitrosation of DEN, a pathway of metabolic detoxification. The addition of SKF-525A, a preferential inhibitor of CYP2B, abrogated the denitrosation of DEN.CONCLUSION： These results suggest that the mechanism of action of CXB involves enhancement of the detoxification of DEN by an increasing denitrosation via CYP2B1/2.     

超声造影分析二乙基亚硝胺诱发性大鼠肝癌模型的实验研究

目的探讨超声造影评估二乙基亚硝胺诱发性大鼠肝癌模型的可行性。方法20只雄性Wistar大鼠采用30mg/kg腹腔注射二乙基亚硝胺。第14周开始每周轮流抽取5只大鼠进行超声检查,发现肝脏结节的大鼠,进行超声造影(CEUS)检查。检查后处死取标本进行病理诊断。结果大鼠超声检查前死亡率45%(9/20),肝癌造模成功率80%(16/20)。肝细胞癌(HCC)病灶超声检出率72.5%(29/40)。55.2%(16/29)HCC病灶呈“高回声增强-无廓清”为主要CEUS增强模式。结论此动物肝癌模型成瘤率较高,CEUS可用于此模型的评估。     

实验性肝癌前期病变的研究及中药复方861对其的影响

目的 研究肝癌前期病变有关指标的表达和中药复方861对癌前期病变形成过程的影响。方法 以二乙基亚硝胺诱发大鼠肝癌前期病变，以复方861对诱癌过程干预，利用免疫组化、蛋白质印迹(Western—blotting)法分别检测胎盘型谷胱甘肽—S—转移酶(GST—P)、增殖细胞核抗原(PCNA)的表达。结果 免疫组化染色结果示第6周时见GST—P阳性细胞，第8周开始普遍见GST—P结节。PCNA免疫组化结果示，第6周开始有阳性细胞表达，部分GST—P阳性结节内PCNA阳性细胞较多。Western—blotting结果示GST—P从诱癌第6周开始有阳性表达，造模组GST—P阳性表达比同期干预组表达量大。结论 PCNA标识的GST—P阳性的细胞是早期癌前期病变的有效标志。复方861可抑制癌前期病变的进程。     

Perindopril sensitizes hepatocellular carcinoma to chemotherapy: A possible role of leptin / Wnt/ β-catenin axis with subsequent inhibition of liver cancer stem cells

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death. The major challenge in managing HCC is the resistance to chemotherapy. Leptin hormone is associated with different oncogenic pathways implicated in drug resistance. Angiotensin II was found to decrease the production and secretion of leptin.ObjectiveThis study investigated the potential role of an ACEI perindopril as a chemosensitizer agent to sorafenib.MethodHCC was induced in mice using a single dose of diethylnitrosamine DENA (200 mg/kg) followed by phenobarbital 0.05% in drinking water for 16 weeks. Mice were then treated with perindopril (1 mg/kg/day), Sorafenib (30 mg/kg/day), or both of them for another four weeks. Leptin, VEGF, MMP-9, Cyclin D1, EpCAM, and β-catenin were measured using immunoassay while Wnt and ALDH1 were assayed using western blotting assay.ResultsPerindopril whether alone or in combination with sorafenib decrease liver enzymes and preserve the liver architecture. Our study revealed that perindopril significantly increased the antineoplastic, antiangiogenic as well as anti-metastatic effects of sorafenib. This effect was correlated with the downregulation of the leptin / Wnt / β-catenin pathway and overexpression of ALDH1 while downregulation of EpCAMConclusionThis study presents perindopril as a potential chemosensitizer agent that works through decreased expression of the leptin / Wnt / β-catenin pathway.