一氧化氮在大鼠创伤局部微循环灌流减少中的作用

目的和方法：用激光多普勒微循环血流仪测定大鼠断肢局部微循环血液灌流量，用比色法测定血浆亚硝酸盐（ＮＯ２－）水平间接反映血浆一氧化氮（ＮＯ）含量。结果：（１）断肢局部微循环血液灌流量在断肢Ｏ、１、２和３ｈ均明显下降，局部股静脉血浆ＮＯ２－含量在断肢２、３ｈ显著降低，断肢２、３ｈ局部股静脉血浆ＮＯ２－水平与局部微循环血液灌流量呈显著正相关。（２）ＮＯ合成底物左旋精氨酸（Ｌ－Ａｒｇ）能改善断肢２、３ｈ的局部微循环血液灌流量，一氧化氮合成抑制剂左旋单甲基精氨酸（Ｌ－ＮＭＡ），可抑制此作用。（３）断肢３ｈ局部股静脉血浆ＮＯ２－水平Ｌ－Ａｒｇ组明显高于生理盐水（ＮＳ）组（Ｐ＜００５），Ｌ－Ａｒｇ合用Ｌ－ＮＭＡ组则明显低于ＮＳ组（Ｐ＜００５）。结论：断肢创伤通过内源性Ｌ－Ａｒｇ－ＮＯ通路而降低ＮＯ生成，参与断肢局部２、３ｈ的血管痉挛和微循环血液灌流量下降     

常压缺氧大鼠血浆和肺组织TXB_2和6－keto－PGF_（1α）的动态变化

本文观察常压缺氧２４ｈ及１，２，４ｗ（６ｈ／ｄ，６ｄ／ｗ）后大鼠血小板数及聚集率和血浆及肺ＴＸＢ＿２和６－ｋｅｔｏ－ＰＧＦ＿（１α）的动态变化。结果表明（１）缺氧大鼠血小板数呈先增加、后恢复、再下降，血小板聚集率呈先减少、后增加再减少的动态变化。（２）血浆ＴＸＢ＿２和６－ｋｅｔｏ－ＰＧＦ＿（１α）均明显升高，（３）肺组织６－ｋｅｔｏ－ＰＧＦ＿（１α）升高早于ＴＸＢ＿２，提示ＰＧＩ＿２参干旱期缺氧肺血管张力调节。（４）缺氧大鼠２周时肺动脉高压达到高峰，右心室明显肥厚，而肺组织ＴＸＢ＿２至缺氧４周后才见增加，表明ＴＸＢ＿２不是缺氧早期肺血管收缩的主要因素。     

先天性多发性关节弯曲、肾功紊乱及胆汁郁积（ARC）综合征之肝组织学改变：3例报告及文献复习

先天性多发性关节弯曲、肾功紊乱及胆汁郁积（ＡＲＣ）综合征之肝组织学改变：３例报告及文献复习（英）／ＨｏｒｓｌｅｎＳＰ…／／ＪＭｅｄＧｅｎｅｔ－１９９４，３１．－６２～６４婴儿先天性多发性关节弯曲伴发胆汁郁积性肝病和肾小管性酸中毒而死亡，自１９７３年Ｌ...     

大脑5－HT的变化及5－HT_2受体拮抗剂赛庚啶对沙土鼠脑缺血的影响

本文研究大脑组织５－ＨＴ及其代谢产物变化以及５－ＨＴ＿２受体拮抗剂赛庚啶对脑缺血损伤的防治作用。实验共分三部分，结果是：（１）在沙土鼠急性前脑缺血模型证明，缺血１０ｍｉｎ，３０ｍｉｎ，及缺血３０ｍｉｎ再灌２ｈ，可见脑组织５－ＨＴ减少，说明脑缺血时脑组织释放５－ＨＴ增加而摄取减少，５－ＨＴ代谢产物５ＨＩＡＡ缺血后也见减少，但再灌后明显增加。（２）５－ＨＬ＿２受体拮抗剂赛庚啶可减轻沙土鼠缺血５ｍｉｎ再灌７天后海马ＣＡ＿１区神经元迟发性损伤，给赛庚啶组ＣＡ＿１区神经元数为１６８．７９±６２．５／ｍｍ，而生理盐水对照组为８４．７２±８７．３１／ｍｍ。（３）赛庚啶对沙土鼠单侧脑缺血能减少脑组织水及钠含量改善脑水肿及减少脑组织Ｃａ￣（２＋）含量。从５－ＨＴ＿２受体拮抗剂的作用说明，５－ＨＴ可通过５－ＨＴ＿２介导加重脑缺血时神经元损伤，加重脑水肿和脑组织Ｃａ＿（２＋）聚积，５－ＨＴ＿２受体拮抗剂可减轻缺血性脑损伤。     

重型乙型肝炎患者β_2－微球蛋白的检测及意义

对３１例重型乙型肝炎患者用放射免疫分析法检测血清中β＿２－Ｍ含量，并用免疫组化ＡＢＣ法检测肝内β＿２－Ｍ、ＨＢＡｇ和Ｔ细胞亚群的表达与分布。结果发现重型肝炎血清中β＿２－Ｍ含量显著增高，且与病情轻重相关；肝内浸润细胞中ＣＤ８￣＋细胞占优势，提示肝内β＿２－Ｍ、ＨＢｓＡｇ表达及ＣＤ８￣＋细胞的浸润与细胞损伤有关。     

中医温病营血分证有关微循环障碍的实验研究

观察中医温病营血分证家兔模型的微循环、血液流变性和炎症介质的变化。结果发现，模型家兔球结膜血管的数目少，直径细，异形多，渗出多，血流慢；全血粘度在低切时增高，红细胞聚集性增加，变形性降低；血小板少，ＴＸＡ２、ＰＡＩ、ＰＡＦ、ＩＬ－６和ＴＮＦ－α等的水平均增高；ｔ－ＰＡ的水平降低；肺、肝、肾组织切片中多见血管内血栓形成。这些指标与正常对照均有显著差异（Ｐ＜０．０５～０．００１）。说明温病营血分证是由多种炎症介质包括细胞因子所介导引起的全身性包括多脏器的严重微循环障碍     

嗜酸性粒细胞IL—2RmRNA表达及IL—2的趋化作用

本文报道从多粘菌素Ｂ诱导的豚鼠腹腔渗出液，分离嗜酸性粒细胞（Ｅｏｓ），提取总ＲＮＡ，以地高辛配基标记的ＩＬ－２ＲａｃＤＮＡ探针进行点杂交，检测出ＥｏｓＩＬ－２ＲａｍＲＮＡ表达。测定ｒｈＩＬ－２对豚鼠Ｅｏｓ的趋化作用，ＥＤ＿（５０）为１０￣（－１３）ｍｏｌ／Ｌ、最大趋化作用浓度为１０￣（－１２）ｍｏｌ／Ｌ。但在１０￣８～１０￣（－１２）ｍｏｌ／Ｌ浓度范围内，ＩＬ－２对４８ｈＥｏｓ存活率及Ｅｏｓ过氧化物酶活性均无显著作用。表明Ｅｏｓ有功能性ＩＬ－２受体表达，其作用主要为极强的趋化反应。     

人醛缩酶A单克隆抗体的制备及应用

纯化并鉴定了人醛缩酶Ａ、Ｂ、Ｃ（ｈＡＬＤ－Ａ、Ｂ、Ｃ）。用ｈＡＬＤ－Ａ免疫Ｂａｌｂ／Ｃ小鼠，将免疫的脾细胞和Ｐ＿３－Ｘ＿（６３）－Ａｇ８．６５３骨髓瘤细胞用ＰＥＧ进行融合，用ＥＬＩＳＡ法筛选，ｈＡＬＤ－Ｂ、Ｃ作阴性对照，将阳性反应的杂交瘤细胞进行克隆，获得３株杂交瘤细胞株。其ＭｃＡｂ的亚类分别为ＩｇＧ２ｂ，ＩｇＧ１、ＩｇＭ，亲合常数分别为７．５×１０￣（１０）、３．５×１０￣９、２．３×１０￣９。３株细胞株分泌的单抗通过Ｉｍｍｕｎｏｂｌｏｔｔｉｎｇ得到证实。用亲合层析法从人肝癌细胞中提纯了ＡＬＤ－Ａ，ＳＤＳ－ＰＡＧＥ显示为单一区带，但其电泳迁移率较ｈＡＬＤ－Ａ滞后。     

抗病毒蛋白MxA的诱导和检测方法的实验研究

目的 研究抗病毒蛋白ＭｘＡ的表达及其活性。方法用ＩＦＮα２ｂ或３型腺病毒（Ａ＿３）分别对ＷＩ－３８细胞或人外周血单个核细胞（ＰＢＭＣｓ）作用 １２或２４ ｈ，并用Ｗｅｓｔｅｍ ｂｌｏｔ法或ＦＡＣＳ法，分别对Ｍ蛋白的表达进行定性和定量检测。采用微量细胞病变抑制法，对重组的ＭｘＡ和ＩＦＮα２ｂ进行抗病毒效应实验。结果 （１）低浓度的 ＩＦＮα２ｂ（＞ １×1０～＃ＩＵ／Ｌ）和Ａｄ＿３（＞２００ ＴＣＩＤ＿５０），均可诱导相应细胞表达 ＭｘＡ；（２）１０μｇ／Ｌ ＭｘＡ可抵抗２０个 ＴＣＩＤ＿（５０）的 ＨＳＶ－Ｉ、Ｐｏｌｉｏ． Ｖ感染 Ｖｅｒｏ细胞、Ａｄ＿３感染ＨｅＬａ细胞，抵抗２００个ＴＣＩＤ＿（５０）的ＶＳＶ感染Ｗｉｓｈ细胞。结论ＭｘＡ只能由干扰素（ＩＮＦα2ｂ）或病毒诱导产生，其具有广谱的抗病毒活性。     

高原对大鼠血浆与组织TXB_2和6－Keto－PGF_（1α）含量的影响及

高原对大鼠血浆与组织ＴＸＢ＿２和６－Ｋｅｔｏ－ＰＧＦ＿（１α）含量的影响及丹参的保护作用青海省中医药研究所（西宁８１０ｏｏ０）王立义青海省红十字医院陈琴音本文观察了平原大鼠进入海拔４４７５ｍ高原后，其血浆及心、肝、脾组织血栓索Ｂ２（ＴＸＢ２）和６－酮...     

The coagulation system contributes to alphaVbeta6 integrin expression and liver fibrosis induced by cholestasis

Chronic injury to intrahepatic bile duct epithelial cells (BDECs) elicits expression of various mediators, including the αVβ6 integrin, promoting liver fibrosis. We tested the hypothesis that tissue factor (TF)-dependent thrombin generation and protease activated receptor-1 (PAR-1) activation contribute to liver fibrosis induced by cholestasis via induction of αVβ6 expression. To test this hypothesis, mice deficient in either TF or PAR-1 were fed a diet containing 0.025% α-naphthylisothiocyanate (ANIT), a BDEC-selective toxicant. In genetically modified mice with a 50% reduction in liver TF activity fed an ANIT diet, coagulation cascade activation and liver fibrosis were reduced. Similarly, liver fibrosis was significantly reduced in PAR-1(-/-) mice fed an ANIT diet. Hepatic integrin β6 mRNA induction, expression of αVβ6 protein by intrahepatic BDECs, and SMAD2 phosphorylation were reduced by TF deficiency and PAR-1 deficiency in mice fed the ANIT diet. Treatment with either an anti-αVβ6 blocking antibody or soluble transforming growth factor-β receptor type II reduced liver fibrosis in mice fed the ANIT diet. PAR-1 activation enhanced transforming growth factor-β1-induced integrin β6 mRNA expression in both transformed human BDECs and primary rat BDECs. Interestingly, TF and PAR-1 mRNA levels were increased in livers from patients with cholestatic liver disease. These results indicate that a TF-PAR-1 pathway contributes to liver fibrosis induced by chronic cholestasis by increasing expression of the αVβ6 integrin, an important regulator of transforming growth factor-β1 activation.     

Quantitative assessment of the rat intrahepatic biliary system by three-dimensional reconstruction

The anatomical details of the biliary tree architecture of normal rats and rats in whom selective proliferation was induced by feeding alpha-naphthylisothiocyanate (ANIT) were reconstructed in three dimension using a microscopic-computed tomography scanner. The intrahepatic biliary tree was filled with a silicone polymer through the common bile duct and each liver lobe embedded in Bioplastic; specimens were then scanned by a microscopic-computed tomography scanner and modified Feldkamp cone beam backprojection algorithm applied to generate three-dimensional images. Quantitative analysis of bile duct geometry was performed using a customized software program. The diameter of the bile duct segments of normal and ANIT-fed rats progressively decreased with increasing length of the biliary tree. Diameter of bile ducts from ANIT-fed rats (range, 21 to 264 microm) was similar to that of normal rats (22 to 279 microm). In contrast, the number of bile duct segments along the major branch reproducibly doubled, the length of the bile duct segments decreased twofold, and the length of the biliary tree remained unchanged after ANIT feeding. Moreover, the total volume of the biliary tree of ANIT-fed rats was significantly greater (855 microl) than in normal rats (47 microl). Compared with normal rats, the total surface area of the biliary tree increased 26 times after ANIT-induced bile duct proliferation. Taken together, these observations quantitate the anatomical remodeling after selective cholangiocyte proliferation and strongly suggest that the proliferative process involves sprouting of new side branches. Our results may be relevant to the mechanisms by which ducts proliferate in response to hepatic injury and to the hypercholeresis that occurs after experimentally induced bile duct proliferation.     

Connection of ductlike structures induced by a chemical hepatocarcinogen to portal bile ducts in the rat liver detected by injection of bile ducts with a pigmented barium gelatin medium.

Newly found metaplastic ductlike structures that form in the liver of rats exposed to carcinogens are connected to preexisting bile ducts. Male Fischer rats fed a diet of N-2-acetylaminofluorene in a choline-deficient diet (CDAAF) develop a massive proliferation of oval cells which appear to differentiate into bile-ductlike structures. However, unlike normal or proliferating bile ducts, these ductlike structures contain alpha-fetoprotein (AFP) and albumin, which are markers for proliferating hepatocytes and some hepatocellular carcinomas. Bile duct injections with a green pigmented barium gelatin medium filled the lumens of the ductlike structures and typical ductlike structures induced by the CDAAF diet, as well as the proliferating bile ducts induced by the noncarcinogenic alpha-naphthylisothiocyanate (ANIT), and the ducts in the normal controls. AFP was present in the ductlike structures in the rats fed AAF, but not in the bile ducts of animals fed ANIT. These studies suggest that most, if not all, of the ductlike structures produced during chemical hepatocarcinogenesis are derived from bile ducts, yet have the capacity to produce AFP and albumin.     

Effect of cycloheximide on the distribution of alpha-naphthylisothiocyanate in rats

Others have shown that in rats the administration of cycloheximide (2 mg/kg) within 2 hr before or after 300 mg/kg of α-naphthylisothiocyanate (ANIT) blocks the development of hyperbilirubinemia and cholestasis. The effects of cycloheximide on the tissue distribution of ANIT labeled with ¹⁴C or ³H were examined. In cycloheximide-treated animals, less ANIT-derived radioactivity was found in blood, liver, fat and several other tissues at various times after ANIT than in controls. Rats given a mixture of (naphthyl-4-³H) ANIT and (isothiocyanate-¹⁴C) ANIT with a ratio of ${}^6\text{H}{}^{14}\text{C}\text{:6.3}$, excreted in their bile during the first 8 hr after ANIT, more ³H compared to ¹⁴C. In rats treated with cycloheximide, the ${}^3\text{H}{}^{14}\text{C}$ ratio did not change. The data suggest that normal animals excrete an ANIT metabolite in their bile from which the ¹⁴C label in the isothiocyanate moiety has been lost, whereas this does not happen in cycloheximide-treated animals. This provides further evidence that biotransformation of ANIT is necessary for the development of hyperbilirubinemia and cholestasis. Bile might be a convenient starting material for the isolation and identification of ANIT metabolites.     

Strukturelle und mikrozirkulatorische Veränderungen in der Rattenleber nach intermittierender Gabe von Alpha-Naphthyl-isothiocyanat (ANIT) über 12 Monate

Zusammenfassung Am Modell der chronisch-intermittierenden Intoxikation der Ratte mit ANIT werden die Bindegewebsneubildung und deren Einfluß auf die Leberstruktur, auf die Mikrovascularisation und Mikrozirkulation und damit die Blutversorgung der geschädigten Parenchymanteile verfolgt. Während eine einmalige Verbreichung von ANIT reversible Veränderungen wie bei intrahepatischer Cholestase hervorruft, bewirkt eine chronische oder chronisch-intermittierende Intoxikation eine Proliferation von Gallengängen und im Gefolge derselben die Bildung von Bindegewebssepten, die zu einem Umbau der Leberstruktur, ähnlich wie bei der primären biliären Cirrhose führt. Die irreversiblen Veränderungen entwickeln sich bei einer Versuchsdauer von 6–8 Monaten. Die Progredienz der Veränderungen ist an die Verabreichung von ANIT gebunden. Anzeichen eines eigenständigen Fortschreitens können nicht nachgewiesen werden.

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Changes in structure and microcirculation of the rat liver after intermittent application of alpha-naphthyl-isothiocyanate (ANIT) for 12 months

Summary Long-Term intermittent intoxication of rats with ANIT was used to study the proliferation of connective tissue and its influence on the microvascularisation and microcirculation and the blood supply of the damaged liver. A single application of ANIT produces reversible lesions similar to intrahepatic cholestasis. Long-term and intermittent intoxication causes proliferation of bile ducts and consequently formation of connective tissue septs. That leads to a change in the architecture of the liver similar to primary biliary cirrhosis. These irreversible changes develop 6 to 8 months after the beginning of intoxication. Further development of the lesion is dependant on application of ANIT. There is no evidence of autonomous progression.

     

Alpha-naphthyl-isothiocyanate-induced cholestasis in the rat: studies of liver plasma membrane enzymes.

Oral administration of a single dose of alpha-naphthyl-isothiocyanate (ANIT) to rats produced a conjugated hyperbilirubinaemia, maximal at 2 days and which subsided by 7. The activities of 3 liver plasma membrane enzymes, Mg-2+-ATPase, (Na-+-K-+)-ATPase and 5-nucleotidase, and serum bilirubin levels were studied for up to 7 days after treatment. Activities of the 3 enzymes were significantly decreased at 2 days after treatment and returned to normal by 7, thus varying inversely with the degree of hyperbilirubinaemia. Enzyme histochemistry used to demonstrate canalicular localization of Mg-2+-ATPase in sections of whole liver and of isolated plasma membrane pellets showed that the reduction in activity was not a uniform partial loss, but represented a range of reductions in most canaliculi with a few retaining normal staining intensity. The results suggest that after ANIT intoxication there is a membrane lesion which may be responsible for the observed hyperbilirubinaemia due to the failure of secretion of biliary constituents into the canaliculus. However, more direct studies are necessary to determine whether any one of these enzymes is directly involved in the transport of biliary constituents across the bile canalicular membrane.     

Serum gamma glutamyl transferase as a specific indicator of bile duct lesions in the rat liver.

Serum gamma-glutamyl transferase (GGT), a marker of hepatic injury used extensively in humans, has been used rarely in rats because its specificity has not been previously defined. Studies were designed for investigation of the specificity of serum GGT activity with the use of cell type specific hepatotoxicants in Fischer 344 rats. Single necrogenic doses of CCl4, allyl alcohol (AA), and alpha-naphthylisothiocyanate (ANIT) were used to produce cell specific injury in centrilobular hepatocytes, periportal hepatocytes, and bile duct cells, respectively. Administration of CCl4 markedly increased serum activities of alanine aminotransferase (ALT), alkaline phosphatase (AP), and serum bile acid concentrations within 24 hours but had no effect on serum GGT activity. ANIT treatment increased serum GGT and AP activities and bile acid concentration 24 hours following administration. Allyl alcohol administration increased serum ALT activity but had no effect on GGT activity. Administration of ANIT in the diet at 0.01%, 0.022%, 0.047%, and 0.1% for 2, 4, and 6 weeks produced dose- and time-dependent increases in serum GGT activity which strongly correlated with quantitative increases in hepatic bile duct volume, which was determined morphometrically. These observations support the use of serum GGT activity in the rat as diagnostic of bile duct cell necrosis when increases are detected shortly after the insult and as an indicator of possible bile duct hyperplasia.     

The effects of cycloheximide, actinomycin D, and ethionine on the biliary excretion of labelled alpha-naphthylisothiocyanate in rats.

It has recently been shown that rats given a mixture of [³H-4-naphthyl] α-naphthylisothiocyanate (ANIT) and [¹⁴C-isothiocyanate] ANIT having a ${}^3\text{H}{}^{14}\text{C}$ ratio equal to 6.3, excreted more ³H than ¹⁴C into bile during the first 8 hours after ANIT administration, the ratio increasing to 7.5. However, in rats pretreated with cycloheximide, the ratio remained similar to that of the ANIT mixture administered (6.3) and data indicated that this inhibitor blocked this increase in ³H. Here we present further data showing that this effect by cycloheximide is dose-dependent, can be observed as late as 16 hours after ANIT administration, and is sensitive to early posttreatment times not to exceed 1 hour after ANIT. Actinomycin D and dl-ethionine also significantly blocked the increase in ${}^3\text{H}{}^{14}\text{C}$ ratio but it was found necessary to give both inhibitors 4 hours before ANIT administration in order to effect the decrease within 8 hours after ANIT administration. If given 30 minutes before ANIT, these inhibitors showed no effect until 9 hours after ANIT administration. These data further support the conclusion that cycloheximide, as well as actinomycin D and dl-ethionine, may act by inhibiting the formation of a toxic moiety of ANIT. The effects of these inhibitors on decreasing the excretion of a ³H moiety of ANIT in bile seem to coincide, on the basis of potency and protection, with their effects against hyperbilirubinemia and cholestasis.