Intravenous bilirubin infusion causes vacuolization of the cytoplasm of hepatocytes and canalicular cholestasis.

To examine whether intravenous bilirubin infusion causes cholestasis and impairs liver metabolism, bile secretion and ethanol clearance were measured in 34 anaesthetized pigs before and after intravenous infusion of 0.5 mumol kg-1 min-1 bilirubin for 4.5 hours. Bilirubin infusion increased plasma bilirubin to 556 +/- 76 mumol l-1 and hepatic tissue bilirubin to 3.5 +/- 1.3 mmol kg tissue weight-1. Bilirubin infusion depressed bilirubin secretion and net hepatic uptake of cholate and taurocholate, and caused a 86 +/- 6% reduction of cholate-induced bile secretion. Bilirubin caused formation of large cytoplasmic vacuoles in hepatocytes and dilatation of bile canaliculi. Ethanol clearance and secretin-dependent ductular bile secretion were unaffected by bilirubin. We conclude that intravenous infusion of unconjugated bilirubin causes accumulation of bilirubin in the liver, vacuolization of the hepatocyte cytoplasm and canalicular but not ductular cholestasis. The canalicular cholestasis is not due to impaired hepatic mitochondrial energy metabolism, but may be due to inhibition of a common pathway for lipid, bilirubin and bile salt secretion from hepatocytes.     

Down-regulation of OATP1B proteins correlates with hyperbilirubinemia in advanced cholestasis

Aim: Organic anion-transporting polypeptides OATP1B1 and OATP1B3 are sinusoidal membrane transporters mediating liver uptake of a wide range of substrates including conjugated and unconjugated bilirubin, xenobiotics and drugs. Absence of OATP1Bs in the liver causes Rotor syndrome. Our aim was to correlate OATP1B expression with hyperbilirubinemia in common liver diseases. Methods: Immunoreactivity of five antibodies against human OATP1Bs was tested on frozen and formalin-fixed paraffin-embedded liver tissue of mouse strains transgenic for SLCO1B1 or SLCO1B3 and on human specimens. The proportion of hepatocytes expressing OATP1Bs was then assessed immunohistologically in formalin-fixed paraffin-embedded liver samples obtained from patients with hepatocellular and primary biliary liver diseases. UGT1A1 promoter TATA-box and SLCO1B1 rs4149056 genotyping was performed to rule out individuals predisposed to hyperbilirubinemia. Results: The most specific detection of OATP1B3 was achieved with the H-52 (sc-98981) antibody. OATP1B1 was specifically recognized with the ESL (ab15441) anti-OATP1B1 antibody, but only in frozen sections. The MDQ (ab15442) anti-OATP1B1 antibody cross-reacted with both OATP1B proteins in liver tissue of the transgenic mouse strains. Expression of the OATP1B proteins was decreased in advanced liver diseases and inversely correlated with serum bilirubin levels. The reduction was more pronounced in advanced primary biliary diseases (1.9±1.1 vs. 2.7±0.6; P=0.009). Conclusions: Down-regulation of OATP1B proteins may contribute to pathogenesis of jaundice accompanying advanced cholestatic liver diseases.     

Orcein positive granules in the hepatocytes in chronic intrahepatic cholestasis

Summary The morphological characteristics of orcein positive granules in hepatocytes from 11 patients with chronic intrahepatic cholestasis and from 2 newborn normal livers were studied. Histochemical investigations revealed their protein nature and many sulphydryl and/or disulphide groups. Copper was demonstrated in the granules by histochemical techniques and electron X-ray micronalysis. No difference was observed in the hepatic distribution and appearance of the granules between the livers of those with chronic cholestasis and the newborn. Ultrastructurally, a variety of electron dense granules were seen at the site of orcein positive granules in the hepatocytes of the patient with primary biliary cirrhosis. Some had a single-layered membrane and seem to be lysosomal derivatives. It is suggested that the copper in lysosomes seen in both chronic cholestasis and normal newborn livers, need not to be cytotoxic.     

Heparin and mast cells in the course of experimental extrahepatic cholestasis

The blood heparin concentration and the number of mast cells in the tissues were studied in rats at various times after ligation of the common bile duct. The blood heparin level rose during cholestasis. The total number of mast cells was slightly reduced on the third day, increased on the seventh day, and back to normal on the 14th day. The ratio between granulated and degranulated forms of cells was sharply altered in favor of the latter. Changes in the number of mast cells and the increase in the number of degranulated forms are regarded as the result of irritation of mast cells by bile acids and pigments which accumulate in the body.Department of Pathological Physiology and Department of Pathological Anatomy, I. P. Pavlov First Leningrad Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR P. N. Veselkin.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 4, pp. 407–409, April, 1976.     

Increase of the total amount of conjugated and unconjugated bilirubin in the normothermic perfused isolated dog kidney system

Summary The metabolism of bilirubin was studied in 36 isolated perfused dog kidney systems. When the plasma haemoglobin binding capacity was not saturated, and even when it was so, the free plasma haemoglobin level being nevertheless lower than 50 mg/100 ml, there were no significant changes in the total amount of bilirubin in 7 male and 6 female isolated kidney perfusion systems during 7 h of perfusion. If the free plasma haemoglobin level was higher than 50 mg/100 ml during perfusion, there was a significant increase of 3.06±0.33 mg bilirubin in all 15 male kidney perfusion systems, and a significant rise of 2.92±1.07 mg bilirubin in 2 of the 11 female kidney perfusion systems. In the other 9 female systems, there was no significant change in the total amount of bilirubin after 6.5 h of perfusion. In the 15 male and the 2 female kidney perfusion systems there was an hourly production of 0.33±0.05 mg bilirubin as opposed to 0.03±0.01 mg in the other 3 groups. In the 15 males 74.0±5.2% of the total amount of conjugated bilirubin was excreted in the urine, in the 2 females only 10±0.7%. It was concluded that all male and a few female kidneys converted significant amounts of free haemoglobin into bilirubin. The male kidney excreted this conjugated bilirubin in significant amounts, the female kidney did not do so.     

血清胆红素在冠心病患者中的变化及意义

目的 探讨血清胆红素水平与冠心病的关系.方法 经冠状动脉造影确诊的冠心痛患者(冠心病组)92例和非冠心痛患者(对照组)69例,冠心病组又分为单支病变亚组(40例)、双支病变亚组(31例)和三支病变亚组(21例),测定血清总胆红素、直接胆红素和间接胆红素水平. 结果 冠心病组血清胆红素水平显著低于对照组(P＜0.01),双支病变亚组和三支病变亚组血清胆红素水平均显著低于单支病变亚组(P＜0.05,P＜0.01).结论 血清胆红素水平与冠心病密切相关,且随冠心痛病情严重程度而降低,低胆红素血症是冠心病的独立危险因素之一.     

Neonatal hyperbilirubinemia and a common mutation of the bilirubin uridine diphosphate-glucuronosyltransferase gene in Japanese

Neonatal hyperbilirubinemia, which is prevalent among Asian peoples, has been considered as a physiological phenomenon, and its metabolic basis has not been clearly explained. Gilbert syndrome is a common inherited disease of unconjugated hyperbilirubinemia due to decreased bilirubin uridine diphosphate-glucuronosyltransferase (B-UGT), and its role in neonatal jaundice has recently been considered. We have previously reported that the Gly71Arg mutation of the B-UGT gene associated with Gilbert syndrome is prevalent in Japanese, Korean, and Chinese populations and was more frequently detected in neonates with severe hyperbilirubinemia than in control subjects. We have studied 159 Japanese full-term neonates, evaluating the relationship between the B-UGT genotype and the severity of jaundice, as assessed with a transcutaneous bilirubinometer. The gene frequency of the Gly71Arg mutation in these neonates was 0.19, and neonates carrying the Gly71Arg mutation had significantly increased bilirubin levels on days 2–4, manifested in a gene dose-dependent manner. The frequency of the Gly71Arg mutation was 0.47 in the neonates who required phototherapy (i.e., those with more severe hyperbilirubinemia), significantly higher than 0.16 in the neonates who did not require the therapy. The gene frequency of the TA repeat promoter polymorphism, the (TA)₇ mutation, was 0.07, and neonates carrying this mutation did not have an increase in bilirubin. These results suggested that the Gly71Arg mutation contributes to the high incidence of neonatal hyperbilirubinemia in Japanese. Received: June 16, 1998 / Accepted: August 5, 1998     

Synergistic effects of polymer adsorbents on the performance of bilirubin hemoperfusion

Neonatal hyperbilirubinemia (jaundice) is a common disease with high incidence. Currently, the clinical inefficiency of adult bilirubin hemoperfusion medical adsorbent is a major technical barrier for the application of hemoperfusion treatment to rescue the severe neonatal jaundice. Based on the well-known principle of synergistic effects, a series of customized bilirubin polymeric compounds, comprised of one or more of the following components (glycidyl methacrylate, sodium acrylate, methacrylic acid isooctyl, hexamethylene diamine, albumin), were designed and fabricated based on molecular design. Their adsorption performances upon bilirubin were investigated and compared under the same conditions, and the compound with the highest adsorption performance was then subject to preliliminary safety assessments and compared with a commercial one (BS330). The results showed that positive synergistic effects appeared on the adsorption performance to adsorb bilirubin based on this study, and the one comprised of glycidyl methacrylate+sodium acrylate+methacrylic acid isoocty+hexamethylene diamine+albumin possesses the highest adsorption performance as well as outome clinical acceptable medical safety assessments, and its adsorption efficiency was up to 46% while the commerical one’s was about 26% under the same conditions. This study sheds a new light on how to design and develop hemoperfusion bilirubin adsorbents with good overall clinical performance, as well as providing a novel idea and experimental referrences for future related topics.     

Qualification for endoscopic retrograde cholangiopancreatography in the diagnosis and treatment of extrahepatic cholestasis caused by choledocholithiasis

Introduction

Choledocholithiasis, being the most common cause of extrahepatic cholestasis, is diagnosed on the basis of clinical symptoms, laboratory findings, and imaging results. An important diagnostic and also therapeutic procedure performed in patients with choledocholithiasis is retrograde cholangiopancreatography (ERCP). However, due to the high rate of complications associated with ERCP, the decision on its implementation should be preceded by a thorough analysis of the case, aimed at confirmation of the diagnosis.

Material and methods

The present study is a retrospective analysis of 86 patients qualified for ERCP due to suspected choledocholithiasis. The diagnosis was based on clinical symptoms and/or laboratory and/or imaging results. The presence of one or more of the three abovementioned criteria was a key to classify patients in one of three risk groups of choledocholithiasis: high, intermediate and low.

Results

In the high-risk group, where choledocholithiasis was confirmed by clinical symptoms, laboratory findings and results of imaging tests, the accuracy of diagnosis was 100%. In the intermediate-risk group – choledocholithiasis diagnosed on the basis of clinical symptoms and laboratory results – the accuracy of diagnosis was approximately 81.5% (p < 0.05). In the low-risk group, in which choledocholithiasis was diagnosed only on the basis of clinical symptoms, diagnosis accuracy was approximately 50% (p < 0.05).

Conclusions

The combination of clinical symptoms, laboratory tests and imaging findings allows patients to be classified into three risk groups of choledocholithiasis: high, intermediate and low. Taking into account the probability of choledocholithiasis and the risk of ERCP complications, only patients from high and intermediate risk groups should be qualified for the procedure.     

Some molecular mechanisms of the antioxidative action of dalargin on the liver in experimental cholestasis

Clinical Biochemical Laboratory and Department of Anesthesiology and Resuscitation, A. V. Vishnevskii Institute of Surgery, Academy of Medical Sciences of the USSR, Moscow (Presented by Academician of the Academy of Medical Sciences of the USSR V. D. Fedorov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 113, No. 1, pp. 38–40, January, 1992.     

Immunological basis in the pathogenesis of intrahepatic cholestasis of pregnancy

Intrahepatic cholestasis of pregnancy poses a great risk to both maternal and fetal health. Despite extensive research, much of the pathogenesis of this disorder is unknown. The increase in bile acids observed in patients with intrahepatic cholestasis of pregnancy has been noted to cause a change in the immune system from the normally mediated TH2 response to one that is more oriented towards TH1. In this literature review, we have critically reviewed the current literature regarding the changes in the immune system and the potential effects of immunological changes in the management of the patient. The current treatment, ursodeoxycholic acid, is also discussed along with potential combination therapies and future directions for research.     

妊娠期肝内胆汁淤积症合并乙型肝炎病毒感染对母婴结局的影响

目的探讨妊娠期肝内胆汁淤积症(ICP)合并乙型肝炎病毒感染的妊娠结局。方法回顾性总结2006年1月-2007年12月陕西省妇幼保健院86例ICP合并乙型肝炎病毒感染者(研究组),同期住院治疗的单纯ICP孕妇118例(对照组)的妊娠结局,比较两组的早产率、新生儿窒息率、产后出血率等并发症的发生情况。结果妊娠期肝内胆汁淤积症合并乙型肝炎病毒感染各并发症的发生情况:早产76例、妊娠期高血压疾病17例、产后出血8例。与对照组并发症的发生差异有显著性(P<0.05)。结论妊娠期肝内胆汁淤积症合并乙型肝炎病毒感染对母婴结局产生更严重的影响。加强母婴监护,药物治疗同时适时终止妊娠可有效改善妊娠结局。     

妊娠期肝内胆汁淤积症中ABCB4基因外显子12突变的研究

目的探讨ABCB4基因外显子12的突变与妊娠期肝内胆汁淤积症（ICP）发病的关系。方法从31名ICP患者外周血标本中提取出基因组DNA,聚合酶链反应（PCR）扩增ABCB4基因外显子12,PCR产物经DNA序列测定检测突变情况。结果31名ICP患者均扩增出ABCB4基因外显子12的靶基因片断,未见外显子12的缺失,随机挑选20例标本测定外显子12的DNA序列,未发现突变。结论ABCB4基因外显子12的突变与中国皖南地区的ICP发生无关或关联很小,皖南地区ICP患者中可能存在其他的ABCB4基因突变热点。ABCB4基因的突变与中国人ICP发病的相关性仍应进行更大样本量的研究和更多外显子的筛查。     

Role of estrogens in regulation of the bile acid composition of the enterohepatic system of rabbits and monkeys with extrahepatic cholestasis

Laboratory of Biochemistry, Research Institute of Pediatrics, Ministry of Health of the RSFSR, Nizhnii Novgorod Laboratory of Biochemical Endocrinology, All-Union Endocrinologic Scientific Center, Academy of Medical Sciences of the USSR, Moscow (Presented by Academician of the Academy of Medical Sciences of the USSR Yu. A. Pankov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 111, No. 6, pp. 599–601, June, 1991.     

Activity of lysosomal enzymes in the bile and serum of mice with intrahepatic cholestasis

Lysosomal enzyme activity in the bile and blood serum was compared in mice with experimental intrahepatic cholestasis induced by α-naphthyl isothiocyanate and Triton WR 1339. Triton WR 1339 increases the synthesis of cholesterol (fatty acid precursor) in liver cells. The development of intrahepatic cholestasis was confirmed by the increase in activities of alkaline phosphatase and γ-glutamyltransferase in blood serum. Administration of Triton WR 1339 in a dose of 100 mg/100 g was followed by a 10-fold increase in β-galactosidase activity (hepatocyte lysosomal enzyme) in the bile, but not in the serum of mice. β-Galactosidase activity significantly increased in the bile, but decreased in the serum of mice after treatment with α-naphthyl isothiocyanate in a dose of 200 mg/kg. Our results indicate that intrahepatic cholestasis is manifested in increased secretion of lysosomal glycosidases into the bile. Bile components can aggravate damage to liver cells by affecting the processes of hepatocyte apoptosis and necrosis. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 145, No. 5, pp. 496–499, May, 2008     

孕鼠肝内胆汁淤积症对母鼠和胎鼠心脏的影响

目的探讨孕鼠肝内胆汁淤积症对母鼠和胎鼠心脏的影响。方法应用雌、孕激素建立妊娠肝内胆汁淤积大鼠模型,光镜和电镜观察母鼠和胎鼠心脏的病理改变。结果（1）胆淤组和对照组比较,孕鼠血清丙氨酸转氨酶（ALT）、门冬氨酸转氨酶（AST）、总胆汁酸（TBA）的差异有显著性（P〈0．01）。（2）胆淤组孕鼠肝脏光镜下见部分肝细胞有颗粒样变性和空泡变性,电镜下见肝组织中毛细胆管扩张,毛细胆管及肝细胞内见高电子致密物沉积。（3）胆淤组孕鼠胎盘光镜下见部分滋养细胞颗粒样变性和空泡变性。（4）胆淤组孕鼠心脏光镜下见心肌组织中局灶性心肌细胞颗粒样变性。（5）胆淤组胎鼠心脏光镜下见心肌组织中心肌细胞广泛空泡变性;电镜下见胎鼠心肌细胞内高电子致密物沉积,部分肌丝断裂,肌节模糊,细胞内线粒体水肿,脱髓鞘样改变。结论孕鼠肝内胆汁淤积症时高胆汁酸血症对母鼠和胎鼠心肌细胞均有明显的毒性作用,尤其以胎鼠心肌细胞病变更为严重。     

胆红素对大鼠急性肺损伤形成过程中氧自由基以及caspase-3表达的影响

目的： 探讨胆红素对抗急性肺损伤（ALI）形成的可能机制。方法: 健康雌性Wistar大鼠(190-210 g) 30只，随机分为生理盐水对照组、脂多糖（LPS）致ALI模型组、胆红素干预组。检测肺组织匀浆中羟自由基（OH^-）、过氧化氢(H₂O₂)和超氧阴离子自由基（O₂^·）含量以及肺组织中caspase-3表达的变化。结果: ①ALI模型组肺组织匀浆OH^-、H₂O₂、O₂^·含量及肺组织中caspase-3表达显著高于生理盐水对照组（均P<0.05）。②胆红素干预组肺组织匀浆OH^-、H₂O₂、O₂^·及肺组织中caspase-3表达明显高于ALI正常大鼠（均P<0.05），但少于ALI模型组（均P<0.05）。结论: ①胆红素能在一定程度上减少肺内凋亡细胞数量。②胆红素能减少ALI大鼠肺组织OH^-、H₂O₂、O₂^·水平。③ Caspase-3表达的变化有促脂多糖性肺损伤细胞凋亡作用。     

Isolation and primary culture of adult pig hepatocytes

Hepatocytes were directly isolated from pig livers obtained from the abattoir and cultivated on a type I collagen-coated polystyrene surface. Digestion of the partial liver lobe resulted in an average yield of 1.4 × 109 cells (9.9 × 106 cells/g liver) with an average viability of 92.5 percent. The yield and viability of cells were improved by dispase/collagenase digestion. The emergence of blebbing cells was blocked by supplying oxygen to the cell isolation buffers. Hepatocytes isolated seeded onto the polystyrene surfaces remained viable and functional at a comparable level to that of rat hepatocytes, though their functions decreased over time. These data indicate that adult pig hepatocytes can be harvested with high yields and can retain viability and differentiated functions through this method.     

地塞米松对乙炔雌二醇诱发的肝内胆汁淤积症孕鼠儿茶酚氧位甲基转移酶活性、胎盘和肝脏组织雌激素受体表达的影响

目的：研究地塞米松对雌激素诱发的肝内胆汁淤积症孕鼠儿茶酚氧位甲基转移酶（catechol-O-methyl-transferase,COMT）活性及肝脏和胎盘雌激素受体（estrogen receptor,ER）表达的影响,探讨地塞米松治疗妊娠期肝内胆汁淤积症（intrahepatic cholestasis of pregnancy,ICP）的作用机制。方法：选择SD孕鼠随机分为ICP对照组（A组）、ICP地塞米松干预组（B组）、非ICP对照组（C组）、非ICP地塞米松干预组（D组）。4组孕鼠均于见到阴道血性分泌物后剖腹取胎,计量胎鼠身长、体质量,计算死胎数量。光镜下观察各组孕鼠胎盘和肝脏组织病理学改变,并采用免疫组织化学法检测其ER的表达;采用高效液相色谱法检测各组孕鼠COMT的活性浓度;采用放射免疫法检测孕鼠血清游离雌三醇（unconjugated estriol,uE3）浓度。结果：（1）4组胎鼠中,死胎率A组高于B组和C组,差异有统计学意义（P〈0.01）。（2）A组胎鼠身长和体质量低于B组和C组,差异有统计学意义（P〈0.01）,D组显著低于C组（P〈0.01）。（3）4组孕鼠肝脏和胎盘组织病变积分结果比较,A组显著高于B组和C组,差异有统计学意义（P〈0.01）,C组与D组比较,差异无统计学意义（P〉0.05）。（4）孕鼠胎盘组织中ER主要分布于细胞浆内,少数在细胞核内表达,肝脏组织中ER主要分布在细胞膜上。4组孕鼠ER表达的平均光密度值比较,A组高于B组和C组,D组低于C组,差异均有统计学意义（均P〈0.01）。（5）孕鼠红细胞COMT活性浓度比较,B组和C组均高于A组,D组高于C组,差异均有统计学意义（均P〈0.01）。（6）孕鼠血清uE3浓度比较,B组和C组均低于A组,D组低于C组,差异均有统计学意义（均P〈0.01）。（7）COMT活性浓度与uE3水平呈负相关（r=-0.381,P〈0.05）。结论：地塞米松对雌激素诱发的肝内胆汁淤积的孕鼠具有保护作用,并能改善胎鼠预后,其机制与对孕鼠胎盘及肝脏细胞ER的表达及肝脏COMT活性浓度的调节作用有关。