Biomedical subjects

G Feldmann

Publications and source records attributed to G Feldmann.

At least 73 records · Page 4Linked to original sources

PubMed1997

Effects of simvastatin, pentoxifylline and spironolactone on hepatic fibrosis and portal hypertension in rats with bile duct ligation.

AIMS/METHODS: Our aim was to study the antifibrotic and hemodynamic effects of simvastatin (SMV), pentoxifylline (PTX) and spironolactone (SPN), three drugs which may have antifibrotic and/or portal hypotensive properties, in a model of hepatic fibrosis and portal hypertension induced in rats by bile duct ligation. A blind study was performed in five groups of 53 Sprague-Dawley rats: sham, placebo (PL), SMV (2.5 mg x kg(-1) x J(-1)), PTX (50 mg x kg(-1) x J(-1)) and SPN (100 mg x kg(-1) x J(-1)). Drugs were administered by daily gavage over a 4-week period as soon as bile duct ligation was performed. At day 28, the following parameters were evaluated: area of hepatic fibrosis by image analysis after staining collagen with picrosirius and plasma concentrations of hyaluronate, splanchnic and systemic hemodynamics (radiolabeled microspheres). RESULTS: Portal venous pressure (PL: 15.5+/-1.5, SMV: 15.8+/-2.5, PTX: 15.9+/-1.8, SPN: 13.5+/-2.1 mmHg, p<0.05) and porto-systemic shunts (PL: 30+/-31, SMV: 18+/-27, PTX: 25+/-24, SPN: 5+/-4%, p<0.05) were significantly reduced in the SPN group; other hemodynamic parameters were not significantly altered. There was a significant correlation between portosystemic shunts and portal pressure (r(s)=0.47, p<0.01). The area of fibrosis was not significantly different among the four groups of bile duct ligated rats (PL: 8.7+/-3.9, SMV: 7.1+/-3.6, PTX: 7.8+/-2.7, SPN: 6.6+/-3.3%) but was higher than in sham rats (1.5+/-0.5%, p<0.001). Hyaluronate was significantly higher in bile duct ligated rats (from 374+/-162 to 420+/-131 microg/l, among the four groups) than in sham rats (52+/-16 microg/l, p<0.0001). CONCLUSIONS: In this model, none of the drugs prevented hepatic fibrosis. On the other hand, spironolactone decreased portal pressure and prevented porto-systemic shunts. Therefore, this drug may have beneficial effects in patients with early portal hypertension.

Animals↗

PubMed1997

Liver apoptosis.

Apoptosis, also called programmed cell death, is a peculiar form of cell death different from cell necrosis in many morphological and biochemical aspects. Like mitosis or differentiation, apoptosis is a normal cell phenomenon which depends on the expression of genes capable of inducing or inhibiting this type of cell destruction. But apoptosis can also be triggered by many external factors and has been described in many diseases. The very different conditions where programmed cell death occurs suggest that the mechanisms leading to the activation of apoptosis-controlling genes are variable. As in other cells, apoptosis occurs in the liver cells, first in the normal state during liver development and then in the adult liver, respectively for liver organogenesis and the renewal of hepatocytes. But apoptosis is also present in various viral, immunological, malignant or drug-induced human liver diseases. In addition, in the animal, hepatocyte apoptosis can be triggered either in vivo or in vitro by many toxic agents. In contrast to other cells, the mechanisms leading to liver cell apoptosis remain poorly investigated. However, two proteins could play an important role in this field, the fas/apo-1 protein present at the surface of hepatocytes and the bcl-2 protein localized in biliary cells. Analysis of the genes controlling the expression of these two proteins could provide essential information on the mechanisms of liver apoptosis.

Animals↗

PubMed1997

Complement activation after ischemia-reperfusion in human liver allografts: incidence and pathophysiological relevance.

BACKGROUND & AIMS: Little is known about the occurrence and possible consequences of local complement activation in human liver transplantation. The aim of this study was to search for signs of complement activation in human liver allografts and evaluate their relationship to cell injury and leukocyte sequestration. METHODS: In 33 postreperfusion biopsy specimens, 9 preoperative specimens, and 10 intraoperative specimens, the cytolytic membrane attack complex of complement was localized, expression of complement inhibitors was evaluated, and intragraft accumulation of leukocytes and platelets was quantitated. RESULTS: In control samples and preoperative biopsy specimens, the membrane attack complex was detected only in extracellular deposits, associated with its soluble inhibitors clusterin and vitronectin. Comparable observations were performed in 14 postoperative specimens. In the remaining 19 postoperative specimens, membrane attack complex decorated a variable proportion of hepatocytes. The extent of membrane attack complex deposition correlated with the increase in postoperative aspartate aminotransferase levels in serum (P = 0.002) and the decrease in postoperative factor V levels in serum (P = 0.002). It also correlated with the number of leukocytes and platelets accumulating within the graft (P < or = 0.001). CONCLUSIONS: Local complement activation is frequent during human liver transplantation and probably contributes to cell injury and leukocyte sequestration in the allograft.

Adult↗

PubMed1997

Diterpenoids from germander, an herbal medicine, induce apoptosis in isolated rat hepatocytes.

BACKGROUND & AIMS: Germander was withdrawn from the market after its use for weight control caused an epidemic of hepatitis. Its toxicity was shown to be caused by diterpenoids and their cytochrome P4503A-mediated metabolic activation into electrophilic metabolites that deplete cellular thiols. The aim of the present study was to determine the mechanisms of cell death. METHODS: Isolated rat hepatocytes were incubated for 2 hours with germander diterpenoids (100 micrograms/mL). RESULTS: Diterpenoids decreased cell glutathione, increased cytosolic [Ca2+], activated Ca(2+)-dependent tissue transglutaminase forming a cross-linked protein scaffold, and caused internucleosomal DNA fragmentation and the ultrastructural features of apoptosis. Cell death was prevented by decreasing metabolic activation (with troleandomycin), preventing depletion of glutathione (with cystine), blocking activation of Ca(2+)-modulated enzymes (with calmidazolium), or inhibiting internucleosomal DNA fragmentation (with aurintricarboxylic acid). Apoptosis was increased and diterpenoids caused overexpression of p53 and interleukin 1 beta-converting enzyme in rats treated with dexamethasone (cytochrome P4503A inducer). Apoptosis was also increased by a diet deficient in sulfur amino acids. CONCLUSIONS: The germander furano diterpenoids cause apoptosis within 2 hours in isolated rat hepatocytes. Electrophilic metabolites may stimulate apoptosis by decreasing thiols, increasing [Ca2+], and activating Ca(2+)-dependent transglutaminase and endonucleases.

Amino Acids, Sulfur↗

PubMed1997

An improved digitonin-collagenase perfusion technique for the isolation of periportal and perivenous hepatocytes from a single rat liver: physiological implications for lobular heterogeneity.

Morphological and functional heterogeneity of hepatocytes according to their position in the liver lobule has been known for many years. The digitonin-collagenase perfusion technique is widely used to study hepatocyte heterogeneity and has yielded reliable data. However, with this procedure, periportal (PP) or perivenous (PV) hepatocytes are isolated from different livers, allowing only comparison between cell populations issued from two separate animals. To overcome this drawback, we have modified this technique by perfusing the two main rat liver lobes of a single animal in succession. The procedure involved alternate clamping of the median and the left lateral lobes, restricting digitonin infusion to one lobe via the portal vein, and to the other via the caudal vena cava. Lobe exclusion during digitonin perfusion, and zonal restriction of digitonin-induced damage, were monitored using macroscopic and histological controls. We compared our results with previous data on PP and PV hepatocytes issued from two different livers using the conventional digitonin-collagenase perfusion technique. First, we found that the cellular sensitivity to angiotensin II, a calcium-mobilizing agonist, was 60% to 80% higher in PV than in PP hepatocytes, whereas, previously, no difference had been recorded. Second, we found that albumin messenger RNAs (mRNAs) were 35% more abundant in PP than in PV hepatocytes, whereas, previously, larger differences had been reported. Our results show that PP and PV hepatocytes may be isolated from a single liver using an improved digitonin-collagenase perfusion technique. Furthermore, we suggest that zonal differences can be artificially masked or amplified when comparing PP and PV cell populations from two different livers, indicating that it is preferable to use a single liver for accurate zonal comparisons between hepatocytes.

Albumins↗

PubMed1997

Glucocorticoids inhibit mitochondrial matrix acyl-CoA dehydrogenases and fatty acid beta-oxidation.

Glucocorticoid administration may produce fatty liver in humans. We investigated the effects of dexamethasone on hepatic mitochondria and lipid metabolism in mice. Dexamethasone 21-phosphate (20 microM) did not inhibit the mitochondrial inner membrane-bound very-long-chain acyl-CoA dehydrogenase but inhibited the matrixlocated long-, medium-, and short-chain dehydrogenases. Dexamethasone 21-phosphate (20 microM) inhibited the first beta-oxidation cycle of [1-(14C)]butyric acid and [1-(14C)]octanoic acid but not that of [1-(14C)]palmitic acid. Administration of dexamethasone 21-phosphate (100 mg/kg) decreased the in vivo oxidation of [1-(14C)]butyric acid and [1-(14C)]octanoic acid into [14C]CO2 but not that of [1-(14C)]palmitic acid and decreased the hepatic secretion of triglycerides. After 5 days of treatment (100 mg/kg daily), hepatic triglycerides were increased and both microvesicular steatosis and ultrastructural mitochondrial lesions were present. In conclusion, glucocorticoids inhibit medium- and short-chain acyl-CoA dehydrogenation and hepatic lipid secretion in mice. These effects may account for their steatogenic effects in humans.

Acyl-CoA Dehydrogenases↗

PubMed1996

Inhibition of liver RNA breakdown during acute inflammation in the rat.

Liver RNA- and protein-degradation rates were measured after the induction of acute inflammation in the rat. A preliminary study determined changes in hepatic RNA and protein content 12, 18 and 24 h after a turpentine oil injection. The RNA content in turpentine-treated rats compared with pair-fed animals increased significantly and sharply from 12 h (+ 11%) to 18 h (+ 32%) and slightly thereafter (+ 37% at 24 h). The liver protein content was significantly enhanced only at 24 h (+ 11%) in response to inflammation. RNA-degradation rates were determined in livers perfused cyclically in situ for 15 min by measuring the accumulation of radioactive cytidine in the medium 60 h after in vivo labelling of RNA by [5-3H]cytidine instead of [6-14C]orotic acid, the most commonly used radioactive marker. Several validation procedures showed that the method employed was a valid alternative to the use of radioactive orotic acid. RNA-degradation rates, which mainly reflect rRNA breakdown, were significantly lower in the turpentine-treated rats than in respective pair-fed animals at 18 and 24 h (57 and 45% decrease respectively). Proteolysis rates measured at 24 h together with RNA breakdown by valine accumulation in the perfusion medium were not modified after turpentine treatment. The main factors known to regulate RNA degradation (amino acids, insulin/glucagon ratio) were measured in the portal blood 24 h after induction of acute inflammation. Of the known regulatory amino acids, only glutamine and to a lesser extent methionine were increased in the turpentine-treated rats as compared with their pair-fed counterparts. The insulin/glucagon molar ratio was similar in both groups. In conclusion, the reduced breakdown of RNA, especially rRNA, is largely responsible for the accumulation of hepatic RNA during acute inflammation. This inhibition of RNA degradation could possibly be related to the increase in glutamine.

Amino Acids↗

PubMed1996

Structural and functional differentiation of sinusoidal endothelial cells during liver organogenesis in humans.

During fetal life, human liver sinusoids, which differentiate between 4 and 12 weeks of gestation from capillaries of the septum transversum, must support an important hematopoietic function and acquire the structural and functional characteristics of adult sinusoids. To gain insight into their differentiation process, we studied the expression of (1) markers of continuous endothelia, absent from adult sinusoidal endothelial cells (PECAM-1, CD34, and 1F10); (2) functional markers of adult sinusoidal endothelial calls (CD4, 1CAM-1, CD32, and CD14); and (3) extracellular matrix components (laminin, tenascin, fibronectin, and thrombospondin) in 37 fetuses of different gestational ages. We identified two successive differentiation events. (1) An early structural differentiation, occurring from 5 to 12 weeks of gestation, was characterized by the loss of continuous endothelial cell markers and a reduction in the perisinusoidal amount of laminin and in the deposition of tenascin, fibronectin, and thrombospondin; at the end of this process, fetal liver sinusoids present structural characteristics comparable to those of the sinuses in adult hematopoietic bone marrow. (2) A later functional differentiation was characterized by the acquisition of the markers of adult sinusoidal endothelial cells, initiating at 10 weeks of gestation and completed by 20 weeks of gestation; this process likely contributes to adapt liver sinusoids to the specific functions of the adult hepatic tissue.

Adult↗

PubMed1996

Interphase cytogenetic studies of human hepatocellular carcinomas by fluorescent in situ hybridization.

Although numerous allelic chromosome losses have been reported in hepatocellular carcinomas (HCC), chromosome analysis by cytogenetic methods has rarely been performed in these tumors, unlike other solid malignant tumors. The purpose of the current study was to analyze primary liver tumors by conventional cytogenetic methods and by a new molecular cytogenetic technique, called fluorescent in situ hybridization (FISH), a technique that has been recently proposed to count the number of chromosome copies in interphase nuclei with chromosome centromeric probes. Primary cultures of tumoral cells were prepared to obtain metaphases. Specific chromosomes probes 7, 17, and 20 were used to perform in situ hybridization on isolated intact tumoral cells. Seven cases of primary liver tumors (six cases of HCC and one case of benign focal hepatic nodular hyperplasia) were investigated. A few metaphases were obtained in five of the seven tumors, and in most cases numerical abnormalities were difficult to interpret. In contrast with in situ hybridization, all cases of HCC showed losses and/or gains of chromosomes. Loss of one to three chromosomes occurred in five tumors. A gain of two chromosomes was observed in two of these five tumors. In only one case, a gain of only three chromosomes occurred. In addition, a loss of chromosome 17 was recorded for the benign tumor. These results demonstrate that FISH with specific probes can provide information on chromosome number in the tumoral cells of primary liver tumors even in the absence of analyzable metaphases. This technique opens new possibilities for the investigation of chromosome abnormalities in HCC.

Adult↗

PubMed1996

Fibrolamellar carcinoma of the liver: composition of the extracellular matrix and expression of cell-matrix and cell-cell adhesion molecules.

We have analyzed the composition of the tumor stroma and the expression of cell-matrix and cell-cell adhesion molecules in 11 cases of fibrolamellar carcinoma of the liver (FLC), in comparison with 34 cases of hepatocellular carcinoma and 8 cases of focal nodular hyperplasia. Fibrolamellar carcinoma was characterized by the presence of large amounts of tenascin in tumor stroma and by the scarce expression of basement membrane components at the contact of neoplastic clusters. Like normal hepatocytes, neoplastic cells constantly expressed the alpha1 integrin chain, lacked the beta4 integrin chain, and coexpressed E-cadherin and the hepatocyte N-related cadherin. Abnormalities in the expression of cell adhesion molecules, including altered cadherin expression, alphaV integrin chain induction, and CD44 expression, were detected in the majority of cases. The composition of the tumor stroma and the pattern of expression of cell adhesion molecules in fibrolamellar carcinoma were reminiscent of those observed in grade III and grade IV hepatocellular carcinomas. Our results therefore show that, despite its slow local growth and good prognosis, fibrolamellar carcinoma expresses many characteristics usually associated with clinically aggressive malignancies. Further studies are needed to identify the factors responsible for the apparent dissociation between clinical behavior and biological characteristics in this tumor.

Adult↗

PubMed1996

Coexpression of periportal and perivenous enzymes in rat hepatocytes after experimental bile duct ligation: comparison with intrasplenically transplanted hepatocytes.

The coexpression of normally periportal and perivenous markers has been described in heterotopically transplanted hepatocytes. To determine whether such a coexpression might also occur in hepatocytes retaining their original intrahepatic location, we compared in bile-duct-ligated livers and intrasplenically transplanted hepatocytes, the expression and distribution of the predominantly periportal glucose-phosphatase, succinate dehydrogenase, and lactate dehydrogenase, the predominantly perivenous glutamate dehydrogenase, NADPH-dehydrogenase, and beta-hydroxybutyrate dehydrogenase, and the strictly perivenous glutamine synthetase. The coexpression of high levels of the two periportal markers glucose-6-phosphatase and lactate dehydrogenase and of the perivenous marker NADPH dehydrogenase was observed in two situations: in clusters of hepatocytes isolated within the ductular proliferation in bile-duct-ligated livers and the majority of intrasplenically transplanted hepatocytes. The expression of glutamine synthetase was different according to the site. The protein was observed in certain intrasplenically transplanted hepatocytes bordering the splenic vessels but was never detected in hepatocyte clusters found in bile-duct-ligated livers. Our study therefore suggests that the coexpression of periportal and perivenous markers in the same hepatocytes is likely to be a non-specific consequence of the loss of the normal connections of hepatocytes with the normal liver microcirculation.

Animals↗

PubMed1996

Effects of insulin, dexamethasone and cytokines on alpha 1-acid glycoprotein gene expression in primary cultures of normal rat hepatocytes.

While the effects of insulin, dexamethasone and cytokines on alpha 1-acid glycoprotein gene expression have been investigated in various hepatoma cell lines, the individual and combined effects of these components on the expression of this gene have been rarely studied in cultured normal rat hepatocytes. In this cell model, we have shown that mRNA levels of alpha 1-acid glycoprotein were not decreased at least during the first 24 h of culture under basal conditions. During these short-term cultures, the expression of alpha 1-acid glycoprotein in normal hepatocytes showed a high degree of responsiveness to dexamethasone alone (20-fold increase) and to dexamethasone associated with various cytokines (interleukin-1 beta, interleukin-6 and tumor necrosis factor alpha) with a 40 to 100-fold increase depending on the cytokine. Insulin alone did not modify alpha 1-acid glycoprotein mRNA; however, this hormone exerted a positive effect (about 50% increase) in the presence of dexamethasone or dexamethasone with cytokines. These results indicate that the regulation of alpha 1-acid glycoprotein in cultured normal rat hepatocytes presents major differences when compared to reported observations in rat hepatoma cell lines.

Albumins↗

PubMed1996

Expression of cadherins and alpha-catenin in primary epithelial tumors of the liver.

BACKGROUND & AIMS: Cadherins and their associated molecules, such as alpha-catenin, have been shown recently to play a pivotal role in epithelial carcinogenesis. METHODS: The expression of E-cadherin, N-cadherin, and alpha-catenin in 10 normal samples, 28 focal nodular hyperplasias, 9 liver cell adenomas, 65 hepatocellular carcinomas, and 9 cholangiocarcinomas was studied by immunohistochemistry and Western blotting. RESULTS: In the normal liver, hepatocytes expressed E-cadherin and a 129-kilodalton cadherin identified by the anti-N-cadherin antibody GC4. The expression level of alpha-catenin was low. Bile duct cells expressed only E-cadherin and showed high levels of alpha-catenin. The expression of cadherins and alpha-catenin was preserved in focal nodular hyperplasia. In liver cell adenomas, cadherins and alpha-catenin were heterogeneously expressed. In hepatocellular carcinomas, cadherin and alpha-catenin expression was frequently reduced or absent. Alterations in cadherin expression correlated with large tumor size, low grade of histological differentiation, and occurrence of capsular and vascular invasion. In cholangiocarcinomas, neoplastic cells inconstantly expressed E-cadherin and alpha-catenin. CONCLUSIONS: Alterations of cadherin and alpha-catenin expression are frequent in liver cell adenomas and primary liver carcinomas. Their incidence in hepatocellular carcinomas is of prognostic significance.

Adenoma, Liver Cell↗

PubMed1996

Uncoupling of rat and human mitochondria: a possible explanation for tacrine-induced liver dysfunction.

BACKGROUND & AIMS: Tacrine administration (1-3 mg/kg) may lead to sinusoidal concentrations in the micromolar range and produce liver dysfunction in 50% of recipients. The aim of this study was to determine the cellular effects of tacrine that account for liver dysfunction. METHODS: The effects of tacrine on mitochondrial function were determined in isolated rat liver mitochondria, cultured rat hepatocytes, and isolated human lymphocytes. RESULTS: In vitro, tacrine was taken up by rat liver mitochondria, decreased their membrane potential, and stimulated their respiration. Ex vivo, respiration was increased in rat mitochondria isolated 30 minutes after the administration of 2 mg of tacrine per kilogram. After 7 days of culture, tacrine (2.5 mumol/L) decreased rat hepatocyte adenosine triphosphate levels. Ten micromolar decreased 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium reduction and neutral red uptake without modifying cell glutathione, the morphology of the endoplasmic reticulum, or protein synthesis. Tacrine (1.25 mumol/L) decreased levels of adenosine triphosphate in human lymphocytes. CONCLUSIONS: The weak base tacrine exerts a protonophoric effect in mitochondria that wastes energy and decreases levels of adenosine triphosphate in rat and human cells. These effects are visible after clinically relevant doses of tacrine and might be involved in tacrine-induced liver dysfunction.

Animals↗

PubMed1996

Retrograde injections of formaldehyde into the biliary tree induce alterations of biliary epithelial function in rats.

Formaldehyde may induce severe lesions of intrahepatic and extrahepatic bile ducts. The purpose of this study was to examine in vivo the functional consequences of an alteration of the biliary epithelium induced by a retrograde intrabiliary injection of formaldehyde in rats. After basal bile collection, a 10% formaldehyde solution was injected into the biliary tree of anesthetized rats, and the cannula was occluded for 30 minutes. Choleresis was then reestablished, and bile flow, bile acid, and bicarbonate secretion were measured both spontaneously and during ursodeoxycholate infusions. Formaldehyde injections induced a significant increase in bile flow and a marked inhibition of ursodeoxycholate-induced increase in biliary bicarbonate concentration and secretion. Biliary glucose secretion, which is normally very low, was increased about 20-fold in animals injected with formaldehyde. Histological and ultrastructural examination of the liver showed alterations of biliary epithelial cells, whereas hepatocytes, bile canaliculi, and canalicular tight junctions remained normal. Hepatocytic excretory function, as assessed by biliary secretion of bile acids, was not affected. It was concluded that short-term formaldehyde intrabiliary injections cause an inhibition of ursodeoxycholate-induced hypersecretion of bicarbonate, an increase in biliary glucose secretion, and selective structural alterations of biliary epithelial cells. These results suggest that formaldehyde retrograde biliary injection may be a useful model to study alterations of biliary epithelial function in vivo.

Animals↗

PubMed1996

Effect of diethylmaleate on bile secretion and ultrastructural appearance of hepatocytes in normal rats and mutant rats with defective organic anion secretion.

Diethylmaleate is an organic anion secreted into bile as a glutathione conjugate. Its transport by the hepatocyte is associated with dilatation of the Golgi apparatus and the appearance of small vesicles in the pericanalicular area. It has been speculated that the Golgi apparatus could play a role in the intracellular transport and/or the biliary canalicular secretion of diethylmaleate. The purpose of this work was to determine whether the alterations in the Golgi apparatus and the pericanalicular vesicles could mediate the canalicular secretion of diethylmaleate. Diethylmaleate biliary secretion and diethylmaleate-induced bile flow were measured in Sprague-Dawley rats, and in TR- rats which have an inherited defect in the excretion into bile of organic anions, including glutathione conjugates. Livers of both Sprague-Dawley and TR-rats were examined by electron microscopy, to characterize the changes in intracellular organelles. In Sprague-Dawley rats, as previously described, diethylmaleate administration was associated with an increase in bile flow, which was parallel in time to the secretion into bile of diethylmaleate conjugates. Electron microscopic examination of the liver after diethylmaleate administration showed dilatation of the Golgi saccules. In contrast, in TR- rats, the increase in bile flow and the secretion of diethylmaleate conjugated were nearly absent. Nevertheless, electron microscopic examination showed a dilatation of the Golgi saccules similar to that observed in Sprague-Dawley rats. TR- rats, in addition to the changes in the Golgi apparatus, had marked dilatation of the endoplasmic reticulum. These results show that biliary secretion of diethylmaleate conjugates was severely impaired in TR- rats, in spite of a dilatation of the Golgi apparatus and of the endoplasmic reticulum. We conclude that it is unlikely that the alterations in the Golgi apparatus (and the endoplasmic reticulum) induced by diethylmaleate play a role in the canalicular secretion of diethylmaleate. We do not exclude the possibility that these organelles could play a role in intracellular transport of this compound. Alternatively, these alterations could be due to a "toxic" effect of diethylmaleate accumulation in hepatocytes.

Animals↗

PubMed1996

Cellular and subcellular localization of acetaldehyde-protein adducts in liver biopsies from alcoholic patients.

Acetaldehyde, the first product of ethanol in hepatocytes, can react with protein to form acetaldehyde-protein adducts (APAs). Because it has been suggested that these adducts could be involved in the pathogenesis of ethanol-induced hepatic lesions and in fibrogenesis, we performed an ultrastructural immunohistochemical study to precisely define the cellular and subcellular localization of APAs. A preembedding technique of indirect immunoperoxidase was performed in liver biopsy specimens from eight patients with alcoholic liver disease, using a specific antiserum against APAs. In all specimens, APAs were detected in the rough endoplasmic reticulum, in some peroxisomes, and in the cytosol of hepatocytes. In four patients with steatofibrosis or cirrhosis, labeling of Ito cells was also observed. In these cases, the same staining pattern was observed in the cytoplasmic processes of myofibroblasts in areas of fibrogenesis. When isolated rat Ito cells were incubated in the presence of acetaldehyde, APAs were also detected in the cytoplasm. These results show that APA formation occurs in hepatocytes at the sites of acetaldehyde production. Detection of APAs in human and rat Ito cells strongly suggests that acetaldehyde can diffuse into Ito cells and bind to cytoplasmic proteins to form local APAs. Because Ito cells are the main effector cells of liver fibrosis, detection of APAs in these cells points to their possible involvement in liver fibrogenesis.

Acetaldehyde↗

PubMed1996

The transcytotic pathway of an apical plasma membrane protein (B10) in hepatocytes is similar to that of IgA and occurs via a tubular pericentriolar compartment.

In hepatocytes, newly synthesized apical plasma membrane proteins are first delivered to the basolateral surface and are supposed to reach the apical surface by transcytosis. The transcytotic pathway of apical membrane proteins and its relationship with other endosomal pathways has not been demonstrated morphologically. We compared the intracellular route of an apical plasma membrane protein, B10, with that of polymeric IgA (pIgA), which is transcytosed, transferrin (Tf) which is recycled, and asialoorosomucoid (ASOR) which is delivered to lysosomes. Ligands and anti-B10 monoclonal IgG were linked to fluorochromes or with peroxidase. The fate of each ligand was followed by confocal and electron microscopy in polarized primary monolayers of rat hepatocytes. When fluorescent anti-B10 IgG and fluorescent pIgA were simultaneously endocytosed for 15-30 minutes, they both uniformly labelled a juxtanuclear compartment. By 30-60 minutes, they reached the bile canaliculi. Tf and ASOR were also routed to the juxtanuclear area, but their fluorescence patterns were more punctate. Microtubule disruption prevented all ligands from reaching the juxtanuclear area. This area corresponded, at least partially, to the localization of the mannose 6-phosphate receptor, an endosomal marker. By electron microscopy, the juxtanuclear compartment was made up of anastomosing tubules connected to vacuoles, and was organized around the centrioles. B10 and pIgA were mainly found in the tubules, whereas ASOR was segregated inside the vacuolar elements and Tf within thinner, recycling tubules. In conclusion, transcytosis of the apical membrane protein B10 occurs inside tubules similar to those carrying pIgA, and involves passage via the pericentriolar area. In the pericentriolar area, the transcytotic tubules appear to maintain connections with other endosomal elements where sorting between recycled and degraded ligands occurs.

Animals↗