Hepatic Lipocytes Research Articles

Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion.

Background & AimsLiver fibrosis arises from long-term chronic liver injury, accompanied by an accelerated wound healing response with interstitial accumulation of extracellular matrix (ECM). Activated hepatic stellate cells (HSC) are the main source for ECM production. MicroRNA29a (miR-29a) is a crucial antifibrotic miRNA that is repressed during fibrosis, resulting in up-regulation of collagen synthesis.MethodsIntracellular and extracellular miRNA levels of primary and immortalized myofibroblastic HSC in response to profibrogenic stimulation by transforming growth factor β (TGFβ) or platelet-derived growth factor-BB (PDGF-BB) or upon inhibition of vesicular transport and autophagy processes were determined by quantitative polymerase chain reaction. Autophagy flux was studied by electron microscopy, flow cytometry, immunoblotting, and immunocytochemistry. Hepatic and serum miR-29a levels were quantified by using both liver tissue and serum samples from a cohort of chronic hepatitis C virus patients and a murine CCl4 induced liver fibrosis model.ResultsIn our study, we show that TGFβ and PDGF-BB resulted in decrease of intracellular miR-29a and a pronounced increase of vesicular miR-29a release into the supernatant. Strikingly, miR-29a vesicular release was accompanied by enhanced autophagic activity and up-regulation of the autophagy marker protein LC3. Moreover, autophagy inhibition strongly prevented miR-29a secretion and repressed its targets’ expression such as Col1A1. Consistently, hepatic miR-29a loss and increased LC3 expression in myofibroblastic HSC were associated with increased serum miR-29a levels in CCl4-treated murine liver fibrosis and specimens of hepatitis C virus patients with chronic liver disease.ConclusionsWe provide evidence that activation-associated autophagy in HSC induces release of miR-29a, whereas inhibition of autophagy represses fibrogenic gene expression in part through attenuated miR-29a secretion.

Translating an Understanding of the Pathogenesis of Hepatic Fibrosis to Novel Therapies

The response to injury is one of wound healing and fibrogenesis, which ultimately leads to fibrosis. The fibrogenic response to injury is a generalized one across virtually all organ systems. In the liver, the injury response, typically occurring over a prolonged period of time, leads to cirrhosis (although it should be pointed out that not all patients with liver injury develop cirrhosis). The fact that many different diseases result in cirrhosis suggests a common pathogenesis. The study of hepatic fibrogenesis over the past 2 decades has been remarkably active, leading to a considerable understanding of this process. It clearly has been shown that the hepatic stellate cell is a central component in the fibrogenic process. It also has been recognized that other effector cells are important in the fibrogenic process, including resident fibroblasts, bone marrow-derived cells, fibrocytes, and even perhaps cells derived from epithelial cells (ie, through epithelial to mesenchymal transition). A key aspect of the biology of fibrogenesis is that the fibrogenic process is dynamic; thus, even advanced fibrosis (or cirrhosis) is reversible. Together, an understanding of the cellular basis for liver fibrogenesis, along with multiple aspects of the basic pathogenesis of fibrosis, have highlighted many exciting potential therapeutic opportunities. Thus, although the most effective antifibrotic therapy is simply treatment of the underlying disease, in situations in which this is not possible, specific antifibrotic therapy is likely not only to become feasible, but will soon become a reality. This review highlights the mechanisms underlying fibrogenesis that may be translated into future antifibrotic therapies and to review the current state of clinical development.

Noninvasive Assessment of Liver Fibrosis and Portal Hypertension With Transient Elastography

Hepatic fibrogenesis, the "final" common result of injury to the liver, is believed to be a critical factor leading to hepatic dysfunction and may be important in the pathogenesis of portal hypertension. Thus, accurate assessment of the degree of fibrosis is important clinically. For many years, examination of hepatic histopathology has been considered to be the "gold standard" tool used to assess fibrosis. However, liver biopsy is invasive, and in many instances not favored by patients or physicians. Thus, alternative approaches to measure liver fibrosis would be extremely attractive. To the extent that transient elastography is able to measure "liver stiffness," which is proportional to the degree of liver fibrosis, this technique holds great promise.

Abrogation of the Antifibrotic Effects of Natural Killer Cells/Interferon-γ Contributes to Alcohol Acceleration of Liver Fibrosis

Chronic alcohol drinking accelerates liver fibrosis in patients with viral hepatitis that cannot be fully explained by ethanol-enhanced liver damage. Here, we identified a novel mechanism by which alcohol accelerates liver fibrosis: inhibition of the antifibrotic effects of natural killer (NK) cells and interferon-gamma (IFN-gamma). Alcohol administration was achieved by feeding mice with a liquid diet containing 5% ethanol for 8 weeks. Liver fibrosis was induced by administration of carbon tetrachloride (CCl(4)) for 2 weeks. Hepatic stellate cells (HSCs) were also isolated and cultured for in vitro studies. CCl(4) treatment induced greater fibrosis and less apoptosis of HSCs in ethanol-fed mice compared with pair-fed mice. Polyinosinic-polycytidylic acid (Poly I:C) or IFN-gamma treatment inhibited liver fibrosis in pair-fed but not in ethanol-fed mice. Poly I:C activation of NK cell cytotoxicity against HSCs was attenuated in ethanol-fed mice compared with pair-fed mice, which was due to reduced natural killer group 2 member D (NKG2D), tumor necrosis factor-related apoptosis-inducing ligand, and IFN-gamma expression on NK cells from ethanol-fed mice. In vitro, HSCs from ethanol-fed mice were resistant to IFN-gamma-induced cell cycle arrest and apoptosis compared with pair-fed mice. Such resistance was due to diminished IFN-gamma activation of signal transducer and activator of transcription 1 (STAT1) in HSCs from ethanol-fed mice caused by the induction of suppressors of cytokine signaling proteins and the production of oxidative stress. Finally, HSCs from ethanol-fed mice were resistant to NK cell killing, which can be reversed by transforming growth factor-beta1 (TGF-beta1) neutralizing antibody. Chronic ethanol consumption attenuates the antifibrotic effects of NK/IFN-gamma/STAT1 in the liver, representing new and different therapeutic targets with which to treat alcoholic liver fibrosis.

Gene Expression Profiles During Hepatic Stellate Cell Activation in Culture and In Vivo

Following hepatic injury, hepatic stellate cells (HSCs) transdifferentiate to become extracellular matrix-producing myofibroblasts and to promote hepatic fibrogenesis. In this study, we determine gene expression changes in 3 different models of HSC activation and investigate whether HSC culture activation reproduces gene expression changes of HSC in vivo activation. HSCs were isolated by density centrifugation and magnetic antibody cell sorting from normal mice, CCl(4)-treated mice, and mice that underwent bile duct ligation (BDL). Gene expression was analyzed by microarray and confirmed by polymerase chain reaction and Western blot analysis. Two thousand seventy-three probe sets were differentially expressed in at least 1 of 3 models of HSC activation, including novel genes that encode proinflammatory and antiapoptotic mediators; transcription factors; cell surface receptors; and cytoskeleton components such as CXCL14, survivin, septin 4, osteopontin, PRX1, LMCD1, GPR91, leiomodin, and anillin. BDL- and CCl(4)-activated HSCs showed highly correlated gene expression patterns, whereas culture activation only partially reproduced the gene expression changes observed during BDL- and CCl(4)-induced activation. Coculture with Kupffer cells or lipopolysaccharide treatment during culture activation shifted the expression of most examined genes toward the pattern observed during in vivo activation, suggesting a role for these factors in the microenvironment that drives HSC activation. The almost identical HSC gene expression patterns after BDL or CCl(4) treatment indicate that HSCs exert similar functions in different types of liver injury. Because culture activation does not properly regulate gene expression in HSCs, in vivo activation should be considered the gold standard for the study of HSC biology.

The dual play of endothelin receptors in hepatic vasoregulation

HepatologyVolume 31, Issue 4 p. 1025-1026 EditorialFree Access The dual play of endothelin receptors in hepatic vasoregulation Chantal Housset M.D., Ph.D., M.D., Ph.D., Corresponding Author Chantal Housset M.D., Ph.D., M.D., Ph.D. [email protected] Faculté de Médecine Saint-Antoine, Paris, FranceINSERM U402, Faculté de médecine Saint-Antoine, 27, rue de Chaligny, 75571 Paris Cedex 12, France. fax: (33) 1-40 01 14 99===Search for more papers by this author Chantal Housset M.D., Ph.D., M.D., Ph.D., Corresponding Author Chantal Housset M.D., Ph.D., M.D., Ph.D. [email protected] Faculté de Médecine Saint-Antoine, Paris, FranceINSERM U402, Faculté de médecine Saint-Antoine, 27, rue de Chaligny, 75571 Paris Cedex 12, France. fax: (33) 1-40 01 14 99===Search for more papers by this author First published: 30 December 2003 https://doi.org/10.1053/he.2000.6708Citations: 4AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL No abstract is available for this article. References 1 Yanagisawa M, Kurihara H, Kimura S, Tomobe Y, Kobayashi M, Mitsui Y, Yazaki Y, et al. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature 1988; 332: 411– 415.MEDLINE 2 Arai H, Hori S, Aramori I, Ohkubo H, Nakanishi S. Cloning and expression of a cDNA encoding an endothelin receptor. Nature 1990; 348: 730– 732.MEDLINE 3 Sakurai T, Yanagisawa M, Takuwa Y, Miyazaki H, Kimura S, Goto K, Masaki T. Cloning of a cDNA encoding a non-isopeptide-selective subtype of the endothelin receptor. Nature 1990; 348: 732– 735.MEDLINE 4 Clozel M, Gray GA, Breu V, Loffler BM, Osterwalder R. The endothelin ETB receptor mediates both vasodilatation and vasoconstriction in vivo. Biochem Biophys Res Commun 1992; 186: 867– 873.MEDLINE 5 Gondo K, Ueno T, Sakamoto M, Sakisaka S, Sata M, Tanikawa K. The endothelin-1 binding site in rat liver tissue: Light- and electron-microscopic autoradiographic studies. Gastroenterology 1993; 104: 1745– 1749.MEDLINE 6 Housset C, Rockey DC, Bissell DM. Endothelin receptors in rat liver: Lipocytes as a contractile target for endothelin-1. Proc Natl Acad Sci U S A 1993; 90: 9266– 9270.MEDLINE 7 Pinzani M, Failli P, Ruocco C, Casini A, Milani S, Baldi E, Giotti A, et al. Fat-storing cells as liver-specific pericytes. Spacial dynamics of agonist-stimulated intracellular calcium transients. J Clin Invest 1992; 90: 642– 646.MEDLINE 8 Kawada N, Tran-Thi TA, Klein H, Decker K. The contraction of hepatic stellate (Ito) cells stimulated with vasoactive substances. Eur J Biochem 1993; 213: 815– 823.MEDLINE 9 Rockey DC, Fouassier L, Chung JJ, Carayon A, Vallée P, Rey C, Housset C. Cellular localization of endothelin-1 and increased production in liver injury in the rat: potential for autocrine and paracrine effects on stellate cells. Hepatology 1998; 27: 472– 480.MEDLINE 10 Rockey DC, Housset CN, Friedman SL. Activation-dependent contractility of rat hepatic lipocytes in culture and in vivo. J Clin Invest 1993; 92: 1795– 1804.MEDLINE 11 Gandhi CR, Stephenson K, Olson MS. Endothelin, a potent peptide agonist in the liver. J Biol Chem 1990; 265: 17432– 17435.MEDLINE 12 Kaneda K, Ekataksin W, Sogawa M, Matsumura A, Cho A, Kawada N. Endothelin-1-induced vasoconstriction causes a significant increase in portal pressure of rat liver: Localized constrictive effect on the distal segment of preterminal portal venules as revealed by light and electron microscopy and serial reconstruction. Hepatology 1998; 27: 735– 747.MEDLINE 13 Bauer M, Zhang JX, Bauer I, Clemens MG. ET-1 induced alterations of hepatic microcirculation: sinusoidal and extrasinusoidal sites of action. Am J Physiol 1994; 267: G143– G149.MEDLINE 14 Rockey DC, Chung JJ. Reduced nitric oxide production by endothelial cells in cirrhotic rat liver: endothelial dysfunction in portal hypertension. Gastroenterology 1998; 114: 344– 351.MEDLINE 15 Reichen J, Gerbes AL, Steiner MJ, Sägesser H, Clozel M. The effect of endothelin and its antagonist Bosentan on hemodynamics and microvascular exchange in cirrhotic rat liver. J Hepatol 1998; 28: 1020– 1030.MEDLINE 16 Kurihara Y, Kurihara H, Suzuki H, Kodama T, Maemura K, Nagai R, Oda H, et al. Elevated blood pressure and craniofacial abnormalities in mice deficient in endothelin-1. Nature 1994; 368: 703– 710.MEDLINE Citing Literature Volume31, Issue4April 2000Pages 1025-1026 ReferencesRelatedInformation

Perinatal expression of genes that may participate in lipid metabolism by lipid-laden lung fibroblasts

Although a morphologically distinct population of lipid-laden interstitial cells (lipofibroblasts, LF), has been identified, the origins and functions of this population during lung development and disease remain undefined. Illumination of the developmental and functional characteristics of two other populations of lipid-laden mesenchymal cells, namely adipocytes and hepatic lipocytes, has fashioned tools that can be used to explore similar properties in pulmonary LFs. As the LF is transiently a very abundant cell in the perinatal lung, we elected to study the perinatal ontogeny of the expression of several genes that are involved in the acquisition of lipids by adipocytes, and may be involved in promoting the triglyceride accumulation that is the morphologic hallmark of the pulmonary LF. We found that the maximal expression of peroxisome proliferator-activated receptor-gamma (PPAR-gamma), at gestational day 21 in the LF, precedes the rise at birth, in the expression of genes that are involved in the hydrolysis of triglycerides at the plasma membrane (lipoprotein lipase, LPL), transport of fatty acids across the plasma membrane (fatty acid transporter, FAT) and in the cytoplasm (adipocyte lipid binding protein, ALBP). The steady-state levels of LPL, FAT, and ALBP mRNAs that were isolated from whole lung tissue showed a similar temporal pattern. The levels of the protein products of the LPL and ALBP genes changed in tandem with those of their precursor mRNAs in the LF, suggesting that these gene products are under pre-translational control. These findings indicate that characteristic adipocyte genes are also expressed in lipid-laden pulmonary fibroblasts and may participate in triglyceride accumulation and metabolism by these cells.

Peroxisome proliferators alter lipid acquisition and elastin gene expression in neonatal rat lung fibroblasts.

During the alveolar stage of lung development, lipid droplet-laden interstitial cells are present at the base of elongating alveolar septa. These cells that have been named lipid interstitial cells or lipofibroblasts (LFs) may supply lipids for surfactant production, the synthesis of membrane phospholipids, and/or energy metabolism. They also have myofibroblastic characteristics and participate in the generation of the interstitial elastic fiber network, that is, in the pulmonary alveolar septum. To understand how this cell regulates its lipid-storing and elastin-producing properties, we have examined the effects of peroxisome proliferators on the expression of the genes that are associated with an elastin-producing myofibroblastic phenotype or an adipocyte-like phenotype. Two known ligands for peroxisome proliferator-activated receptors, 5,8,11,14-eicosatetraynoic acid (ETYA) and 15-deoxy-delta-12,14-prostaglandin J2 (15-dPGJ2), decrease elastin gene transcription and the steady-state levels of tropoelastin (TE) and alpha-smooth muscle actin mRNAs in cultured LFs. Concurrently, cultured LFs increase the expression of adipocyte lipid binding protein, which is regarded as an adipocyte-specific protein, and accumulate lipid droplets. Their abilities to store lipids and express desmin intermediate filaments, alpha-smooth muscle actin, and smooth muscle myosin heavy chain in contractile filaments in vitro illustrate similarities among the pulmonary LF, the hepatic lipocyte, and the contractile interstitial cell, which contribute to the repair reaction in the lung after pulmonary injury.

Retinoids may increase fibrotic potential of TGF-β: Crosstalk between two multi-functional effectors

HepatologyVolume 26, Issue 4 p. 1067-1068 EditorialFree Access Retinoids may increase fibrotic potential of TGF-β: Crosstalk between two multi-functional effectors R Blomhoff, R BlomhoffSearch for more papers by this author R Blomhoff, R BlomhoffSearch for more papers by this author First published: 30 December 2003 https://doi.org/10.1002/hep.510260439Citations: 1AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. References 1 Wake K. Perisinusoidal stellate cells (fat-storing cells, interstitial cells, lipocytes), their related structure in and around the liver sinusoids, and vitamin A-storing cells in extrahepatic organs. Int Rev Cytol 1980; 66: 303– 353. 2 Blomhoff R, Wake K. Perisinusoidal stellate cells of the liver: important roles in retinol metabolism and fibrosis. FASEB J 1991; 5: 271– 277. 3 Friedman SL. The cellular basis of hepatic fibrosis. Mechanisms and treatment strategies. N Engl J Med 1993; 328: 1828– 1835. 4 Davis BH, Kramer RT, Davidson NO. Retinoic acid modulates rat Ito cell proliferation, collagen, and transforming growth factor beta production. J Clin Invest 1990; 86: 2062– 2070. 5 Davis BH, Vucic A. The effect of retinol on Ito cell proliferation in vitro. Hepatology 1988; 8: 788– 793. 6 Sato T, Kato R, Tyson CA. Regulation of differentiated phenotype of rat hepatic lipocytes by retinoids in primary culture. Exp Cell Res 1995; 217: 72– 83. 7 Senoo H, Wake K. Suppression of experimental hepatic fibrosis by administration of vitamin A. Lab Invest 1985; 52: 182– 194. 8 Seifer WF, Bosma A, Hendriks HF, van Leeuwen RE, van Thil-de Ruiter GC, Seifert-Bock I, Knock DL, et al. Beta-carotene (provitamin A) decreases the severity of CCl4-induced hepatic inflammation and fibrosis in rats. Liver 1995; 15: 1– 8. 9 Seifer WF, Bosma A, Brouwer A, Hendriks HF, Roholl PJ, van Leeuwen RE, Ruiter GC van Thil-de, et al. Vitamin A deficiency potentiates carbon tetrachloride-induced liver fibrosis in rats. Hepatology 1994; 19: 193– 201. 10 Okuno M, Moriwaki H, Imai S, Muto Y, Kawada N, Suzuki Y, Kojima S. Retinoids exacerbate rat liver fibrosis by inducing the activation of latent TGF-β in liver stellate cells. Hepatology 1997; 26: 918– 926. 11 Blomhoff R. Vitamin A in Health and Disease. New York: Marcel Dekker, Inc., 1994: 1– 677. 12 Blomhoff R, Green MH, Berg T, Norum KR. Transport and storage of vitamin A. Science 1990; 250: 399– 404. 13 Blomhoff R, Helgerud P, Rasmussen M, Berg T, Norum KR. In vivo uptake of chylomicron retinyl ester by rat liver: evidence for retinol transfer from parenchymal to nonparenchymal cells. Proc Natl Acad Sci U S A 1982; 79: 7326– 7330. 14 Blomhoff R, Rasmussen M, Nilsson A, Norum KR, Berg T, Blaner WS, Kato M, et al. Hepatic retinol metabolism, distribution of retinoids, enzymes, and binding proteins in isolated rat liver cells. J Biol Chem 1985; 260: 13560– 13565. 15 Blaner WS, van Bennekum AM, Brouwer A, Hendriks HF. Distribution of lecithin-retinol acyltransferase activity in different types of rat liver cells and subcellular fractions. FEBS Lett 1990; 274: 89– 92. 16 Mangelsdorf DJ, Thummel C, Beato M, Herrlich P, Schütz G, Kastner P, Mark M, et al. The nuclear receptor superfamily: the second decade. Cell 1995; 83: 835– 839. 17 Ohata M, Lin M, Satre M, Tsukamoto H. Diminished retinoic acid signaling in hepatic stellate cells in cholestatic liver fibrosis. Am J Physiol 1997; 272: G589– G596. 18 Takahashi N, Bretiman TR. Retinoylation of proteins in mammalian cells, pp. 257– 273. In: R Blomhoff, ed. Vitamin A in Health and Disease. New York: Marcel Dekker, Inc., 1994: 257–273. 19 Koga H, Fujita I, Miyazaki S. Effects of all-trans retinoic acid on super-oxide generation in intact neutrophils and a cell-free system. Br J Haematol 1997; 97: 300– 305. 20 Buck J, Derguini F, Levy E, Nakanishi K, Hmmerling U. Intracellular signalling by 14-hydroxy-4, 14-retro-retinol. Science 1991; 254: 1654– 1656. 21 Achkar CC, Derguini F, Blumberg B. 4-oxoretinol, a new natural ligand and transactivator of the retinoic acid receptors. Proc Natl Acad Sci USA 1996; 93: 4879– 4884. Citing Literature Volume26, Issue4October 1997Pages 1067-1068 ReferencesRelatedInformation

The pulmonary lipofibroblast (lipid interstitial cell) and its contributions to alveolar development.

The pulmonary lipofibroblast is located in the alveolar interstitium and is recognizable by its characteristic lipid droplets. During alveolar development it participates in the synthesis of extracellular matrix structural proteins, such as collagen and elastin, and as an accessory cell to the type II pneumocyte, in the synthesis of surfactant. The lipofibroblast contains cortical contractile filaments and is thereby related to the contractile interstitial cells that are normally found at the alveolar septal tips and after lung injury. The morphologic, immunologic, and biochemical characteristics of the lipofibroblast and its probable physiologic functions are reviewed. The retinoid and lipid metabolism of the lipofibroblast is compared with that of the hepatic lipocyte and the adipocyte. Although the functions of the lipofibroblast remain incompletely characterized, this cell type is emerging as an important contributor to pulmonary alveolar septal development.

Ethanol metabolism, cirrhosis and alcoholism

Alcohol-induced tissue damage results from associated nutritional deficiencies as well as some direct toxic effects, which have now been linked to the metabolism of ethanol. The main pathway involves liver alcohol dehydrogenase which catalyzes the oxidaation of ethanol to acetaldehyde, with a shift to a more reduced state, and results in metabolic disturbances, such as hyperlactacidemia, acidosis, hyperglycemia, hyperuricemia and fatty liver. More severe toxic manifestations are produced by an accessory pathway, the microsomal ethanol oxidizing system involving an ethanol-inducible cytochrome P450 (2E1). After chronic ethanol consumption, there is a 4- to 10-fold induction of 2E1, associated not only with increased acetaldehyde generation but also with production of oxygen radicals that promote lipid peroxidation. Most importantly, 2E1 activates many xenobiotics to toxic metabolites. These include solvents commonly used in industry, anaesthetic agents, medications such as isoniazid, over the counter analgesics (acetaminophen), illicit drugs (cocaine), chemical carcinogens, and even vitamin A and its precursor β-carotene. Furthermore, enhanced microsomal degradation of retinoids (together with increased hepatic mobilization) promotes their depletion and associated pathology. Induction of 2E1 also yields increased acetaldehyde generation, with formation of protein adducts, resulting in antibody production, enzyme inactivation, decreased DNA repair, impaired utilization of oxygen, glutathione depletion, free radical-mediated toxicity, lipid peroxidation, and increased collagen synthesis. New therapies include adenosyl- l-methionine which, in baboons, replenishes glutathione, and attenuates mitochondrial lesions. In addition, polyenylphosphatidylcholine (PPC) fully prevents ethanol-induced septal fibrosis and cirrhosis, opposes ethanol-induced hepatic phospholipid depletion, decreased phosphatidylethanolamine methyltransferase activity and activation of hepatic lipocytes, whereas its dilinoleoyl species increases collagenase activity. Current clinical trials with PPC are targeted on susceptible populations, namely heavy drinkers at precirrhotic stages.

Antifibrogenic effect of a deletion variant of hepatocyte growth factor on liver fibrosis in rats

Hepatic fibrosis, which may lead to cirrhosis, is associated with most chronic liver diseases. Current therapies for hepatic fibrosis are, however, generally ineffective. In this report we assessed the efficacy of the treatment of hepatic fibrosis with a naturally occurring deletion variant of hepatocyte growth factor (dHGF). The administration of dHGF increased liver weight and suppressed the increase of hepatic collagen content in rats treated with dimethylnitrosamine (DMN) to induce hepatic fibrosis. Furthermore, dHGF exerted its mitogenic and antifibrogenic activities even after the liver fibrosis had been established with DMN. Northern blot analysis showed that dHGF suppressed the increase of messenger RNA (mRNA) levels of procollagen alpha 2(I), alpha l(III), alpha 1(IV), transforming growth factor beta1 (TGF-beta1), desmin (a marker of hepatic lipocytes), and alpha-smooth muscle (sm)-actin (a marker of activated hepatic lipocytes). In addition to suppressing the elevated TGF-beta1, mRNA level in hepatic fibrosis, dHGF had a potent ability to decrease TGF-beta1 mRNA level even in a normal liver. Immunohistochemical analysis revealed that desmin-positive cells and alpha-sm-actin-positive cells were increased in the hepatic fibrosis, whereas neither cells were seen in livers of DMN-treated rats given dHGF. We conclude that dHGF prevents and improves the DMN-induced hepatic fibrosis in rats by reducing mRNA levels of procollagens and TGF-beta1, by inhibiting an activation of hepatic lipocytes, and by stimulating liver regeneration. dHGF may be useful for and applicable to the treatment of fibrosis in chronic liver diseases. (Hepatology 1996 Sep;24(3):636-42)

Antifibrogenic Effect of A Deletion Variant of Hepatocyte Growth Factor on Liver Fibrosis in Rats

Hepatic fibrosis, which may lead to cirrhosis, is associated with most chronic liver diseases. Current therapies for hepatic fibrosis are, however, generally ineffective. In this report we assessed the efficacy of the treatment of hepatic fibrosis with a naturally occurring deletion variant of hepatocyte growth factor (dHGF). The administration of dHGF increased liver weight and suppressed the increase of hepatic collagen content in rats treated with dimethylnitrosamine (DMN) to induce hepatic fibrosis. Furthermore, dHGF exerted its mitogenic and antifibrogenic activities even after the liver fibrosis had been established with DMN. Northern blot analysis showed that dHGF suppressed the increase of messenger RNA (mRNA) levels of procollagen α2(I), αl(III), α1(IV), transforming growth factor β1 (TGF–β1), desmin (a marker of hepatic lipocytes), and α–smooth muscle (sm)–actin (a marker of activated hepatic lipocytes). In addition to suppressing the elevated TGF–β1, mRNA level in hepatic fibrosis, dHGF had a potent ability to decrease TGF–β1 mRNA level even in a normal liver. Immunohistochemical analysis revealed that desmin–positive cells and α–sm–actin–positive cells were increased in the hepatic fibrosis, whereas neither cells were seen in livers of DMN–treated rats given dHGF. We conclude that dHGF prevents and improves the DMN–induced hepatic fibrosis in rats by reducing mRNA levels of procollagens and TGF–β1, by inhibiting an activation of hepatic lipocytes, and by stimulating liver regeneration. dHGF may be useful for and applicable to the treatment of fibrosis in chronic liver diseases.

Osteonectin gene expression in fibrotic liver

Osteonectin is an extracellular matrix (ECM) protein which is secreted by various cell types, and regulates tissue remodeling and cell proliferation. In the present study, we have examined the expression of osteonectin in fibrotic liver. Osteonectin transcripts were undetectable in normal liver, however, the abundant expression of the osteonectin gene was detected in fibrotic liver. The transcripts of osteonetin were only detected in hepatic lipocytes and the number of lipocytes was increased in fibrotic liver. These results suggested that in fibrotic liver, enhanced osteonectin expression may play an important role in liver fibrosis.

Influence of acetaldehyde and ethanol on rat hepatic lipocyte characteristics in primary culture

Influence of acetaldehyde and ethanol on rat hepatic lipocyte characteristics in primary culture

Complement C4 protein gene expression by rat hepatic lipocytes: [formula omitted]. Saint Louis University Health Sciences Center and St. Louis VA Medical Center, St. Louis, MO, and ^Alton Ochsner Medical Foundation, New Orleans, LA

Complement C4 protein gene expression by rat hepatic lipocytes: [formula omitted]. Saint Louis University Health Sciences Center and St. Louis VA Medical Center, St. Louis, MO, and ^Alton Ochsner Medical Foundation, New Orleans, LA

Increase of lipocyte number and lipid-storage in early stages of primary biliary cirrhosis and study of the effect of ursodeoxycholic acid on lipocytes in PBC: [formula omitted]. Depts. of Pathology and Medicine, The Toronto Hospital, and Division of Clinical Pharmacology, Sunnybrook Health Science Centre, Toronto, Ontario, Canada

Activation of hepatic lipocytes to myofibroblastlike cells observed in cell culture and during liver fibrogenesis is characterized by an increase in collagen formation and cell proliferation. These changes appear to be associated with the loss of intracellular retinoid in lipocytes, the principal storage site for vitamin A in the body. To evaluate whether retinoids have the capability to suppress lipocyte activation, we exposed cultured lipocytes in both native and myofibroblastlike states to retinoids and determined their effects on collagen production, intracellular retinoid level, and cell proliferation. Retinol (1 μM) and retinoic acid (1 μM) supplementation of primary rat lipocyte cultures inhibited the spontaneous increase in collagen synthesis associated with lipocyte activation; lower concentrations of retinol (10 or 100 nM) were also effective. Simultaneously, retinol addition prevented a precipitous decline in intracellular retinoid content in the absence of added retinoid. These retinoid effects were reversed by a change to unsupplemented control medium. When cells in the myofibroblastlike state were exposed to retinol (⩾ 1 μM), a significant increase in intracellular retinoid levels and reduction in collagen synthesis occurred. Lipocytes in both native and myofibroblastlike states secreted four to five times higher amounts of type I collagen than type III collagen, but retinol and retinoic acid particularly inhibited production of type I collagen. Cell proliferation measured by [3H]thymidine incorporation was also inhibited by retinol. These results demonstrate that extracellular retinoids suppress lipocyte-activated collagen synthesis and cell proliferation and support the interpretation that retinoids themselves are regulatory factors in maintenance of the lipocyte in its native, differentiated state.

Rat hepatic lipocytes synthesize and secrete transin (Stromelysin) in early primary culture

Background & Aims : Hepatic lipocyte proliferation and activation are pivotal in liver fibrosis. Disruption of normal lipocyte-matrix interactions may contribute to this process. The synthesis of transin, which degrades normal liver matrix, by culture-activated hepatic lipocytes was investigated. Methods : Lipocytes were isolated by pronase/collagenase perfusion, density gradient centrifugation, and centrifugal elutriation. Transin messenger RNA in lipocytes was analyzed by Northern blotting. Transin activity was analyzed by zymography, Western blotting, immunocytochemistry, and quantitative [ 14C]β-casein degradation assay. Results : Transin messenger RNA was detected in early primary culture (3–5 days) but not in freshly isolated lipocytes or late primary culture. Zymography of lipocyte medium showed caseinolytic activity (relative molecular weight, 57 kilodaltons and 60 kilodaltons) inhibited by ethylenediaminetetraacetic acid but not thiol or serine protease inhibitors. Immunoblotting and immunocytochemistry confirmed the presence of transin in media and cells. Quantitative transin activity decreased progressively with increasing duration of primary lipocyte culture and myofibroblastic transformation. Conclusions : Rat hepatic lipocytes express the transin gene and secrete its product during the early phase of lipocyte activation in primary culture. Because this enzyme degrades a wide spectrum of normal basement membrane proteins and activates progelatinase B and interstitial collagenase, it may have an important role in liver injury and fibrosis.

Induction of β-platelet-derived growth factor receptor in rat hepatic lipocytes during cellular activation in vivo and in culture: Wong L, Yamasaki G, Johnson RL, Friedman SL. J Clin Invest 1994;94:1563–1569

In hepatic fibrogenesis, stellate cells are activated leading to production and deposition of extracellular matrix. To clarify the role of PDGF-B in liver fibrogenesis, we overexpressed PDGF-B in the liver of transgenic mice.Transgenic mice for the conditional overexpression of PDGF-B in the liver under control of an albumin promoter were generated utilising the Cre/loxP system. Constitutive PDGF-B expression was achieved after breeding with mice expressing Cre-recombinase under actin promoter control. Tamoxifen inducible expression was achieved after breeding with mice expressing Cre under transthyretin receptor promoter control. Levels of fibrosis were assessed and the expression of regulators of matrix remodelling was measured.PDGF-B expression caused hepatic stellate cell and myofibroblast activation marked by alpha-smooth muscle actin and PDGFR-β expression. Liver fibrosis was verified macroscopically, histologically and by collagen I mRNA quantification in 4–6 week-old animals. MMP-2, MMP-9 and TIMP-1 were upregulated whereas TGF-β expression was unchanged.We identified PDGF-B as a proliferative and profibrogenic stimulus and potential inducer of stellate cell transdifferentiation in vivo. PDGF-B overexpression causes liver fibrosis without significantly upregulating TGF-β1, suggesting a TGF-β-independent mechanism. The established model provides a tool for testing anti-PDGF-B therapeutic strategies in liver fibrosis in vivo.

Tissue inhibitor of metalloproteinase-I and interstitial collagenase expression in autoimmune chronic active hepatitis and activated human hepatic lipocytes.

1. Activated hepatic lipocytes are central to the pathogenesis of liver fibrosis as the principal source of both interstitial collagens and matrix-degrading metalloproteinases. In progressive fibrosis there is a failure to degrade interstitial collagens with a reported decrease in collagenase activity. In these studies we investigate expression of the potent collagenase inhibitor, tissue inhibitor of metalloproteinase-1, and interstitial collagenase in end-stage autoimmune chronic active hepatitis and activated human hepatic lipocytes in culture. 2. Messenger RNA transcripts for interstitial collagenase and tissue inhibitor of metalloproteinase-1 in explanted human liver were quantified by ribonuclease protection assay and densitometric analysis. This indicated that tissue inhibitor of metalloproteinase-1 and interstitial collagenase expression in autoimmune chronic active hepatitis were also coordinately up-regulated. 3. Using Northern analysis of RNA from human lipocytes in primary culture on plastic, mRNA for interstitial collagenase could not be detected in unstimulated cells but was present after stimulation with tumour necrosis factor alpha. Tissue inhibitor of metalloproteinase-1 mRNA was present in unstimulated lipocytes and up-regulated fivefold in response to tumour necrosis factor alpha. Using activity assay of serum-free conditioned media, interstitial collagenase could not be detected in unstimulated primary cultures, primary cultures stimulated with tumour necrosis factor alpha or transforming growth factor beta-1 (n = 3 and n = 4 respectively) or in passaged lipocytes (n = 6). In contrast, free tissue inhibitor of metalloproteinase-1 activity was present in unstimulated and passaged cultures and this was increased in response to tumour necrosis factor alpha and transforming growth factor beta-1.(ABSTRACT TRUNCATED AT 250 WORDS)