Background Doses of ethanol (EtOH) that are not overtly cytotoxic inhibit mitogen‐induced hepatocyte proliferation and delay liver regeneration after 70% partial hepatectomy (PH). The mechanisms for this are poorly understood. This study evaluates the hypothesis that EtOH inhibits hepatocyte proliferation after PH by inducing redox‐sensitive factors, such as p38 mitogen‐activated protein kinase (MAPK) and p21 (WAF1/CIP1), that protect cells from oxidative stress but prevent cell‐cycle progression by inhibiting cyclin D1. Methods Mechanisms that regulate the transition from the prereplicative G 1 phase of the cell cycle into S phase were compared in EtOH‐fed mice and normal pair‐fed mice after PH. Results Prior EtOH exposure significantly increases p38 MAPK and p21 after PH. This is accompanied by reduced expression of cyclin D1 messenger RNA and protein, increases in other cell‐cycle regulators (such as signal transducer and activator of transcription‐3 and p27) that are normally inhibited by cyclin D1, and hepatocyte G 1 arrest. Conclusions EtOH amplifies G 1 checkpoint mechanisms that are induced by oxidative stress and promotes hepatic accumulation of factors, including p38 MAPK, p21, and signal transducer and activator of transcription‐3, that enhance cellular survival after oxidant exposure. Therefore, cell‐cycle inhibition may be an adaptive response that helps EtOH‐exposed livers survive situations, such as PH, that acutely increase reactive oxygen species in hepatocytes.
Background and Aims Liver disease is prevalent in the United States, and as the population ages, an increasing number of patients are anticipated to present for care. The state of the current hepatology workforce and future demand for hepatology providers is not known. The aim of this study was to model future projections for hepatology workforce demand. Approach and Results A workforce study of hepatology providers in the United States was completed using primary and secondary data sources. An integrated workforce framework model was used that combined socioeconomic factors that drive economic demand, epidemiological factors that drive need, and utilization rates of health care services. Supply and demand projections were calculated for adult and pediatric hepatology professionals. Sensitivity analyses were conducted to cover the feasible range of these assumptions. An electronic survey of American Association for the Study of Liver Diseases (AASLD) members whose practice included 50% or more hepatology was conducted. In 2018, the adult and pediatric workforce included 7,296 and 824 hepatology providers, respectively, composed of hepatologists, gastroenterologists, and advanced practice providers whose practice was ≥50% hepatology. The modeling analysis projects that in 2023, 2028, and 2033, there will be shortages of 10%, 23%, and 35% adult hepatology providers, respectively, and 19%, 20%, and 16% pediatric hepatology providers, respectively. In sensitivity analyses, a shortage of hepatology providers is predicted even under optimistic assumptions. Among the respondents to the survey, the median age was higher among gastroenterologists and general hepatologists compared with transplant hepatologists. The most common category treated by transplant hepatologists was general hepatology. Conclusions There is an impending critical shortage of adult and pediatric hepatology providers. Strategies are needed to encourage clinicians to pursue hepatology, especially in areas outside of transplant centers.
Although ethanol is known to sensitize hepatocytes to tumor necrosis factor (TNF) lethality, the mechanisms involved remain controversial. Recently, others have shown that adding TNFα to cultures of ethanol-pretreated hepatocytes provokes the mitochondrial permeability transition, cytochrome crelease, procaspase 3 activation, and apoptosis. Although this demonstrates that ethanol can sensitize hepatocytes to TNF-mediated apoptosis, the hepatic inflammation and ballooning hepatocyte degeneration that typify alcohol-induced liver injury suggest that other mechanisms might predominate in vivo. To evaluate this possibility, acute responses to lipopolysaccharide (LPS), a potent inducer of TNFα, were compared in mice that had been fed either an ethanol-containing or control diet for 5 weeks. Despite enhanced induction of cytokines such as interleukin (IL)-10, IL-15, and IL-6 that protect hepatocytes from apoptosis, ethanol-fed mice exhibited a 4–5-fold increase in serum alanine aminotransferase after LPS, confirming increased liver injury. Six h post-LPS histology also differed notably in the two groups, with control livers demonstrating only scattered apoptotic hepatocytes, whereas ethanol-exposed livers had large foci of ballooned hepatocytes, inflammation, and scattered hemorrhage. No caspase 3 activity was noted during the initial 6 h after LPS in ethanol-fed mice, but this tripled by 1.5 h after LPS in controls. Procaspase 8 cleavage and activity of the apoptosis-associated kinase, Jun N-terminal kinase, were also greater in controls. In contrast, ethanol exposure did not inhibit activation of cytoprotective mitogen-activated protein kinases and AKT or attenuate induction of the anti-apoptotic factors NF-κB and inducible nitric oxide synthase. Consistent with these responses, neither cytochrome c release, an early apoptotic response, nor hepatic oligonucleosomal DNA fragmentation, the ultimate consequence of apoptosis, was increased by ethanol. Thus, ethanol exacerbates TNF-related hepatotoxicity in vivo without enhancing caspase 3-dependent apoptosis. Although ethanol is known to sensitize hepatocytes to tumor necrosis factor (TNF) lethality, the mechanisms involved remain controversial. Recently, others have shown that adding TNFα to cultures of ethanol-pretreated hepatocytes provokes the mitochondrial permeability transition, cytochrome crelease, procaspase 3 activation, and apoptosis. Although this demonstrates that ethanol can sensitize hepatocytes to TNF-mediated apoptosis, the hepatic inflammation and ballooning hepatocyte degeneration that typify alcohol-induced liver injury suggest that other mechanisms might predominate in vivo. To evaluate this possibility, acute responses to lipopolysaccharide (LPS), a potent inducer of TNFα, were compared in mice that had been fed either an ethanol-containing or control diet for 5 weeks. Despite enhanced induction of cytokines such as interleukin (IL)-10, IL-15, and IL-6 that protect hepatocytes from apoptosis, ethanol-fed mice exhibited a 4–5-fold increase in serum alanine aminotransferase after LPS, confirming increased liver injury. Six h post-LPS histology also differed notably in the two groups, with control livers demonstrating only scattered apoptotic hepatocytes, whereas ethanol-exposed livers had large foci of ballooned hepatocytes, inflammation, and scattered hemorrhage. No caspase 3 activity was noted during the initial 6 h after LPS in ethanol-fed mice, but this tripled by 1.5 h after LPS in controls. Procaspase 8 cleavage and activity of the apoptosis-associated kinase, Jun N-terminal kinase, were also greater in controls. In contrast, ethanol exposure did not inhibit activation of cytoprotective mitogen-activated protein kinases and AKT or attenuate induction of the anti-apoptotic factors NF-κB and inducible nitric oxide synthase. Consistent with these responses, neither cytochrome c release, an early apoptotic response, nor hepatic oligonucleosomal DNA fragmentation, the ultimate consequence of apoptosis, was increased by ethanol. Thus, ethanol exacerbates TNF-related hepatotoxicity in vivo without enhancing caspase 3-dependent apoptosis. tumor necrosis factor lipopolysaccharide TNF receptor pair-fed Jun N-terminal kinase electrophoretic mobility shift assay interleukin inducible nitric oxide synthase heat shock protein-70 mitogen-activated protein kinase extracellular signal-regulated kinase protein kinase B inhibitor κβ kinase Although ethyl alcohol has been recognized as a significant hepatotoxin for centuries, the mechanisms involved in alcohol-induced liver disease remain uncertain. It is known that chronic alcohol ingestion potentiates liver injury inflicted by many other toxins (1.Seeff L.B. Cuccherini B.A. Zimmerman H.J. Alder E. Benjamin S.B. Ann. Intern. Med. 1986; 104: 399-404Crossref PubMed Scopus (369) Google Scholar), viral hepatitis (2.Marsano L.S. Pena L.R. Hepatogastroenterology. 1998; 45: 331-339PubMed Google Scholar, 3.Serfaty L. Chazouilleres O. Poujol-Robert A. Morand-Joubert L. Dubois C. Chretien Y. Poupon R.E. Petit J.C. Poupon R. Hepatology. 1997; 26: 776-779Crossref PubMed Scopus (116) Google Scholar), and hepatic hypoxia-reoxygenation (4.Thurman R.G. Ji S. Matsumura T. Lemasters J.J. Fundam. Appl. Toxicol. 1984; 4: 125-133Crossref PubMed Google Scholar, 5.Israel Y. Orrego H. Recent Dev. Alcohol. 1984; 2: 119-133Crossref PubMed Scopus (28) Google Scholar) or ischemia-reperfusion (6.Gao W. Connor H.D. Lemasters J.J. Mason R.P. Thurman R.G. Transplantation. 1995; 59: 674-679Crossref PubMed Scopus (60) Google Scholar) and yet fails to cause serious liver damage in most healthy adults (7.Lelbach W.K. Prog. Liver Dis. 1976; 5: 494-513PubMed Google Scholar, 8.Pequignot G. Tuyns A.S. Berta J.L. Int. J. Epidemiol. 1978; 7: 113-120Crossref PubMed Scopus (191) Google Scholar, 9.Johnson R.D. Williams R. Alcohol Alcohol. 1985; 20: 137-142PubMed Google Scholar). These clinical observations suggest that ethanol exposure enhances hepatic vulnerability to a secondary inflammatory or oxidative stress, such that serious liver injury is most likely to occur when proinflammatory/pro-oxidant factors are superimposed onto a background of ethanol use (10.Day C.P. James O. Gastroenterology. 1998; 114: 842-845Abstract Full Text Full Text PDF PubMed Scopus (3406) Google Scholar). Because tumor necrosis factor-α ((TNFα)1 and cytokines that potentiate TNF activity are the common, proximal mediators of inflammatory liver injury (11.Vassalli P. Annu. Rev. Immunol. 1992; 10: 411-452Crossref PubMed Scopus (1798) Google Scholar, 12.Tracey K.J. Cerami A. Annu. Rev. Cell Biol. 1993; 9: 317-343Crossref PubMed Scopus (760) Google Scholar, 13.Baker S.J. Reddy E.P. Oncogene. 1996; 12: 1-9PubMed Google Scholar), several laboratories have been evaluating the possibility that ethanol may sensitize hepatocytes to TNF-related toxicity (14.Honchel R. Ray R.B. Marsano L. Cohen D. Lee E. Shedlosky S. McClain C.J. Alcohol. Clin. Exp. Res. 1992; 16: 665-669Crossref PubMed Scopus (107) Google Scholar, 15.Zeldin G. Rai R. Yang S.Q. Lin H.Z. Yin M. Diehl A.M. Alcohol. Clin. Exp. Res. 1996; 20: 1639-1645Crossref PubMed Scopus (55) Google Scholar, 16.Fernandez-Checa J.C. Kaplowitz N. Garcia-Ruiz C. Colell A. Miranda M. Marie M. Ardite E. Morales A. Am. J. Physiol. 1997; 273: G7-G17Crossref PubMed Google Scholar, 17.Yin M. Wheeler M.D. Kono H. Bradford B.U. Gallucci R.M. Luster M.I. Thurman R.G. Gastroenterology. 1999; 117: 942-952Abstract Full Text Full Text PDF PubMed Scopus (634) Google Scholar, 18.Pastorino J.G. Hoek J.B. Hepatology. 2000; 31: 1141-1152Crossref PubMed Scopus (175) Google Scholar). The evidence supporting this concept is growing. For example, animal studies demonstrate that ethanol enhances liver injury caused by lipopolysaccharide (LPS) (14.Honchel R. Ray R.B. Marsano L. Cohen D. Lee E. Shedlosky S. McClain C.J. Alcohol. Clin. Exp. Res. 1992; 16: 665-669Crossref PubMed Scopus (107) Google Scholar), a process that requires TNFα (19.Mayeux P.R. J. Toxicol. Environ. Health. 1997; 51: 415-435Crossref PubMed Scopus (193) Google Scholar). Moreover, disruption of the gene that encodes the TNF type 1 receptor (TNFR-1) completely protects mice from the liver injury that results from chronic intragastric infusions of ethanol (17.Yin M. Wheeler M.D. Kono H. Bradford B.U. Gallucci R.M. Luster M.I. Thurman R.G. Gastroenterology. 1999; 117: 942-952Abstract Full Text Full Text PDF PubMed Scopus (634) Google Scholar). The latter finding demonstrates that TNFα is required for ethanol-induced liver disease in mice. Assuming that similar mechanisms operate in humans who habitually consume alcohol-containing beverages, then is it important to delineate the cellular mechanisms that mediate ethanol-related sensitization to TNF lethality.Healthy hepatocytes are typically resistant to TNFα toxicity unless they have been pretreated with agents that inhibit protein synthesis. Under such circumstances, TNFα induces the mitochondrial permeability transition, cytochrome c release, caspase 3 activation, and eventually causes death by apoptosis (20.Leist M. Cantner F. Bohlinger I. Germann P.C. Tiegs G. Wendel A. J. Immunol. 1994; 153: 1778-1788PubMed Google Scholar). Recently, Hoek and colleague (18.Pastorino J.G. Hoek J.B. Hepatology. 2000; 31: 1141-1152Crossref PubMed Scopus (175) Google Scholar) showed that ethanol mimics these effects of protein synthesis inhibitors. That is, when HepG2 cells or primary rat hepatocytes that have been pretreated with ethanol are exposed to TNFα in vitro, mitochondrial permeability transition, cytochrome c release, caspase 3 activation, and apoptosis ensue. Because treatment of these cultures with mitochondrial permeability transition inhibitors rescues them from TNF lethality, these authors (18.Pastorino J.G. Hoek J.B. Hepatology. 2000; 31: 1141-1152Crossref PubMed Scopus (175) Google Scholar) concluded that ethanol sensitizes the liver to injury by potentiating TNF-induced apoptosis.The purpose of the present study is to determine whether similar mechanisms operate in vivo. To accomplish this, ethanol and pair-fed mice were given a single intraperitoneal injection of LPS, and subsequent liver damage as well as antecedent activation of mechanisms that promote and inhibit apoptosis were compared. Surprisingly, although ethanol exposure markedly enhanced LPS-induced liver injury under these experimental conditions, it actually inhibited caspase 3 activation. Moreover, the increased liver damage in the ethanol-fed group was not associated with increased cytochrome c release or nuclear oligonucleosomal DNA fragmentation. Taken together, these findings demonstrate a discordance between the previously reported in vitro mechanisms for ethanol hepatotoxicity and those that operate to sensitize ethanol-exposed livers to TNF toxicity in vivo. In vivo, ethanol-related potentiation of TNF-mediated liver injury does not require caspase 3 activation, and hepatocyte death appears to result more from lysis rather than from increased, classical apoptosis.DISCUSSIONDecades of research have culminated in relatively recent insights that are widely believed to explain paradoxical inter- and intra-individual differences in the vulnerability to alcohol-induced liver damage. Namely, several lines of evidence suggest that serious alcohol-related liver damage requires the introduction of an inflammatory or oxidant stress to ethanol-exposed livers that have become partially depleted of various survival factors that normally protect hepatocyte viability (58.Tilg H. Diehl A.M. N. Engl. J. Med. 2000; 343: 1467-1476Crossref PubMed Scopus (830) Google Scholar). Thus, moment-to-moment differences in the balance between cytotoxic and cytoprotective factors dictate the severity of alcohol-related liver damage at any given point in time. Several laboratories have exploited this general concept to develop cell culture and animal models that can be studied to characterize molecular targets for future therapeutic interventions. Work from many different groups identifies TNFα as a critical mediator of alcohol-related liver damage (14.Honchel R. Ray R.B. Marsano L. Cohen D. Lee E. Shedlosky S. McClain C.J. Alcohol. Clin. Exp. Res. 1992; 16: 665-669Crossref PubMed Scopus (107) Google Scholar, 15.Zeldin G. Rai R. Yang S.Q. Lin H.Z. Yin M. Diehl A.M. Alcohol. Clin. Exp. Res. 1996; 20: 1639-1645Crossref PubMed Scopus (55) Google Scholar, 16.Fernandez-Checa J.C. Kaplowitz N. Garcia-Ruiz C. Colell A. Miranda M. Marie M. Ardite E. Morales A. Am. J. Physiol. 1997; 273: G7-G17Crossref PubMed Google Scholar, 17.Yin M. Wheeler M.D. Kono H. Bradford B.U. Gallucci R.M. Luster M.I. Thurman R.G. Gastroenterology. 1999; 117: 942-952Abstract Full Text Full Text PDF PubMed Scopus (634) Google Scholar, 18.Pastorino J.G. Hoek J.B. Hepatology. 2000; 31: 1141-1152Crossref PubMed Scopus (175) Google Scholar). Recent studies of hepatocytes that were exposed to ethanol and then treated with TNFα in vitro demonstrated an enhanced induction of several events that are involved in apoptotic cell death (18.Pastorino J.G. Hoek J.B. Hepatology. 2000; 31: 1141-1152Crossref PubMed Scopus (175) Google Scholar), raising the exciting possibility that agents that prevent TNF-dependent activation of caspase 3 might be effective as treatments for alcoholic liver disease. However, although there is evidence of increased hepatocyte apoptosis in some patients and experimental animals with alcohol-related liver damage (59.Galle P.R. J. Hepatol. 1997; 27: 405-412Abstract Full Text PDF PubMed Scopus (83) Google Scholar, 60.Goldin R.D. Hunt N.C. Clark J. Wickeramasinghe S.N. J. Pathol. 1993; 171: 73-76Crossref PubMed Scopus (90) Google Scholar, 61.Baroni G.S. Marucci L. Benedetti A. Mancini r. Jezequel A.-M. Orlandi F. J. 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Moreover, we find that increased liver damage in ethanol-fed mice occurs despite robust, acute induction of various survival responses, including MAPKs, AKT, NF-κB, and iNOS, which usually protect hepatocytes from apoptosis following LPS exposure (30.Kim Y.M. Kim T.H. Chung H.T. Talanian R.V. Yin X.M. Billiar T.R. Hepatology. 2000; 32: 770-778Crossref PubMed Scopus (209) Google Scholar,47.Iimuro Y. Nishiura T. Hellerbrand C. Behrns K.E. Schoonhoven R. Grisham J.W. Brenner D.A. J. Clin. Invest. 1998; 101: 802-811Crossref PubMed Scopus (418) Google Scholar, 55.Kim Y.M. Lee B.S. Yi K.Y. Paik S.G. Biochem. Biophys. Res. Commun. 1997; 236: 655-660Crossref PubMed Scopus (144) Google Scholar, 65.Cross T.G. Scheel-Toellner D. Henriquez N.V. Deacon E. Salmon M. Lord J.M. Exp. Cell Res. 2000; 256: 34-41Crossref PubMed Scopus (616) Google Scholar). 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These factors are produced primarily by hepatic macrophages (68.Decker K. Eur. J. Biochem. 1990; 192: 245-261Crossref PubMed Scopus (880) Google Scholar) and thus are not abundant in hepatocyte cultures. Our ribonuclease protection analyses of liver RNA demonstrate that in vivo exposure to ethanol amplifies basal and LPS-related induction of several cytokines that inhibit apoptosis. At least one of these (i.e. IL-15) inhibits the recruitment of adaptor molecules to the cytosolic domains of Fas and TNFR-1, blocking the propagation of apoptotic signals that are normally initiated by these receptors (57.Waldman T.A. Dubois S. Tagaya Y. Immunity. 2001; 14: 105-110PubMed Google Scholar). Therefore, in intact mice, IL-15 and other cytokines may abrogate apoptotic responses that proceed unchecked in isolated hepatocytes cultured without these factors. Additional experiments are needed to evaluate this possibility directly. Chronic ethanol exposure also modifies the availability of intracellular factors that regulate apoptosis. For example, chronic consumption of ethanol inhibits hepatic mitochondrial respiration and ATP synthesis in patients and experimental animals (69.Cunningham C.C. Coleman W.B. Spach P.I. Alcohol Alcoholism. 1990; 25: 127-136Crossref PubMed Scopus (147) Google Scholar, 70.Hoek J.B. Lee C.P. Current Topics Bioenerg.17. Academic Press Limited, London1994: 197-241Google Scholar). This influences the zonal distribution of alcohol-related liver injury, which typically clusters in the most hypoxic parts of liver lobules around terminal hepatic venules, where we observed the most severe injury after LPS. However, hepatocytes in more well oxygenated areas of the liver usually are less ATP-depleted and survive (4.Thurman R.G. Ji S. Matsumura T. Lemasters J.J. Fundam. Appl. Toxicol. 1984; 4: 125-133Crossref PubMed Google Scholar, 5.Israel Y. Orrego H. Recent Dev. Alcohol. 1984; 2: 119-133Crossref PubMed Scopus (28) Google Scholar). Indeed, moderate reductions in cellular ATP content might even be somewhat protective (71.Lemasters J.J. Qian T. Bradham C.A. Brenner D.A. Cascio W.E. Trost L.C. Nishimura Y. Nieminen A.-L. Herman B. J. Bioenerg. Biomembr. 1999; 31: 305-318Crossref PubMed Scopus (338) Google Scholar) because the final stages of caspase 3 activation do not occur when ATP levels dip below a certain threshold (72.Leist M. Single M. Castoldi A.F. Kuhnle S. Nicotera P. J. Exp. Med. 1997; 185: 1481-1486Crossref PubMed Scopus (1636) Google Scholar). However, any situation that causes extreme ATP depletion impairs the functioning of ion transporters that regulate membrane permeability, and eventually this leads to organelle and cell lysis, i.e. necrosis (5.Israel Y. Orrego H. Recent Dev. Alcohol. 1984; 2: 119-133Crossref PubMed Scopus (28) Google Scholar,73.Eguchi Y. Shimizu S. Tsujimoto Y. 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Cell Res. 1999; 246: 221-232Crossref PubMed Scopus (52) Google Scholar), it is unlikely that ethanol-exposed hepatocytes with marginal ATP stores can survive this process. Thus, only the healthiest liver cells with relatively normal ATP contents are likely to be cultured from ethanol-treated animals. This methodological obstacle is not overcome by exposing primary hepatocytes from ethanol-naive animals to ethanol in vitrobecause brief periods of ethanol treatment are probably not sufficient to reproduce the full spectrum of ethanol-related mitochondrial toxicity that inhibits hepatic ATP synthesis during chronic in vivo exposure to ethanol (84.Bailey S.M. Pietsch E.C. Cunningham C.C. Free Radic. Biol. Med. 1999; 27: 891-900Crossref PubMed Scopus (181) Google Scholar). Even efforts to substitute hepatocyte cell lines for primary hepatocytes are potentially problematic because, like other neoplastic cells, HepG2 cells have induced nonmitochondrial pathways for ATP synthesis (38.Capuano F. Guerrieri F. Papa S. J. Bioenerg. Biomembr. 1997; 29: 379-384Crossref PubMed Scopus (74) Google Scholar) and thus might not experience ATP depletion despite ethanol-related mitochondrial toxicity. The fact that cells with relatively normal ATP stores are over-represented in the previously mentioned in vitro models introduces a bias that favors the apoptotic (rather than necrotic) death pathway.Thus, because in vitro hepatocyte culture systems do not fully reproduce the extra- or intracellular microenvironment of hepatocytes in intact livers, it is somewhat risky to extrapolate the resultant in vitro data to more clinically relevant in vivo models of chronic ethanol exposure (69.Cunningham C.C. Coleman W.B. Spach P.I. Alcohol Alcoholism. 1990; 25: 127-136Crossref PubMed Scopus (147) Google Scholar, 70.Hoek J.B. Lee C.P. Current Topics Bioenerg.17. Academic Press Limited, London1994: 197-241Google Scholar). This, in turn, has potentially important therapeutic implications that merit future evaluation because the in vivo data suggest that pharmacologic inhibition of caspase 3 may prove to be unnecessary (because it occurs naturally) in subjects who habitually consume alcohol. If so, then this intervention will not protect alcohol abusers from alcohol-related liver injury, and alternative therapeutic strategies will need to be developed for this important cause of chronic liver disease. Although ethyl alcohol has been recognized as a significant hepatotoxin for centuries, the mechanisms involved in alcohol-induced liver disease remain uncertain. It is known that chronic alcohol ingestion potentiates liver injury inflicted by many other toxins (1.Seeff L.B. Cuccherini B.A. Zimmerman H.J. Alder E. Benjamin S.B. Ann. Intern. Med. 1986; 104: 399-404Crossref PubMed Scopus (369) Google Scholar), viral hepatitis (2.Marsano L.S. Pena L.R. Hepatogastroenterology. 1998; 45: 331-339PubMed Google Scholar, 3.Serfaty L. Chazouilleres O. Poujol-Robert A. Morand-Joubert L. Dubois C. Chretien Y. Poupon R.E. Petit J.C. Poupon R. Hepatology. 1997; 26: 776-779Crossref PubMed Scopus (116) Google Scholar), and hepatic hypoxia-reoxygenation (4.Thurman R.G. Ji S. Matsumura T. Lemasters J.J. Fundam. Appl. Toxicol. 1984; 4: 125-133Crossref PubMed Google Scholar, 5.Israel Y. Orrego H. Recent Dev. Alcohol. 1984; 2: 119-133Crossref PubMed Scopus (28) Google Scholar) or ischemia-reperfusion (6.Gao W. Connor H.D. Lemasters J.J. Mason R.P. Thurman R.G. Transplantation. 1995; 59: 674-679Crossref PubMed Scopus (60) Google Scholar) and yet fails to cause serious liver damage in most healthy adults (7.Lelbach W.K. Prog. Liver Dis. 1976; 5: 494-513PubMed Google Scholar, 8.Pequignot G. Tuyns A.S. Berta J.L. Int. J. Epidemiol. 1978; 7: 113-120Crossref PubMed Scopus (191) Google Scholar, 9.Johnson R.D. Williams R. Alcohol Alcohol. 1985; 20: 137-142PubMed Google Scholar). These clinical observations suggest that ethanol exposure enhances hepatic vulnerability to a secondary inflammatory or oxidative stress, such that serious liver injury is most likely to occur when proinflammatory/pro-oxidant factors are superimposed onto a background of ethanol use (10.Day C.P. James O. Gastroenterology. 1998; 114: 842-845Abstract Full Text Full Text PDF PubMed Scopus (3406) Google Scholar). Because tumor necrosis factor-α ((TNFα)1 and cytokines that potentiate TNF activity are the common, proximal mediators of inflammatory liver injury (11.Vassalli P. Annu. Rev. Immunol. 1992; 10: 411-452Crossref PubMed Scopus (1798) Google Scholar, 12.Tracey K.J. Cerami A. Annu. Rev. Cell Biol. 1993; 9: 317-343Crossref PubMed Scopus (760) Google Scholar, 13.Baker S.J. Reddy E.P. Oncogene. 1996; 12: 1-9PubMed Google Scholar), several laboratories have been evaluating the possibility that ethanol may sensitize hepatocytes to TNF-related toxicity (14.Honchel R. Ray R.B. Marsano L. Cohen D. Lee E. Shedlosky S. McClain C.J. Alcohol. Clin. Exp. Res. 1992; 16: 665-669Crossref PubMed Scopus (107) Google Scholar, 15.Zeldin G. Rai R. Yang S.Q. Lin H.Z. Yin M. Diehl A.M. Alcohol. Clin. Exp. Res. 1996; 20: 1639-1645Crossref PubMed Scopus (55) Google Scholar, 16.Fernandez-Checa J.C. Kaplowitz N. Garcia-Ruiz C. Colell A. Miranda M. Marie M. Ardite E. Morales A. Am. J. Physiol. 1997; 273: G7-G17Crossref PubMed Google Scholar, 17.Yin M. Wheeler M.D. Kono H. Bradford B.U. Gallucci R.M. Luster M.I. Thurman R.G. Gastroenterology. 1999; 117: 942-952Abstract Full Text Full Text PDF PubMed Scopus (634) Google Scholar, 18.Pastorino J.G. Hoek J.B. Hepatology. 2000; 31: 1141-1152Crossref PubMed Scopus (175) Google Scholar). The evidence supporting this concept is growing. 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Assuming that similar mechanisms operate in humans who habitually consume alcohol-containing beverages, then is it important to delineate the cellular mechanisms that mediate ethanol-related sensitization to TNF lethality. Healthy hepatocytes are typically resistant to TNFα toxicity unless they have been pretreated with agents that inhibit protein synt
Hepatic steatosis may have a generally benign prognosis, either because most hepatocytes are not significantly injured or mechanisms to replace damaged hepatocytes are induced. To determine the relative importance of these mechanisms, we compared hepatocyte damage and replication in ethanol-fed and ob/ob mice with very indolent fatty liver disease to that of healthy control mice and PARP-1(-/-) mice with targeted disruption of the DNA repair enzyme, poly(ADP-ribose) polymerase. Compared to the healthy controls, both groups with fatty livers had significantly higher serum alanine aminotransferase values, hepatic mitochondrial H(2)O(2) production, and hepatocyte oxidative DNA damage. A significantly smaller proportion of the hepatocytes from fatty livers entered S phase when cultured with mitogens. Moreover, this replicative senescence was not reversed by treating cultured hepatocytes with agents (i.e., betaine or leptin) that improve liver disease in intact ethanol-fed or leptin-deficient mice. Hepatocytes from PARP1(-/-) mice also had more DNA damage and reduced DNA synthesis in response to mitogens. However, neither mice with fatty livers nor PARP-1-deficient mice had atrophic livers. All of the mice with senescent mature hepatocytes exhibited hepatic accumulation of liver progenitor (oval) cells and oval cell numbers increased with the demand for hepatocyte replacement. Therefore, although hepatic oxidant production and damage are generally increased in fatty livers, expansion of hepatic progenitor cell populations helps to compensate for the increased turnover of damaged mature hepatocytes. In conclusion, these results demonstrate that induction of mechanisms to replace damaged hepatocytes is important for limiting the progression of fatty liver disease.
Doses of ethanol (EtOH) that are not overtly cytotoxic inhibit mitogen-induced hepatocyte proliferation and delay liver regeneration after 70% partial hepatectomy (PH). The mechanisms for this are poorly understood. This study evaluates the hypothesis that EtOH inhibits hepatocyte proliferation after PH by inducing redox-sensitive factors, such as p38 mitogen-activated protein kinase (MAPK) and p21 (WAF1/CIP1), that protect cells from oxidative stress but prevent cell-cycle progression by inhibiting cyclin D1.Mechanisms that regulate the transition from the prereplicative G1 phase of the cell cycle into S phase were compared in EtOH-fed mice and normal pair-fed mice after PH.Prior EtOH exposure significantly increases p38 MAPK and p21 after PH. This is accompanied by reduced expression of cyclin D1 messenger RNA and protein, increases in other cell-cycle regulators (such as signal transducer and activator of transcription-3 and p27) that are normally inhibited by cyclin D1, and hepatocyte G1 arrest.EtOH amplifies G1 checkpoint mechanisms that are induced by oxidative stress and promotes hepatic accumulation of factors, including p38 MAPK, p21, and signal transducer and activator of transcription-3, that enhance cellular survival after oxidant exposure. Therefore, cell-cycle inhibition may be an adaptive response that helps EtOH-exposed livers survive situations, such as PH, that acutely increase reactive oxygen species in hepatocytes.
