tion of TGF-� such as growth inhibition and apoptosis of hepatocytes.For example, Bcl-2 repression by TGF-� requires C-terminal phos-phorylation of Smad3.6 To evade apoptosis, HCC might acquire Bcl-2overexpression induced by pSmad3L pathway. Escaping apoptosis is acritical step in the progression to full malignancy of cancers, whichovercome multiple fail-safe genetic controls.

From the viewpoint of TGF-� signaling, a key therapeutic aim incancer would be restoration of the lost tumor suppressor functionobserved in normal hepatocytes at the expense of effects promotingaggressive behavior in HCC. Our model suggests that specific inhibi-tors of the pSmad3L-mediated oncogenic/fibrogenic signaling shouldinhibit progression of HCC.

KOICHI MATSUZAKI

Departments of Gastroenterology and Hepatology,Kansai Medical University, Osaka, Japan

IWATA OZAKI

Health Administration Center,Saga Medical School,Saga University, Saga, Japan

References1. Matsuzaki K, Murata M, Yoshida K, Sekimoto G, Uemura Y, Sakaida N, et

al. Chronic inflammation associated with hepatitis C viral infection perturbs

hepatic TGF-� signaling, promoting cirrhosis and hepatocellular carci-noma. HEPATOLOGY 2007;46:48-57.

2. Zavadil J, Bottinger EP. TGF-� and epithelial-to-mesenchymal transitions.Oncogene 2005;24:5764-5774.

3. Weng HL, Ciuclan L, Liu Y, Hamzavi J, Godoy P, Gaitantzi H, et al.Profibrogenic transforming growth factor-�/activin receptor-like kinase 5signaling via connective tissue growth factor expression in hepatocytes.HEPATOLOGY 2007;46:1257-1270.

4. Kaimori A, Potter J, Kaimori JY, Wang C, Mezey E, Koteish A. Trans-forming growth factor-beta1 induces an epithelial-to-mesenchymal tran-sition state in mouse hepatocytes in vitro. J Biol Chem 2007;282:22089-22101.

5. Sekimoto G, Matsuzaki K, Yoshida K, Mori S, Murata M, Seki T, et al.Reversible Smad-dependent signaling between tumor suppression and on-cogenesis. Cancer Res 2007;67:5090-5096.

6. Yang YA, Zhang GM, Feigenbaum L, Zhang YE. Smad3 reduces suscepti-bility to hepatocarcinoma by sensitizing hepatocytes to apoptosis throughdownregulation of Bcl-2. Cancer Cell 2006;9:445-457.

Copyright © 2008 by the American Association for the Study of Liver Diseases.Published online in Wiley InterScience (www.interscience.wiley.com).DOI 10.1002/hep.22199Potential conflict of interest: Nothing to report.

Disease-Specific Autoantibodies in Patients with Acute Liver Failure: The King’s College LondonExperience

To the Editor:

We read with interest the article by Leung et al.,1 who investigated thepresence of antimitochondrial antibodies (AMAs), the hallmark of pri-mary biliary cirrhosis, in patients with acute liver failure (ALF). This is acondition frequently characterized by oxidative stress,2 which is a possibletrigger for AMA production3; this is the rationale of the study. As controls,antibodies to glycoprotein 210 kDa (gp210), speckled protein 100 kDa(sp100), centromere, chromatin, soluble liver antigen (SLA), tissue trans-glutaminase, and deaminated gliadin peptides were also studied.

AMAs were sought by a highly sensitive and specific triple-hybrid(mitochondrial antigen 3 [MIT3]) enzyme-linked immunosorbent assay(ELISA) containing the immunodominant mitochondrial epitopes of E2subunits of pyruvate dehydrogenase, branched chain 2-oxo-acid dehydro-genase, and 2-oxo-glutarate dehydrogenase and were found in 28/69(40.6%) patients, 9 (13%) of whom had acetaminophen-related ALF.The only additional antibody reactivity present in a significant percentage(57.1%) was that against anti–tissue transglutaminase, whereas other au-toantibodies such as those against SLA were practically absent when test-ing was performed with a commercially available ELISA.1

Investigating an autoimmune component of ALF, we recently de-scribed autoantibody reactivity in an ALF series in the United King-dom comprising 73 patients4 and, in apparent contrast to Leung etal.,1 found virtually no AMAs (1/73, 1%) through testing with immu-nofluorescence but did find anti-SLA antibodies in 22% through test-ing with a sensitive radioligand assay.5 Because discrepancies betweenthe two studies may be due to methodological differences, we retested47 of our ALF patients in whom sera were available both by conven-tional indirect immunofluorescence6 and by an MIT3 ELISA similarto that used by Leung et al. Although all sera remained negative forAMA by immunofluorescence, 13/47 (28%) were positive by theMIT3 assay. Conversely, when for the detection of anti-SLA we usedan ELISA kit similar to that used by Leung et al. which is less sensitivethan a radioligand assay, we also found practically no antibody reac-

tivity. Our results indicate the influence of the methodological ap-proach on the results obtained in comparable patient cohorts. We cananticipate anti-SLA to be present in the American series if a sensitiveradioligand assay is used for their detection. We found no relationbetween the presence of anti-MIT3 AMAs and patient age, sex, bio-chemical parameters of liver injury, immunoglobulin response modeof presentation of ALF, or outcome of illness.

Both Leung et al.’s study1 and our study show that autoantibodyreactivity is frequent in a condition such as ALF characterized by extensiverelease of autoantigens subsequent to massive liver damage. This autore-activity is, however, transient because, on repeat testing, AMAs in Leung etal.’s study and anti-SLA in our own cohort disappear.

In conclusion, reactivities to antigens specific to autoimmune liverdiseases are present in the course of ALF, although their detectiondepends on the methodology used. These specific autoimmune reac-tivities are likely to arise from the release of antigens from damagedhepatocytes and stimulation of lymphocytes that have escaped negativedeletion, but they do not, however, progress to chronic autoimmuneliver disease, probably because of the intervention of regulatory mech-anisms after the acute phase of the disease is over. A deficiency of suchmechanisms characterizes both autoimmune hepatitis and primary bil-iary cirrhosis.6-8

In view of the discrepancy in results when different techniques areused, standardization of methodological approaches for autoantibodydetection is warranted to avoid confusing the clinician.

WILLIAM BERNAL

FRANCESCA MEDA

YUN MA

DIMITRIOS P. BOGDANOS

DIEGO VERGANI

Institute of Liver StudiesKing’s College London School of Medicine at King’s College HospitalLondon, United Kingdom

1096 CORRESPONDENCE HEPATOLOGY, March 2008

References1. Leung PS, Rossaro L, Davis PA, Park O, Tanaka A, Kikuchi K, et al.

Antimitochondrial antibodies in acute liver failure: implications for pri-mary biliary cirrhosis. HEPATOLOGY 2007;46:1436-1442.

2. Ferret PJ, Hammoud R, Tulliez M, Tran A, Trebeden H, Jaffray P, et al.Detoxification of reactive oxygen species by a nonpeptidyl mimic of super-oxide dismutase cures acetaminophen-induced acute liver failure in themouse. HEPATOLOGY 2001;33:1173-1180.

3. Wu CT, Eiserich JP, Ansari AA, Coppel RL, Balasubramanian S, BowlusCL, et al. Myeloperoxidase-positive inflammatory cells participate in bileduct damage in primary biliary cirrhosis through nitric oxide-mediatedreactions. HEPATOLOGY 2003;38:1018-1025.

4. Bernal W, Ma Y, Smith HM, Portmann B, Wendon J, Vergani D. Thesignificance of autoantibodies and immunoglobulins in acute liver failure:a cohort study. J Hepatol 2007;47:664-670.

5. Ma Y, Okamoto M, Thomas MG, Bogdanos DP, Lopes AR, Portmann B, etal. Antibodies to conformational epitopes of soluble liver antigen define asevere form of autoimmune liver disease. HEPATOLOGY 2002;35:658-664.

6. Longhi MS, Ma Y, Bogdanos DP, Cheeseman P, Mieli-Vergani G, Ver-gani D. Impairment of CD4(�)CD25(�) regulatory T-cells in autoim-mune liver disease. J Hepatol 2004;41:31-37.

7. Longhi MS, Ma Y, Mitry RR, Bogdanos DP, Heneghan M, Cheeseman P, etal. Effect of CD4� CD25� regulatory T-cells on CD8 T-cell function inpatients with autoimmune hepatitis. J Autoimmun 2005;25:63-71.

8. Longhi MS, Hussain MJ, Mitry RR, Arora SK, Mieli-Vergani G, VerganiD, et al. Functional study of CD4�CD25� regulatory T cells in healthand autoimmune hepatitis. J Immunol 2006;176:4484-4491.

Copyright © 2008 by the American Association for the Study of Liver Diseases.Published online in Wiley InterScience (www.interscience.wiley.com).DOI 10.1002/hep.22179Potential conflict of interest: Nothing to report.

Reply:

Unless we are both studying the same sera, there is no basis ofcomparison. In our clinical cases and I assume their clinical series,many types of ALF were studied and we both found AMAs when weused the same method. It is also obvious that no international stan-dardization exists, and measurements of all these antibodies will not bestandardized until the release of the newer multiphasic kits in 2009 andbeyond.

M. ERIC GERSHWIN, M.D.Division of Rheumatology/AllergyUniversity of California, Davis Medical CenterSacramento, CA

Copyright © 2008 by the American Association for the Study of Liver Diseases.Published online in Wiley InterScience (www.interscience.wiley.com).DOI 10.1002/hep.22198Potential conflict of interest: Nothing to report.

Adoptive Transfer of Regulatory T Cells in an Animal Model of a Diet-Induced Fatty Liver

To the Editor:

I read with interest an excellent study by Ma et al. that clarifiedthe mechanisms of progression from hepatic steatosis to nonalco-holic steatohepatitis (NASH) in a mouse model of a high-diet-induced fatty liver.1 The authors revealed that CD4� CD25�

forkhead box P3� hepatic regulatory T cells (T regs) decreasedduring the administration of a high-fat diet, and the levels of T regswere less than half of those in control mice at the end of the 8-weekexperiment. Marked reduction of T regs, which was caused byapoptosis due to oxidative stress, was associated with enhancedexpression of tumor necrosis factor (TNF) alpha and its intracellu-lar mediators in liver. Adoptive-transferred T regs suppressed high-fat diet-induced intrahepatic TNF alpha signaling. Mice in whichT regs were depleted or T regs were transferred, both of which werefed diets containing a high amount of fat, were given a singlerelatively low dose of lipopolysaccharide (LPS) (100 �g/mouse) toinduce hepatic injury. Six hours after injection of LPS, LPS-in-duced liver damage was compared between the groups. A pretreat-ment with adoptive transfer of T regs significantly attenuatedhepatotoxicity of LPS in mice with hepatic steatosis. The authorsspeculate that enhanced LPS signaling followed by proinflamma-tory cytokine production as a second hit necessary for the progres-sion from steatosis to nonalcoholic steatohepatitis can be reducedby restoring the function of T regs.

T regs are engaged in the regulation of organ-specific immuneresponses to pathogens and self-proteins. A recent in vitro studyindicates that activated T regs rapidly inhibit induction of Th1cytokine mRNA, whereas suppression of anti-inflammatory Th2cytokines is delayed.2 This article by Ma and colleagues may be thecornerstone for a better understanding of a variety of immune-related liver diseases in addition to nonalcoholic liver disease. Iwould like to ask the authors whether animal models with chronic

endotoxemia, which are described in a recent article,3 are moresuitable than those with acute endotoxemia for the study to inves-tigate the role of T regs in liver disease. Mild chronic endotoxemiahas been observed in chronic liver disease,4 chronic kidney disease,5

and chronic cardiovascular disease.6

TETSUJI FUJITA, M.D.Department of SurgeryJikei University School of MedicineTokyo, Japan

References1. Ma X, Hua J, Mohamood AR, Hamad ARA, Ravi R, Li Z. A high-fat diet

and regulatory T cells influence susceptibility to endotoxin-induced liverinjury. HEPATOLOGY 2007;46:1519-1529.

2. Oberle N, Eberhardt N, Falk CS, Krammer PH, Suri-Payer E. Rapidsuppression of cytokine transcription in human CD4�CD25� T cells byFoxp3� regulatory T cells: independence of IL-2 consumption, TGF-�,and various inhibitors of TCR signaling. J Immunol 2007;179:3578-3587.

3. Cani PD, Amar J, Iglesias MA, Poggi M, Knauf C, Bastelica D, et al.Metabolic endotoxemia initiates obesity and insulin resistance. Diabetes2007;56:1761-1772.

4. Fukui K. Relation of endotoxin, endotoxin binding proteins and macro-phages to severe alcoholic liver injury and multiple organ failure. AlcoholClin Exp Res 2005;29:172S–179S.

5. Goncalves S, Pecoits-Filho R, Perreto S, Barberato SH, Stinghen AEM,Lima EGA, et al. Association between renal function, volume status andendotoxaemia in chronic kidney disease patients. Nephrol Dial Transplant2006;21:2788-2794.

HEPATOLOGY, Vol. 47, No. 3, 2008 CORRESPONDENCE 1097