The immune system is responsible for defending the body against other organisms. It must recognize the presence of an invader, discriminating in the process between self and nonself. Disorders in which the immune system reacts against self-components of the body are called autoimmune diseases. In children, the liver may be the target of an autoimmune reaction. In patients with autoimmune hepatitis, the main target is the hepatocyte. In sclerosing cholangitis, the prime targets are the bile duct cells. Overlapping syndromes have been described in which this kind of differentiation is less evident. Other immune-mediated processes with characteristics similar to autoimmune liver diseases have been observed in transplanted livers and in the livers of patients who received bone marrow transplants and have graft-versus-host disease (GvHD). In addition, in several cases of chronic viral liver infection, an autoimmune reaction may develop, as is shown by the presence of autoantibodies in sera and the aggravation of the disease when a patient receives treatment with interferon (an immune response stimulant). Altogether, these facts suggest that studies concerning liver autoimmunity could have wide applications in hepatology.
AREAS OF EMPHASIS
Autoimmune Hepatitis (AIH)
Autoimmune hepatitis is a disease of unknown origin in which progressive destruction of the hepatic parenchyma occurs (1–3). Although the peak incidence is in adolescent females, AIH has been diagnosed in infants as young as 6 months of age (2). The disease carries a high mortality rate if untreated with immunosuppressants; however, the drugs currently used for treatment have undesired side effects (2,3). Even with treatment, progression to cirrhosis and end-stage liver disease may occur in 10% to 20% of cases and liver transplantation may be necessary(2,3). However, the disease may recur after transplantation, and perhaps because of the underlying immune dysregulation, rejection may be more severe and problematic. This disease follows a chronic but fluctuating course and is associated with marked hypergammaglobulinemia, circulating autoantibodies, low levels of complement factor 4, and a high prevalence of HLA B8 and DR3 and DR4 haplotypes. Other autoimmune disorders are found frequently in patients with autoimmune hepatitis and in their first-degree relatives (1–3). Autoimmune-associated disorders are of B- and T-lymphocyte origin, and other anomalies of the immune system such as low levels of circulating IgA are common. Different subgroups of AIH can be distinguished according to the autoantibody found in the serum. Autoimmune hepatitis type 1 is characterized by the presence of anti–smooth muscle antibodies in patients' sera (2,4). This autoantibody recognizes specifically the actin filaments. Antinuclear antibodies are found in sera of 70% of patients with AIH type 1 but are rarely the only marker in children. Autoimmune hepatitis type 2 is defined by the detection of liver–kidney microsomal antibody type 1. This autoantibody is directed against the cytochrome P450 2D6. In 30% of patients with AIH type 2, an autoantibody against a liver cytosolic (LC1) protein of 60 kd is found. The LC1 autoantibodies recognize a liver-specific protein called formimino-transferase cyclodeaminase (5). Autoimmune hepatitis type 1 is the most frequent subgroup in North American children.
These clinical and laboratory observations led to the hypothesis that AIH is a multifactorial disease. In attempting to explain the pathogenesis of this disorder, it is possible to establish four groups of factors: 1) genetic background, 2) age and sex, 3) modification of the immunologic system, and 4) environmental factors that may trigger the autoimmune process. For some time, the pathogenesis of autoimmune hepatitis has been analyzed primarily by using human material from patients and their families. This type of analysis will continue to be useful in the years to come but is limited by the availability of material. Application of the knowledge obtained from patients could lead to the development of an animal model that would give new insights into the relative importance of the pathogenic factors listed above.
Sclerosing Cholangitis (SC)
Sclerosing cholangitis is a chronic inflammatory disease affecting mainly bile ducts of different sizes (6). Sclerosing cholangitis afflicts boys twice as frequently as it affects girls (6–8). No medication used to date has been effective in stopping the progression to biliary cirrhosis. If liver transplantation is not performed, bile duct cancer (cholangiocarcinoma) may develop (9). The incidence of SC in childhood is unknown; about 75% of cases in adults are associated with inflammatory bowel diseases (IBD) (6). The diagnosis of this chronic liver disease can precede the clinical onset of IBD. In children, poor outcome (death or liver transplantation) has been observed in 35% to 56% of patients with IBD-associated or idiopathic SC (7,8).
For both diseases, more precise and thorough definition of the liver-specific antigens that provoke the immune response and the stimuli that promote the expression of those antigens is necessary, as is better means of inhibiting an immune attack on the liver without the deleterious consequences of general host immunosupression. A better definition of the type of liver-specific antigen should allow a more precise and earlier diagnosis. At present, no good animal model exists for either disease; thus development of appropriate models is a priority. Finally, although a number of large-scale multicenter clinical trials have been performed in adults with autoimmune hepatitis and sclerosing cholangitis, only a few anecdotal experiences have been published regarding the treatment of either disease in children. The congressional mandate to include children in therapeutic trials should stimulate the organization of appropriate large-scale, randomized, prospective trials of new therapies for these diseases in children and adolescents.
Mechanisms responsible for the onset, the development, and the evolution of autoimmune hepatitis and sclerosing cholangitis must be characterized more thoroughly.
An animal model will allow the study of 1) the immunologic mechanisms of the autoimmune reaction in the liver; 2) the influence of age, sex, and genetic background in the level and persistence of the liver-specific autoimmune response; 3) the possibility of bringing about a transition phase from an acute (virus or drug induced) to a chronic disease; and 3) the role of cytokines in the liver injury.
Research on circulating autoantibodies will 1) explain why specific and nonspecific autoantibodies develop in the sera of patients with AIH, 2) identify liver-specific antigens recognized by B lymphocytes, and 3) allow design of new diagnostic tests.
Isolation and characterization of liver-specific T-cell clones will help 1) identify liver-specific antigens recognized by T lymphocytes, and 2) allow design of new forms of treatment.
Clinical trials with recently developed immunosuppressive drugs will 1) decrease the harmful consequences of the high doses of corticosteroids currently used for treatment, and 2) avoid the development of cirrhosis and hepatic failure observed in patients with partial or no response to currently available treatments.
Genetic analysis will help 1) define and confirm the contribution of the MHC loci to the development of AIH, and 2) identify candidate genes (non-MHC) that may be involved in the pathogenesis of AIH.
An animal model will allow study of 1) the effect on the bile duct epithelium of different cytokines and bacterial antigens produced or absorbed during inflammatory colitis, 2) the influence of sex and genetic background in the development of SC, and 3) the immunologic mechanisms of bile duct epithelium injury.
Development of bile duct epithelium in culture will allow analysis of 1) the expression of MHC proteins on the cell surface, 2) the response of these cells to an immune-mediated aggression, and 3) the modification of both processes by different drugs.
Identification of specific serological markers will permit 1) early diagnosis and 2) improved monitoring of the response to therapy.
Isolation and characterization of T-cell clones from SC livers, transplanted livers, and biopsied tissues of patients with GvH will allow the identification of specific bile duct cell antigens.
Genetic profiles (MHC and non-MHC) will be an important tool for identifying patients with IBD at risk for SC.
Several models of acute, subacute, and chronic hepatitis could be tested, using drugs, viruses, antibodies, or liver subcellular fractions as triggers (10). Transgenic mice expressing particular cytokines develop hepatitis (11,12). Two mechanisms of cell death could be explored in these models, the cell lysis and cell apoptosis induced by an immune attack of the liver tissue. Antigenic sites recognized by B or T cells involved in the autoimmune response have shown sequence homology with viral proteins. This process is termed molecular mimicry and can be well studied using a transgenic mice model in which viral proteins are expressed and in which a viral infection triggers the autoimmune process.
Organ-specific autoantibodies, such as anti–liver cytosol antibodies (5) anti-Asialoglycoprotein receptor antibodies (13), and anti–liver-soluble protein (14) should be the focus of research. The diagnostic and prognostic usefulness of these autoantibodies should be assessed in children. New, easier, and cheaper diagnostic methods should be developed. Identification and isolation of autoantigens and of specific antigenic sequences or structures should clarify the mechanisms of autoantibody development.
Characterization of the T lymphocytes proliferating in the presence of liver-specific antigens would be a major step in the comprehension of the disease. Analyses of the T-cell type and repertoire and of the specific epitopes that stimulated the cell proliferation are necessary steps in the search for new therapies.
Clinical Trials of Treatment
Several immunosuppressive agents (cyclosporine, tacrolimus, rapamycin, mycophenolate mofetil, anti–IL2-R antibodies) have been studied recently in patients after liver transplantation. These medications could produce a rapid and complete regression of the autoimmune process in most patients while avoiding the consequences of long-term administration of high doses of corticosteroids in growing children (15).
Studies should be performed in patients and their relatives (16). Fine mapping of the susceptibility MHC-linked loci should facilitate genetic screening of individuals at high risk for developing AIH and identification of non-MHC genes that intervene in the development of AIH. Analysis of genetic and serologic markers could allow early diagnosis in susceptible individuals.
In an experimental model of ulcerative colitis in rats, a progressive SC developed. Histologic lesions similar to those observed in the livers of patients with SC have appeared in animals after chemical-induced colitis in models of small bowel bacterial overgrowth, in which bacteria-derived proinflammatory peptides seem to play a major role. Cytokine-induced bile duct injury also should be explored.
Bile Duct Cells in Culture
Primary cultures and bile duct cell lines have been obtained. The expression of MHC molecules and the regulation of its gene transcription and translation by foreign and self-products should be analyzed. These cells could be used as a target for the selection and activation of T-cell clones, and to identify specific autoantibodies.
Three types could be sought: 1) bile duct–specific molecules liberated in the blood stream during cell lysis; 2) antibodies against specific bile duct cell antigens; and 3) antineutrophil antibodies with different specificities.
T-cell Clone Isolation From the Liver of Patients With Bile Duct Immune-mediated Injury
Comparison of T-cell repertoire and types could clarify the mechanisms of immune system activation and selection.
MHC and Non-MHC Genes
Comparison of genotypes should be made between patients with IBD with and without SC, and between children with and without signs of overlapping AIH.
PROJECTED TIMETABLE AND FUNDING REQUIREMENTS
The development of animal models for autoimmune hepatitis and sclerosing cholangitis will help answer many of our current questions about the pathogenesis of these diseases. The advantages of an animal model include 1) possibly dissecting each step of the autoimmune process, 2) testing different endogenous and exogenous factors in separate sets of experiments, 3) availability of material, and 4) high reproducibility of the results. The production of an animal model is expensive and requires a long period of time to obtain the necessary number of individuals. Yet once the colony has been established, several studies can be performed at the same time. Because of the low frequency of these diseases, obtaining enough material from children would require the collaboration of several specialized centers. The same arguments could be used for clinical trials, especially when treatments are proposed to newly diagnosed patients. These multicenter studies are expensive and take approximately 4 to 6 years to be completed and analyzed.
The funding decisions concerning AIH and SC should take into account the fact that these results also will contribute to a better understanding of 1) other autoimmune extrahepatic diseases, 2) immune mechanisms of viral aggression of the liver, and 3) liver disorders observed in transplant patients. Most patients with AIH are infants and children younger than 18 years of age, and the disease is usually more severe during this period of life. New therapies developed for this population also will be useful for adult patients.
Funding for the development of an animal model should be provided through the RO1 grant mechanism at $150,000 to $200,000 per grant per year.
The definition of relevant antigens and antibodies should take 5 years, with two groups working on this problem. The required funding is estimated at $150,000/year for an ROI × two investigators × 5 years = $1,500,000. Isolation and characterization of liver-specific T-cell clones is predicted to take a similar amount of time and money, as will the work on genetic susceptibility. Thus, $4,000,000 total funding would be required for these three areas of investigation.
Organization of prospective, randomized, controlled trials of new immunosuppressive therapies for autoimmune hepatitis in children and adolescents is predicted to cost $300,000/year for 5 years. It is anticipated that part of the funding for the clinical trial(s) should come from the pharmaceutical industry.
Development of an animal model should occur through the RO1 funding mechanism, $150,000 to $200,000 per grant per year.
Study of bile duct epithelium in culture, identification of specific serologic markers, isolation and characterization of T-cell clones from livers, and studies of genetic susceptibility = $1,500,000 (as for autoimmune hepatitis above) × four areas = $6,000,000 total.
HEALTH AND ECONOMIC OUTCOMES
Any progress made in understanding a liver autoimmune disease probably could be applied to other organ-specific and non–organ-specific autoimmune diseases. Although liver rejection and GvHD do not strictly fill the definition of autoimmune disease, the mechanisms of these immune attacks on the liver could be similar to those of autoimmune hepatitis and especially to those of sclerosing cholangitis. Any advance in understanding the pathogenesis of a liver immune disease will therefore be applied to the prevention or treatment of other similar disorders.
Although no data are available on health care costs for SC and AIH, an estimate is possible using data from different sources, such as the Studies of Pediatric Liver Transplantation (SPLIT) Register that accumulates data from 28 pediatric centers in North America, and the Clinical Transplantation Register (reports from UNOS and Canada registers). According to these sources, every year 40 children and 420 adults receive transplants in North America because of SC or AIH. In a considerable number of adult patients, the onset of these diseases occurred in childhood. Considering that the cost of liver transplantation averages $150,0000 to $200,000 in the first year, at least $69,000,000 will be spent every year on transplantation alone. This does not take into account the medical costs of those patients not requiring transplant or the consequences of these diseases on patients' quality of life. In addition, patients with SC and IBD have a 50% risk for cholangiocarcinoma in the 25 years after the diagnosis of the disease (8). Occasionally, hepatocarcinoma develops in patients with AIH. Long-term follow-up of SC and AIH for early diagnosis and treatment of these serious complications adds to substantial health care costs.
The pathogenesis of autoimmune liver diseases is poorly understood, and consequently treatment is empirical. Most patients with autoimmune hepatitis are pubertal girls. At this age, the consequences of the current practice of corticosteroid administration on body self-image are devastating. In addition, complete remission is not obtained in all patients, and liver transplantation is indicated in 10% to 20% of cases. Trials of new immunosuppressive drugs should be performed. In patients with sclerosing cholangitis, no form of treatment prevents the evolution toward liver cirrhosis and hepatic failure. Efforts leading to early diagnosis, to understanding the mechanisms responsible for the destruction of the liver, and to the development of new therapies will improve the quality of life and the survival of patients with immune-mediated disorders of the liver.
1. Johnson PJ, McFarlane IG. Meeting report: International Autoimmune Hepatitis Group. Hepatology 1993; 18:998–1005.
2. Alvarez F. Autoimmune hepatitis. In: Suchy FJ, Sokol RJ, Balistreri WF, eds. Liver Disease in Children. Philadelphia: Lippincott, Williams & Wilkins; 2001:429–41.
3. Krawitt EL. Autoimmune hepatitis. N Engl J Med 1996; 334:897–903.
4. McFarlane IG. The relationship between autoimmune markers and different clinical syndromes in autoimmune hepatitis. Gut 1998; 42:599–602.
5. Lapierre P, Hajoui O, Homberg JC, et al. Formiminotransferase cyclodeaminase is an organ-specific autoantigen recognized by sera of patients with autoimmune hepatitis. Gastroenterology 1999; 116:643–9.
6. Lee Y-M, Kaplan MM. Primary sclerosing cholangitis. N Engl J Med 1995; 332:924–33.
7. Debray D, Pariente D, Urvoas E, et al. Sclerosing cholangitis in children. J Pediatr 1994; 124:49–56.
8. Wilschanski M, Chait P, Wade JA, et al. Primary sclerosing cholangitis in 32 children: clinical, laboratory, and radiographic features, with survival analysis. Hepatology 1995; 22:1415–22.
9. Broome U, Lofberg R, Veress B, et al. Primary sclerosing cholangitis and ulcerative colitis: evidence for increased neoplastic potential. Hepatology 1995; 22:1404–8.
10. Lohse AW. Experimental models of autoimmune hepatitis. Semin Liver Dis 1991; 11:241–7.
11. Toyonaga T, Hino O, Sugai S, et al. Chronic active hepatitis in transgenic mice expressing interferon-gamma in the liver. Proc Natl Acad Sci USA 1994; 91:614–8.
12. Gorham JD, Lin JT, Sung JL, et al. Genetic regulation of autoimmune disease: BALB/c background TGF beta 1-deficient mice develop necroinflammatory IFN-gamma-dependent hepatitis. J Immunol 2001; 166:6413–22.
13. Hajoui O, Debray D, Martin S, et al. Auto-antibodies to the asialoglycoprotein receptor in sera of children with auto-immune hepatitis. Eur J Pediatr 2000; 159:310–3.
14. Herkel J, Heidrich B, Nieraad N, et al. Fine specificity of autoantibodies to soluble liver antigen and liver/pancreas. Hepatology 2002; 35:403–8.
15. Alvarez F, Ciocca M, Canero-Velasco C, et al. Short-term cyclosporine induces a remission of autoimmune hepatitis in children. J Hepatol 1999; 30:222–7.
16. Djilali-Saiah I, Ouellette P, Caillat-Zucman S, et al. CTLA-4/CD 28 region polymorphisms in children from families with autoimmune hepatitis. Hum Immunol 2001; 62:1356–62.