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Serological diagnosis of anti-SLA positive Autoimmune hepatitis
Reply to Volkmann et al.
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Martin E.P. Volkmann, M.D. Zentrallabor, Universitätsklinikum Heidelberg, Jochen Ludwig, Julia Galler-Weber, Heidi Rossmann, Andrea Bäurle, Walter Fiehn, Christian Strassburg*, Michael P. Manns*
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Martin_Volkmann{at}med.uni-heidelberg.de Martin E.P. Volkmann, et al.
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To the editor: Ballot and colleagues describe an interesting proteomic approach in which rat targets of anti-soluble-liver antigen antibodies are characterized by two dimensional gel electrophoresis and MALDI-TOF analysis. They present a sulfotransferase, isoforms of alpha enolases and of catalase as possible target antigens for anti-SLA antibodies. They conclude that their approach serves to improve diagnostic tools and the understanding of immune processes. Anti-Soluble Liver Antigen autoantibodies do indeed play an important role in the diagnosis of autoimmune hepatitis due to their high specificity. Recently three cDNAs of a putative target antigen have been described (1, 2, 3) by independent investigators. The molecules are thought to be associated with an UGA suppressor serine t-RNA carrying selenocystein (tRNPser/sec; 2). It could be shown through preabsorption experiments that the recombinant target is able to compete with reactivity to the wild type antigen efficiently in a dose dependent manner. In the first study 63/69 SLA/LP positive sera were found reactive (90%; 1), in a subsequent study 67/85 (78,8%; 3) of the sera showed a positive reaction. Specificities of or close to 100% were described for both candidate proteins, matching the results in the polyclonal inhibition anti SLA/LP ELISA. Also, conformation dependent epitopes have been defined on the putative SLA/LP molecule (5) as well as three linear immunodominant regions (6). Furthermore a highly efficient commercially available ELISA based on one of the recombinant isolates (1) has been developed. In contrast to these findings Ballot and colleagues present three possible antigens identified by MALDI-TOF analysis of seroreactive spots in 2-D immunoblot corresponding to bands detected by 1-D immunoblotting found also in 4 to 8.5% of controls without homology to tRNPser/sec. The most sensitive of these targets showed 68% positive sera and is implicated in several other autoimmune conditions as the authors rightly state. This is in contrast to the results obtained by the original anti-SLA inhibition ELISA over years by different groups, to which the authors also refer. Furthermore no experimental evidence for the newly described antigens in a purified form has been presented. Due to these findings we would like to caution against expectations for an essential improvement of anti-SLA testing by the new putative target antigens. The existing clinical data obtained with the recombinant SLA/LP/tRNPser/sec molecule are clearly superior to those described by Ballot and colleagues, both regarding numbers and results concerning diagnostic accuracy. Clearly not all anti-SLA/LP positive sera are positive in the SLA/LP ELISA. Also,we do not question the fact that additional antigens may be involved in anti-SLA positive autoimmune hepatitis which is evident from the frequent occurrence of anti-nuclear antibodies and anti-smooth-muscle antibodies in these samples. However, the existing clinical data clearly indicate that 1) the polyclonal inhibition ELISA remains the gold standard of anti-SLA testing and 2) the recombinant SLA/LP ELISA is a highly efficient readily available diagnostic tool. In the particular case of anti SLA/LP where over a period of more than ten years several targets have been proposed, any newly proposed target should be clinically evaluated before highly efficient existing serological tests are questioned. The existing data are clearly not sufficient to justify considering the molecular nature of SLA an open question again and claiming diagnostic uncertainty where valuable diagnostics are available. References 1. Wies I, Brunner S, Henninger J, Herkel J, Kanzler S, Mayer zum Büschenfelde KH. Identification for SLA/LP autoantibodies in autoimmune hepatitis. Lancet 2000;355:1510-15 2. Costa M, Rodriguez-Sanchez JL, Czaja AJ, Gelpi C. Isolation and characterization of cDNA encoding the antigenic protein of the human tRNA(ser)sec complex recognized by autoantibodies from patients with type- 1 autoimmune hepatitis. Clin Exp Immunol 2000;121:364-74 3. Volkmann M, Bäurle A, Heid H, Strassburg CP, Trautwein C, Fiehn W and Manns M. Soluble Liver Antigen: Isolation of a 35-kd recombinant protein (SLA-p35) specifically recognizing sera from patients with autoimmune hepatitis. Hepatology 2001;33: 591-6 4. Ma Y, Okamoto M, Thomas MG, Bogdanos DP, Lopes AR, Portmann B, Underhill J, Durr R, Mieli-Vergani G, Vergani D Antibodies to conformational epitopes of soluble liver antigen define a severe form of autoimmune liver disease. Hepatology 2002; 35: 658-64 5. Herkel J, Heidrich B, Nieraad N, Wies I, Rother M, Lohse AW. Fine specificity of autoantibodies to soluble liver antigen and liver/pancreas. Hepatology 2002; 35: 403-8 6. Martin L, Bäurle A, Fiehn W, Volkmann M. Immunoreactivity to cytokeratin 8/18 in patients with soluble liver antigen (SLA) positive autoimmune hepatitis (AIH type 3) is not sufficient for diagnostic use. Clin Lab 2000; 46: 339-344 |
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Catherine Johanet, PhD Saint-Antoine Hospital, Paris, France, Eric Ballot, Arnaud Bruneel
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catherine.johanet{at}sat.ap-hop-paris.fr Catherine Johanet, et al.
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The authors of the letter seem unhappy with the report of antigens for anti-SLA antibodies other than the recombinant 50 kDa UGA serine tRNA- associated protein (tRNP)(ser)(sec) used in a commercially available ELISA. But, as suggested by the replying authors themselves (1), this tRNA-associated protein remains to be formally identified in the liver. Indeed, we have not found this protein as an immunoreactive spot on our 2D -immunoblotting experiments, which investigated proteins according to their level of expression in the cell. As described by other studies using immunoblotting (2, 3) it seems that anti-SLA positive sera are immunoreactive with various antigens. We insist on the fact that the 50 kDa antigenic protein we described has a pI value (i.e. 6.0 – 6.5) in the same range that the pI value of the 50 kDa immunoreactive spot found by others (i.e. 6.0 – 7.5) (2, 4). This value is much lower than the theoretical pI of 9.5 for the (tRNP)(ser)(sec) related protein. The replying authors claim that valuable diagnostic information is available using this recombinant protein and agree to consider the inhibition ELISA as the “gold standard” for anti-SLA testing. Using inhibition techniques, several authors (5, 6) found anti-SLA antibodies only in AIH-1 or in cryptogenic hepatitis and never in AIH-2. Furthermore, it is now established that SLA and liver pancreas (LP) are the same antigen. Stechemesser et al. (7) have shown that anti-LP antibodies were preferentially associated with AIH-1 and never with anti-LKM1 antibodies, the marker of AIH-2. By contrast, using recombinant (tRNP)(ser)(sec) related protein, two distinct studies (8, 9) reported a high prevalence of anti-SLA antibodies in AIH-2 and in autoimmune sclerosing cholangitis. Why such discrepancies about the 50,000 antigen pI values and the anti-SLA prevalence in AIH-2 according to the nature of target antigen ? The antigen origin (i.e., human recombinant or rat/rabbit cytosol) cannot explain these discrepancies, because it has been shown that the (tRNP)(ser)(sec) related protein was highly conserved through evolution (10). We believe that SLA is a generic term regrouping many cytosolic antigenic targets in AIH-1 or in cryptogenic hepatitis. Our paper as well as an additional recent study discussing GST as putative anti-SLA target (11) incite us to claim that the molecular nature of SLA, although a commercially diagnostic ELISA is available, still remains an open question. References. 1. Volkmann M, Martin L, Bäurle A, Heid H, Strassburg C P, Trautwein C, Fiehn W, Manns M. Soluble liver antigen: isolation of a 35-kd recombinant protein (SLA-p35) specifically recognizing sera from patients with autoimmune hepatitis. Hepatology 2001;33:591-6. 2. Wächter B, Kyriatsoulis A, Lohse AW, Gerken G, Meyer zum Büschenfelde KH, Manns M. Characterization of liver cytokeratin as a major target antigen of anti-SLA antibodies. J Hepatol 1990;11:232-9. 3. Wesierska-Gadek J, Grimm R, Hitchman E, Penner E. Members of the glutathione S-transferase gene family are antigens in autoimmune hepatitis. Gastroenterology 1998;114:329-35. 4. Martin L, Bäurle A, Fiehn W, Volkmann M. Immunoreactivity to cytokeratin 8/18 in patients with soluble liver antigen (SLA) positive autoimmune hepatitis (AIH type 3) is not sufficient for diagnostic use. Clin Lab 2000;46:339-44. 5. Ballot E, Homberg JC, Johanet C. Antibodies to soluble liver antigen: an additional marker in type 1 auto-immune hepatitis. J Hepatol 2000;33:208-15. 6. Manns M, Gerken G, Kyriatsoulis A, Staritz M, Meyer zum Büschenfelde KH. Characterization of a new subgroup of autoimmune chronic active hepatitis by autoantibodies against a soluble liver antigen. Lancet 1987;1:292-4. 7. Stechemesser E, Klein R, Berg PA. Serological definition of new subgroup of patients with autoimmune chronic active hepatitis. Hepatology. 1993;18:1-9. 8. Ma Y, Okamoto M, Thomas MG, Bogdanos DP, Lopes AR, Portmann B, Underhill J, Durr R, Mieli-Vergani G, Vergani D. Antibodies to conformational epitopes of soluble liver antigen define a severe form of autoimmune liver disease. Hepatology. 2002;35:658-64. 9. Vitozzi S, Djilali-Saiah I, Lapierre P, Alvarez F. Anti-soluble liver antigen/liver-pancreas (SLA/LP) antibodies in pediatric patients with autoimmune hepatitis. Autoimmunity. 2002;35:485-92. 10. Herkel J, Heidrich B, Nieraad N, Wies I, Rother M, Lohse AW. Fine specificity of autoantibodies to soluble liver antigen and liver/pancreas. Hepatology. 2002;35:403-8. 11. Wesierska-Gadek J, Lindner H, Hitchman E, Sarg B, Penner E. Anti-SLA seropositive auto immune hepatitis sera recognize distinct subunits of glutathione S-transferase : high prevalence of the Ya autoantigen. Cell Mol Biol (Noisy-le-grand). 2002;48:301-7. |
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