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The dominant self and the cryptic self: shaping the autoreactive T-cell repertoire
The dominant self and the cryptic self:
shaping the autoreactive T-cell repertoire Guy Gammon, Eli E. Sercarz and Gilles Benichou Deletion of autoreactive T cells during the induction of self tolerance has been directly demonstrated. However, it is still relatively easy to detect self reactivity in normal healthy animals. In this article, Guy Gammon, Eli Sercarz and Gilles Benichou speculate on which T cells may escape toleranceinduction and discuss how these cells could subsequently be involved in autoimmunity. It is now widely accepted that self proteins are continuously processed and presented as peptides in association with self MHC molecules for interaction with T cells 1,2. Recognition of these self ligands has profound effects, both positive and negative, on the development of the Tcell repertoire. In recent years a much clearer picture of these events has emerged but many questions remain unanswered. One of the major problems still to be clarified is how crippling deletion of the T-cell repertoire is avoided, since there are so many self proteins, potentially yielding so many more peptides. Overwhelming T-cell deletion would result if every peptide could induce tolerance, but this does not happen. In fact, although deletion of some self-reactive T cells has been demonstrated 3,4, peripheral reactivity to many self peptides in the context of both major histocompatibility complex (MHC) class I and class II molecules has been repeatedly observed s-7. This reactivity is not restricted to proteins sequestered from the immune system but includes reactivity to certain determinants on widely expressed and readily accessible molecules. In this situation it appears that individual determinants, rather than the whole self antigen, are sequestered. This article examines the rules guiding whether or not a particular determinant region on a self molecule can induce tolerance. !
Factors determining tolerance induction In undertaking a study of self tolerance, it is important to choose self antigens that are readily available in the thymus at the time of the acquisition of the repertoire during development. For this reason we examined tolerance/reactivity to a set of determinants on self MHC class I and II molecules, bound as peptides within the MHC class-II-binding groove 6. Five class II peptides, three from the helical regions and two from the floor of the groove of the A k molecule, were examined for immunogenicity in syngeneic mice. Immunogenicity would indicate that tolerance had not been induced, that the peptide could bind MHC and that self-reactive T cells were present in the adult mouse. B10.A mice (MHC haplotype Kk, A k, E k, D a, L d) showed a vigorous response to two of the five A k peptides. Likewise, of the three class I peptides from the polymorphic helical regions of D d or Ld tested for immunogenicity, one induced a response. Bind-
ing studies performed in vitro showed that one of the nonimmunogenic peptides, D a 61-85, bound to the A k molecule with high affinity; presumably it failed to stimulate a response because it had induced T-cell deletion. This was confirmed by showing that D a 61-85 was immunogenic in B10.BR and CBA mice which express A k, but D k instead of D a. Thus tolerance to a protein must be considered at the level of reactivity to single determinant regions, as animals can be tolerant to certain determinants on a self protein but not to others. What factors determine the fate of developing lymphocytes that recognize a specific determinant region? For B cells the relevant factors are the concentration of the ligand, the affinity of the surface immunoglobulin receptor and the associated signals that influence whether the cell is activated or deleted upon contact with the ligand (reviewed in Ref. 8). Which B cells will undergo tolerization can be predicted on the basis of (1) their affinity for antigen- higher affinity B cells are tolerized in preference to those of lower affinity and (2) the concentration of antigen in the system. The situation for T cells is similar but more complex since, unlike B cells which recognize free native antigen, T cells recognize a complex ligand composed of a pepti& fragment of the antigen bound to a cell surface MHC molecule. At present it is impossible to measure either the concentration of a T-cell ligand or the affinity of the T-cell antigen receptor. Consequently it is impossible to predict with any certainty whether a specific T cell will be deleted.
Factors determining response magnitude Some information on the concentration of the tolerogenic ligand, or at least on the relative levels of the different determinants displayed at the cell surface, can be obtained by examining T-cell responses since the same ligand is involved in both activation and tolerization 9-1~. Detailed analysis of T-cell reactivity to nonself proteins has shown that T cells recognize discrete sites on the antigen. Usually one or two dominant (major) determinant region(s) induce most of the T-cell reactivity observed, while a greater number of subdominant (minor) regions induce the remainder of the response 12. Several factors contribute to this hierarchy of T-cellinducing regions. First, antigenic molecules undergo
© 1991, Elsevier Science Publishers Ltd, UK. 0167- 4919/91/$02.00
Immunology Today
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vo . 12 No. 6 1991
extensive processing before their component peptide fragments are presented to T cells. As the antigen unfolds and is partially degraded, certain determinants will be ex.posed preferentially, whereas other areas may undergo extensive degradation, destroying potential T-celt determinants. Second, competition can occur among peptides for association with an MHC molecule13. Consequently, the amount of each peptide at the cell surface will be highly variable, even for peptides derived from the same protein. Data from several experimental systems have suggested that this is the most important parameter in determining the relative magnitude of response to each region 12. A third factor that may influence the magnitude of the response to a determinant region is the precursor frequency of specific T cells. However, it is extremely unlikely that the average receptor affinity of the available repertoire is different for each determinant on the antigen given the diversity of T-cell populations; affinity should, therefore, not be a major factor in determining the hierarchy of determinant regions. The exception would be the situation where a foreign antigen mimics a self determinant so that all the high-affinity T cells were deleted. What effect will these factors that influence the magnitude of a response have on the induction of tolerance? Dominant determinant regions that are processed and presented efficiently will act as good inducers of tolerance, while determinant regions that are inefficiently processed and presented will be poor tolerogens. Thus, there are two functional sets of self determinants: those that readily tolerize, and comprise the dominant self, and those that do not tolerize but spare members of a potentially autoimmune repertoire, and comprise the cryptic self. Inefficient processing may to some extent be overcome by the presence of large amounts of protein and the differential effect of processing efficiency will be greatest in the case of self molecules present at low concentration. The cryptic self, therefore, will include all the determinants on antigens in immunologically privileged sites as well as many determinants on antigens accessible to the immune system. Experimental support for the hypothesis that T cells that recognize poorly presented determinants can escape tolerance has been obtained in an experimental model of tolerancei4. Adult mice that are rendered tolerant to chicken lysozyme by a single intravenous injection of antigen do not respond to immunization with whole lysozyme or peptides containing dominant determinant regions. However, immunization with peptides containing certain subdominant determinant regions does stimulate T-cell reactivity. The data on reactivity to MHC peptides can also be explained on the basis of differential efficiency of processing and presentation among individual determinant regions. Thus the D a peptide 61-85, which binds Akwith high affinity but is nonimmunogenic in B10.A mice, is a highly visible determinant and hence a good tolerogen. The two class II peptides that were immunogenic are poorly processed determinant regions. Both of these class II peptides were shown to bind to MHC with high affinity, suggesting that the block to efficient presentation occurs during degradation of the native molecule rather than, as has been proposed, as a result of competition with other peptides for the MHCbinding sitesis.
Immunology Today
Determinant response hierarchy and autoimmunity The threshold escape hypothesis described above implies that, since animals are functionally tolerant to self, any remaining anti-self activity, for example to self MHC peptides as described above, must be directed towards poorly presented determinants. These self-reactive T cells are not activated by ambient self antigen because they recognize cryptic determinants that do not reach the threshold concentration on the cell surface that is necessary for engaging T cells. This resolves the apparent paradox of how animals thrive but possess so much potential self reactivity. Self tolerance depends on the stability of the hierarchy of presentation of determinant regions and anything that upsets this balance may cause autoimmunity. One direct way of destabilizing the hierarchy is to immunize with peptides but other, more subtle, ways of altering the response hierarchy, such as priming with homologous proteins, may also be effective. Because homologous proteins have some sequence differences, they may be processed differently, generating an altered pattern of peptide products and consequently an altered hierarchy of response specificity12. For example, the fine specificity of the rat T-cell response to self myelin basic protein (MBP) varies, depending on whether guinea-pig or rat MBP is used as the immunogen~6. The observation that guinea-pig MBP is much more effective than rat MBP in the induction of experimental allergic encephalomyelitis (EAE) may be explained by the ability of homologous proteins to activate T cells directed against the cryptic self. Subdominant determinants have been shown to be important in this disease17. In human pathology, the initial priming event could be exposure to a dominant determinant on a bacterial or viral protein which induces a response that is crossreactive with a cryptic self determinant for which a sizeable repertoire of T cells could exist. Possible examples of this are antigens, such as the heat shock proteins, that show strong sequence preservation during evolution. Once an autoimmune response has been initiated, will it be perpetuated ? The threshold concentration for recall of reactivity should be lower than that required for the initial activation of naive cells. The responses generated by peptide priming of lysozyme-tolerant mice can be recalled both by the peptide and, to a variable extent, by the whole antigen14. However, some peptide-induced responses require very high levels of native antigen to recall any in vitro response and are, therefore, essentially peptide-specific determinants. Additional factors may help perpetuate reactivity in vivo: for example the release of additional antigen from damaged tissue and the local production of cytokines (such as turnout necrosis factor and gamma-interferon) that upregulate expression of MHC and adhesion molecules. Such conditions of enhanced antigen presentation may lead to the generation of responses to other previously cryptic determinant regions rendered visible during these inflammatory events. Moreover, increased levels of ligand will activate low-affinity T cells that will also contribute to the autoimmune response. This broadening of the target specificity may be the critical step in the pathological process resulting in substantial tissue damage. In summary, there is a growing awareness of the
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iii!ii!i!iiiii!!iii!i!iii!i!iii!i!iiiiiiiiiiiiiiii!iiiiiiii!iii!iiiiiiiiii !i i!iiiii!iiiiiiiiiiiiiiiiiiiiiiii iiiiiiiiii!iiiiii!iiiiiiii!iiii i!iiiiiiiiiiiiiiliiiiiiiliiiiiii i i extensive T-cell repertoire directed against the cryptic self Immunopathol. 56, 287-297 in healthy individuals. The activation and recruitment of 6 Benichon, G., Takizawa, P.A., Ho, P.T. et al. (1990) these cells in autoimmune disease and secondary mech- J. Exp. Med. 172, 1341-1346 anisms that contribute to their immunoregulation re- 7 Schild, H., Rotzschke, O., Kalbacher, H. and Rammensee, quire fuller understanding and may reveal new targets for H. (1990) Science 247, 1587-1589 8 Goodnow, C.C. (1989) Curr. Opin. Immunol. 2, 226-236 therapeutic intervention. 9 Gammon, G., Dunn, K., Shastri, N. et al. (1986) Nature 319,413-415 Guy Gammon is at Xenova Ltd, 545 Ipswich Rd, Slough 10 Groves, E.S. and Singer, A. (1983)].Exp. Med. 158, SL1 4EQ, UK; Eli Sercarz and Gilles Benichou are at the 1483-1497 Dept of Microbiology and Molecular Genetics, University 11 Matzinger, P., Zamoyska, R. and Waldmann, H. (1984) Nature 308, 738-739 of California, Los Angeles, CA 90024, USA. 12 Gammon, G., Shastri, N., Cogswell, J. et al. (1987) Immunol. Rev. 98, 53-71 References 13 Adorini, L., Muller, S., Cardinaux, F. et al. (1988) Nature 1 Kourilsky, P., Chaouat, G., Rabourdin-Combe, C. and 334, 623-625 14 Gammon, G. and Sercarz, E.E. (1989) Nature 342, Claverie, J. (1987) Proc. Natl Acad. Sci. USA 84, 3400-3404 2 Lorenz, R.G. and Allen, P.M. (1988) Proc. Natl Acad. Sci. 183-185 15 Waldmann, H., Cobbold, S., Benjamin, R. and Qin, S. USA 85, 5220-5223 3 Kappler, J.W., Roehm, N. and Marrack, P. (1987) Cell 49, (1988) J. Autoimmun. 1,623-629 273-280 16 Happ, M.P. and Heber-Katz, E. (1988) J. Exp. Med. 167, 4 Kisielow, P., Bluthmann, H., Staerz, U.D., Steinmetz, M. 502-513 and Von Boehmer, H. (1988) Nature 333,742-746 17 Clayton, J.P., Gammon, G., Ando, D.G. et al. (1989) 5 Gammon, G. and Sercarz, E.E. (1990) Clin. Immunol. J. Exp. Med. 169, 1681-1691
Parasitol. T o d a y 7 (1), 5 - 1 1
;mmuno;ogy today
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No. 6 1991
shaping the autoreactive T-cell repertoire Guy Gammon, Eli E. Sercarz and Gilles Benichou Deletion of autoreactive T cells during the induction of self tolerance has been directly demonstrated. However, it is still relatively easy to detect self reactivity in normal healthy animals. In this article, Guy Gammon, Eli Sercarz and Gilles Benichou speculate on which T cells may escape toleranceinduction and discuss how these cells could subsequently be involved in autoimmunity. It is now widely accepted that self proteins are continuously processed and presented as peptides in association with self MHC molecules for interaction with T cells 1,2. Recognition of these self ligands has profound effects, both positive and negative, on the development of the Tcell repertoire. In recent years a much clearer picture of these events has emerged but many questions remain unanswered. One of the major problems still to be clarified is how crippling deletion of the T-cell repertoire is avoided, since there are so many self proteins, potentially yielding so many more peptides. Overwhelming T-cell deletion would result if every peptide could induce tolerance, but this does not happen. In fact, although deletion of some self-reactive T cells has been demonstrated 3,4, peripheral reactivity to many self peptides in the context of both major histocompatibility complex (MHC) class I and class II molecules has been repeatedly observed s-7. This reactivity is not restricted to proteins sequestered from the immune system but includes reactivity to certain determinants on widely expressed and readily accessible molecules. In this situation it appears that individual determinants, rather than the whole self antigen, are sequestered. This article examines the rules guiding whether or not a particular determinant region on a self molecule can induce tolerance. !
Factors determining tolerance induction In undertaking a study of self tolerance, it is important to choose self antigens that are readily available in the thymus at the time of the acquisition of the repertoire during development. For this reason we examined tolerance/reactivity to a set of determinants on self MHC class I and II molecules, bound as peptides within the MHC class-II-binding groove 6. Five class II peptides, three from the helical regions and two from the floor of the groove of the A k molecule, were examined for immunogenicity in syngeneic mice. Immunogenicity would indicate that tolerance had not been induced, that the peptide could bind MHC and that self-reactive T cells were present in the adult mouse. B10.A mice (MHC haplotype Kk, A k, E k, D a, L d) showed a vigorous response to two of the five A k peptides. Likewise, of the three class I peptides from the polymorphic helical regions of D d or Ld tested for immunogenicity, one induced a response. Bind-
ing studies performed in vitro showed that one of the nonimmunogenic peptides, D a 61-85, bound to the A k molecule with high affinity; presumably it failed to stimulate a response because it had induced T-cell deletion. This was confirmed by showing that D a 61-85 was immunogenic in B10.BR and CBA mice which express A k, but D k instead of D a. Thus tolerance to a protein must be considered at the level of reactivity to single determinant regions, as animals can be tolerant to certain determinants on a self protein but not to others. What factors determine the fate of developing lymphocytes that recognize a specific determinant region? For B cells the relevant factors are the concentration of the ligand, the affinity of the surface immunoglobulin receptor and the associated signals that influence whether the cell is activated or deleted upon contact with the ligand (reviewed in Ref. 8). Which B cells will undergo tolerization can be predicted on the basis of (1) their affinity for antigen- higher affinity B cells are tolerized in preference to those of lower affinity and (2) the concentration of antigen in the system. The situation for T cells is similar but more complex since, unlike B cells which recognize free native antigen, T cells recognize a complex ligand composed of a pepti& fragment of the antigen bound to a cell surface MHC molecule. At present it is impossible to measure either the concentration of a T-cell ligand or the affinity of the T-cell antigen receptor. Consequently it is impossible to predict with any certainty whether a specific T cell will be deleted.
Factors determining response magnitude Some information on the concentration of the tolerogenic ligand, or at least on the relative levels of the different determinants displayed at the cell surface, can be obtained by examining T-cell responses since the same ligand is involved in both activation and tolerization 9-1~. Detailed analysis of T-cell reactivity to nonself proteins has shown that T cells recognize discrete sites on the antigen. Usually one or two dominant (major) determinant region(s) induce most of the T-cell reactivity observed, while a greater number of subdominant (minor) regions induce the remainder of the response 12. Several factors contribute to this hierarchy of T-cellinducing regions. First, antigenic molecules undergo
© 1991, Elsevier Science Publishers Ltd, UK. 0167- 4919/91/$02.00
Immunology Today
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vo . 12 No. 6 1991
extensive processing before their component peptide fragments are presented to T cells. As the antigen unfolds and is partially degraded, certain determinants will be ex.posed preferentially, whereas other areas may undergo extensive degradation, destroying potential T-celt determinants. Second, competition can occur among peptides for association with an MHC molecule13. Consequently, the amount of each peptide at the cell surface will be highly variable, even for peptides derived from the same protein. Data from several experimental systems have suggested that this is the most important parameter in determining the relative magnitude of response to each region 12. A third factor that may influence the magnitude of the response to a determinant region is the precursor frequency of specific T cells. However, it is extremely unlikely that the average receptor affinity of the available repertoire is different for each determinant on the antigen given the diversity of T-cell populations; affinity should, therefore, not be a major factor in determining the hierarchy of determinant regions. The exception would be the situation where a foreign antigen mimics a self determinant so that all the high-affinity T cells were deleted. What effect will these factors that influence the magnitude of a response have on the induction of tolerance? Dominant determinant regions that are processed and presented efficiently will act as good inducers of tolerance, while determinant regions that are inefficiently processed and presented will be poor tolerogens. Thus, there are two functional sets of self determinants: those that readily tolerize, and comprise the dominant self, and those that do not tolerize but spare members of a potentially autoimmune repertoire, and comprise the cryptic self. Inefficient processing may to some extent be overcome by the presence of large amounts of protein and the differential effect of processing efficiency will be greatest in the case of self molecules present at low concentration. The cryptic self, therefore, will include all the determinants on antigens in immunologically privileged sites as well as many determinants on antigens accessible to the immune system. Experimental support for the hypothesis that T cells that recognize poorly presented determinants can escape tolerance has been obtained in an experimental model of tolerancei4. Adult mice that are rendered tolerant to chicken lysozyme by a single intravenous injection of antigen do not respond to immunization with whole lysozyme or peptides containing dominant determinant regions. However, immunization with peptides containing certain subdominant determinant regions does stimulate T-cell reactivity. The data on reactivity to MHC peptides can also be explained on the basis of differential efficiency of processing and presentation among individual determinant regions. Thus the D a peptide 61-85, which binds Akwith high affinity but is nonimmunogenic in B10.A mice, is a highly visible determinant and hence a good tolerogen. The two class II peptides that were immunogenic are poorly processed determinant regions. Both of these class II peptides were shown to bind to MHC with high affinity, suggesting that the block to efficient presentation occurs during degradation of the native molecule rather than, as has been proposed, as a result of competition with other peptides for the MHCbinding sitesis.
Immunology Today
Determinant response hierarchy and autoimmunity The threshold escape hypothesis described above implies that, since animals are functionally tolerant to self, any remaining anti-self activity, for example to self MHC peptides as described above, must be directed towards poorly presented determinants. These self-reactive T cells are not activated by ambient self antigen because they recognize cryptic determinants that do not reach the threshold concentration on the cell surface that is necessary for engaging T cells. This resolves the apparent paradox of how animals thrive but possess so much potential self reactivity. Self tolerance depends on the stability of the hierarchy of presentation of determinant regions and anything that upsets this balance may cause autoimmunity. One direct way of destabilizing the hierarchy is to immunize with peptides but other, more subtle, ways of altering the response hierarchy, such as priming with homologous proteins, may also be effective. Because homologous proteins have some sequence differences, they may be processed differently, generating an altered pattern of peptide products and consequently an altered hierarchy of response specificity12. For example, the fine specificity of the rat T-cell response to self myelin basic protein (MBP) varies, depending on whether guinea-pig or rat MBP is used as the immunogen~6. The observation that guinea-pig MBP is much more effective than rat MBP in the induction of experimental allergic encephalomyelitis (EAE) may be explained by the ability of homologous proteins to activate T cells directed against the cryptic self. Subdominant determinants have been shown to be important in this disease17. In human pathology, the initial priming event could be exposure to a dominant determinant on a bacterial or viral protein which induces a response that is crossreactive with a cryptic self determinant for which a sizeable repertoire of T cells could exist. Possible examples of this are antigens, such as the heat shock proteins, that show strong sequence preservation during evolution. Once an autoimmune response has been initiated, will it be perpetuated ? The threshold concentration for recall of reactivity should be lower than that required for the initial activation of naive cells. The responses generated by peptide priming of lysozyme-tolerant mice can be recalled both by the peptide and, to a variable extent, by the whole antigen14. However, some peptide-induced responses require very high levels of native antigen to recall any in vitro response and are, therefore, essentially peptide-specific determinants. Additional factors may help perpetuate reactivity in vivo: for example the release of additional antigen from damaged tissue and the local production of cytokines (such as turnout necrosis factor and gamma-interferon) that upregulate expression of MHC and adhesion molecules. Such conditions of enhanced antigen presentation may lead to the generation of responses to other previously cryptic determinant regions rendered visible during these inflammatory events. Moreover, increased levels of ligand will activate low-affinity T cells that will also contribute to the autoimmune response. This broadening of the target specificity may be the critical step in the pathological process resulting in substantial tissue damage. In summary, there is a growing awareness of the
194
Vo112 No. 6 1991
iii!ii!i!iiiii!!iii!i!iii!i!iii!i!iiiiiiiiiiiiiiii!iiiiiiii!iii!iiiiiiiiii !i i!iiiii!iiiiiiiiiiiiiiiiiiiiiiii iiiiiiiiii!iiiiii!iiiiiiii!iiii i!iiiiiiiiiiiiiiliiiiiiiliiiiiii i i extensive T-cell repertoire directed against the cryptic self Immunopathol. 56, 287-297 in healthy individuals. The activation and recruitment of 6 Benichon, G., Takizawa, P.A., Ho, P.T. et al. (1990) these cells in autoimmune disease and secondary mech- J. Exp. Med. 172, 1341-1346 anisms that contribute to their immunoregulation re- 7 Schild, H., Rotzschke, O., Kalbacher, H. and Rammensee, quire fuller understanding and may reveal new targets for H. (1990) Science 247, 1587-1589 8 Goodnow, C.C. (1989) Curr. Opin. Immunol. 2, 226-236 therapeutic intervention. 9 Gammon, G., Dunn, K., Shastri, N. et al. (1986) Nature 319,413-415 Guy Gammon is at Xenova Ltd, 545 Ipswich Rd, Slough 10 Groves, E.S. and Singer, A. (1983)].Exp. Med. 158, SL1 4EQ, UK; Eli Sercarz and Gilles Benichou are at the 1483-1497 Dept of Microbiology and Molecular Genetics, University 11 Matzinger, P., Zamoyska, R. and Waldmann, H. (1984) Nature 308, 738-739 of California, Los Angeles, CA 90024, USA. 12 Gammon, G., Shastri, N., Cogswell, J. et al. (1987) Immunol. Rev. 98, 53-71 References 13 Adorini, L., Muller, S., Cardinaux, F. et al. (1988) Nature 1 Kourilsky, P., Chaouat, G., Rabourdin-Combe, C. and 334, 623-625 14 Gammon, G. and Sercarz, E.E. (1989) Nature 342, Claverie, J. (1987) Proc. Natl Acad. Sci. USA 84, 3400-3404 2 Lorenz, R.G. and Allen, P.M. (1988) Proc. Natl Acad. Sci. 183-185 15 Waldmann, H., Cobbold, S., Benjamin, R. and Qin, S. USA 85, 5220-5223 3 Kappler, J.W., Roehm, N. and Marrack, P. (1987) Cell 49, (1988) J. Autoimmun. 1,623-629 273-280 16 Happ, M.P. and Heber-Katz, E. (1988) J. Exp. Med. 167, 4 Kisielow, P., Bluthmann, H., Staerz, U.D., Steinmetz, M. 502-513 and Von Boehmer, H. (1988) Nature 333,742-746 17 Clayton, J.P., Gammon, G., Ando, D.G. et al. (1989) 5 Gammon, G. and Sercarz, E.E. (1990) Clin. Immunol. J. Exp. Med. 169, 1681-1691
Parasitol. T o d a y 7 (1), 5 - 1 1
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No. 6 1991