Graft-versus-host disease: the need for a new terminology

Immunology Today, v'oL I I. No. 12 1990

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For some decades, graft-versus-host (GVH) reactions seemed readily comprehensible. It was generally accepted that after introduction of immunocompetent, histoincompatible lymphocytes into an immunodeficient host the grafted lymphocytes would start a 'rejection' response against their host..,Asa result, GVH diseasedeveloped. Acute diseasemight result in death and su~ivors became chimaeras with varying pathology - chronic GVH disease. In recent years, however, some intriguing exceptions to the general rules for the development of GVH disease have been reported. Theseexceptions, particularly the activatior of GVH diseaseby stimuli other than histocompatibility barriers prompted Gerard Bos and colleagues to propose this change in terminology. History of GVH reaction In 1953 the term GVH reaction was introduced into the literature simultaneously by Simonsen and Dempster, as an explanation for the 'pyroninophilic' interstitial infiltrate observed in kidney allografts in dogs T!ley considered these cells to be of graft origin and to react against host alloantigens. The phenomenon was, however, due to a host-versus-graft reaction and therefore the term GVH reaction was improperly used ~.2. In 1954 Billingham et al. were the first to demonstrate that parental grafts transplanted to F~ hybrid mice were geneticalhj qualified to react against their hosts 3. In 1957 Simonsen, and Billingham and Brent described a clinical picture of diarrhoea, skin lesions, retarded growth and death in r ewborn animals injected with allogeneic blood, leukocytes or spleen cells; Billingham and Brent . . . . . . . . . . . . y.... ,.,,,,= runt u , ~ a ~ -,-. At the same time, similar syndromes occurring after transfer of allogeneic bone marrow cells to lethally irradiated mice and rats were documented. This syndrome was called secendary disease (the initial symptoms due to the X-irraci~ation were referred to as primary disease) or homologous disease6.7. Although no experimental evidence was available at that time, the tissue damage observed in the experiments described above was thought to be secondary to interactions between effector (donor) lymphocytes and histoincompatible target cells8. Indeed, small lymphocytes (obtained from thoracic duct lymph) were identified as being responsible for the induction of disease in newborn rats 9, F~ hybrid rats ~° and lethally X-irradiated mice ~. By 1962 runt disease, secondary disease or homologous disease and the pathology in F~ animals were interpreted as the consequence of GVH reactions and were called GVH disease ~2. On the basis of the experiments described above, Billingham formulated the essential requirements to elicit a GVH reaction ~3. (1) The graft must contain immunologically competent cells. (2) The host must possess important transplantation isoantigens that are lacking in the graft donor, so that the host appears foreign to it and is therefore capable of stimulating it antigenically. (3) The

Dept of Immunology, Universit7 of Limburg, Maastricht, The Netherlands. 1Presentaddress:Deptof InternalMedicine,AcademicHospital, Maastticht, 1heNetherlands. ~ ) I )90, Elseviel Science Publishers Lid, U K. 0167--49191901502 00

disease:lhe need for a new terminology Gerard M.J. BOS1, Gerard D. Majoor and Peter J.C. van Breda Vriesman

host itself must be incapable of mounting an effective immunological reaction against the graft, at least for sufficient time for the latter to manifest its immunological capabilities, i.e. the graft must have some security of tenure. These rules, particularly the second, requiring a difference between graft and host transplantation isoantigens (later called histocompatibility antigens (Ha)), has become a principal rule of GVH disease. In general, within one sp -cies a severe GVH reaction will occur when donors and recipients of bone mawrow transplantation (BMT) are mismatched for major histocompatibility (MHC) antigens. However, ,c should be ~mphasized that after BMT between fully MHC-matched donors and recipients, a GVH reaction may still occur 14,~s.GVH reactions against minor histocompatibility antigens (MiHa) have been suggested to play a role in these cases16. However, these ubservations might h-- interpreted as indicating that histocompatibility barriers are not essential for the induction of GVH pathology. Furthermore, the observation that 'GVH' disease can occur even after autologous BMT strongly suggests that there are exceptions to the second rule of Billingham 17.

Acute graft-versus-hostdisease To understand the immunological mechanisms in GVH disease it is necessary to briefly discuss cellular events. T cells are generally thought to be the effector cells of a GVH reaction 18. The relative importance of T helper (TH) cells and T killer/suppressor (TK/s) cells has been extensively studied in experimental models 19. However, there is still no consensus on the precise contribution of T-cell subsets in the induction of acute GVH disease. For example, GVH reactions have been reported to occur after transfer of spleen cells from nude (athymic) mice. Although no mature T cells are present in these mice, bonemarrow-derived cells of these animals elicit significant acute GVH reactions2°.21~ Other investigators have suggested a role for natural killer (NK) cells, large granular !ymphocytes (LGL) and tumor necrosis factor (TNF) in acute GVH reactions22 25 How effcctor cells finally evoke the pathology associated with a GVH reaction is not yet clear. Lymphocytes might display several different anti-host activities that may contribute to the GVH pathology, including a!loantibody formation, anti-host T-cell cytotox,city2'' and delayedtype hypersensitivity27. Common in vitro d:,~aysto monitor immunological parameters, such as donor anti-recipient cytotoxic T-cell assays and mixed-lymphoo/te cultures have, however, yielded inconsistent results and proved not to be predictive for the occurrence of acute GVH disease28. NK cell involvement, possibly via interleukin 433

-rostrum p-oduct=on, m=ght exp;a~n ~be organ preponderance in acute GV~, d~sease, since NK cells are believed to react against "fetai-hke' antigens:~ ~. These antigens may prevail ~no~9ans with a high cell turnover, such as the skin and the gastrointestinal tract, and may explain the strong ~nvolvement of these organs in GVH disease~2. Another explanation for the preferential involvement of ce~ain organs in GVH disease might be the selective expression of pertinent transplantation antigens. Recently. H-Y antigens on human keratinocytes have been demonstrated to play a role in GVH disease~. Nevertheless. this important observation cannot explain how GVH disease can develop in the absence of an H-Y barrier (female to female transplantation). !n conclusion, th£ current knowledge of the cellular mechanisms involved in acute GVH d~sease identifies neither the crucial e f f e ~ r cell(s), nor the precise role of histocompatibility a!ioantigens in their activation. Chronicgraft-vers~s-hostdisease The (cellular) mechanisms of chronic GVH disease appear to be even more complex than those involved in acute GVH disease. Two explanations of the etiology of chronic GVH disease have been suggested in the literature. According to the first, chronic GVH disease is caused by long-living lymph.~cytes of donor origin that have been sensitized to unknown antigens, probably MiHa of the host ~.3s. According to the second hypothesis, chronic GVH disease is elicited by immunocompetent donor cells that differentiate in the recipient from the bone-marrow inoculum :2. Tsoi et al. 36 ~8 have extensive!y studied both humoral and cellular immunopathology in patients with chronic GVH disease. Their studies showed that increased antihost cytotoxic activity, increased anti-host proliferative activity and decreased suppressor activity may exist in patients with chronic GVH disease. The obsewation that a proliferative response against non-HLA antigens is relevant for chronic GVH disease could not, however, be confirmed by others 39. In concordance with the observation of iSoi, Goulmy et aL~° also suggested a role for persisting cytotoxic T-cell (CTL) activity in chronic GVH disease. On the other hand, more recent work from this group did not corroborate an association between CTL activity and chronic GVH disease4°. Evidence that suppressor mechanisms might play a role in the control of GVH reaction seems, therefore, to be the only cellular mechanism that has not been refuted 4~. tn conclusion, in vitro studies have provided no evidence that chronic GVH disease is due to persistence of alloreactive, long-living donor lymphocytes. Therefore, the term chronic GVH disease is an operational one, merely descriptive of the p ~thology observed in the different models. Only if chronic GVH disease could be demonstrated to be due to persisting donor lymphocytes would the term 'chronic GVH disease' be correct in the immunological sense. On the other hand, if chronic GVH disease could be shown to be the consequence of rea~ions by donor cel!s that originate from the donor marrow inoculum and are educated in the host, the term chronic GVH disease would be misleading. How such donor cells should be designated is not debated ir~ the literature. Therefore, donor lymphocytes educated in recipient thymuses with both donor bone-marrow-derived MHC class,-positive cells and recipient-derived class-II-positive edi434

Immunology Today, Vol 1 I. No. 12 1990

~helial cells should b~ redefined in terms of the discussion on tolerance: are they 'donor' or 'host' lymphocytes? If the latter cells are responsible for the pathology because of a fai!ure of the thymus to educate them to tolerate recipient antigens, the term autoimmunity might be more a~ropriate tc describe the immunological processes in chronic GVH disease, as we suggested earlier~2. A new model In the light of the uncertainty about the GVH component, especially in chronic GVH disease, the recent observation by Glazier et el. 43 on the occurrence of 'syngeneic GVH disease' after a combination of X-irradiahon, syngeneic or autologous bone marrow transplantatio,'~ and the administration of cyclosporin A (CsA) becomes very interesting. Although similar observations have been made before, as reviewed by van Bekkum44. the work cf Glazier and colleagues has restimulated the discussion on this topic. Since this model does not fit with the rules of Billingham as described above (there is no histocom-. patibili~ barrier between donor and reciloient), a GVI-I reaction cannot be responsible for the pathology observed in this model. CsA-~nduced disease, as we prefer to designate the pathology observed in this model, clinically resembles 'classical' GVH disease and may be lethal to the majority of animals. The use of specific pathogen-free animals and their age appear to be important factors in predicting the severity of disease4s.46. After the acute phase of the disease, the clinical symptoms do not decline in all animals but - as after aliogeneic BMT - chronic pathology may develop. We have described the develooment of a scleroderma-like syndrome in the chronic Iohase of this model that is very similar to chronic GVH disease47. The oescription of this model as recently presented by Parkman was not, however, ful!., co.~rect48. CsA-induced disease is not a mild, self-limited disease as stated by ........... ~,,~ u,,~,, severe and progress;re as indicated above. Therefore, the model is of use for the study of both acute and chronic disease. Ihe cellular mechanisms in acute CsA-induced disease are intriguing. Hess e t a L 49 demonstrated the oresence of c~/totoxic T lymphocytes directed against public MHC class II antigens which they held responsible for the observed pathology. If an important part of the response is diT~cted to MHC class II antigen then the organ s0ecificity is hard to understand: in the rat model, the skin is most clearly affected 47. Moreover, in our laboratory we observed one rat with histologically proven CsA-induced disease and myasthenia gravis (with the presence of anti-acetylcholine receptor autoantibodies) ~,authors' unpu~J%l~ed observation), indicating that other antigens can be involved in this model. Othersso have demonstrated autoantibodies of different specificities in a comparable model in mice. In the rat model the development of acute patho!ogy is associated with the repopulation of the periphery with CD4 ~ (TH) lymphocytess~ and adoptive transfer experiments have demonstrated the importance of TH cells p e r se 46. Perhaps these TH cells are involved in the activation of the cytotoxic T cell.• described by Hess et aL 49. Apart from the induction of autoaggressive cells, a recent study describing transfer experiments has demonstrated that either X-irradiation or chemotherapy is necessary,to overrule suppressor mechanisms. Autoimmune disease can be transferred only by lymphocytes to secondary recipients after X-irradiation or chemotherapy s2.

Immunology Today, Vol. I 1, No 12 1990

ros , In the CsA model, class !I antigen expression in the thymus is reduced early after BM[ ~3. It is well known that class II antigens in the thymus, both on bone-marrowderived cells and on epithelial celk, play an important role in tolerance induction and repertoire sele~ion s4. Therefore, reduced expression of class II antigens in the CsA model might play a crucial role in explaining the pathophysiology of the disease in terms of autoimmunity. The thymus may also play a role in classical GVH disease; indeed, at this point, differences between classical GVH di,~ease and the CsA-induced autoimmunity may be quite small 4:.'~. It is known that after BMT, 'education' of pre-T cells to tolerate 'self antigens' occurs v~ithin the thymus; after lethal X-irradiation and BMT a recapitulation of ontogeny occurs 56. Therefore, if donor T ceils differentiate in the thymus of the recipient, but still attack the host, this implies that tolerance induction has failed. If so, the target antigens can either be host-specific antigens (i.e. MiHa) or antigens shared by host and donor ('autoantigens'). In this respect the function of the ihymus after BMT is also important in classical GVH disease. The thymus is known to be a target organ of the acute GVH reaction 5~.5s, and damage to the thymus may lead to a failure to induce tolerance to (auto)antigens in T cells developing from the bone marrow inoculum~'L Therefore, we proposed that the similarity of CsA-induced disease and classical chronic GVH disease could be the consequence of comparable damage to the thymus 42. Thus we agree with the idea of Parkman that the CsA model might shed new light on the mechanisms of classical GVH disease48. The similarities in pathology between acute and chronic CsA-induced and GVH disease suggest similar cellu!ar mechanisms, implicating the involvement of 'autoimmune' reactions in GVH disease. A new terminology

This paper proposes that the te-minology of (syngeneic) GVH disease and GVH leaction is confusing and no longer adequate for all situations. We therefore suggest that the term 'GVH reaction' is abandoned to describe the cellular mechanisms of GVH-like pathology, with the possible exception of the acute phase after allogeneic transplantation A better term for both chronic GVH disease after allogeneic BMT and GVH like disease afmr syngeneic or autologous BMT (with or without the use of CsA) is 'bone marrow transplantation-associated immune disease (BMT-ID)'. We do not support "ihe recent proposal by Fischer et al. 52to revise the rules of Billingham. The observation that, in the absence of a hlstocompatibility barrier between donor and recipient, 'GVH'-like pathology can occur in the CsA model is not correct since this syndrome is not the result of a proven GVH reaction. What should be revised is the idea that chronic GVH-like disease is always due to a GVH reaction. The term Br,4T-ID would cover all syndromes occurring after BMT that may be due to altoimmune reactivities and/or autoimmune reactivities. References

1 Simonsen, M., Bullman, J., Gammeltoft, A., Jensen, F. and Jorgenson, K. (1953) Acta PathoL MicrobioL Stand. 32, 1- 16 2 Simonsen, M. (i985) immunoL Rev. 88, 5-24 3 Bil!ingham, R.E., Brent, L. and Medawar, P.B. (1954) P/.-c Roy. Soc. 143, 43-58 4 Simonsen, M. (1957)Acta PathoL Microbiol. Stand. 40, 480-500

ruea -

5 Billingham, R.E. and Brer% L (1959) Phdos Trans. R Soc London 242,439 477 6 Bekkum van, D.W. and Vos~ O. (1957)J. Cell. Comp. Physiol. 50, 139 7 Congdon, C.C. and Urso, I.S. (1957)J. Exp Meal 33, 749--762 8 Rosenau, VV. and Moon, HD. (1961)J ,'Cat/Cancerlnst. 27, 47 I-483 9 Biqingham R.E., Defendi, V., Silvers, V I.K. and Steinmullef, D. (1962)J. Nat!Cancerlnst 28, 365 404 10 Gowans, J.L. (1962) Ann. NYAcaJ. 5o 99, 432 455 11 Gowans, I.L., McGregor, D.D., C=wen, DM. and Ford, C.I-. (1962) Nature 196, 651 655 12 Simonsen, M. (1962)Prog. Allergy 6, 349 461 13 Billingham, R.E. (;967 1968)Harveyl.ect. 62, 21 78 14 Thomas, E.D., Storb. R., Clift, R A. etal.(1975) NewEngl. J. Med. 292,832 843 15 Storb, R. (1985) l-okaiJ. Exp. Clin. Med. 10, 75 83 16 Goulmy, E., Gratama, J-W., Blok!anc~, E., Zwaan, F.E. and van Rood, J.J. (1983) Nature 302, 159 161 17 Hood, A.F., Vogelsang, GB., Black, LP, Farmer, E R. and Santos, G.W. (1987)Arch. Derrnatol. 123. 745 750 18 Sullivan, K.M. (1986) Int. J. Cell Cloning 4, ($I) 42 93 19 Ko,mgold, R. and Sprent, J. (1987) Transplantation 44, 335 339 20 O'Kunewick, J., Meredith, R.F., Raikow, R.B., Brozovich, B.J. and Maglier, K. (1981) Transplantation =11,201 204 2! O'Kunewick, J.P., Beschorner, W.E., Burro, M.J. and Kociban, D.L. (1985) Transplantation 39, 447 448 22 Parkman, R., RappepoR, J. and Rosen, F. (1980)J. Invest Dermatol. 74,276 279 23 Dokhelar, M-C., Wells, J., Lipinski, M. etal. (1981) Transplantation 31, 61 65 24 Ghayur, T., Seemayer, T.A, Konshavn, P.A.L., Gartner, _LG. and Lapp, W.S. (1987) Transplantation 44, 261 267 25 Piguet, P-F., Grau, G.L, Allet, B. and Vassalli, P. (1987) J. Exp. Med. 166, 1280 1289 26 Cerottini, J-C., Nordin, A.A. and Brunner, K.T. (i 971) J. Exp. Med. 134, 533 564 ~7 UDE.I. .II. .I E.' I , R. ( .I .~.O.J .l Immunol. Rev. 88, 25 58 ~[.I 28 Gale, R.P. (1985) Imrnunol. Rev. 88, 193 214 29 Roder, (1982)J. C;';n. Irnmunol. 2,249 263 30 Peters, P.M., Ortaldo, J.R., Sh~]laby, M.R. eral. (1986) ]. Immunol. 137, 2592 2598 31 Gui!lien, F.J., Ferrara, W.W., Hanock, W.W. etal. (1986) Lab. Invest. 55. 35-42 32 Sale, G.E., Shulman, H.M., Gaiucci, B.B. and Thomas, E.D. (1985) Am. J. Pathol. 118, 278 282 ]] EIs van, C.A.C.M., de Bueger, M.M., Kempenaar, J., Ponec, M. and Goulmy, E. (1989)J. Exp. Med. 170, 1469 1471 34 Graze, P.R. and Gale, R.P. (1979)Am J. Med. 66, 611 620 35 Rood, J.J. van, Goulmy, E. and van Leeuwen, A. (1987) Cellular Immunotherapy of Cancer Alan R Liss Inc. 36 Tsoi, M.S., Storb, R., Jones, E. etal. (1978)J Immunol. 120, 1485 1492 37 Tsoi, M.S., Storb, R., ~bbs, S., Med I L. and Thomas, E.D. (1980)J. Immu~:J/. 12%. 2258 2262 38 Tsoi, M.S., Storb, R., Dobbs, S. and Thomas, E.D. (1981) Nature 292, 355 357 39 EIs van, C.A.C.M., Bakker. A., Zwmderman, A.H. et al. Transplantation (in press) 40 EIs van, C.A.C.M., Bakker, A., Zwindelman, A.H. et al. Transplantation (in press) 41 Tutschka, P.J. (1987) Transplant. Proc 17, ($7) 69 74 42 Bos, G.M.J., Majoor, G.D., Slaaf, D.W. etal. (1989) Transplant. Proc. 21,362- 363 43 Glazier, A, Tutschka, P.J., Farmer, E.R. and Santos, G.W. (1983)3. Exp. Med. 158, 1-8 van Bekkum, D.W. (1985)in Bone Ma',ow Transplantation (van Bekkum, D.W. and Lowenberg, B., eds). pp. 147 2!2, Marcel Dekker Inc. 435

Immunology Today, Vol. 11, No. 12 1990

-ros ru F~scher, AC. Beschomer, W.E. and Hess, A.D. (1989)

Transplant Proc. 21, 3033-3035 46 Sorokin. R.. Kimura. H.. Schroder, K., Wilson, D.H. and Wilson. D B (1986)J. Exp. Med. 164, 1615-1625 47 Bos. G.M.J., Maloor. G.D., Willighagen, R.G.J. and van Breda VriesmaP. P.J.C.(1989)J. Invest. DermatoL 93, 6t0-615 Parkman. R. (1989)Irnmunol. Today 10, 362-364 49 Hess, A,D., Horwitz. L., Beschorner, W.E. and Santos, W. (1985) J. Exp. Med. 161. 718-730 50 Sakaguchi. S. and Sakaguchi. N. (1989)J. ImmunoL ~42, 471-480 51 Bos, GMJ., Majoer, G.D. and van Breda Vriesman, P.J.C. ......,.-,(1988) Clin. Exp. ImmunoL 74, 443-448

Fischer, A.C., l~eschorner, W.E. and Hess, A.D, (1989)

J. Exp. Med. 169, 1031-1041

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Antigen processingand presentation in vivo: the microenvironmentas a crucialfactor

Antigen processing and presentation in vitro is an increasingly well understood phenomenon. However, in wvo, a large number of variables conspire to obscure and confuse. In this article, Nico van Roo~en attempts to bring order to events that occur in the spleen after antigenic challenge: starting with the large body of reliable in vitro data he incorporates information on splenic anatemy, cell trafficking and the ceflular mkroenvironment to a rive at a physiological model for antigel~ handling in wvo.

Marc H.V. van Regenmortel has expressed the feelings of many immunologists - that the science of immunology is replete with inductive inferences that are far from deductively certain 1. 1he author referred to the philosopher of science, D.L. Hull, for his theorem "Science is not just the making of observations: it is the making of inferences on the basis of observation.: within the kamework of a theory. -2. The above opirion seems to be particularly apt for ~mmunolJgical research on antigen processing and its presentation to T cells. ,, ,e list of cell types that have this capability in vitro is growing continuously. Clearly, an in vivo framework i- needed to understand how different cells and their microenvironment might coeperate in the inductive phase of ~n imt~une response. A role for m~crophages in tile processing of antigens as an initial step in immunity" n~s been the subject of numerous studie'~ since the ear!y 1960,- (Ref. 3). In the 1970s, dendritic cells were shown to be able to present antigens to T cells4 and the last decade has seen a rapidly increasing body of literature en 3 cells as the most likely candidates for both processing and presentation c," antigens to T cellss 7 In spite of the latter developments, macrophages are still be!ieved by many authors to trigger immune reactions since intracellular processing of antigers followed by presentation of the processed antigen to T cells has been demonstrated unequivocally for these cells8--~°. Moreover, small albumin antigens, which failed to elicit a

Dept of Histology, Medical Facdty, Vrije Universiteit, ~d. Boechcrststraat7, 1081 BTAmsterdam, The Netherlands. 436

NJcovan Rooijen detectable antibody response when injected intravenously in mice, did induce a substantial response when they were targeted to macrophages by incorporation in !iposomes. Elimination of the macrophages in the spleen reduced the response significantly 11. From these studies it appears that B cells and dendritic cells cannot handle certain antigens without pr~processing by macrophages. Most recent evidence .'avouring a role for macrophages, dendritic cells and B cells in the processing and presentation of antigens to T cells has been obtained from in vitro studies; that is, in Lhe absence of certain normal cell populations and their microenvironment. Under such conditions several tell populations obtained from various organs in the body can initiate an antigen-:pecific ~,timulation of T cells8-1°. These in vitro studies form an indispensable contribution i0 the understanding of the mechanism of intracellular antigen processing and major histocompatibility complex (MHC)-restricted presentation to T cells. However, the question of which of these cell types are involved in the early events in the immune response in vivo can only be answered if the microenvkonment of the cells is taken into account. Ta~ing the example of the spleen, a lymphoid organ that receives antigens from the circulation, B cells, T cells, macrophages and dendritic cells all have characteristic locations within one or other of the splenic compartments 12. Here, a hypothesis is presented on how B cells, T cells, macrophages and dendritic cells may cooperate in the induction of immune reactions in the spleen. It is a speculative attempt to link the results of in vitro and in vivo studies and is based on the assumption that the microenvironment plays a crucial role in determining which cells have which function in immunity. It is also based on a model in which antigen uptake and processing on the one hand, and antigen presentation and promotion of T-cell (~'~ 1990. Elsevier Science Publishers Ltd. UK 0167--4919/90/502,00