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Inductive and differentiative signals delivered by C8-substituted guanine ribonucleosides Michael G. Goodman The C8-substituted guanine ribonucleosides are a recently described class of immunomodulator which among a variety of pleiotropic effects have T-cell-replacing activity in vitro and immunopotentiating activity in immunocompromised hosts. In this article Michael Goodman discusses the mechanism of nucleoside transport, the immunobiologic activities of these compounds, and the nature of the cellular interactions involved. The mechanisms by which B lymphocytes are triggered to enter the cell cycle, proliferate, and subsequently differentiate into immunoglobulin secreting cells have been the focus of considerable recent study. Activation appears to be initiated when information-bearing molecules interact with specific receptors located in the plasma membrane. Such interactions are thought to perturb the receptor, causing new associations between intramembranous proteins and ultimately transducing the signal across the lipid bilayer. The nature of the events occurring during and subsequent to signal transduction as well as the identity of the molecular species responsible for these events are not clearly understood. A considerable number of candidates have been proposed as potential mediators of lymphocyte proliferation and differentiation, carrying the signal from the cytoplasmic surface of the cell membrane to the cellular replicative machinery. For nearly a decade, the role of guanosine 3'-5'-cyclic monophosphate (cGMP) as the intracellular mediator of B-lymphocyte activation has been controversial. In 1973, it was first suggested that conjoint actions of adenosine 3'-5'-cyclic monophosphate (cAMP) and cGMP might mediate the induction of B cells to antibody secretion. Induction of B-cell tolerance, in contrast, appeared to be mediated by cAMP acting alone 1. Soon thereafter it was observed that lymphocyte stimulation with the B-cell specific mitogen bacterial lipopolysaccharide (LPS) caused significant rise in intracellular content of cGMP. Moreover, exogenous cGMP was able to stimulate mitogenic responses in spleen cells derived from normal or congenitally athymic (nu/nu) mice 2-4. Weinstein and colleagues 5,6made the interesting observation that cGMP when derivatized at C8 with bromine was considerably more stimulatory than cGMP in native form. This was ascribed to the increased lipophilicity of the brominated analogue. However, reports from Parker's group in St. Louis 7'8 demonstrated clearly that stimulation of human lymphocytes with a variety of agents, although causing a transient elevation of cAMP, did not alter the intracellular content of cGMP. Moreover, Burleson and Sage 9 were unable to document mitogenicity for exogenous cGMP in the guinea pig system. Department of Immunology IMM9, Scripps Clinic and Research Foundation 10666 N. Torrey Pines Road, LaJolla, CA 92037, USA.

Watson later reported that of a variety of B cell mitogens tested, only LPS appeared capable of elevating cGMp10, and furthermore, that exogenous cGMP was incapable of inducing B cells to secrete immunoglobulin 11. Finally, Burleson and Sage 9 and Weber and Goldberg '2 failed to observe any correlation between changes in cAMP or cGMP with the mitogenic or nonmitogenic nature of the probe to which lymphocytes were exposed. In re-evaluating the role of cGMP in B-lymphocyte activation, we directly compared the mitogenicity of exogenously added cGMP and its analogues 13. These studies clearly indicated that whereas native cGMP induced low level proliferation in lymphocytes and dibutyryl cGMP was entirely ineffective in this regard, 8 bromo-cGMP, however, consistently exhibited 3-5 fold stimulation. These unanticipated results suggested that perhaps the nature of the substituent on the cyclic nucleotide was of greater importance than the cyclic nature of the molecule itself. Therefore, 8Br-cGMP was analysed in successive stages, and the C8 bromiriated forms of the cyclic nucleotide, the noncyclic mononucleotide, and the nonphosphorylated nucleoside were compared directly. These studies indicated that the brominated nucleoside (8BrGuo) was approximately 1.5-2 orders of magnitude more potent as a B lymphocyte stimulant per molecule than was the derivatized cyclic nucleotide (Fig. 1, left). The brominated noncyclic mononucleotide (8BrGMP) was intermediate in potency. Kinetic pulse analysis similarly indicated that D N A synthesis occurred most rapidly and to the highest degree in cells incubated with the nucleoside analogue, whereas the mononucleotide and cyclic nucleotide activated DNA synthesis at a slower rate and to lesser degrees (Fig. 1, right). These observations suggest the existence of a hierarchy from brominated nucleoside to mononucleotide to cyclic nucleotide. Other studies, examining the effectiveness of 8 bromoguanine, lacking the D-ribose at N9, substantiated that the derivatized base was without eft~ct 14. Other guanine nucleosides and nucleotides act as immunomodulators. Deoxyguanosine (dGuo) inhibits T suppressor (Ts) cell generation, allowing a suppressed B or T cell response to return to uninhibited levels 15. In lethally irradiated and reconstituted mice, interference with Ts development by dGuo led to the occurrence of a transient spike of homogeneous immunoglobulins in the serum 16. A detailed examination of all immunomoda© 1984, Elsevier Science Publishers B.V., Ams'tcrdam

0167 - 4919/84/$02.04)

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Immunology Today, vol. 5, No. 11, 1984 24 50

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Fig. 1. Comparison of [SH] TdR u~take in splenic lymphocyte cultures stimulate with 8BrGuo, 8BrGMP or 8Br-cGMP. Left. Spleniclymphocytes(4 x 10 ) from CBA/CaJ mice were cultured with incremental concentrationsof I"18BrGuo (--), 8BrGMP (---), 8BrGMP ( . . . . ) for two days. Cultures were pulsed with 1 ~Ci [3H]-TdR for the final 24 hr of incubation. Right. Splenic lymphocytes(4 x 10s) from CBA/CaJ mice were cultured with lmM 8BrGuo, 10raM 8BrGMP, or 10 mM 8Br-cGMP. At varioustimes 1/~Ci[3H]-TdR was added to each set of culturesand 2 hr later this set was harvested.(Reprintedfrom Goodmanand Weigle13.)The key is the same as in the left hand panel. latory nucleosides is beyond the scope of this review; for the effects of a thymidine analogue 17, inosiplex le, adenine arabinoside ~9, and nicotinamide adenine dinucleotide ~° the reader is referred to other sources. Intracellular m e d i a t i o n of l y m p h o c y t e a c t i v a t i o n

That the C8 substituted guanine ribonucleosides are analogues of natural building blocks for cellular nucleic acid suggested that rather than triggering the cell at the cell membrane these lymphocyte activators might he transported into the cell by the purine nucleoside transport system and subsequendy activate the cell from an intracellular ske. In studies designed to evaluate uptake of radiolabelled 8 bromoguanosine, a short initial period of linear uptake was followed by a plateau phase suggesting that a period of unidirectional flux was followed shortly thereafter by bidirectional flux. The magnitude of uptake was dependent upon the concentration ofnucleoside with which the cells were incubated 22. Evaluation of the rate of uptake as a function of concentration indicated that two saturable systems existed for the uptake of these nucleosides. The high affinity system has a K m of 8ttlvI while the low affinity system has a K m of 480/~M. These values are quite analogous to those found experimentally for native guanosine in our laboratory and those reported for adenosine by Strauss and colleagues 2~. Evaluation of the same parameters during the period of bidirectional flux failed to provide evidence of saturation kinetics, strongly suggesting that uptake of these nucleosides occurs by a carrier-mediated transport mechanism.

The foregoing evidence, however, in no way indicates that the site at which triggering occurs is intracellular. Therefore, 8 mercaptoguanosine (8MGuo), a C8 substituted nucleoside with activity identical to 8BrGuo, was conjugated to activated CH-sepharose 4B via a six carbon spacer arm to limit the access of the nucleoside to the interior of the cell 2~, Interposition of the spacer arm between the active nucleoside and the sepharose matrix was done to maximize interaction potential between the nucleoside and the cell and to minimize sequestration in and steric hindrance by the bead. This approach clearly indicated that immobilization of 8 M G u o on sepharose beads entirely abrogated its stimulatory activity. Another approach to immobilization was to offer cells the brominated nucleoside in polymerized form. To this end, polyguanylic acid (Poly G), average molecular weight in excess of 400,000, was brominated at C8 according to the protocol of Lafer et al. 23. Subsequent digestion and analysis of the residue indicated that virtually all guanosine had been brominated. In its polymerized form, 8 bromo G M P , mitogenic in its own right, was immobilized only through 3' and 5' phosphodiester linkages analogous to those of the cyclic nueleotide which was also stimulator,/13. However, the polymerized nucleotide was entirely ineffective as a B cell stimulant in contrast with the high degree of activity of monomeric 8-bromo G M P . Thus again, restriction of access to the interior of the cell appears to correlate with loss of biologic activity. This situation contrasts markedly with that reported by Gregory and Kern for stimulation of thymo-

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TABLE I. Evidencethat C8 substituted guanine ribonucleosidesact at an intracellularsite Observation Consistent with intracellularactivation 1 Carrier mediated transport 2 8MGuo sepharoseconjugatefails to activate 3 Poly8Br guanylieacid failsto activate 4 Membrane rigidificationfailsto impede activation 5 Crosslinkingsurface IgM with wholeAb fails to impede activation

Consistent with plasma membrane activation

cytes by agarose-bound mononucleotides 24. These investigators studied the mitogenic effects of A T P and G T P on thymocytes and found that these nucleoside triphosphates induced a single round of D N A synthesis in two different classes of immature thymic (but not splenic) T cells through a putative initiation site at the cell membrane. Other work from our laboratory has tested the hypothesis that transduction of signals across the plasma membrane should be far more susceptible to interference with plasma membrane function than signals exerted at an intracellular location. Rigiditication of the plasma membrane by incubation with high concentrations of cholesterol 25 did in fact significantly impair the responses to bacterial lipopolysaccharide and purified protein derivative of tuberculin, while leaving the response to 8BrGuo entirely intact. Moreover, crosslinking of membrane immunoglobulin receptors by anti-IgM antibodies was highly inhibitory for the responses to LPS and staphylococcal protein A but not for that to 8 M G u o ~2. The evidence that C8-substituted guanine ribonucleosides act at an intracellular site is summarized in Table I. The foregoing approaches did not deal with the possibility that these nucleosides might ultimately be substrates for guanylate cyclase. Therefore cells were incubated with radiotabelled 8BrGuo or native Guo. The nucleoside pools were extracted and metabolites analysed by thin layer chromotography (TLC) on polyethyleneimine cellulose plates. The resultant data revealed that 8BrGuo was neither degraded nor phosphorylated and did not appear in the cyclic nucleotide fraction. Underivatized radiolabelled Guo however, was degraded and extensively phosphorylated within 15 minutes 26. Other experiments examined the possibility that these derivatized nucleosides might in some manner stimulate the activity of guanylate cyclase, serving as a stimulus for increased production of endogenous cGMP. Careful evaluation of cyclic nucleotide levels by radioimmunoassay, however, indicated clearly that this was not the case 26. Further work on the mechanism of action of these derivatized ribonucleosides will investigate the existence of cytoplasmic or nuclear receptor molecules for these agents.

Effects of C8 d e r i v a t i z e d g u a n i n e ribonucleosides on humoral immunity In other studies, the immunobiologic properties of these compounds have been examined. First, the ability of the C8 derivatized guanine ribonucleosides to induce nonspecific secretion of immunoglobulin was assessed 1~. B cell cultures containing either 8 M G u o or 8BrGuo produced high levels of immunoglobulin-secreting cells as assayed on the test antigens SRBC or trinitrophenyl. As in the case for proliferation, underivatized guanosine was entirely ineffective, and again the same hierarchy of activity from brominated nucleoside to mononucleotide to cyclic nucleotide was in evidence. Moreover, when agents that elevate intracellular levels of c G M P were assessed for their capacity to induce polyclonal immunoglobulin secretion, they were entirely ineffective. Onodera et al. 29 found that 8BrGuo was able to induce resting B cells to polyclonal synthesis of ~1, )t2, and ),3 light chain subtypes. These investigators used the nucleoside as a probe to determine the rates at which each Jt chain subtype was synthesized as well as the precursor frequencies for cells making each subtype. Additionally, when 8BrGuo was tested as a polyclonal stimulus for B cells from h u m a n tonsil, peripheral blood, and bone marrow, 8BrGuo induced the generation of polyclonal IgM, IgG and IgA in amounts equal to or greater than pokeweed-mitogen induced responses 3°. Polyclonal activation, however, occurred in the absence of consistently detectable proliferation, in the hands of

Cellular parameters Normal splenic B cells and spleen cells from congenitally athymic (nu/nu) mice respond to the proliferative properties of 8MGuo, whereas thymocytes and splenic T cells do not. Two subpopulations of B cells appear to respond to this nucleoside 27. The predominant one is a mature population that bears surface b-chains, Ia antigens, C receptors, and (by indirect evidence) the Lyb 3, 5, and 7 antigens. These cells also bear/a-chain and Fc receptors. In addition, a second, minor subpopulation of

TABLE II. Characterizationof 8MGuo responsiveand unresponsive lymphocytes 8MGuo responsive 8MGuo unresponsive nu/nu spleen cells thymocytes splenicB cells(Thy 1.2 - ) splenicT cells(NW passed) FcR ÷ cells FcR- cells /a÷ cells W cells 6+ cells most 6 cells Ia ÷ cells most Ia- cells CR + cells most CR- cells Lyb 3/5/7÷ cells most Lyb 3/5/7- cells

less mature cells that bear only/a-chain and Fc receptors also appears to be reactive to 8 M G u o (Table II). The existence of this immature, reactive B cell subset was confirmed by observation of 8 M G u o responsiveness in lymphocytes from four day old mice whose cells do not yet exhibit these later-appearing markers 2s. Accessory cells appear to play a minimal, if any, role in the 8 M G u o response. These results establish two distinct B cell subpopulations as the major and minor cellular targets of C8-derivatized nucleosides, and suggest that activation results from a direct interaction between nucleoside and cell.

322 TABLE III. Effectsof C8 derivatizedguanine ribonucleosideson humoral immunity I Polyclonalactivationof B cellsto secreteimmunoglobulin II Antigen specificenhancement of: A Primary in vitro responseto T-dependent antigens B Secondaryin vitro IgM and IgG responsesto T-dependent antigen C Primary in vitro responseto T-independent antigens (TI2>TI-1) D Primary in vivv responseto T-dependent antigens III T cell-likeinductivesignal for B cellspresented with T-dependent antigens A Occursindependentlyof T cellsand T cell-derived lymphokines B Is enhanced synergisticallyby T cellsand T cell-derived lymphokines Osundwa et al. so as well as ourselves (Goodman, M. G. et al., unpublished observations). This human polyclonal response was found to be essentially T ceU-independent. The C8-derivatized guanine ribonucleosides exert potent adjuvant effects both in vitro and in vivo (Table III). The in vitro effects are easily observed when 8 M G u o is added to murine spleen cells cultured with sheep erythrocytes 3~. Moreover, the adjuvant effect is not attributable to polyclonal activation in so far as the magnitude of the antigen-specific response in cultures containing antigen and nucleoside is far in excess of the sum of the polyclonal and antigen-specific responses assessed independently. Adjuvanticity is exerted not only on the primary antibody response but also upon the activity of memory ceils challenged in vitro with antigen to mount a secondary IgM and IgG response. Adjuvanticity is always greatest at optimal antigenic concentrations. W h e n the immunoenhancing properties of these compounds are tested on the responses to thymus-independent (TI) antigens, reactivity to TI-2 antigens is enhanced far more than that to TI-1 antigens, This suggests that there may perhaps be some element ofT-cell contribution to the adjuvant response since TI-2 responses are far more T-dependent than are TI-1 responses 32'~3.Examination of the kinetics ofnucleoside addition to culture indicates that supplementation of cultures can be delayed two or even three days after initiation of four-day cultures with retention of full immunoenhancement. This suggests not only that these derivatized nucleosides can enhance an ongoing immune response, but also that clonal expansion of antigen-reactive cells may not be critical to the mechanism of adjuvanticity since addition of nucleoside can be delayed so long. In vivo adjuvanticity of 8MGuo and 8BrGuo is observed when uptake of the nucleosides into the circulation is retarded by presentation of the nucleosides as an insoluble suspension 34. Administration of the insoluble material on the day of immunization results in the requirement for doses on the order of 25-30 mg per animal. When administration of nucleoside is delayed until three or four days after immunization, however, optimal immunoenhancement is observed at concentrations approximately two orders of magnitude lower. While these effects are observed at all concentrations of antigen tested, maximal immunoenhancement occurs at optimally immunogenic concentrations. Still other experiments demonstrate that oral administration of deri-

Immunology Today, vol. 5, No. 11, 1984

vatized nucleoside is effective over a broad dose range. In vivo responses tested to date include the IgM response to SRBC and the IgG response to H G G . Finkleman et al. have found that administration of 8 M G u o in vivo increases the Ab response to goat IgG in both IgG~ and IgG 2 subclasses (F. Finkleman, personal communication). In addition, Ahmed and M o n d have evaluated the effects of 8 M G u o on the humoral response to the TI-2 antigen TNP-ficoll. In normal mice the IgG 2 response was markedly enhanced, with a lesser degree of IgG~ enhancement. I g M and IgG 3 responses were unaltered (J. Mond, personal communication). T cell replacing activity of C8 d e r i v a t i z e d guanine ribonucleosides Further study of the role played by T cells in the immunoenhancing effect of these nucleosides led to the striking observation that 8 M G u o and 8BrGuo are capable of transmitting T cell-like inductive signals for B cells in vitro 3s. Thus, cultures of spleen cells that had been rigorously depleted of T lymphocytes and were entirely unable to respond to T-cell dependent antigens in vitro, acquired the capability of mounting high level responses to such antigens in the presence of 8MGuo or 8BrGuo. The antigen-specific component of this type of response far exceeded the nonspecific (polyclonal) response also initiated in the presence of these agents in vitro. Likewise, cultures of spleen cells from congenitally athymic mice, which are unable to respond to T D antigens, became capable of sizeable responses to such antigens when cultures were supplemented with derivatized nucleosides. B cell-enriched lymphocyte populations generated from normal spleen cells or from congenitally athymic animals contain precursor T cells which can be expanded under stimulation by lymphokines 36'37'38.Therefore, the potential influence of such lymphokines in vitro was abrogated by adding the immunosuppressive agent cyclosporin A to culture in the minute concentrations shown previously to inhibit production of I L I and IL2 (Ref. 39). At concentrations of cyclosporin A as low as 100 ng/m the response of spleen cells to the T-dependent antigen SRBC was entirely inhibited. However, antigen stimulated cultures containing optimal concentrations of 8MGuo retained not only their full degree of antigen reactivity but also immunoenhancement comparable to that shown by uninhibited cultures 35. These experiments indicated clearly that the immunoenhancing effects of 8 M G u o are independent not only of mature T cells but also o f T cellderived lymphokines. These data, however, should not be misconstrued to suggest that there is no role for T cells in immunoenhancement by 8MGuo. W h e n cultures containing B cells, rnacrophages, and antigen were supplemented with optimal concentrations of 8MGuo, the otherwise T-dependent response again became T-independent. However, when such cultures additionally received T cells equal to the number of B cells cultured, the magnitude of the resultant response was increased another 2- to 4-fold. The magnitude of this increment suggested the existence of a synergistic relationship between the T-independent response and the effect of T cells. Moreover, a similar synergistic relationship could be demon-

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strated for cultures of B cells and macrophages containing SRBC and derivatized nucleosides in the presence oflymphokines generated by mixed lymphocyte cultures. These effects are summarized in Table III. W h e n the mechanism by which these derivatized nucleosides transmit T cell-like inductive signals was compared to the T cellreplacing activity contained in lymphokine preparations, it was found that the T cell-replacing activity of IL-2-associated lymphokines acted optimally at an early time in the culture period and was totally ineffective when added three days after initiation of cultures 35. In contrast, optimal concentrations of 8BrGuo acted efficiently when added three days after initiation, and in fact when cultures received nucleoside at this time and IL-2 associated lymphokines at day 0, an additive response was observed. This evidence strongly suggests that T cell-like signals from these lymphokines act by a mechanism entirely distinct from that of the C8-derivatized nucleosides. A hitherto unknown defect in the responsiveness of B lymphocytes from SoIL mice to activation by C8-substituted guanine nucleosides has enabled us to distinguish definitively two qualitatively distinct classes of signal delivered to B cells by C8-substituted guanine ribonucleosides. This defect renders B cells from SoIL mice unresponsive to the inductive (early acting) signal of 8 M G u o that culminates in nonspecific B cell activation. Unresponsiveness is not attributable to a shift in either the dose-response or kinetic profiles, nor can the presence of suppressor cells be demonstrated. In striking contrast, SJL B cells exhibit normal responsiveness to the differentiative (T cell-like, or late acting) signal provided by 8MGuo. This signal enables SJL B cells, depleted of T cells and adherent cells, to respond to T cell-dependent antigens, and synergizes with T cell-derived lymphokines. These data suggest that nonspecific secretion of immunoglobulin is dependent upon both inductive and differentiative signals, that antigen alone can supply an effective inductive signal for antigen-specific responses, and that the SJL mouse will provide a useful model for selective study of inductive v. differentiative events. Experiments examining the effects of these nucleosides on populations of purified T cells have failed to provide any evidence of direct T cell activation. Splenic T cells do not proliferate when stimulated with these derivatized nucleosides 27, nor do they produce IL2. The magnitude of primary mixed lymphocyte reactions is not amplified by these agents, nor have we been able to augment helper T cell activity generated in vivo by administration of these nucleosides as insoluble suspensions. Effects on murine models of immunodeficiency The potential benefit of an agent with potent immunoenhancing or T cell-replacing activities lies in the possibility of restoring a significant degree of immune function to an immunocompromised host. The CBA/N mouse, carrying the xid defect, an x-linked deficit in the function of a B-cell subpopulation bearing Lyb 3/5/7 surface antigens 4°'4~'42,was chosen as the first subject for these experiments. Immunodeficient F1 male mice generated a poor response to the T-dependent antigen SRBC under both in vitro and in vivo conditions. This deficient response could be restored to nearly normal levels in vitro by addition of

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8MGuo to antigen-activated cultures, and in vivo by administration of the nucleoside as an insoluble suspension 34(Fig. 2). Not only were the responses restored to the level generated by phenotypically normal F t female mice, but the magnitude of the responses was in fact comparable to the nucleoside-augmented responses observed in normal animals. Ahmed and M o n d have observed that the inability of CBA/N mice to respond to TNP-ficoll can be reversed in the presence of 8 M G u o (Ahmed and Mond, unpublished observations). Another model of immunodeficiency, that provided by the congenitally athymic (or nude) mouse, has been discussed earlier. Finally, the immunodeficiency model provided by the ageing mouse was studied. These mice are unresponsive to T-dependent antigens 43, do not make IL-2 responses to lectin stimulation 4~, and exhibit immune deficits in both B-cell and T-cell compartments 45. The responsiveness of spleen cells from these animals to T-dependent antigens can be restored to normal levels by supplementation of cultures with optimal concentrations of 8-mercaptoguanosine, but not of LPS. This correlates with the demonstrated inability of LPS to activate polyclonal immunoglobulin secretion in cells from these animals ~, whereas we have observed that they are quite responsive to the polyclonal effect of 8MGuo. The biochemical basis for this dichotomy is not yet clear, but it may perhaps relate to a defect in plasma membrane function 47,48, serving to limit the ability of LPS and similar ligands to transduce differentiative signals across the membrane. Many potential applications exist for these compounds in the clinic. The adjuvant activity of these substituted nucleosides may make them useful adjuvants for use with synthetic vaccines, in order to maximize induction of immunity to a variety of pathogenic agents. Moreover, the ability of these compounds to function as immunomodulators may be used to advantage in a number of situations. First, they may allow lower dosage schedules of antibiotics to be used for patients with bacterial (or fungal) infections, thereby lowering the incidence of

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untoward reactions. Second, they may be useful in a variety of immunodeficiency diseases in which B cell inactivity leads to significant morbidity, such as common variable hypogammaglobulinemias, aging, and immunodeficiency associated with chronic disease. Finally, these nucleosides may find application in the context of pharmacologically- or radiologicaUy-induced immunosuppression. ~I] I thank Ms. Alice Bruce Kay for excellent secretarial work in preparation of the manuscript. This is publication number 3423 I M M from the Department of Immunology, Scripps Clinic and Research Foundation, La Jolla, California. This work was supported in part by United States Public Health Service Grants AI 15284 and AI 07007, and Research Career Development Award AI 00374.

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Graft-versus-host reactions: clues to the etiopathology of a spectrum of immunological diseases Ernst Gleichmann*, Steven T. Pals, Anton G. Rolink¢, Thadd~ius Radaszkiewicz* and Helga Gleichmann§ Parental strain T lymphocytes injected into adult FI mice respond to allogeneic M H C antigens and so induce the symptoms of systemic graft v. host disease (GVHD). GVHD has two forms, stimulato~y and suppressive, and in each there are pathological changes which resemble those seen in a variety of human disorders of immune regulation. In this article Ernst Gleichmann and his colleagues review the pathogenesis of these disorders and their induction by G V H reactions. Central Laboratory of the Netherlands Red Gross Blood Transfusion Service, incorporating the Laboratory of Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, The Netherlands. *Division of Immunology, Medical Institute of Environmental Hygiene, 1)-4000, Dfisseldorf 1, FRG.; *Basel Institute for Immunology, Basel, Switzerland; *Institute of Pathology, University of Vienna, Vienna, Austria; SDiabetes Research Institute at the Universky of Dfisseldorf, D/.isseldorf, FRG. © 1984,ElsevierSciencePublishersB.V.,Amsterdam 0167- 4919/84/$02.00

Graft-versus-host reactions (GVHR) When parental strain lymphocytes are injected into adult F1 hybrid (i.e. semi-allogeneic) mice, the recipients are genetically tolerant of parental strain donor ceils but the grafted T lymphocytes on the other hand react against allogeneic histocompatibiIity antigens of the host and thus induce a spectrum of pathological alterations (Fig. 1A). The generation of some of these alterations depends on