Endocardial infiltrates in human heart transplants: A serial biopsy analysis comparing four immunosuppression protocols

Endocardial Infiltrates in Human Heart Transplants: A Serial Biopsy Analysis Comparing Four lmmunosuppression Protocols R, D. CLARKE FORBES, MD, FRCP(C), REED A. ROWAN, PtiD, AND MARGARET E. BILLINGHAM, MB, E&, FRCPATH Endocardial mononuclear cell infiltrates were studied in 2,350 consecutive biopsies from 172 patients over a period ranging from 4 to 16 months post cardiac transplantation. The patients, otherwise unselected, were equally subdivided into four groups based upon the specific type of maintenance immunosuppression used. This was to allow for comparison of the effects of four separate commonly used recipient immunosuppression protocols, which could potentially influence the characteristics of the infiltrates. With azathiaprine-corticosteroid immunosuppression, endocardial infiltrates in otherwise normal biopsies were exceedingly rare, very minor, and invariably unifocal. Mild and moderate rejection were associated with a highly significant stepwise increase in incidence, prominence, and multifocality of endocardial infiltrates. In contrast, with each of the three cyclosporine-based recipient immunosuppression protocols which were evaluated, approximately 15% of biopsies with no evidence of rejection were associated with endocardial infiltrates. There was a wide range of variation in the prominence of the endocardial infiltrates present. Multifocal infiltrates were frequently encountered, the incidence of which was exclusively dependent upon the specific cyclosporine-based immunosuppression protocol used. With mild and with moderate rejection there was a significant stepwise increase in overall biopsy incidence of all endocardial infiltrates in each of the three groups, although there was no variation in relative prominence of the infiltrates, or in incidence of multifocality when biopsies without rejection were compared. The presence of conspicuous vascular spaces within endocardial infiltrates and significant extension of the infiltrates into the adjacent myocardium, with or without associated myofiber necrosis, were characteristic features of the most prominent endocardial infiltrates in all three cyclosporine-based immunosuppression groups. This constellation of changes has sometimes been referred to as “Quilty” effect. The relative incidence with which these particular features were encountered in association with endocardial infiltrates did not vary with rejection category of the biopsies. This study has shown that the presence of all forms of endocardial infiltrates, in the absence of concomitant

From the Departments of Pathology, McGill University, Montreal, Quebec, Canada; and Stanford University School of Medicine, Stanford, CA. Accepted for publication December 4, 1989. Dr Forbes is a visiting scholar in Cardiac Pathology, Stanford University School of Medicine, Stanford, CA. Dr Forbes was supported by the Medical Research Council of. Canada Grant No. MT-6083; Drs Billingham and Rowan are supported by a National Institutes of Health Transplantation Grant No. HL-13108. Key words: endocardial infiltrates, heart transplantation, cyclosporine. Address correspondence and reprint requests to R. D. Clarke Forbes, MD, FRCP(C), Department of Pathology, McGill University, Lyman Duff Medical Sciences Bldg, 3775 University St, MontrCa1, Quebec H3A 2B4 Canada. 0 1990 by W.B. Saunders Company. 0046-8177/90/2108-0011$5.00/0

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rejection, is a characteristic manifestation of cyclosporine-based recipient immunosuppression, regardless of the specific protocol and cyclosporine dosage schedule. Under azathiaprine-based immunosuppression, endocardial infiltrates are almost invariably associated with rejection. It is postulated that cyclosporinerelated endocardial mononuclear cell infiltration, in the absence of overt rejection, may result from a low level alloimmune response secondary to fluctuations in cyclosporine drug levels or related factors, and that the incidence with which these infiltrates occur can be augmented during acute rejection episodes when the strength of the recipient immune response is magnified. HUM PATHOL 21:850-855. 0 1990 by W.B. Saunders Company.

Focal endocardial mononuclear cell infiltrates are frequently encountered in the absence of underlying rejection in endomyocardial biopsies of human heart transplants when cyclosporine is used for recipient immunosuppression. The most prominent of these infiltrates, which has been referred to as “Quilty” e ff ec t , may be confused with significant rejection, particularly when there is infiltration of the adjacent myocardium with associated myofiber damage. ‘Z Similar infiltrates, in the absence of rejection, have been said not to occur in association with the earlier azathiaprine-based immunosuppression protocols, although no formal documentation of comparative data has been published.* The cause of cyclosporine-related endocardial infiltrates is unknown. This systematic histopathologic study provides a comprehensive comparative analysis of the characteristics of endocardial infiltrates in both normal biopsies and in biopsies with rejection from patients maintained on four separate commonly used recipient immunosuppression protocols. The resultant data provide further insight into the nature and possible underlying causes of endocardial mononuclear cell infiltration in transplanted human hearts. MATERIALS AND METHODS The study consisted secutive endomyocardial

of a histologic biopsies from

examination

of con-

172 patients over a period ranging from 4 to 16 months after cardiac transplantation. All patients underwent heart transplantation and were subsequently followed at the Stanford University Medical Center. Biopsies of forty-three patients maintained on each of the following immunosuppression protocols were studied: (1) azathiaprine-corticosteroids (azathiaprine group); (2) cyclosporine-corticosteroids (cyclosporine group); (3) azathiaprine-cyclosporine-corticosteroids (triple therapy group); and (4) prophylactic early posttransplant

ENDOCARDIAL INFILTRATES [Forbes et al]

0KT3 combined with the triple therapy protocol (OKT3 group). The last three groups are representative of patients maintained on the three most commonly used cyclosporinebased immunosuppression protocols currently in use. In each separate group, all biopsies of 10 unselected patients were studied up to 4 months, all biopsies of 10 unselected patients were studied up to 8 months, all biopsies of nine unselected patients were studied up to 1 year, and all biopsies of 14 unselected patients were studied up to 16 months. A total of 2,350 biopsies were analyzed. The azathiaprine group consisted of 641 biopsies, the cyclosporine group of .599 biopsies, the triple therapy group of 619 biopsies, and the OKT3 group of 491 biopsies. An average of 3.3 fragments from each biopsy was assessed in all three cyclosporine-based treatment groups. In the azathiaprine group an average of 2.6 fragments were evaluated. The vast majority of the biopsies were performed using the Stanford bioptome with !JF sleeve.‘,’ Endocardial mononuclear cell infiltrates (El), when present, were graded as follows: (1) Grade 1 (Gl). infiltrate with a depth of five lymphoid cells or less: (2) Grade 2 (G2), infiltrate with a depth of six to 10 lymphoid cells: (3) Grade ((;3), infiltrate with a depth of greater than 10 lymphoid cells. For determination of EI grade, the number of mononuclear cell layers was counted directly at the site of greatest El depth. The method of measurement of EI depth in terms of. cell layers was based on the observation that EI in biopsies OF paiients under cyclosporine immunosuppression consist primarily of monomorphic lymphoid cells.‘,? Endocardial infiltrates in all biopsies were further classified in terms 01‘ the presence or absence of multifocal infiltrates, extension (of infiltrates into the adjacent myocardium, associated myofiber degeneration or necrosis, prominent vascular spaces within the infiltrates, and relation of infiltrates to remote previous biopsy sites. Remote previous biopsy sites were identified as generally irregular craterform fibrous scars associated with local compensatory myofiber hypertrophy and/or disarray.‘.2 Only G2 and G3 EI in direct relation to remote biopsy sites were enumerated. The rejection status of all biopsies was determined and correlated with EI incidence and with all of the above EI parameters. Acute rejection was graded according to the Stanford Classification,‘~’ except that ongoing, resolving, and resolvled rejection categories were omitted and the relevant biopsies were classified as moderate, mild, or no evidence of rejection (NER) according to the histologic features. Also. severe rejection, being very infrequent, was combined and categorized with moderate rejection. Endocardial infiltrates in biopsies showing acute rejection were further subdivided on the basis of whether or not focal EI were exclusively confined to the same fragment(s) in which acute rejection was present. The incidence of diffuse continuous involvement of the entire endocardial surface of two or more biopsy fragments by EI in association with rejection was also documented. All statistical analysis was done using the x2 method.

well as the incidence of all El and various El grades in each category are summarized graphically in Fig 1. Only three of 480 NER biopsies in the azathiaprine group were associated with EI (0.07%). In all three biopsies EI were Gl, unifocal, and without infiltration of the adjacent myocardium (Fig 2). In biopsies with rejection, (1) overall incidence of all El (P < O.OOl), (2) relative grade distribution of EI (P < 0.025), and (3) incidence of multifocal El (P < 0.05) all showed a highly significant positive stepwise increase with rejection category, when NER, mild rejection and moderate rejection biopsies were compared. One-third of all mild rejection biopsies showed associated EI, all of which were Gl or G2. The Gl:G2 ratio was 11:3. Sixty percent of moderate rejection biopsies showed concomitant El. The Gl :G2:G3 ratio was 4:3: 1. Focal EI confined exclusivelv to specific biopsy fragment(s) showing acute rejection were present in 12% of mild rejection biopsies and 7% of moderate rejection biopsies with El. Diffuse continuous EI involving the entire endocardial surface of two or more biopsy fragments were present in < 1% of mild rejection biopsies and 7% of moderate rejection biopsies. Diffuse El with mild rejection were all Gl; G2- or G3-diffuse El were associated with moderate rejection. In biopsies with moderate rejection showing this feature, rejection tended to be extensive, involving most if not all of the biopsy fragments. In two biopsies from one patient in this group in which extensive toxoplasma myocarditis was present, there was also an associated diffuse continuous G3 El. These two biopsies were otherwise formally excluded from the study. Infiltration of the underlying myocardium by El was confined to three biopsies, all of which showed moderate rejection. The El were all G3. There was no

RESULTS

100

m

Endocardlial Infiltrates in Biopsies of Patients in the Azathiaprine-Corticosteroid Immunosuppression Group [Azathiapirine Group).

mild

The numerical rejection, and

l-

distribution moderate

MILD

rm)DmATE

FIGURE 1. The distribution of endocardial infiltrates in biopsies with no evidence of rejection (NER), mild rejection (MILD], and moderate reiection (MODERATE1 in the azathiaorine immunosuppression g&up. To&l number oi biopsies in ea& cateaoty are represented by black bars. El + represents the biopsy incidence of all endocardial infiltrates in each cateaorv. Gl. 62. and 63 represent the biopsy incidence of grades 1.2, dnd 3 endocardial infiltrates in each category, as defined in the Materials and Methods section.

of biopsies in NER, rejection categories as

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Endocardial Infiltrates in Biopsies of Patients in the Three Cyclosporine-based lmmunosuppression Groups (Cyclosporine, Triple Therapy, OKr3 Groups) The numerical distribution of biopsies in NER, mild rejection, and moderate rejection categories and the incidence of all EI and various EI grades in each category are summarized graphically for each separate group in Fig 3. In striking contrast to the azathiaprine group, approximately 15% of NER biopsies in the three cyclosporine-based treatment groups showed EI: 6% Gl, 3% G2, 6% G3. There was a significantly higher incidence of Gl EI in the cyclosporine group (9%). There were no significant intergroup differences in relative incidence of other EI grades. There was a highly significant positive association between overall biopsy incidence of EI, regardless of grade, and rejection category of the biopsies in all three groups (moderate rejection > mild rejection > NER, P < 0.001). This relationship did not significantly differ from that observed in the azathiaprine group. In contrast to the azathiaprine group, none of the three cyclosporine-based immunosuppression groups showed a significant variation in relative incidence of any EI grade(s) when NER, mild rejection, and mod-

FIGURE 2. Minor localized grade 1 endocardial infiltrate five cells in depth in an endomyocardial biopsy from a patient in the azathiaprine immunosuppression group. There was no evidence of rejection (Hematoxylin-eosin stain; magnification x 300.)

associated myofiber damage. Conspicuous vascular spaces within EI were not present in any of the biopsies. Endocardial infiltrates specifically related to remote previous biopsy sites were not observed.

600 600 W

5oc -

q

El+ Gl

0

1004 MILD

600

NR

M3DERATE

,

n q

MILD

M3DEFlATE

1 11OKT3

Biopsies

El+ Gl

FIGURE 3. The distribution of endocardial infiltrates in biopsies with no evidence of rejection (NER), mild rejection (MILD), and moderate rejection (MODERATE) in the (top left] cyclosporine, [top right] triple therapy, and [bottom) prophylactic OKT34riple therapy immunosuppression groups. Total number of biopsies in each category are represented by black bars. El + represents the biopsy incidence of all endocardial infiltrates in each category. Gl, 62, and 63 represent the biopsy incidence of grades 1, 2 and 3 endocardial infiltrates in each category, as defined in the Materials and Methods section.

100 m

MILD

MXDEFWTE

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ENDOCARDIAL INFILTPATES (Forbes et al)

erate rejection biopsies were compared. However, overall EI grades in each of the three groups were higher than those encountered in the azathiaprine group in association with both mild (P < 0.0001) and moderate (P < 0.025) rejection. This was also clearly the case rfor NER biopsies (P < 0.0001). In contrast to the azathiaprine group, in which all El in NER biopsies were unifocal, multifocal El were present in NER biopsies in all of the three cyclosporine-based immunosuppression groups. The biopsy incidence of multifocal EI was greater in the triple therapy and 0KT3 groups (40%‘) than in the cyclosporine group (20%) (P < 0.025), and this was unrelated to other variables. The incidence of multifocal EI did not vary with rejection category of the biopsies in all three groups, in contrast to the azathiaprine group. However, multifocal EI were significantly more common in the three cyclosporine-based treatment groups than in the azathiaprine group in association with both mild (P < 0.0012). and moderate (P < 0.025) rejection. The incidence of focal EI exclusively confined to specific fragment(s) which showed concomitant acute rejection, and the incidence of diffuse continuous EI involving the entire endocardial surface of fragments in association with rejection, showed no significant intergroup differences and did not significantly difler in any way from the azathiaprine group. As in the diffuse continuous El were azathiaprine group, largely associated with extensive multifocal acute re,jection. Extension of EI into the adjacent myocardium showed a highly significant positive correlation with EI grade in all three cyclosporine-based immunosuppression groups (G3 > G2 > Cl, P < 0.005). There were no intergroup differences. A total of 12%) of all EI showed intramyocardial extension, SO% of which were G3 EI (Fig 4). In about 20% of biopsies with intramyocardial extension of EI, localized degenera-

tion or necrosis of adjacent cardiac myofibers was also present (Fig 5). Prominent vascular spaces within EI were almost exclusively associated with G3 EI; less than 1% of G2 EI contained conspicuous vascular spaces (Fig 6). There were no intergroup differences. There was also no significant variation in biopsy incidence of any of these parameters in association with EI when NER, mild rejection, and moderate rejection biopsies were compared. The overall findings were in contrast to the azathiaprine group, in which only 3% of EI showed intramyocardial extension, all in association with moderate rejection, and neither myofiber necrosis nor prominent vascular spaces within EI were observed. Approximately 6% of all G2 or G3 EI present in each of the three cyclosporine-based treatment groups was directly related to remote previous biopsy sites, about 20%’ of which also contained conspicuous vascular spaces (Fig 7). The overall incidence of EI in relation to previous biopsy sites was independent of rejection category. As previously noted, biopsy site-related EI were not observed in the azathiaprine group. DISCUSSION This study has shown that in endomyocardial biopsies of heart transplants from patients maintained on azathiaprine-corticosteroid immunosuppression, EI were exceedingly rare in NER biopsies, and when present were extremely minor and invariably unifocal. Significant EI were exclusively confined to biopsies with rejection. Incidence, grade, and multifocality of EI showed a strong stepwise positive correlation with mild and with moderate rejection. Thus, with azathiaprine-based immunosuppression, EI are a direct reflection of the acute rejection process, and are rarely observed and are of trivial significance in NER biopsies.

FIGURE 4. Prominent grade 3 endocardial infiltrate with extensive infiltration of the underlying myocardium in an endomyocardial biopsy frorn a patient in the triple therapy immunosuppression group. There was no evidence of rejection. (Hematoxylin-eosin stain; magnification x 300.)

FIGURE 5. Extension of endocardial infiltrate into underlying myocardium with prominent intermyocyte extensions and myofiber damage. Endocardial infiltrate is depicted in Figure 4. (Hematoxylin-eosin stain; magnification x 450.)

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FIGURE 6. Prominent grade 3 endocardial infiltrate with conspicuous vascular spaces, There is minor infiltration of the adjacent myocardium. Endomyocardial biopsy is from a patient in the cyclosporine immunosuppression group. There was no evidence of rejection, (Hematoxylin-eosin stain; magnification x 250.)

In the three cyclosporine-based immunosuppression groups, EI of all grades were present in about 15% of NER biopsies. There was no striking intergroup variation. Thus, the presence of EI in NER biopsies in these three groups is not directly related to maintenance cyclosporine dosage levels, or to the specific cyclosporine-based immunosuppression protocol used. In all three groups, as in the azathiaprine group, there was a highly significant stepwise increase in overall EI biopsy incidence when NER, mild rejection, and moderate rejection biopsies were compared. In contrast to the azathiaprine group, however, mild and moderate rejection in all three cyclosporinebased immunosuppression groups were unassociated with significant differences in relative incidence of

FIGURE 7. Prominent grade 3 endocardial infiltrate in area of irregular dense fibrosis containing lipocytes and surrounded by cardiac myofibers showing compensatory hyperlrophy and disarray. This is characteristic of a remote previous biopsy site. Endomyocardial biopsy is from a patient in the cyclosporine immunosuppression group. There was no evidence of rejection. (Hematoxylin-eosin stain: magnification x 250.)

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the various EI grades, or of multifocal EI, when compared with NER biopsies. The results of this study indicate that EI are an integral component of the acute rejection process in human heart transplants, regardless of the nature of the immunosuppression protocol used. With azathiaprine-based recipient immunosuppression, NER biopsies are unassociated with significant EI, and the incidence, grade, and degree of multifocality of EI show a direct correlation with rejection severity. In contrast, with all cyclosporine-based immunosuppression protocols, NER biopsies are associated with a significant incidence of EI of all grades, as well as multifocal EI, and rejection is reflected only by increased biopsy incidence of all EI, without statistically significant variations in relative incidence of EI grade distribution or in incidence of multifocality. It has also been shown that diffuse continuous EI involving the entire endocardial surface of a significant number of biopsy fragments are closely associated with extensive acute rejection regardless of the nature of the immunosuppression protocol. However, prominent diffuse EI may also occur with extensive myocarditis regardless of etiology as indicated by the biopsy findings in association with toxoplasma myocarditis in the current study, and by the observations of others in non-transplant settings.“*4 Prominent endocardial infiltrates unrelated to acute rejection were first described in heart transplants shortly after the introduction of cyclosporine for maintenance immunosuppression. The term “Quilty” e ff ec t is commonly used to denote substantial focal EI which may be present in this setting, and is named after the first patient in which the phenomenon was observed. l.* “Quilty” effect has been shown to occur in 10% of biopsies under cyclosporinecorticosteroid immunosuppression; it does not in itself require augmentation of immunosuppression, and tends to disappear in subsequent biopsies, but may recur. “Quilty” effect is generally described as a prominent localized mononuclear cell EI, characteristically containing conspicuous vascular spaces, and although often confined to the endocardium, frequent cellular infiltration of the adjacent myocardium has also been reported, sometimes in association with myofiber necrosis. “Quilty” effect can be present in otherwise normal biopsies, but can also coexist in biopsies with acute rejection.‘J The current study indicates that all significant EI in the absence of rejection, regardless of prominence, multifocality, or specific associated morphologic features, are characteristic of endomyocardial biopsies from patients maintained on all current forms of cyclosporinebased immunosuppression. Since the presence of conspicuous vascular spaces and adjacent intramyocardial extension with or without myofiber necrosis shows a strong correlation with the prominent G3 EI, “Quilty” effect represents a particularly distinctive manifestation of this general phenomenon. This study has also shown that all of the specific features frequently associated with “Quilty” effect do not vary in relative incidence as a feature of EI when NER,

ENDOCARDIAL INFILTRATES (Forbes et al)

of this hypothesis, manipulative studies in an appropriately controlled experimental model are required. This comparative biopsy analysis has characterized the features of EI under four separate immunosuppression protocols based upon a combination of several specific morphologic parameters. The data define and clearly delineate the characteristics of EI which are associated with azathiaprine and with cyclosporine-based recipient immunosuppression, and provide further insight into the nature of the possible causative factors involved in the development of EI in human heart transplants.

mild rejection, and moderate rejection biopsies are compared. The cause of El in human heart transplants associated with cyclosporine-based immunosuppression is unknown. It has been suggested that EI may represent a lymphoproliferative process related to posttransplant immunosuppression.5 However, neither lymphoma nor infection has been shown to be implicated in the development of cyclosporine-related EI.’ Alth.ough acyclovir-responsive endocardial lymphomatous infiltrates associated with Epstein-Barr virus infection have been reported, the morphologic features are said to be readily distinguishable from cyclosporine-associated EI.” Also, the current data clearly indicate that a putative lymphoproliferative origin must be specifically related to cyclosporinebased immunosuppression, since EI do not significantly occur in the azathiaprine treatment group in the absence of rejection. This is in contrast to the evidence that development of posttransplant lymphoproliferative disease, in general, is unrelated to the specific nature of the immunosuppression regimen used.7 The possibility has also been raised that EI may represent a direct drug effect of cyclosporine itself.” If this is the case, EI are clearly not cyclosporine dose-related. The current study has shown that there is al high incidence of EI in all forms of cyclosporine-based immunosuppression, regardless of the specific protocol and the cyclosporine dosage schedule. Finally, it has been postulated that El associated with cycllosporine-based immunosuppression may represenr a chronic or recurrent form of indolent rejection., which is probably unrelated to graft outcome.* Endocardial mononuclear cell infiltration is a characteristic early feature of unmodified acute rejection in experimental models, and is never observed in fully syngeneic heart graft controls.g.10 Early rejection in azathiaprine-immunosuppressed recipients is also frequently associated with EI, as demonstrated by this and other previous studies,‘.2 while EI are very rarely present in NER biopsies. It has been shown that cyclosporine drug levels may undergo va.riable degrees of fluctuation dependent upon the degree of absorption as well as other factors.x.l ‘-I9 Cyclosporine does not alter T cell allorecognition, but acts primarily through specific depression of lymphokine generation, which is absolutely required for alloactivation and generation of a signiflcan t recipient alloimmune response. l 1,12Thus, the heart transplant could be subject to constant but generally ineffective probing by recipient allospecific T cells. Fluctuations in cyclosporine levels could lead to variabile degrees of T cell escape from the lymphokine inhibitory effect. This could result in a variable recurrent indolent alloimmune response, which could manifest itself primarily as EI. The overall incidence of EI could be further augmented during overt rejection episodes, to a degree reflective of the strength of the magnified recipient immune response, as indicated by this study. To test the validity

Acknowledgment. The authors are grateful for the assistance of Graciela Rodriguez, Marilyn Masek, and Eileen Mackay in typing and arranging the manuscript, and thank Pauline Zera for the preparation of the figures.

ADDENDUM Since submission of this manuscript, Suit et al have reported a significant negative correlation between whole blood cyclosporine levels and the presence of endocardial infiltrates-that is, a lower whole blood cyclosporine level was associated with an increased probability of endocardial infiltrates.” I‘hese data provide further evidence for the hypothesis developed in this study.

REFERENCES 1. Billingham ME: The postsurgical heart. The pathology of cardiac transplantation. Am J Cardiovasc Path01 I :3 19-334, 1988 2. Billingham ME: Endocardial biopsy detection of acute rejection in car&ac allograft recipients. Hea;t Vessels 1:86-90, 1985 3. Aretz HT. Billingham ME. Edwards WD. et al: Mvocarditis. A histopathologic defi&ion and’ classification. Am J (‘tardiovasc Pathol 1:3-14. 1986 4. Marcus LC. Steer AC, Duray AE. et al: Fatal pancarditis in a patient with co-existing Lyme disease and habesiosis. Ann Intern Med 103:374-376, 1985 5. Kemnitz 1. Cohnert T, Shafers Hi. et al: A classification of cardiac allograft-rejection. A mod&cat&n of the classification by Billingham. Am J Surg Pathol 11:503-515. 1987 6. Stovin PGI: Monitoring of cardiac rejection by endomyocardial biopsy, in Wallwork J (ed): Heart and Heart Lung Transplantation. Philadelphia, PA, Saunders, 1989, pp 313-340 7. Penn I: Malignant lymphoma in organ transplantation. Transplant Proc 13:736-738, 1981 8. Billingham ME: The diagnosis of acute cardiac rejection by endomyocarc%al biopsy, in Woln>r E. Laczkovics A. Ha&l M (edsj: New Trends in Heart TransDlantation. Bib1 Cardiol. Basel. Switzerland, Karger Basel, 1988, ‘pp 83-102 9. Forbes RDC, Guttmann RD: Pathogeneric studies of cardiac allograft rejection using inbred rat models. Immunol Rev 775-29, 1984 10. Westra AL, Romaniu KA, Ruffa ‘T. et al: Infiltration pattern of rat heart allografts during rejection. Transplant Proc 19:374, 1987 t 1. Kahan BD: Cyclosporine: The agent and its actions. Transplant Proc 27:5-16, 1985 12. Shevach EM: The effects of cyclosporine A on the immune system. Ann Rev Immunol 3:397-423. 1985 13. de Groen PC: Cyclosporine, low-density lipoprotein, and cholesterol. Mayo Clin P;oc 65:1012-1021, 1988 . ’ 14. Suit PF. Kottke-Marchant K, Ratliff NB, et al: Comparison of whole-blood cyclosporine levels and the frequency of endocardial infiltrates (the Quilty lesion) in cardiac transplantation. Transplantation 48:6 18-62 1, 1989

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