Hypogammaglobulinemia following cardiac transplantation: a link between rejection and infection

Hypogammaglobulinemia Following Cardiac Transplantation: A Link Between Rejection and Infection Mohamad H. Yamani, MD,a Robin K. Avery, MD,b Steven D. Mawhorter, MD,b James B. Young, MD,a Norman B. Ratliff, MD,c Robert E. Hobbs, MD,a Patrick M. McCarthy, MD,d Nicholas G. Smedira, MD,d Marlene Goormastic, MPH,e David Pelegrin, RN,a and Randall C. Starling, MD, MPHa Background: Hypogammaglobulinemia (HGG) has been reported after solid organ transplantation and is noted to confer an increased risk of opportunistic infections. Objectives: In this study, we sought to assess the relationship between severe HGG to infection and acute cellular rejection following heart transplantation. Methods: Between February 1997 and January 1999, we retrospectively analyzed the clinical outcome of 111 consecutive heart transplant recipients who had immunoglobulin G (IgG) level monitoring at 3 and 6 months post-transplant and when clinically indicated. Results: Eighty-one percent of patients were males, mean age 54 ⫾ 13 years, and the mean follow-up period was 13.8 ⫾ 5.7 months. Patients had normal IgG levels prior to transplant (mean 1137 ⫾ 353 mg/dl). Ten percent (11 of 111) of patients developed severe HGG (IgG ⬍ 350 mg/dl) post-transplant. The average time to the lowest IgG level was 196 ⫾ 125 days. Patients with severe HGG were at increased risk of opportunistic infections compared to patients with IgG ⬎ 350 mg/dl (55% [6 of 11] vs 5% [5 of 100], odds ratio ⫽ 22.8, p ⬍ 0.001). Compared to patients with no rejection, patients who experienced three or more episodes of rejection had lower mean IgG (580 ⫾ 309 vs 751 ⫾ 325, p ⫽ 0.05), and increased incidence of severe HGG (33% [7 of 21] vs 2.8% [1 of 35], p ⫽ 0.001). The incidence of rejection episodes per patient at 1 year was higher in patients with severe HGG compared to patients with IgG ⬎350 (2.82 ⫾ 1.66 vs 1.36 ⫾ 1.45 episodes/patient, p ⫽ 0.02). The use of parenteral steroid pulse therapy was associated with an increased risk of severe HGG (odds ratio ⫽ 15.28, p ⬍ 0.001). Conclusions: Severe HGG after cardiac transplantation may develop as a consequence of intensification of immunosuppressive therapy for rejection and hence, confers an increased risk of opportunistic infections. IgG level may be a useful marker for identifying patients at high risk. J Heart Lung Transplant 2001;20:425–430.

From the Departments of aCardiology, bInfectious Diseases, c Anatomic Pathology, dCardiothoracic Surgery, and eTransplant Center, Kaufman Center for Heart Failure, Cleveland Clinic Foundation, Cleveland, Ohio. Submitted March 9, 2000; accepted November 6, 2000. Reprint requests: Randall C. Starling, MD, Heart Transplant Medical Services, Cleveland Clinic Foundation, Department of

Cardiology/F25, 9500 Euclid Avenue, Cleveland, Ohio 44195. Telephone: 216-444-2268. Fax: 216-444-7155. Copyright © 2001 by the International Society for Heart and Lung Transplantation. 1053-2498/01/$–see front matter PII S1053-2498(00)00331-4

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nfectious complications are an important cause of morbidity and mortality in solid organ transplantation. The risk of opportunistic infection is determined by the interaction between the epidemiologic exposures that the patient encounters and the patient’s net state of immunosuppression.1 A multiinstitutional study has identified certain major risk factors for cumulative infections after heart transplantation.2 These risk factors include older recipient age, ventilator support at transplant, ventricular assist device at transplant, black donor, female donor, and OKT3 induction therapy. Recently, hypogammaglobulinemia (HGG) has been identified as a significant factor associated with increased risk of opportunistic infections and poorer outcome after renal and lung transplantation.3,4 Similarly, Avery et al5,6 reported an increased risk of opportunistic infections in heart transplant recipients who were profoundly hypogammaglobulinemic. The two major barriers to successful transplantation, rejection and infection, are closely tied together by the current requirement for lifelong immunosuppressive therapy.7 Grossi et al8 noted that almost 33% of infectious episodes in heart transplant recipients were preceded by supplementary treatment for rejection with steroid pulses or cytolytic therapy. We postulate that HGG may serve as a common factor in these processes of rejection and infection, possibly reflecting the underlying intensity of the immunosuppressive therapy. Some of the newer efficacious purine antagonists, such as mycophenolate mofetil (MMF), possess an anti–B-cell effect9; however, MMF’s potential relationship to significant HGG is unknown. To the best of our knowledge, this is the first report to describe the relationship of HGG to both rejection and infection, identifying the use of parenteral steroid pulse therapy as a major risk factor, and unraveling the importance of IgG level monitoring in identifying high-risk patients.

METHODS Between February 1997 and January 1999, we retrospectively analyzed the clinical outcome of 111 heart transplant recipients at the Cleveland Clinic Foundation. Patients underwent IgG level monitoring at baseline (pre-transplant), at 3 and 6 months post-transplant, and when clinically indicated. HGG was defined as IgG ⬍ 700 mg/dl. IgG was classified as mild (501 to 700 mg/dl), moderate (350 to 500 mg/dl), or severe (⬍ 350 mg/dl). Mean follow-up period was 13.8 ⫾ 5.7 months.

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Acute cellular rejection was defined as Grade 3A (moderate) or greater using International Society Heart Lung Transplant (ISHLT) criteria. Moderate rejection was treated with pulse steroid therapy, oral prednisone 100 mg/day, or intravenous methylprednisolone 1 g/day, for 3 consecutive days. Eight patients received OKT3 and 11 patients were switched from cyclosporine to tacrolimus for the treatment of recurrent rejection. The maintenance immunosuppressive therapy included three possible regimens: azathioprine/cyclosporine (AZA/CsA; n ⫽ 23), mycophenolate mofetil/cyclosporine (MMF/CsA; n ⫽ 77), or mycophenolate mofetil/ tacrolimus (MMF/FK; n ⫽ 11). All patients were maintained on oral prednisone. Infectious outcomes were described as asymptomatic cytomegalovirus (CMV) viremia, and clinical infectious diseases. The latter category consisted of opportunistic (including symptomatic CMV disease) and non-opportunistic infections.

Statistical Analysis Data are presented as mean ⫾ standard deviation. The chi-square and Fisher exact tests were used to test for the association between IgG level group and the corresponding variables: immunosuppressive regimen; infection outcome; and rejection episodes. The Kruskal–Wallis test was used to test for differences in levels between groups and Spearman’s correlation was used to test for significant association between two continuous factors. Logistic regression analysis was used to determine the odds ratio of developing severe HGG following the use of steroids. p ⬍ 0.05 was considered statistically significant.

RESULTS The baseline characteristics of the patient population are described in Table I. The mean pre-transplant IgG level of the patient population was 1137 ⫾ 353 mg/dl. Fifty-seven percent (63 of 111) of the patient population developed HGG (IgG ⬍ 700 mg/dl). Fifty-one percent (34 of 63) were classified as mild (IgG 501 to 700 mg/dl), 32% (18 of 63) as moderate (IgG 350 to 500), and 17% (11 of 63) as severe HGG (IgG ⬍ 350 mg/dl). The average elapsed time post-transplant to the lowest IgG level recorded was 196 ⫾ 125 days.

Correlation Between IgG Level and Risk of Infection During follow-up, 41 of 111 (37%) patients developed asymptomatic CMV viremia. Patients with

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TABLE I Baseline characteristics Patients (n) Gender (M) Donor age (years) Recipient age (years) Etiology Dilated cardiomyopathy Ischemic cardiomyopathy Mean follow-up period (months) Ischemic time (minutes) LVAD, n (%) OKT3 induction, n (%)

111 81% 35.7 ⫾ 4 54.3 ⫾ 13 45% 55% 13.8 ⫾ 5.7 174.7 ⫾ 51.0 21 (19%) 9 (8%)

LVAD, left ventricular assist devie (as a bridge to transplant).

severe HGG were noted to have a higher incidence of this complication compared with the other IgG level groups (Table II), which was independent of the CMV donor/recipient serostatus among the different IgG level groups. The design of the study precludes us from knowing the exact time relationship between the onset of CMV viremia and onset of

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HGG due to the lack of simultaneous IgG level determination. During the same follow-up period, 29 of 111 (26%) patients experienced a total of 32 clinical infection episodes, resulting in an infection rate of 0.29/patient. Eighty-one percent (26 of 32) of the infection episodes occurred within 6 months of transplant. The spectrum of infectious diseases included 19 bacterial (Streptococcus pneumonia, 1; Staphylococcus epidermidis, 2; Staphylococcus aureus, 7; Klebsiella pneumonia, 1; Enterococcus spp, 1; Acinetobacter spp, 1; Pseudomonas aeruginosa, 3; Serratia marcesens, 1; Nocardia spp, 2), 11 viral (influenza A, 1; herpes zoster, 2; respiratory syncitial virus, 1; and tissue-invasive and symptomatic CMV syndrome, 7), and 2 fungal (Candida and Aspergillosis) pathogens. The time relationship between HGG and subsequent development of clinical infections is shown in Table II. Patients with severe HGG had a higher incidence of opportunistic infections (Table II) (odds ratio ⫽ 22.8, p ⬍ 0.001). However, there was

TABLE II Risk of infection in relation to IgG level IgG Category (mg/dl)

n ⫽ 111 (%) Mean IgG level (mg/dl) Tx-lowest IgG* (days) Lowest IgG clinical infaction† Asymptomatic cytomegalovirus viremia, n (%) Clinical infections,‡ n (%) Opportunistic Non-opportunistic Total

<350

350 to 500

501 to 700

>700

11 (10%) 217 ⫾ 70 172 ⫾ 91 86 ⫾ 106 7 (64%)

18 (16%) 414 ⫾ 34 217 ⫾ 144 288 ⫾ 288 5 (28%)

34 (31%) 599 ⫾ 52 176 ⫾ 103 99 ⫾ 55 7 (21%)

48 (43%) 908 ⫾ 219 208 ⫾ 138 85 ⫾ 123 22 (46%)

0.66 0.46 0.02

6 (55%) 1 (9%) 7 (64%)

0 (0%) 1 (6%) 1 (6%)

1 (3%) 2 (6%) 3 (9%)

4 (8%) 3 (6%) 7 (15%)

⬍0.0001 0.67 ⬍0.001

p-Value

*Represents the average time interval (days) from transplant date to development of hypogammaglobulinemia (lowest igG level) in each category. The IgG ⬎700 category is considered normal. † Represents the average time interval (days) from the lowest igG level in each category to the development of clinical infection. ‡ Infections that occurred after the development of hypogammaglobulinemia.

TABLE III Maintenance immunosuppressive therapy in relation to IgG level IgG (mg/dl)

AZA/Csa MMF/Csa MMF/FK Total (n)

<350

350 to 500

501 to 700

>700

Total, n (%)

2 (9) 4 (5.2) 5 (45.5)* 11 (10)

4 (17) 13 (17) 1 (9) 18 (16)

5 (22) 26 (33.8) 3 (27.3) 34 (31)

12 (52) 34 (44) 2 (18.2) 48 (43)

23 77 11 111 (100%)

AZA, azathioprine; MMF, mycophenolate mofetil; Csa, cyclosporine; FK, tacrolimus. All patients were on oral maintenance prednisone. *p - 0.001 when comparing MMF/FK group to the other immunosuppressive regimens.

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TABLE IV Cumulative steroid dose for the treatment of acute rejection in relation to IgG level IgG (mg/dl)

n ⫽ 111 (%) Oral prednisone (mg) Intravenous methylprednisolone (g)

<350

350 to 500

501 to 700

>700

p-Value

11 (10) 409 ⫾ 336 5.18 ⫾ 4.04

19 (16) 366 ⫾ 300 1.33 ⫾ 2.35

34 (31) 388 ⫾ 358 0.79 ⫾ 1.70

48 (43) 293 ⫾ 347 0.68 ⫾ 1.54

NS ⬍0.001

no significant difference in the risk of non-opportunistic infections among the different IgG groups.

Correlation Between IgG Level and Immunosuppressive Therapy There was no significant difference in the use of OKT3 (induction, n ⫽ 8 patients; and treatment of rejection, n ⫽ 9 patients) among the different IgG level groups. A greater percentage of patients in the tacrolimus group had severe HGG than in those on cyclosporine-based therapy (45.5% [5 of 11] vs 6% [6 of 100], p ⫽ 0.001) (Table III). These patients were switched from cyclosporine to tacrolimus because of recurrent rejection, and hence the development of severe HGG was related to the underlying severity of rejection rather than due to the tacrolimus itself. Using logistic regression analysis, the parenteral use of pulse steroid therapy for the treatment of rejection was associated with an increased risk of developing severe HGG (odds ratio ⫽ 15.28, p ⬍ 0.001). There was no significant difference in the long-term maintenance or cumulative oral prednisone dose used for the treatment of acute rejection among the different IgG level groups (Table IV). However, the cumulative parenteral steroid dose used for the treatment of rejection was much higher in the profoundly hypogammaglobulinemic patients (Table IV).

FIGURE 1 Mean IgG level in relation to frequency of rejection episodes.

Correlation Between IgG Level and Acute Rejection Seventy-four percent (82 of 111) of patients experienced a total of 168 episodes of rejection, 85% of which occurred within the first 3 months of transplant. Patients who experienced three or more episodes of rejection had a significantly lower mean IgG level (Figure 1) and higher incidence of severe HGG (Figure 2) compared to patients with less frequent episodes. The 1-year incidence of rejection per patient was also significantly higher in patients with severe HGG compared with the other IgG groups (Figure 3).

Time Relationship Between Rejection and Infection The mean time interval from transplant to the first episode of rejection was 31 ⫾ 40 days, and from the last episode of treated rejection to infection was 54 ⫾ 76 days. Fifty-three percent (17 of 32) of the infectious episodes were preceded by rejection. Almost half of these episodes (8 of 17) occurred within 2 weeks of treating a rejection episode. Infection was noted to occur simultaneously with rejection in 19% (6 of 32) of the cases. However, it preceded rejection in 12% of the cases (4 of 32), with a mean time interval of

FIGURE 2 Incidence of severe

hypogammaglobulinemia in relation to frequency of rejection episodes.

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FIGURE 3 Incidence of rejection episodes per patient at 1 year in relation to IgG level.

49 ⫾ 35 days. Also, in 16% (5 of 32) of the cases, infection occurred in the absence of rejection.

DISCUSSION HGG has recently been shown to be associated with increased risk of opportunistic infections following heart transplantation.5 This is the first report to elucidate the relationship of HGG to both rejection and infection in the heart transplant population. Our major findings include: (i) patients with severe HGG are at increased risk of clinical infections, a finding similar to what has been described before in renal3 and lung4 transplant recipients; (ii) the clinical infection rate in our study, 0.29/patient, falls within the reported range (0.09 to 0.92 infections/ patient) of a multi-institutional study for infections after heart transplantation2; and (iii) the increased risk of infection in our study was mainly due to opportunistic infections, which reflects the immunosuppressed state of the severely HGG patient. HGG would traditionally be more associated with bacterial infections due to encapsulated organisms such as Streptococcus pneumonia and Hemophilus influenza. The absence of a significant difference of non-opportunistic infections among the different IgG groups may be related to the small sample size of our patient population, and hence the low event rate. It is also possible that the absolute total IgG level may not be accurate enough in predicting such complications, but rather the IgG subclass determination would have been a better indicator. Second, patients with recurrent episodes of rejection are at increased risk of developing severe HGG, and hence patients with severe HGG had a higher 1-year incidence of rejection. Thus, HGG may develop as a consequence of intensification of immunosuppressive therapy for rejection, and subsequently lead to infection.

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The nature and intensity of the underlying immunosuppressive therapy may provide a possible explanation as to the identity of such a link, considering that over half of the infection episodes (53%) were preceded by a treatment for rejection. These immunosuppressant agents affect primarily T-cell function (ie, lymphokine production), which in turn may alter B-cell function indirectly.10 Alternatively, some of the newer agents, such as mycophenolate mofetil, may affect B-cell function directly.9 It is thus possible that immunoglobulin production may be altered as a result of these B-cell effects (indirect and/or direct pathways). The relatively high incidence of CMV (both asymptomatic viremia and CMV disease) in our profoundly hypogammaglobulinemic patients and the relative absence of encapsulated organisms may suggest that the lymphokine-dependent T-cell mechanism (indirect pathway) is the operational one. Identification of the IgG subclasses would be helpful in further clarifying the underlying mechanism, because it has been previously shown that antibodies against CMV antigens belong primarily to the IgG1 and IgG3 subclasses,11 which are known to be T-cell dependent.12 The lack of IgG subclass determination is a limitation of this study, but it did not affect our results, because we were able to show increased incidence of CMV viremia and opportunistic infection in correlation with the reduction of total IgG level, an easier parameter to measure. This is in contrast to a recently reported experience in a renal transplant population in which selective low IgG1 level, but not reduced total IgG, was associated with severe infections.13 We were unable to identify a particular immunosuppressive regimen responsible for the high incidence of severe HGG, although it may appear from the results in Table III that tacrolimus-based therapy is more associated with severe HGG. No relationship of causality between the two could be proven, especially because 11 patients who were originally on cyclosporine were switched to tacrolimus due of recurrent rejection. The development of severe HGG was more likely related to the increased frequency of rejection episodes rather than to tacrolimus itself. Therefore, the MMF/tacrolimus group is inherently biased due to its selectivity, and comparison between these regimens may be unjustifiable. This represents another limitation of the study, however, it was not our primary objective to pinpoint a specific culprit regimen. It would be worthwhile to compare the different immunosuppressant regimens in a future, larger scale trial. Our report clearly describes the time relationship

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between infection and rejection. Fifty-three percent of the infection episodes were preceded by rejection and almost half of these episodes occurred within 2 weeks of treatment. Grossi et al8 reported a similar observation where supplementary treatment for rejection preceded the infection episodes by ⬍15 days in 55% of the cases, again illustrating the importance of this rejection–infection time relationship. Yet, nearly half of the infection complications did not have an event heralding their potential occurrence. This makes the role of a screening mechanism, such as IgG determination, more relevant. A third major finding of this study is that the parenteral use of steroid pulse therapy was associated with a significant risk of developing severe HGG (odds ratio ⫽ 15.28, p ⬍ 0.001). However, the maintenance prednisone dose and the cumulative oral prednisone dose used for the treatment of rejection did not differ significantly between the different IgG level groups. It is well known that large doses of glucocorticoids are lympholytic,14 and that corticosteroids may affect leukocyte migration15 and activation.16 Wieneke et al13 reported a reduction of T-helper cells in renal transplant recipients treated with steroid pulses for acute rejection—thus affecting T-cell– derived lymphokines, which in turn may alter B-cell function and hence IgG subclass composition. In conclusion, the intensified immunosuppressive therapy coupled with the parental use of pulse steroid therapy for the treatment of acute rejection predisposes heart transplant recipients to profound HGG that is associated with increased risk of CMV and opportunistic infections. IgG level monitoring may be helpful in identifying high-risk patients who may be considered potential candidates for immunoglobulin therapy, close monitoring, and enhanced prophylaxis for infection.

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early, cumulative, and fatal infections after heart transplantation: a multiinstitutional study. J Heart Lung Transplant 1996;15:329 – 41. Braun WE, Avery R, Gifford RW Jr, Straffon RA. Life after 20 years with a kidney transplant: redefined disease profiles and an emerging nondiabetic vasculopathy. Transplant Proc 1997;29:247–9. Goldfarb NS, Stillwell PC, Mehta AC, et al. Hypogammaglobulinemia as a possible cause of deterioration in lung transplant patients. J Heart Lung Transplant 1998;17:86. Avery RK, Mawhorter SD, Starling RC, et al. Hypogammaglobulinemia in heart transplant recipients treated with tacrolimus, mycophenolate, and prednisone. Transplantation 1999;67(suppl):S95. Corales R, Chua J, Mawhorter SD, et al. Significant hypogammaglobulinemia in six heart transplant recipients: an emerging clinical phenomenon? Transplant Infect Dis. In press. Kontoyiannis DP, Rubin RH. Infection in the organ transplant recipient. Infect Dis Clin N Am 1995;9:811–22. Grossi P, De Maria R, Caroli A, Zaina MS, Minoli L. Infections in heart transplant recipients: the experience of the Italian Heart Transplantation Program. Italian Study Group on Infections in Heart Transplantation. J Heart Lung Transplant 1992;11:847– 66. Kobashigawa JA. Mycophenolate mofetil in cardiac transplantation. Curr Opin Cardiol 1998;13:117–21. Carpenter CB. Immunosuppression in organ transplantation. N Engl J Med 1990;322:1224 – 6. Mathiesen T, Brattstrom J, Anderson A, Linde P, Ljungman P, Wahren B. Immunoglobulin G subclasses and lymphocyte responses to cytomegalovirus in transplant patients with cytomegalovirus infections. J Med Virol 1992;36:65–9. Rosenberg YT, Chiller JM. Ability of antigen-specific helper cells to effect a class-restricted increase in total Ig-secreting cells in spleen after immunization with the antigen. J Exp Med 1979;150:517–30. Wieneke H, Otte B, Lang D, Heidenreich S. Predictive value of IgG subclass levels for infectious complications in renal transplant recipients. Clin Nephrol 1996;45:22–28. Kroemer G, Martinez-AC. Pharmacological inhibition of programmed cell death. Immunol Today 1994;15:235– 42. Sackstein R, Borenstein M. The effects of corticosteroids on lymphocyte recirculation in humans: analysis of the mechanism of impaired lymphocyte migration to lymph node following methylprednisolone administration. J Invest Med 1995;43:68 –77. Kelso A, Munk A. Glucocorticoid inhibition of lymphokine secretion by alloreactive T lymphocyte clones. J Immunol 1984;133:784 –91.