Objectives: The purpose of this study was to investigate the frequency and prognostic importance of acute cellular rejection after heart transplant.

Materials and Methods: All 84 heart transplant patients at our center from January 1993 to January 2014, including all 576 endomyocardial biopsies, were evaluated with retrospective review of clinical records and endomyocardial biopsies. Routine and clinically indicated endomyocardial biopsies after heart transplant were graded for acute cellular rejection (2005 International Society for Heart and Lung Transplantation Working Formulation). Survival analysis was performed using Kaplan-Meier method.

Results: There were 61 male (73%) and 23 female recipients. Median age at heart transplant was 29 years (range, 1-62 y). Posttransplant early mortality rate was 17.9% (15 patients). In the other 69 patients, 23 patients died and 46 patients (66.7%) were alive at mean 69.3 ± 7.2 months after heart transplant. Mean follow-up was 35.4 ± 29.8 months (range, 0.07-117.5 mo). Mean 8.4 ± 4.2 endo­myocardial biopsies (range, 1-19 biopsies) were performed per patient. Median first biopsy time was 7 days (range, 1-78 d). The frequency of posttransplant acute cellular rejection was 63.8% (44 of 69 patients) by histopathology; 86% patients experienced the first episode of acute cellular rejection within 6 months after transplant. There were 18 patients with acute cellular rejection ≥ grade 2R on ≥ 1 endomyocardial biopsy in 44 patients with acute cellular rejection. No significant difference was observed between survival rates of patients with grade 1R or ≥ grade 2R acute cellular rejection, or between survival rates of patients with or without diagnosis of any grade of acute cellular rejection. Acute cellular rejection was not related to any prognostic risk factor.

Conclusions: Acute cellular rejection had no negative effect on heart recipient long-term survival, but it was a frequent complication after heart transplant, especially within the first 6 months.

Key words : Acute cellular rejection, Cardiac, Outcome, Survival

Introduction

Acute cellular rejection (ACR) is an inflammatory response, comprised mainly of lymphocytes, directed against the transplanted organ. Recognition of rejection in the transplanted heart is based on direct histologic examination of allograft tissue samples obtained by endomyocardial biopsy. An international histopathologic grading system for cardiac allograft biopsies was adopted by the International Society for Heart Transplantation in 1990.¹ The revision of the 1990 Working Formulation for the standardization of nomenclature in the diagnosis of heart rejection was reported by the International Society for Heart and Lung Transplantation (ISHLT) Working Group in 2005.² This report summarized the revised consensus classification for cardiac allograft rejection with a multidisciplinary review of the cardiac biopsy grading system. The revised (R) categories of cellular rejection were: Grade 0R, no rejection (no change from 1990); Grade 1R, mild rejection (1990 Grades 1A, 1B, and 2); Grade 2R, moderate rejection (1990 Grade 3A); and Grade 3R, severe rejection (1990 Grades 3B and 4). The ISHLT Working Group recommended that grade 2R should be the threshold for treatment.

The effect of ACR episodes on the survival of heart transplant recipients is controversial. Previous studies supported the notion that recurrent ACR episodes increase the probability of allograft coronary artery disease and possibly decrease long-term survival when identified after the first year, particularly in pediatric heart transplant recipients.^3-6 However, another study group found that ≥ 3 episodes of acute antibody-mediated rejection resulted in a statistically significant increase in cardiovascular mortality. In contrast, ACR episodes did not increase the risk of cardiovascular mortality.⁷ The frequency of acute rejection has declined with new maintenance immunosuppression options.⁸

This study was performed to investigate the frequency of ACR after heart transplant in our center and its prognostic importance in consideration of the current histopathologic grading system for cardiac allograft biopsies.

Materials and Methods

Patients
A retrospective clinicopathologic evaluation was performed of all endomyocardial biopsies of patients who underwent first-time allograft heart transplant at Başkent University Hospital in Ankara from January 1993 to January 2014. In the 21 years, 84 patients had heart transplant at the Department of Cardiovascular Surgery, and 15 patients died of early posttransplant complications.

The other 69 patients were followed by routine or additional clinically indicated endomyocardial biopsies. Routine endomyocardial biopsies were performed weekly for the first 2 weeks, monthly between the second and tenth week, and every 3 months between 10 weeks and 1 year. This study included 576 endomyocardial biopsies in 69 recipients. All endomyocardial biopsies that were diagnosed with any grade of ACR were reviewed and graded according to the 2005 ISHLT Working Formulation. The assessments of endomyocardial biopsies were performed only by light microscopy. All endomyocardial biopsy specimens included 2 or 3 tissue pieces. Light microscopy was performed with 5-μm sections from paraffin-embedded tissue using hematoxylin-eosin and histochemical staining methods including Masson trichrome. All biopsy specimens were evaluated with ≤ 6 microscopic sections per tissue piece.

Clinical data about presentation and follow-up were retrieved from patient and hospital records. Patients were followed by echocardiography and dobutamine stress echocardiography.

Statistical analyses
Continuous variables were presented as mean ± standard deviation, and categorical variables were presented as percentage and compared using chi-square test. Nonparametric tests (Wilcoxon rank sum test) were used for the relation between prognostic parameter and existing ACR. Multiple logistic regression was used to compute the risk of experiencing ACR and the relative odds ratios for 8 main prognostic groups. These groups were identified as demographic or clinical features that possibly had prognostic importance for heart transplant recipients (Table 1). The survival analysis was performed using Kaplan-Meier method and log-rank test. Grade 1R ACR and ACR ≥ grade 2R were labeled as degree 1 and degree 2, respectively, in Kaplan-Meier curves (Figure 1 and 2). Statistical significance was set at P ≤ .05. The analyses were performed using statistical software (SPSS, Version 15.0, SPSS Inc., Chicago, IL, USA).

Results

Demographic and clinical characteristics
There were 61 male (73%) and 23 female (27%) recipients. Median age at time of heart transplant was 29 years (range, 1 to 62 y). There were 2 patients who had combined cardiac and renal transplant. The 2 most frequent reasons for heart transplant were idiopathic dilated cardiomyopathy (39 patients) and ischemic heart disease (16 patients) (Table 2). Biatrial anastomosis was performed for orthotopic heart transplant in all patients. Hemofiltration was used routinely during the operations. In all cases, standard triple immunosuppressive therapy without induction therapy was used; the patients received corticosteroids, cyclosporine, and mycophenolate mofetil. Sirolimus was employed in some cases due to impaired renal function. For patients who had ACR ≥ grade 2R rejection, treatment consisted of pulse intravenous corticosteroids. Cyclosporine was replaced with tacrolimus for the treatment of grade 1R rejection.

Complications after heart transplant in 69 recipients were shown in Table 3. Mean follow-up was 35.4 ± 29.8 months (range, 0.07-117.5 mo). There were 15 of 84 recipients (17.9%) who died of early posttransplant complications. The other 69 patients were followed; 23 patients died and the other 46 patients (66.7%) were alive at mean 69.3 ± 7.2 months after heart transplant. In the posttransplant midterm or late phases, the known causes of death were sudden death (3 patients), sepsis (3 patients), and heart failure (2 patients). Other clinical data of the 69 recipients with follow-up were summarized (Table 4).

Endomyocardial biopsy
During follow-up, mean 8.4 ± 4.2 endomyocardial biopsies (range, 1-19 biopsies) were performed per patient. First biopsy was performed at median 7 days (range, 1 to 78 d). The frequency of posttransplant ACR diagnosed with histopathology was 63.8% (44 of 69 patients). Eighty-six percent of patients experienced their first episode of ACR in the early phase after transplant (first 6 months). There were 18 patients who had ACR ≥ grade 2 on ≥ 1 endomyocardial biopsy. Thus, the frequency of patients who experienced moderate or high grade ACR was 26% of 69 recipients.

Survival
The survival rates in the fifth and tenth years were 58.3% and 54.8% for all 84 recipients. The 5- and 10-year survival rates were 71% and 66.7% for 69 surviving recipients after exclusion of patients who had early posttransplant mortality. No significant difference was observed between survival rates of patients with grade 1R ACR and ACR ≥ grade 2R (Figure 1) or between survival rates of patients who had diagnosis of any grade of ACR and patients who did not have ACR (Figure 2) on their endomyocardial biopsies. Experience of having ACR was not significantly related to any prognostic risk factor (P > .05). No statistical difference was observed by logistic regression among the prognostic groups for risk of existence of ACR (Table 1).

Discussion

In this study, we observed higher frequency of ACR (63.8%) and ACR ≥ grade 2R (26%) than other similar studies in the current literature, especially those reported since 2003.² Some authors suggested that the frequency of histologic rejection has decreased with current immunosuppression.⁸ Several studies showed that acute rejection is less frequent with a tacrolimus, than cyclosporine-based regimen.⁹ Our study included the 10 years before 2003, when new immunosuppressive regimens were not used. Standard posttransplant triple immunosuppressive therapy not including tacrolimus have been used in our center since 2003. Thus, no recipient included in this study initially had a tacrolimus-based regimen; cyclosporine was replaced with tacrolimus in cases of grade 1R rejection diagnosed with histopathology. Moreover, we estimated that noncompliance with intake of immunosuppressive drugs contributed slightly to the high ACR rate in the Turkish population. Another report from our center included 13 heart transplant patients from 2003-2007, and the acute rejection rate was lower (31%) but the rate of ACR ≥ grade 2R was high; all 4 patients with ACR experienced grade 3A (2R) ACR.¹⁰ However, the mean follow-up of this study was only 18.6 months (range, 1 to 38 mo), and the number of cases was very small.

A current similar study from Skane University Hospital in Sweden showed proportionally more first-year ACR ≥ grade 2 in endomyocardial biopsies in heart transplants performed during 1988-1999 (9.6%) than 2000-2010 (5.5%).¹¹ This study showed low frequency and severity of first-year ACR with 6.5% routine endomyocardial biopsies and 14.1% additional clinically indicated endomyocardial biopsies showing ACR ≥ grade 2. They studied only posttransplant first year biopsies and they searched the percentage of ACR ≥ grade 2-positive biopsy specimens in all first year biopsies.

In contrast, we researched the ACR rate on the basis of recipients who had the diagnosis of any grade ACR on ≥ 1 endomyocardial biopsy. We observed that ACR was a frequent complication in heart transplant recipients in our center, especially within 6 months after heart transplant. There were 86% recipients who had ACR, with their first episode of ACR within 6 months after surgery. This can be explained by frequent routine endomyocardial biopsies that may allow early detection of ACR in this period and the use of a standard triple immunosuppressive therapy, cyclosporine-based, without induction therapy in all recipients and with improved immunosuppressive adjustments.

The 5- and 10-year survival rates were 71% and 66.7% for 69 surviving recipients, when post­transplant early mortality was excluded in our study. These results, as an experience of a single center, were compatible with others in the current literature.^8,12

There are many different factors that possibly have an effect on survival of allograft organ recipients. Some demographic and clinical factors were recorded and evaluated for a relation to ACR experience, but no relevant factors were noted statistically. Söderlund and associates observed that proportionally more (P < 0.05) first-year cases of ACR ≥ grade 2 in endomyocardial biopsies were observed in heart transplants with pediatric (11.3%) than adult (7.1%) donors, and in sex-mismatched (10.4%) than sex-matched (6.3%) heart transplants.11 They did not observe a statistically significant relation with first-year ACR ≥ grade 2 and other risk factors such as age of recipient, difference in age between recipient and donor, sex of recipient, sex of donor, ABO-matching between recipient and donor, and maintenance immunosuppression in patients alive at discharge. None of these risk factors had a statistically significant relation to ACR experience in our study (Table 1). They also found that 5- and 10-year survival was lower in heart transplant patients with ≥ 1 compared with 0 first-year ACR ≥ grade 3A/3B. We did not observe a significant difference between survival rates of patients with ACR ≥ grade 2R and patients who had no diagnosis of any grade of ACR (Figure 1 and 2). Chin and associates identified a progressive decrease in survival with more frequent rejection episodes and late rejection (even when asymptomatic) after transplant in pediatric heart transplant recipients.3 They suggested that recurrent rejection is a risk factor for mortality after pediatric heart transplant, particularly when complicated by hemodynamic compromise. Our study included 27 pediatric recipients (aged < 18 y) that comprised 39.1% of all 69 surviving recipients. We studied pediatric and adult recipients together, and we observed that ACR experience had no negative effect on heart recipient survival. However, pediatric recipients may be the topic of a further research study with clinico­pathologic parameters.

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