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J Heart Lung Transplant. Author manuscript; available in PMC 2008 May 1. Published in final edited form as: doi: 10.1016/j.healun.2007.02.003. | PMCID: PMC1950728 NIHMSID: NIHMS22539 |
ALTERATIONS IN THE FIBRINOLYTIC CASCADE POST TRANSPLANT: EVIDENCE OF A BIMODAL EXPRESSION PATTERN 1Raymond L. Benza,2,9 Matthew A. Cavender,3 Joseph Barchue,4 Jose A. Tallaj,5 Robert C. Bourge,6 James K. Kirklin,7 and Christopher S. Coffey8 2School of Medicine, Department of Medicine, Division of Cardiovascular Disease, University of Alabama at Birmingham (UAB), Birmingham, Alabama 35294 3School of Medicine, Department of Medicine, Division of Cardiovascular Disease, University of Alabama at Birmingham (UAB), Birmingham, Alabama 35294 4School of Medicine, Department of Medicine, Division of Cardiovascular Disease, University of Alabama at Birmingham (UAB), Birmingham, Alabama 35294 5School of Medicine, Department of Medicine, Division of Cardiovascular Disease, University of Alabama at Birmingham (UAB), Birmingham, Alabama 35294 6School of Medicine, Department of Medicine, Division of Cardiovascular Disease, University of Alabama at Birmingham (UAB), Birmingham, Alabama 35294 7Department of Surgery, Division of Cardiothoracic Surgery, University of Alabama at Birmingham (UAB), Birmingham, Alabama 35294 8School of Public Health, Department of Biostatistics University of Alabama at Birmingham (UAB) Birmingham, Alabama 35294 Changes in the fibrinolytic system that occur after cardiac transplantation (CTx) and the factors which influence such changes are poorly described, yet may be ultimately important in determining the varying morphologic features of transplant related coronary artery disease (Tx CAD). Baselines demographic, as well as serial clinical information and plasma fibrinolytic levels were prospectively recorded preCTx and multiple times postCTx in 110 de novo cardiac transplant recipients. We noted a biphasic change in fibrinolytic activity over the first year of CTx with an early immediate decline in PAI-1 activity (p > 0.001) matched with stable PAP (plasmin) activity corresponding to an “enhanced” fibrinolytic state early post CTx followed by a significant increase at 6 months (p = 0.004) and 1 year (p < 0.001) in PAI-1 activity concomitant with a significant decline in PAP after 3 months (p = 0.005 at 3 months, p < 0.001 at 6 months, and p < 0.001 at 1 year) corresponding to an “impaired” fibrinolytic state late post CTx. This biphasic nature of the fibrinolytic system could account for the varying morphologic features of Tx CAD. Keywords: Cardiac Transplantation, Chronic Rejection, Fibrinolysis, Clinical Transplantation, Allograft Coronary Disease The development of an accelerated form of coronary artery disease (Tx CAD) significantly limits the long-term success of cardiac transplantation ( 1, 2). Tx CAD is common and detected angiographically in 44-79% of recipients by 5 years ( 1, 3- 7) and nearly universal by IVUS by one year ( 8). Despite the advancements in clinical immunosuppression, the incidence and prevalence of Tx CAD remains constant ( 3- 7, 9). Limitations in donor availability and the diminished survival associated with retransplantation ( 10, 11)underscore the need for further insight and research. The development of Tx CAD particularly intimal proliferation is due to a complicated interplay between immune and nonimmune factors. Among the nonimmune factors, alterations in fibrinolytic activity (FA) appear to be important ( 12- 16). Impaired FA usually results from diminished levels of the plasminogen activators (PAs), t-PA and urokinase (u-PA) and/or the presence of excess PAI-1, the inhibitor of these PAs. This results in decreased plasmin production and fibrin deposition, which is a common feature of grafts with angiographically detectable Tx CAD (atheromatous form). The effects of enhanced fibrinolysis or increased plasmin activity on Tx CAD are less known, but may be important in the development of the early intimal response. The purposes of this observational study are to characterize the changes in components of the fibrinolytic system in transplant recipients over time and determine whether these changes could feasibly influence the sequential changes in morphology seen typically in Tx CAD. Patients Between 06/01/1997 and 12/01/2001, 110 denovo cardiac transplants were prospectively enrolled. Informed consent was obtained from all patients. This study was approved by the Institutional Review Board at UAB. Serial plasma t-PA, PAI-1, u-PA, fibrinogen levels, PAI-1 activity, and plasmin/alpha-2-anti-plasmin (PAP) assays (plasmin activity) were recorded preCTx and postCTx (1 week; 1, 3, 6 , and 12 months). Donor and Recipient Demographics Data extracted at baseline (preCTx) included specific information on donor/recipient age, gender, race, recipient BMI, smoking status, blood pressure (mmHg), and presence of diabetes mellitus. In addition to serial plasma fibrinolytic levels, we collected serial recipient BMIs, blood pressures, creatinine levels, immunosuppressants (dosing/levels), CMV reactivity, rejection episodes, lipoproteins, as well as lipid-lowering and anti-hypertensive drug use. Statistical Analysis The percentage change in fibrinolytic protein levels and activity from baseline was computed for each time point. Due to the skewness of the distributions, we applied a logarithmic transformation to all values. We utilized a t-test to test for a significant change from baseline at each time point, using a Bonferroni correction to adjust for multiple comparisons. For those serial levels that were of most interest (PAI-1 activity and PAP), we also fit a repeated measures mixed model to examine change from baseline during the first year postCTx. These mixed models examined linear, quadratic, and cubic trends over time, adjusting for baseline values. In order to reduce the problem of multi-collinearity often present in polynomial models, we subtracted the integer value closest to the mean values of time for each individual value. The models also utilized a random intercept for each patient to account for the fact that measurements observed over time within a patient were correlated. The models with change in PAP as the outcome also included a random slope for each patient to account for the fact that the variation in measurements tended to increase over time. We also considered a set of predictors of PAI-1 activity and PAP changes in the first year postCTx consisting of clinical variables (BMI, blood pressure, lipid fractions, creatinine, glucose, diabetes); use of tacrolimus, atorvastatin, pravastatin, ACE inhibitors, cyclosporine, mycophenolate mofetil, OKT3, prednisone; cumulative dosage for ACE inhibitors, cyclosporine, mycophenolate mofetil, prednisone; and the presence of rejection or CMV infection. Due to the fact that this was an exploratory analysis with a small sample size, we made no adjustment for multiple comparisons and used a less stringent significance level of 10% for determining statistical significance. All variables reaching this level of significance in the univariate analysis were entered into a multivariable model using backwards elimination procedures to determine whether the effect remained after adjustment. PAI-1, t-PA, u-PA, and Fibrinogen Levels Fibrinogen and t-PA levels were increased slightly after transplant, but the increases were not significant, except for the 3 month value of t-PA (p < 0.001). u-PA levels decreased significantly within the first month postCTx (p = 0.001) but returned to baseline levels within three months. PAI-1 antigen levels decreased significantly postCTx (p < 0.001 one week post) and remained below baseline values for the first year. PAI-1 Activity and PAP Complexes () There was an initial significant reduction in PAI-1 activity immediately postCTx (p < 0.001 one week post). However, as opposed to PAI-1 antigen levels, PAI-1 activity then began to increase for the remainder of the first year postCTx. As a consequence, there was a significant increase in PAI-1 activity at 6 months (p = 0.004) and 1 year (p < 0.001) postCTx. PAP levels, a marker of plasmin activity, were not significantly different from baseline for the first 3 months postCTx. However, from 3 months on, there was a significant reduction in PAP levels (p = 0.005 at 3 months, p < 0.001 at 6 months, and p < 0.001 at 1 year). Determinants of PAI-1 Activity and PAP Levels Post Transplant ( Tables 1 and 2) By univariate analyses, > 1 rejection episode, a lower ACE inhibitor dose, and a higher cyclosporine (CsA) dose were significantly associated with greater increases in PAI-1 activity over time. In the multivariable model, however, only rejection episodes, and a higher dose of CsA remained independent predictors for increased PAI-1 activity over time ( Table 1). A similar analysis was performed to assess predictors of serial PAP levels. By univariate analyses, a greater L(p)a change from baseline, not taking tacrolimus or mycophenolate mofetil were significantly associated with greater increases over time in PAP levels. In the multivariable model not taking mycophenolate remained an independent predictor ( Table 2). | Table 1Predictors associated with increased change in percentage of baseline PAI-1 activity over time |
| Table 2Predictors associated with increased change in percentage of baseline PAP levels over time |
Tx CAD is characterized by a unique, overlapping biphasic pattern of vascular remodeling starting with a diffuse intimal proliferation early followed by a more classical and coexistent atheromatous component ( 2, 17- 20). The fibrinolytic system is likely important in both developing patterns. The question we sought to answer was whether changes in FA could provide a plausible mechanism to explain the sequential morphologic changes seen in progressive Tx CAD. This study shows that two phases of FA exists post CTx that may explain these morphologic patterns and that these phases may be influenced by clinical factors unique to the transplant environment. The first indication of Tx CAD is the formation of the neointima, which is characterized by intense cell movement and proliferation and requires enhanced, not impaired FA. This is the first key piece of information needed to understand and appreciate how the unique biphasic nature of the fibrinolytic system post transplant could lead to the varying morphologic features of Tx CAD. As noted in this study, PAI-1 activity decreases immediately postCTx with a relative preservation of plasmin activity, which suggests that a plasmin-dominated, profibrinolytic state is present in the short period immediately post transplantation. Plasmin is pivotal to the development of the neointima via the proliferation and migration of SMCs ( 16, 21- 23). Plasmin’s nonproteolytic properties also promote the neointima by acting as a chemo attractant for macrophages and lymphocytes ( 24). It appears that the net result of these cellular mechanisms is an adaptive intimal remodeling process that correlates with the intimal proliferation seen during the first months in allograft vessels ( 8). The atheromatous phase of Tx CAD occurs later postCTx and takes advantage of and builds upon the already expanding neointima ( 17- 19, 25). This phase, however, is propagated by impaired, not enhanced FA marked by heightened PAI-1 activity. This prothrombotic state change during the later part of year 1 correlates well with the microvascular fibrin deposition characteristic of mature Tx CAD ( 14, 15), as well as the increase in angiographically evident Tx CAD by 3 years ( 20). Thus, two distinct fibrinolytic states exist postCTx and these sequential changes in FA could theoretically influence the morphologic shift in Tx CAD. Our analysis also found several clinical factors and medications that may be important in predicting overall fibrinolytic and thrombotic activity. Given that some of these same factors also are known to either promote or mitigate both neointimal proliferation and atheromatous CAD, it is possible that these clinical factors could also influence the formation and/or progression of Tx CAD via mechanisms involving shifts in FA ( 26- 34). This study is limited by the small, single center nature of the cohort and that some time points contain missing values, which could skew the significance of the time course. In addition the independent, albeit transient, effect of cardiopulmonary bypass on the early profibrinolytic state via its effect on t-PA, PAI-1 and plasmin must be considered as a contributor to this process ( 35, 36). In the future, we hope to expand this study to multiple transplant centers in an attempt to solidify the relationship between clinical variables and FA as well as to use changes in FA to predict the development of Tx CAD. | | UAB | University of Alabama at Birmingham | | | CTx | Cardiac transplantation | | | Tx CAD | Transplant-related coronary artery disease | | | IVUS | Intracoronary ultrasound | | | MIT | Maximal intimal thickness | | | FA | Fibrinolytic activity | | | SMCs | Smooth muscle cells | | | t-PA | Tissue plasminogen activator | | | scu-PA | Single chain urokinase type plasminogen activator | | | PAI-1 | Plasminogen activator inhibitor-1 | | | PAs | Plasminogen activators | | | u-PA | Urokinase plasminogen activator | | | PAP | Plasmin/alpha-2-anti-plasmin assays |
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