Cardiac Allograft Failure Research Articles

Lumen loss in transplant coronary artery disease is a biphasic process involving early intimal thickening and late constrictive remodeling: results from a 5-year serial intravascular ultrasound study.

Coronary artery disease is the major cause of late cardiac allograft failure. However, few data exist regarding the natural history of changes in intimal and external elastic membrane (EEM) areas after heart transplantation. In 38 transplant recipients, serial intravascular ultrasound examinations were performed 3.7+/-2.2 weeks after transplantation and annually thereafter for 5 years. In 59 coronary arteries, we compared 135 matched segments among serial studies. In each segment, intravascular ultrasound images were digitized at 1-mm intervals, and mean values of EEM and lumen and intimal areas were analyzed. In the first year after transplantation, the intimal area increased significantly from 1.8+/-1.6 to 3.0+/-2.1 mm(2) (P<0.001). Subsequently, the annual increase in intimal area decreased. EEM area did not change during the first year; however, between years 1 and 3, significant expansion of EEM area occurred (15.4+/-4.6 to 17.2+/-5.4 mm(2), P<0.001). Thereafter, EEM area decreased significantly from 17.2+/-5.4 mm(2) (year 3) to 15.1+/-4.9 mm(2) (year 5, P=0.01). Different mechanisms of lumen loss were observed during 2 phases after transplantation: early lumen loss primarily caused by intimal thickening and late lumen loss caused by EEM area constriction. This serial ultrasound study revealed that most of the intimal thickening occurred during the first year after heart transplantation. Changes in the EEM area showed a biphasic response, consisting of early expansion and late constriction. Thus, different mechanisms of lumen loss were observed during the early and late phases after transplantation.

Nonimmunologic rejection of the heart

Incompatibility between recipient immune cells and donor cells can cause rejection of solid organ transplants. Recently the importance of the microvasculature for transplant survival has been revealed. Evidence indicates that organ transplants fail, not because of cellular rejection, but from loss of donor vessels within the allograft. We asked whether increasing numbers of nonimmunologic changes in the microvasculature such as depletion of vascular antithrombin and arteriolar tissue plasminogen activator and presence of arterial endothelial intercellular adhesion molecule-1 (ICAM-1) and microvascular fibrin, as well as increased levels of soluble ICAM-1 and detectable serum levels of cardiac troponin I, augment the risk of developing coronary artery disease (CAD) and graft failure. We followed up 141 heart transplant recipients 44.9 +/- 2.5 months after transplantation. CAD was evaluated angiographically. Microvascular antithrombin, tissue plasminogen activator, ICAM-1, and fibrin were evaluated immunohistochemically in serial biopsies obtained during the first 3 months after transplantation. Serum soluble ICAM-1 and cardiac troponin I were studied using an enzyme-linked immunosorbent assay. After adjusting for potentially confounding variables, we found that the risk of CAD increases 2.0 times (95% confidence intervals [CI]: 1.6-2.6; p<0.001) and the risk of failure increases 1.9 times (95% CI: 1.3-2.8; p<0.001) with each additional nonimmunologic risk factor. The absence of nonimmunologic risk factors soon after transplantation was associated with a 24-month period free of CAD. Each of these changes, and not cellular rejection, was independently associated with the development of transplant CAD and failure. The greater the number of nonimmunologic risk factors present within the first 3 months after transplantation, the greater the association with increasing development of CAD and failure. We propose that a prothrombogenic microvasculature during the peritransplant period, and not cellular rejection, may cause chronic rejection of the cardiac allograft and subsequent failure.

Expression of NF-kappaB and cytokines in chronic rejection of transplanted murine heart.

The heart transplantation-associated accelerated graft arteriosclerosis (AGAS) is one of the major causes of cardiac allograft failure. We investigated the early time-course of expresssion patterns of cytokines, transcription factor, and its inhibitor in the intraabdominally transplanted mice hearts that differed only in the D locus of class I histocompatibility antigen. The allograft hearts were harvested at 1-3, 5, 7, 14, 28, and 42 days after the transplantation, and the expressions of NF-kappaB/I-kappaB and cytokines (TNF-alpha, INF-gamma) were examined in these specimens. The expressions of TNF-alpha and INF-gamma were observed on day 1, peaking on day 5 and 7, respectively. Activated NF-kappaB (p65) expression was present on the cytoplasm and perinuclear area in the endothelial cells of coronary arteries on day 1. The peak of translocation of NF-B from cytoplasm to nucleus appeared on day 5 in the endothelial cells, myocytes, and leukocytes within the vessels, and remained elevated until day 42. The I-kappaB expression gradually increased from day 1 until day 5, but a remarkable decrease was detected on day 7. Our data suggest that the increased expressions of NF-kappaB/I-kappaB and cytokines (TNF-alpha, INF-gamma) play an important role in inducing immune responses in the donor allograft heart and hence the blockage of the expressions might be mandatory to avoid a potential graft failure.

Simvastatin attenuates vascular hypercoagulability in cardiac transplant recipients.

3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have been shown to reduce cardiac allograft failure and to lower the incidence of transplant coronary artery disease. These effects result from as yet unknown mechanisms not clearly attributable to lipid lowering. We here report that low-dose simvastatin treatment inhibits excessive expression of monocyte tissue factor (TF) and reduces the persistent hypercoagulability state seen in cardiac transplant recipients. Fifteen consecutive heart transplant recipients receiving standard oral immunosuppression were newly assigned to a 10 mg daily simvastatin therapy. Levels of TF activity in both unstimulated and lipopolysaccharide-stimulated peripheral blood mononuclear cells drawn from transplant recipients before and under simvastatin therapy were evaluated by one-stage clotting assay. Monocyte TF activity was found to be significantly increased in cardiac transplant recipients when compared with healthy controls. Excessive monocyte procoagulant activity was reduced in cardiac transplant recipients during simvastatin treatment. This effect occurred independently of the reduction of serum low-density lipoprotein cholesterol. As demonstrated by reverse transcriptase-polymerase chain reaction, monocyte TF reduction by simvastatin, observed in 13 of the 15 transplant recipients investigated, could be ascribed to an inhibition of monocyte TF gene transcription. The reduction of monocyte TF activity during treatment with simvastatin paralleled with the normalization of elevated levels of thrombin-antithrombin complex, prothrombin fragment F1+2, and D-dimer, which are markers of thrombin and fibrin formation indicating coagulation activation after cardiac transplantation. Inhibition of monocyte TF expression and attenuation of the persistent hypercoagulable state observed in cardiac transplant recipients during treatment with simvastatin may represent an important mechanism by which HMG-CoA reductase inhibitors protect against the development of transplant coronary artery disease.

Clinico-pathologic, Immunohistochemical, and TUNEL Study in Early Cardiac Allograft Failure

Early cardiac allograft failure (ECAF) was defined as acute allograft failure in the early transplant period. The aim of this study is to elucidate the clinicopathological and immunohistochemical characteristics and the role of apoptosis in ECAF in nine patients. We reviewed preoperative clinical data and morphological data at the time of autopsy or retransplantation. We also performed TUNEL assay and immunohistochemistry to study fibronectin and tubulin β-II. The average recipient and donor age was 48 ± 10.3 and 28 ± 7.11 respectively. Seven patients died at a mean time of 26 hours. The remaining two patients underwent retransplantation and are alive. The mean cold ischemic time was 124.1 ± 44.5 minutes. No patient had a panel reactive antibody >15% and lymphocytic crossmatch was positive in one case. All cases had grade 2-3 of coagulative necrosis, which correlated positively with fibonectin accumulation in myocyte cytoplasm, and cytoplasmic tubulin loss (p < 0.05). TUNEL technique showed in all cases some degree of DNA strand breaks in cardiomyocytes. Endothelium DNA strand breaks were seen in seven cases. Patients transplanted because of idiopathic dilated cardiomyopathy had a significantly higher degree of DNA strand breaks in cardiomyocytes and endothelial cells (p = 0.03 and p = 0.02) than those transplanted because of ischemic cardiomyopathy. These results indicate that ECAF may be caused by ischemic-reperfusion damage to the donor heart assessed by myocyte coagulative necrosis, fibronectin accumulation in myocytes, tubulin loss, and DNA strand breaks of cardiomyocytes and endothelium. The use of a combination of these techniques might be appropriate in the diagnosis of ECAF in endomyocardial biopsies when it is suspected clinically.

Subunit expression of the cardiac L-type calcium channel is differentially regulated in diastolic heart failure of the cardiac allograft.

Left ventricular diastolic dysfunction is a major cause of cardiac allograft failure. Multimeric L-type calcium channels (alpha1-, alpha2/delta-, and beta-subunits) are essential for excitation/contraction coupling in the heart. Their gene expression was studied in allografts that developed diastolic heart failure. mRNA levels of calcium channel subunits were measured by competitive reverse transcriptase-polymerase chain reaction in microbiopsy samples from the interventricular septum. Size and tissue variabilities between biopsy samples were assessed by determination of cardiac calsequestrin mRNA levels. In the cardiac allografts studied, mRNA levels in microbiopsy samples were considered to represent left ventricular gene expression, because septal and left ventricular gene expression in Northern blots was equivalent, and left ventricles contracted homogeneously. Biopsy samples (n=72) were taken from allografts with normal left ventricular end-diastolic pressure (LVEDP; 8 to 13 mm Hg; n=30), moderately elevated LVEDP (14 to 18 mm Hg; n=26), and elevated LVEDP (19 to 28 mm Hg; n=16). Increased LVEDP was related to slowed diastolic relaxation determined by the time constant tau (r2=0.86), whereas systolic performance (dP/dt; ejection fraction) was preserved. With increasing LVEDP, mRNA levels of the pore-forming alpha1c-subunit (n=15) and of the regulatory alpha2/delta-subunit (n=17) remained unchanged but decreased exponentially (r2=-0.83) for the regulatory beta-subunit (n=40). Compared with cardiac allografts with normal LVEDP (n=15), beta-subunit mRNA level was reduced by 75% at elevated LVEDP (n=9; P=0.012). In an explanted, diastolically failing cardiac allograft, beta-subunit expression was reduced correspondingly by 72% and 76% on the mRNA level in septal and left ventricular myocardium and by 80% on the protein level. The downregulated expression of the calcium channel beta-subunit might contribute to altered calcium handling in diastolically failing cardiac allografts.

Contribution of IFN-γ to acute allograft failure in murine cardiac transplants: Role in vascular thrombosis

390 We investigated the role of IFN-γ in acute cardiac allograft failure using IFN-γ deficient (GKO) mice. Without immunosuppression, BALB/c[H-2d] into wild type C57BL/6 [B6, H-2b] total allogeneic mismatched cardiac grafts ceased beating at 7.2 ± 0.2 days (n=9); BALB/c hearts transplanted into B6GKO ceased beating at 6.6 ± 0.1 days(n=6). Although grafts in both wild type and GKO recipients showed comparably severe mononuclear cell infiltration (rejection), allografts in GKO recipients also showed profound acute coagulation necrosis with vascular thrombosis demonstrable by fibrin staining. By immunohistochemical staining, plasminogen activator inhibitor-1 (PAI-1) was strikingly increased in allografts transplanted into B6GKO hosts relative to B6 wild types; no difference was seen in tissue plasminogen activator staining for allografts in wild type vs GKO hosts. Without immunosuppression, MHC I-mismatched allografts, characterized by a predominant CD8 T cell response (bml into B6), functioned for at least 12 weeks (n=10) and showed only moderate acute rejection. In contrast, bml into B6GKO allografts ceased beating at 7.7 ± 0.7 days(n=7), and showed rejection with coagulative necrosis and vascular thrombosis. Without immunosuppresiosion, MHC II-mismatched allografts, characterized by a predominant CD4 T cell response (bml2 into B6), ceased beating at 31.5± 6.2 days (n=6), and showed severe rejection. In contrast, bml 2 into B6GKO allografts functioned for at least 12 weeks and showed only moderate rejection without coagulative necrosis or vascular thrombosis. To specifically evaluate the role of T cell subpopulations, CD4 and/or CD8 T cells were selectively depleted in recipients by appropriate mAb administration pre-transplant, as shown below. Graft survival times were: BALB/c into B6(anti-CD4): 38.3 ± 6.7 days (n=3), BALB/c into B6GKO (anti-CD4): 10.7± 5.0 days (n=3), BALB/c into B6 (anti-CD8): 12.0 ± 2.3 days(n=5), BALB/c into B6GKO (anti-CD8): 14.3 ± 1.5 days (n=3), BALB/c into B6 (anti-CD4 and CD8): 31.7 ± 3.2 days (n=9), BALB/c into B6GKO(anti-CD4 and CD8): 38.2 ± 7.3 days (n=5). All grafts in the above demonstrated comparably severe rejection; notably, coagulative necrosis and thrombosis were not seen. The results suggest that acute cardiac allograft failure, particularly in GKO, may include a component of vascular thrombosis due to increased PAI-1 activity. Increased thrombosis is particularly evident in the setting of MHC I mismatches, but cannot be wholly attributed to an effect of CD8T cells.

Blockade of very late antigen-4 integrin binding to fibronectin in allograft recipients. II. Treatment with connecting segment-1 peptides prevents chronic rejection by attenuating arteriosclerotic development and suppressing intragraft T cell and macrophage activation.

Chronic rejection remains the leading obstacle to long-term allograft survival. We have shown that treatment of sensitized rats with rapamycin (RPM) does not prevent progressive chronic-type cardiac allograft failure. Having documented the role of fibronectin (FN) in the allograft rejection cascade, we hypothesized that treatment with synthetic peptides that specifically block adhesive interactions between the connecting segment-1 (CS1)-binding domain of FN and alpha4beta1 integrin on circulating cells may prevent the development of chronic rejection in transplant recipients. Lewis rats were sensitized with Brown Norway skin grafts (day -7), followed by transplantation of LBNF1 hearts (day 0). Experimental animals were treated with RPM (day -7 to -1; 0.25 mg/kg/day i.p.), or RPM + CS1 peptides (day +7 to +13; 4 mg/kg/day i.v.), and euthanized at day 60. Unlike cardiac allografts in rats undergoing RPM monotherapy, those after adjunctive CS1 peptides had well preserved myocardial architecture and were free of arteriosclerotic lesions. Moreover, reverse transcription-polymerase chain reaction-based intragraft expression of transcripts for CD3, interferon-gamma, interleukin-12, monocyte chemoattractant protein-1, and transforming growth factor-beta were diminished in the CS1 group when compared with levels in the RPM group. The corresponding expression of cytokine proteins, as determined by immunoperoxidase labeling, was also depressed and correlated with decreased infiltration by T cells and macrophages. CS1 peptide-facilitated blockage of alpha4beta1-FN interactions prevents the development of chronic rejection and depresses the expression of key T cell- and macrophage-associated cytokines/chemoattractants. Hence, local synthesis of FN is an ongoing feature of, and adhesive FN-alpha4beta1 associations are critical for, the development of chronic transplant rejection.

Immediate Cardiac Allograft Failure

Optimal support for immediate cardiac allograft failure is unknown. With the introduction of heparin bonded extracorporeal membrane oxygenated circuits, prolonged cardiopulmonary support is possible. The authors report a case that involved 2 days of right atrial to ascending aorta extracorporeal membrane oxygenated support after immediate donor organ failure prevented the patient from exiting bypass. Continued deterioration in cardiac function led to an attempt at conversion to a total artificial heart as a bridge to retransplant. However, this procedure resulted in transbronchial exsanguination and recipient death. The autopsy showed pulmonary thrombosis with infarction and hemorrhage. The authors recommend caution in the use of extracorporeal membrane oxygenated support for patients with immediate and profound graft failure because of the increased risk of stasis thrombosis and pulmonary infarction due to sluggish pulmonary and left atrial blood flow. Instead, institution of total artificial heart or biventricular support may be preferable as the risks of thrombus and infarction are less.

Primary Graft Failure after Heart Transplantation. Successful Recovery with Pneumatic Biventricular Assistance

A 41-year-old male underwent orthotopic heart transplantation complicated by intraoperative acute allograft failure. The patient required immediate placement of a pneumatic biventricular assist device which was kept for 49 days until graft recovery resulted in successful explantation of the device. The patient was discharged from hospital on postoperative day 112. Management of primary cardiac allograft failure with mechanical ventricular assistance is discussed.

Clinical experience with BioMedicus centrifugal ventricular support in 172 patients.

One hundred and seventy-two patients at The Methodist Hospital in Houston, Texas, were placed on BioMedicus centrifugal ventricular support. One hundred thirty-nine patients were male and 33 were female with a mean age of 59.7 years. Reasons for support were postcardiotomy cardiac failure (129 patients), cardiac allograft failure (17 patients), bridge to transplantation (10 patients), resuscitation (7 patients), postpercutaneous transluminal coronary angioplasty emergent (2 patients), and other (7 patients). Support was by left ventricular assist device in 108 patients, right ventricular assist device in 20 patients, and biventricular assist device in 44 patients. Eighty-four patients (48.8%) were weaned from the ventricular assist device, and 88 patients (51.2%) were not weaned. Thirty-four patients (20.0%) were discharged from the hospital. Complications included coagulopathy, renal insufficiency/failure, respiratory insufficiency/failure, neurological deficits, sepsis, arrhythmias, and device-related complications. Overall causes of death were ventricular failure (55.1%), triage (13.0%), arrhythmias (9.4%), graft failure (5.9%), coagulopathy (4.3%), sepsis syndrome (2.9%), device-related (0.7%), and other (0.7%). BioMedicus centrifugal ventricular support can be implemented rapidly and easily. Device-related complications are few (1.2%), and it is relatively inexpensive when compared with other ventricular assist systems. This series demonstrates that a substantial number of patients may benefit from temporary centrifugal ventricular support.

Blockade of very late antigen-4 integrin binding to fibronectin with connecting segment-1 peptide reduces accelerated coronary arteriopathy in rabbit cardiac allografts.

Graft arteriopathy, a leading cause of cardiac allograft failure, is associated with increased intimal smooth muscle cells, inflammatory cells, and accumulation of extracellular matrix. We hypothesized that cellular fibronectin plays a pivotal role in the progression of the allograft arteriopathy by directing the transendothelial trafficking of inflammatory cells through interaction of the connecting segment-1 (CS1) motif with the very late antigen-4 (VLA-4) integrin, and tested this in vivo using a blocking peptide. Cholesterol-fed rabbits underwent heterotopic cardiac transplantation without immunosuppression. The treatment group (n = 7) received a synthetic CS1 peptide (1 mg/kg per d, subcutaneously), and the controls (n = 7) received an inactive peptide (1 mg/kg per d, subcutaneously). At 7-8 d after transplantation, hearts were harvested and sectioned for morphometric analysis and immunohistochemical studies. We observed a > 50% decrease in the incidence (P < 0.001) and severity (P < 0.001) of donor coronary artery intimal thickening in the CS1-treated compared with the control group. These findings correlated with reduced infiltration of T cells (P < 0.05), a trend toward decreased expression of adhesion molecules (P < 0.06), and less accumulation of fibronectin (P < 0.03). Our data suggest that the VLA-4-fibronectin interaction is critical to the progression of the allograft arteriopathy by perpetuating the immune-inflammatory response in the vessel wall.

Implantation techniques for the Hemopump

Techniques for implantation of the Hemopump, an intraarterial, axial-flow circulatory assist device, are described. The Hemopump, which is currently undergoing clinical investigation, has been used successfully to treat patients experiencing profound left ventricular failure in a variety of clinical situations, including postcardiotomy shock, acute myocardial infarction, cardiac allograft rejection, and cardiac allograft failure.