Robert L. Yowell

Allograft coronary artery disease: clinical correlations with circulating anti-HLA antibodies and the immunohistopathologic pattern of vascular rejection ☆

Despite advances in immunosuppression over the past ten years, the overall longevity of the cardiac allograft has only minimally increased. The average allograft half-life has increased only 6 months (8.8 years to 9.4 years) between the time periods of 1986–1990 and 1991–1997.1 The single major cause of allograft loss after one year remains allograft coronary artery disease (ACAD).1 This entire issue of the Journal of Heart and Lung Transplantation is devoted to the potential etiologic factors as well as the clinical manifestations of ACAD. Traditionally, the immune response to allotransplantation has been divided into the cellular (T-cell) and humoral (B-cell) processes. While it is likely that both processes are activated and ongoing simultaneously in most allograft recipients, patients who appear to have a predominately humoral process have been reported to have worse long-term outcomes.2–10 Patients developing circulating anti-HLA antibodies or the immunohistopathologic pattern of vascular rejection early after cardiac transplantation have demonstrated poorer survival and increased risk for the development of ACAD.4–10

Immunohistochemical analysis of platelet-derived growth factor and basic fibroblast growth factor in cardiac biopsy and autopsy specimens of heart transplant patients☆

The purposes of this study were to examine 250 heart biopsy specimens and 20 autopsy specimens from heart transplant patients for the presence and localization of platelet-derived growth factor (PDGF)and basic fibroblast growth factor (bFGF) and to correlate these findings with the histologic features of rejection and the autopsy findings of graft coronary vasculopathy and global ischemia. Positive specimen staining was significantly more prevalent for PDGF (78% of specimens) than for bFGF (54% of specimens) (p< 0.001). PDGF was distributed more in an interstitial (53%) than a vascular (28%) pattern and was associated with macrophages, whereas bFGF was distributed more in a vascular (50%) than an interstitial (12%) pattern. The prevalence of PDGF (but not bFGF) staining was significantly greater in biopsy specimens with at least grade 2 vascular rejection changes (81%) than in those without vascular rejection changes (58%) (p<0.001). In autopsy specimens, PDGF staining was present in the hearts of all 5 patients (100%) who died of graft failure from coronary vasculopathy and was present in all 11 hearts (100%) with global ischemic changes, but in only 4 of 9 (44%) of the hearts without global ischemia (p<0.01). PDGF staining was absent in nontransplanted heart specimens, whereas bFGF staining in nontransplanted heart specimen was similar to that in transplanted hearts. We conclude that PDGF is increased in transplanted hearts, is distributed more in an interstitial pattern, and is associated with macrophages. Furthermore, PDGF staining is increased in transplanted hearts with evidence of vascular rejection, coronary vasculopathy, or global ischemia.

Vascular rejection in cardiac transplantation: Histologic, immunopathologic, and ultrastructural features

Although the majority of rejection found in cardiac transplant biopsies is cellular in type, a variety of vascular alterations occur in cardiac biopsies, constituting different forms of rejection that can be recognized using light microscopic and immunopathologic criteria. In this report, pathologic aspects of the vascular alterations associated with vascular and mixed rejection of cardiac allografts are described in detail. Methods and controls used in this report are identical to those previously reported. The histologic, immunopathologic, and ultrastructural findings associated with vascular rejection and other vascular processes in cardiac allografts are discussed. The relationship of these findings to chronic allograft rejection and potential pathogenetic mechanisms of these vascular changes are also detailed.

The repetitive histologic pattern of vascular cardiac allograft rejection. Increased incidence associated with longer exposure to prophylactic murine monoclonal anti-CD3 antibody (OKT3).

While vascular cardiac allograft rejection increases morbidity and mortality following transplantation, factors predisposing to its development have not been completely elucidated. To evaluate the influence of the duration of early rejection prophylaxis with the murine monoclonal anti-CD3 antibody (OKT3) on the development of a repetitive histologic pattern of vascular cardiac allograft rejection, endomyocardial biopsies from 344 heart transplant recipients were prospectively evaluated. The influence of clinical characteristics was assessed. Eighty-three patients (24%) developed and 261 patients (76%) did not develop a repetitive histologic pattern of vascular cardiac allograft rejection. The vascular rejection pattern was more common in patients with a positive crossmatch (89% versus 11%, P<0.0001) and OKT3 sensitization (73% versus 27%, P<0.0001), and was positively correlated with the duration of OKT3 treatment (P<0.0001). The correlation persists even after excluding patients with a positive crossmatch or OKT3 sensitization. Patients developing a repetitive histologic pattern of vascular cardiac allograft rejection early after transplantation had decreased allograft survival (P=0.0008). The development of a repetitive histologic pattern of vascular cardiac allograft rejection is positively correlated with the duration of OKT3 treatment. Judicious use of OKT3 in early rejection prophylaxis in cardiac transplantation is warranted.

The Role of Electron Microscopy in Evaluating Ciliary Dysfunction: Report of a Workshop

This report summarizes the proceedings of a workshop organized with the purpose of bringing together many of those with substantial experience in this troublesome area of pathology for an active interchange of ideas, opinions, problems, and solutions. Recognition was given the fact that current knowledge and technical capabilities are woefully inadequate for dealing with the diagnostic questions now being asked. Until such time as these inadequacies can be remedied, a very conservative approach to the interpretation of ultrastructural studies is advocated.

Efficacy of OKT3 retreatment for refractory cardiac allograft rejection.

Although OKT3 monoclonal antibody is a useful therapy for refractory cardiac allograft rejection, the use of OKT3 for prophylaxis may be limited by the potential of sensitization and subsequent loss of efficacy on retreatment. OKT3 was required for refractory rejection in 21 of 165 recipients transplanted between March 1985 and August 1988. Twelve of these patients had previously been exposed to OKT3, and the retreatment efficacy was evaluated. The study population averaged 42.1 +/- 15.3 years of age (mean +/- SEM) and had experienced 2 +/- 1 previous episodes of rejection. The prior episodes of rejection had been treated with pulse methylprednisolone and antithymocyte globulin, and in addition 3 patients (25%) also required a course of antilymphoblast globulin. Retreatment OKT3 for refractory rejection was required 120 +/- 94 days following transplantation. CD3+ lymphocytes were eliminated from the circulation within 24-48 hr in 11 of 12 patients, all of whom showed histologic improvement within the first week. Total resolution on the initial follow-up biopsy was noted in 9 (75%) during the course of therapy. Subsequent rejection episodes occurred in 9 (82%) of the survivors at 71 +/- 64 days. One-year survival was 83% in this vigorously rejecting patient population. Serious infections occurred within 3 months of therapy in 4 (36%). The side effects of OKT3 retreatment were similar to those seen with first exposure and did not require OKT3 discontinuation. Thus OKT3 may be administered with success in most patients who have previously been exposed to it.

Prevention of adverse clinical outcome by monitoring of cardiac transplant patients for murine monoclonal CD3 antibody (OKT3) sensitization

We have previously reported that patients sensitized to murine monoclonal CD3 antibody (OKT3) and maintained on such therapy for induction of immunosuppression have a high mortality and/or allograft loss. In this follow-up study, we retrospectively reviewed all patients routinely and serially monitored by flow cytometry for plasma levels of OKT3 during a 21-month period beginning 1/90. A total of 112 patients were monitored during this period. We retrospectively tabulated the incidence of OKT3 sensitization, rejection pattern and impact on survival of withdrawal of OKT3 at the time of sensitization as compared with the previous study in which withdrawal was not done. Nine patients were excluded from analysis because of withdrawal for reasons other than sensitization: cytokine encephalopathy, infection, postoperative complications, or severe rejection. Twelve patients had OKT3 therapy aborted because of failure to achieve steady-state OKT3 levels or because of decline in levels while on therapy. These patients were thus defined as being sensitized to OKT3. No patient was aborted because of return of CD3 cells in the blood. Only one of the 12 patients sensitized to OKT3 died. Of 91 patients with steady-state OKT3 levels, 6 had high plasma levels (>1000 ng/ml) and 6 had low plasma levels (<500 ng/ml). None of these patients had OKT3 therapy aborted and all are alive. Twelve of these 91 patients had successful retreatment with OKT3 for refractory rejection, indicating that absence of sensitization on induction predicts safety of retreatment with OKT3. We also examined the frequency of associated human antimouse antibody (HAMA) production using the blocking assay modified from Jaffers and Mayes. Only the sensitized patients exhibited a significant association with HAMA production (6/7 tested, P=0.05) Classification of the rejection pattern of the sensitized patients confirmed our previous results: eight of 12 had vascular rejection and 4/12 had mixed rejection. These patterns were prospectively determined. We conclude that serial monitoring of patients for plasma levels of OKT3 is an effective strategy to prevent adverse outcomes of induction with this agent.

Dehydroepiandrosterone protects muscle flap microcirculatory hemodynamics from ischemia/reperfusion injury : An experimental in vivo study

This study evaluated the potential for dehydroepiandrosterone (DHEA) to protect skeletal muscle from reperfusion injury using intravital microscopic observations of isolated rat cremaster muscle flaps. The flaps were subjected to warm ischemia followed by reperfusion in three groups of rats. In group 1 (control, n = 14), muscle flaps were subjected to 6 hours of ischemia and then evaluated after either 90 minutes (n = 8) or 24 hours (n = 6) of reperfusion. Group 2 animals (propylene glycol pretreatment, n = 8) were pretreated with a propylene glycol vehicle, then underwent 6 hours of ischemia and were evaluated after 90 minutes reperfusion. Group 3 animals (DHEA pretreatment, n = 12) were pretreated with DHEA dissolved in propylene glycol, subjected to 6 hours of ischemia, and then evaluated after either 90 minutes (n = 6) or 24 hours (n = 6) of reperfusion. Red blood cell velocity in the flaps main arteriole, functional capillary density, venular constriction index (the ratio of internal to external diameter of postcapillary venules), and microemboli formation were measured. Muscle samples were evaluated by electron microscopy. Control animals showed a 61% reduction in red blood cell velocity (p < 0.05) accompanied by a 69% reduction in functional capillary density (p < .05) acutely and total cessation of flow by 24 hours. No differences between control and propylene glycol treated animals were noted. In DHEA-pretreated animals, reflow occurred in 100% of the flaps, there was a temporary 39% reduction (p < 0.05) in functional capillary density, and all flaps remained viable at 24 hours. In this study, DHEA pretreatment markedly improved muscle flap microcirculatory hemodynamics and protected flaps against ischemia/reperfusion injury.

Neuroendocrine carcinomas: Role of immunocytochemistry and electron microscopy

Neuroendocrine tumors occur in many sites of the body and can present significant diagnostic problems when poorly differentiated. To identify a tumor as neuroendocrine, pathologists commonly use either immunocytochemistry or electron microscopy. In this report, the various immunocytochemical reagents are reviewed along with the ultrastructural features of neuroendocrine tumors. Site-specific variations in neuroendocrine tumors are discussed. A cost-effectiveness evaluation was performed on tumors from one laboratory which showed that electron microscopy was a less expensive diagnostic modality if more than three antibodies were necessary to arrive at the correct pathological diagnosis.

Ultrastructural Findings in Cardiac Transplant Recipients

Ultrastructural findings in 350 endomyocardial biopsy specimens and 59 autopsy or explanted hearts from cardiac transplant recipients are reviewed. Myocyte degeneration can be readily distinguished from necrosis by the technique described. Vascular changes of endothelial activation, endothelial cell damage, and basement membrane reduplication can be readily identified. In addition, the myofilament composition of ischemic hearts in patients with allograft coronary artery disease is distinctive: There is a disproportionate loss of actin over myosin, giving a coarse appearance to the myofilaments. These changes are useful in further defining the morphologic features associated with rejection and ischemia in cardiac transplant recipients.