p53 status and prognosis of locally advanced prostatic adenocarcinoma: a study based on RTOG 8610.

BACKGROUND: The p53 tumor suppressor gene (also known as TP53) is one of the most frequently mutated genes in human cancer. Several studies have shown that p53 mutations are infrequent in prostate cancer and are associated with advanced disease. PURPOSE: We assessed the prognostic value of identifying abnormal p53 protein expression in the tumors of patients with locally advanced prostate cancer who were treated with either external-beam radiation therapy alone or total androgen blockade before and during the radiation therapy. METHODS: The study population consisted of a subset of patients entered in Radiation Therapy Oncology Group protocol 8610 ("a phase III trial of Zoladex and flutamide used as cytoreductive agents in locally advanced carcinoma of the prostate treated with definitive radiotherapy"). Immunohistochemical detection of abnormal p53 protein in pretreatment specimens (i.e., needle biopsies or transurethral resections) was achieved by use of the monoclonal anti-p53 antibody DO7; specimens in which 20% or more of the tumor cell nuclei showed positive immunoreactivity were considered to have abnormal p53 protein expression. Associations between p53 protein expression status and the time to local progression, the incidence of distant metastases, progression-free survival, and overall survival were evaluated in univariate (logrank test) and multivariate (Cox proportional hazards model) analyses. Reported P values are two-sided. RESULTS: One hundred twenty-nine (27%) of the 471 patients entered in the trial had sufficient tumor material for analysis. Abnormal p53 protein expression was detected in the tumors of 23 (18%) of these 129 patients. Statistically significant associations were found between the presence of abnormal p53 protein expression and increased incidence of distant metastases (P = .04), decreased progression-free survival (P = .03), and decreased overall survival (P = .02); no association was found between abnormal p53 protein expression and the time to local progression (P = .58). These results were independent of the Gleason score and clinical stage. A significant treatment interaction was detected with respect to the development of distant metastases: Among patients receiving both radiation therapy and hormone therapy, those with tumors exhibiting abnormal p53 protein expression experienced a reduced time to the development of distant metastases (P = .001); for patients treated with radiation therapy alone, the time to distant metastases was unrelated to p53 protein expression status (P = .91). CONCLUSIONS: Determination of p53 protein expression status yield significant, independent prognostic information concerning the development of distant metastases, progression-free survival, and overall survival for patients with locally advanced prostate cancer who are treated primarily with radiation therapy. IMPLICATIONS: The interaction of radiation therapy plus hormone therapy and abnormal p53 protein expression may provide a clinical link to experimental evidence that radiation therapy and/or hormone therapy act, at least in part, by the induction of apoptosis (a cell death program) and suggests that this mechanism may be blocked in patients whose tumors have p53 mutations.

Consensus statements on radiation therapy of prostate cancer: guidelines for prostate re-biopsy after radiation and for radiation therapy with rising prostate-specific antigen levels after radical prostatectomy. American Society for Therapeutic Radiology and Oncology Consensus Panel.

PURPOSE: To develop evidence-based guidelines for (1) prostate re-biopsy after radiation and (2) radiation therapy with rising prostate-specific antigen (PSA) levels after radical prostatectomy in the management of patients with localized prostatic cancer. DESIGN: The American Society of Therapeutic Radiology and Oncology (ASTRO) challenged a multidisciplinary consensus panel to address consensus on specific issues in each of the two topics. Four well-analyzed patient data sets were presented for review and questioning by the panel. The panel sought criteria that would be valid for patients in standard clinical practice as well as for patients enrolled in clinical trials. Subsequent to an executive session that followed these presentations, the panel presented its consensus guidelines. RESULTS AND CONCLUSIONS: Based on the data presented, the prostate re-biopsy negative rates ranged from 62% to 80% for patients with stage T1-2 tumors. The panel judged that prostate re-biopsy is not necessary as standard follow-up care and that the absence of a rising PSA level after radiation therapy is the most rigorous end point of total tumor eradication. Further, the panel judged that re-biopsy may be an important research tool. Based on the data presented, the long-term (5 years or more) PSA remission rate after salvage radiation therapy ranges from 27% to 45%. The panel requested results from prospective randomized trials to evaluate optimally this information. The panel judged that the total dose of radiation should be 64 Gy or slightly higher and that, in patients with or without radiation therapy, there is no standard role for androgen suppressant therapy for rising PSA values after prostatectomy.

Prostate cancer DNA ploidy and response to salvage hormone therapy after radiotherapy with or without short-term total androgen blockade: an analysis of RTOG 8610.

PURPOSE: DNA ploidy has consistently been found to be a correlate of prostate cancer patient outcome. However, a minority of studies have used pretreatment diagnostic material and have involved radiotherapy (RT)-treated patients. In this retrospective study, the predictive value of DNA ploidy was evaluated in patients entered into Radiation Therapy Oncology Group protocol 8610. The protocol treatment randomization was RT alone versus RT plus short-course (approximately 4 months) neoadjuvant and concurrent total androgen blockade (RT+TAB). PATIENTS AND METHODS: The study population consisted of 149 patients, of whom 74 received RT alone and 75 received RT+TAB. DNA content was determined by image analysis of Feulgen stained tissue sections; 94 patients were diploid and 55 patients were nondiploid. Kaplan-Meier univariate survival, the cumulative incidence method, and Cox proportional hazards multivariate analyses were used to evaluate the relationship of DNA ploidy to distant metastasis and overall survival. RESULTS: DNA nondiploidy was not associated with any of the other prognostic factors in univariate analyses. In Kaplan-Meier analyses, 5-year overall survival was 70% for those with diploid tumors and 42% for nondiploid tumors. Cox proportional hazards regression revealed that nondiploidy was independently associated with reduced overall survival. No correlation was observed between DNA ploidy and distant metastasis. The diminished survival in the absence of an increase in distant metastasis was related to a reduction in the effect of salvage androgen ablation; patients treated initially with RT+TAB and who had nondiploid tumors had reduced survival after salvage androgen ablation. CONCLUSIONS: Nondiploidy was associated with shorter survival, which seemed to be related to reduced response to salvage hormone therapy for those previously exposed to short-term TAB.

Left ventricular mass increases during cardiac allograft vascular rejection.

OBJECTIVES: This study evaluated whether left ventricular mass increases during cellular or vascular (humoral) cardiac allograft rejection. BACKGROUND: An increase in left ventricular mass during cellular cardiac allograft rejection has been described by other investigators, although controversy has existed over the validity of these findings. Left ventricular mass changes have not been evaluated in the setting of vascular (humoral) cardiac allograft rejection. METHODS: To determine the effect of allograft rejection on left ventricular mass, we retrospectively reviewed endomyocardial biopsy results and corresponding echocardiograms in 41 cardiac transplant recipients undergoing treatment for allograft rejection. Left ventricular mass was assessed by two-dimensional echocardiography using the method of Schiller. Maintenance immunosuppression included cyclosporine in all patients. RESULTS: Although significant changes in left ventricular wall thickness, mass and dimensions were not observed in patients experiencing moderate or severe cellular allograft rejection (International Society for Heart and Lung Transplantation grades III and IV, n = 27), marked changes were noted in patients with vascular (humoral) rejection (n = 14). Patients with vascular rejection demonstrated an echocardiographic mean (+/- SEM) increase in left ventricular wall mass (from 109 +/- 17 to 151 +/- 17 g), and left ventricular wall thickness (from 1.3 +/- 0.1 to 1.6 +/- 0.1 cm) during the rejection episode. Additionally, vascular rejection was associated with a trend toward an increase in left ventricular systolic dimension (from 2.6 +/- 0.1 to 3.0 +/- 0.2 cm) and a decrease in left ventricular fractional shortening and increased incidence of hemodynamic compromise with rejection (50% for vascular vs. 11% for cellular rejection). CONCLUSIONS: Left ventricular mass increases during episodes of vascular (humoral) rejection, but there is no significant change in left ventricular mass during cellular cardiac allograft rejection.

Increased incidence of Chlamydia species within the coronary arteries of patients with symptomatic atherosclerotic versus other forms of cardiovascular disease.

OBJECTIVES: The objectives of this study were to test prospectively for an association between Chlamydia and atherosclerosis by comparing the incidence of the pathogen found within atherosclerotic plaques in patients undergoing directional coronary atherectomy with a variety of control specimens and comparing the clinical features between the groups. BACKGROUND: Previous work has suggested an association between Chlamydia pneumoniae infection and coronary atherosclerosis, based on the demonstration of increased serologic titers and the detection of bacteria within atherosclerotic tissue, but this association has not yet been regarded as established. METHODS: Coronary specimens from 90 symptomatic patients undergoing coronary atherectomy were tested for the presence of Chlamydia species using direct immunofluorescence. Control specimens from 24 subjects without atherosclerosis (12 normal coronary specimens and 12 coronary specimens from cardiac transplant recipients with subsequent transplant-induced coronary disease) were also examined. RESULTS: Coronary atherectomy specimens were definitely positive in 66 (73%) and equivocally positive in 5 (6%), resulting in 79% of specimens showing evidence for the presence of Chlamydia species within the atherosclerotic tissue. In contrast, only 1 (4%) of 24 nonatherosclerotic coronary specimens showed any evidence of Chlamydia. The statistical significance of this difference is a p value < 0.001. Transmission electron microscopy was used to confirm the presence of appropriate organisms in three of five positive specimens. No clinical factors except the presence of a primary nonrestenotic lesion (odds ratio 3.0, p = 0.057) predicted the presence of Chlamydia. CONCLUSIONS: This high incidence of Chlamydia only in coronary arteries diseased by atherosclerosis suggests an etiologic role for Chlamydia infection in the development of coronary atherosclerosis that should be further studied.

Correlation of survival with quantitative tissue staining of prostate specific acid phosphatase in patients with prostate carcinoma by using microscopic image analysis: a preliminary report of correlative studies on RTOG protocols 75-06, 77-06, and 83-07.

PURPOSE: We have previously shown that the intensity (graded semiquantitatively as 1-4+) of tissue prostate-specific acid phosphatase (PSAP) staining determined immunocytochemically in a cohort of prostate carcinoma patients from Radiation Therapy Oncology Group Protocols (RTOG) protocols 75-06 and 77-06 correlated with survival. The extent of this staining was heterogeneous and was estimated. The extent of staining was not found to be significantly associated with survival. We undertook the present quantitative study to see if the improved precision and reliability of measurement of the intensity and extent of prostate specific acid phosphatase staining would confirm and extend our previous observations. METHODS AND MATERIALS: Patient cohorts representative of the entire group were obtained from RTOG 75-06 plus 77-06 and 83-07. The RTOG 77-06 plus 75-06 patients (No-Hormone population) did not receive preradiation hormonal therapy. RTOG 83-07 patients (Prehormone population) received one of two types of preradiation chemical androgen ablation. In this study, histologic slides of tumors were immunocytochemically stained for PSAP by the peroxidase-antiperoxidase (PAP) technique using diaminobenezidene (DAB) as a substrate and hematoxylin as a nuclear counterstain. The intensity and extent of immunocytochemical PSAP staining (IPSAP stain) was quantified using our dual wavelength and batch mode image process technique. RESULTS: Our study of 151 cases confirmed that overall survival of patients in both populations was positively correlated with the intensity and extent of IPSAP stain. Results of the two studies were similar. The statistical significance of the relationship of both extent and intensity was greater in the cohort from protocol 83-07, which was the patient group receiving pretreatment with hormones. In a Cox multiple regression analysis including clinical stage, Gleason and M. D. Anderson grades, and the cohort of patients (Prehormone or No-Hormone group) as covariables, both the intensity and extent of the IPSAP stain significantly correlated with survival along with M. D. Anderson grade of the tumor. CONCLUSION: Quantitative image analysis of the IPSAP stain predicts survival in patients treated with external beam radiotherapy with and without prior hormonal therapy.

Detection of allograft endothelial cells of recipient origin following ABO-compatible, nonidentical cardiac transplantation.

Endothelial cells serve an important role in augmenting immune responses through enhanced expression of MHC class II antigens. Immune-mediated vascular injury associated with rejection requires reendothelialization to restore vascular integrity. The origin of the reparative endothelial cells can be determined when ABO antigens expressed on these cells differ in the donor and recipient. To assess the frequency and significance of reendothelialization by recipient endothelial cells, we stained serial endomyocardial biopsies for ABO antigens in 34 (13%) compatible, nonidentical cardiac allograft recipients of 268 cardiac transplant procedures. In ten (30%) the allograft endothelial cells expressed the characteristics of the recipient (five partial and five complete) within 7.5 +/- 1.0 months (mean +/- SEM) after transplantation. Over 26.3 +/- 2.5 months follow-up no differences could be detected in pretransplant characteristics, allograft survival, survivor rejection morbidity, long-term allograft function, and presence of coronary vasculopathy between those whose endothelial cells expressed recipient blood group antigens and those who did not, which may merely be a reflection of the small sample size. This study indicates that recipient reendothelialization occurs frequently following cardiac transplantation and may result from immune-mediated vascular injury. The effect of recipient reendothelialization on allograft tolerance requires further investigation.

Prevention of Pneumocystis carinii pneumonia in cardiac transplant recipients by trimethoprim sulfamethoxazole.

Pneumocystis carinii pneumonia (PCP) continues to cause significant morbidity in recipients of solid-organ transplants. While some programs administer trimethoprim-sulfamethoxazole (TMP-SMX) prophylactically following transplantation, a prospective determination of the safety and efficacy of TMP-SMX in cardiac transplant recipients has not previously been reported. We therefore prospectively randomized 58 cardiac transplant recipients to receive TMP (160 mg)-SMX (800 mg) twice daily either three days per week (group B), or seven days per week (group C), or to receive no treatment (group A). Treatment began 14 days after transplantation and continued for four months. Age, sex, preexisting pulmonary pathology and immunosuppressive protocols did not differ among the groups. Of 17 patients in the control group (A), 7 developed a clinical syndrome compatible with PCP, with the diagnosis histologically confirmed by bronchoalveolar lavage during the first four months following transplantation. In contrast, no patients in either the daily or intermittent therapy groups developed PCP during the study period (P < 0.005). Both doses of TMP-SMX were well tolerated, and discontinuation of therapy was not necessary in any patient. Total white blood cell count, azathioprine dose, and number of treated episodes of rejection per patient did not differ among the three groups. We conclude that TMP-SMX can safely and effectively be administered to prevent the occurrence of P carinii pneumonia during the first four months following cardiac transplantation.

A prospective, randomized comparison of cyclophosphamide and azathioprine for early rejection prophylaxis after cardiac transplantation. Decreased sensitization to OKT3.

Humoral immune responses have been implicated in the pathogenesis of vascular rejection, allograft coronary artery disease, and sensitization to OKT3. Because cyclophosphamide (CP) is a potent suppressor of humoral immunity, we postulated that substituting cyclophosphamide for azathioprine (AZA) would be associated with a decrease in acute vascular rejection and sensitization to OKT3 in cardiac transplant recipients also receiving cyclosporine, corticosteroids, and perioperative OKT3. We prospectively randomized 119 patients to receive azathioprine (n = 61) or cyclophosphamide (n = 58) from the time of transplantation. Dosage was adjusted to target white blood cell (WBC) counts. At six weeks posttransplantation, cyclophosphamide was converted to azathioprine. Patients were followed for a mean of 321 +/- 16 days. At four weeks WBC (1000/mm3) was 9.2 +/- 0.4 (SEM) in the AZA group and 9.7 +/- 0.6 for the CP group (P = 0.4). No differences were noted between the CP and AZA groups in mean cellular grades of rejection (1.8 +/- 0.1 vs. 1.7 +/- 0.1), mean vascular grades of rejection (2.0 +/- 0.1 vs. 1.8 +/- 0.1), early treated rejection episodes (1.9 +/- 0.1 vs. 2.2 +/- 0.1) days to first treated cellular rejection (38 +/- 3 vs. 41 +/- 3), or the number of patients manifesting primarily vascular rejection (18 vs. 19). Major infections and survival did not differ between the two groups. Eight patients in the AZA group developed anti-OKT3 antibodies, whereas only one patient in the CP group did (P = 0.04). In the early posttransplant period cyclophosphamide decreases the incidence of sensitization to OKT3 and appears to be as effective as azathioprine in preventing both cellular and vascular rejection.

Outcome of cardiac transplant recipients with a positive donor-specific crossmatch--preliminary results with plasmapheresis.

To assess the influence of a positive T or B cell IgG crossmatch on the development of rejection and mortality following cardiac transplantation, we reviewed all cardiac transplants performed in Utah between March 1985 and October 1990. Of the 328 cardiac allograft recipients, 11 (3.4%) had an IgG positive crossmatch. Actuarial survival at 24 months in the positive crossmatch group was 57.3% +/- 0.02 while that of the controls was 86.1% +/- 2.1 (P < 0.05). Allograft rejection occurred earlier in recipients with a positive crossmatch (10.0 +/- 5.8 days versus 34.0 +/- 2.3 days, P < 0.001). The first allograft rejection episode in patients with a positive crossmatch was characterized by immunoglobulin and complement deposition in small blood vessels and interstitial edema and endothelial cell activation in the absence of a lymphocytic infiltrate. Furthermore, the allograft rejection in the positive crossmatch group was accompanied by hemodynamic compromise in a large proportion of the patients (73%). In addition to augmentation of immunosuppression, plasma exchange therapy was performed within the first week following transplantation in 8 of the 11 positive crossmatch patients. Survival in the patients treated with plasma exchange (75%) appears to be better than in those not receiving plasma exchange (33%) within one week of transplantation. While immunosuppressive therapy aimed at the humoral arm of the immune system and plasma exchange therapy may improve survival in recipients with a positive donor-specific crossmatch, survival is worse in patients with a positive crossmatch than in patients with a negative crossmatch. Thus, it would appear prudent to prospectively crossmatch cardiac transplant candidates with a greater risk of developing a positive crossmatch, such as those potential recipients with an elevated level of panel-reactive antibodies.

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.

Relationship of OKT3 sensitization and vascular rejection in cardiac transplant patients receiving OKT3 rejection prophylaxis.

We prospectively and serially monitored plasma levels of OKT3 in 20 patients who were receiving 14- or 21-day rejection prophylaxis with OKT3. We retrospectively compared plasma OKT3 levels with biopsy scores assessed by light microscopy and immunofluorescence, clinical findings, human antimouse antibody (HAMA) production assessed by a blocking assay and by ELISA, and circulating immune complex levels assessed by a flow cytometric Raji cell assay. Using these methods, we evaluated the relationship of OKT3 sensitization, a humorally mediated immune response, to the development of vascular rejection in these patients. We found that 6 of 20 patients had declines in plasma OKT3 levels to less than 50% of their steady-state value before the conclusion of therapy (OKT3 consumption). This fall in plasma OKT3 preceded a significant rise in the CD 3 lymphocyte level by up to 3 days. All 6 patients showed HAMA production by either blocking or ELISA assay (P = less than 0.02) and developed vascular rather than cellular rejection (P = less than 0.01). OKT3 sensitization was significantly more common in patients treated with 21-day rejection prophylaxis (4 of 6 patients, P = less than 0.01). Only 4 of 14 other patients showed vascular rejection; 2 of these 4 also developed HAMA without OKT3 consumption and both had been treated with 21-day rejection prophylaxis with OKT3. None of the 20 patients showed significant levels of circulating immune complexes. This study demonstrates that OKT3 sensitization is strongly associated with vascular rejection. Vascular rejection was usually demonstrated 7 days after OKT3 consumption was seen and was coincident with HAMA production. By contrast, 4 patients without OKT3 sensitization had vascular rejection demonstrable in the early posttransplant period; in such patients, prospective immunofluorescence of biopsies was the only reliable indicator of this rejection type. The higher incidence of vascular rejection in these 20 patients was definitely related to the use of 21-day OKT3 rejection prophylaxis. Overall, 7 of the 12 patients treated with this regimen developed vascular rejection. Allograft and patient survival among patients with vascular rejection was significantly worse than in patients with cellular rejection (P = less than 0.01). Prospective monitoring of patients treated with OKT3 by serial plasma levels and by biopsy immunofluorescence will identify patients at risk for these types of humoral rejection.

The use of mycophenolate mofetil (RS-61443) in human heart transplant recipients.

Mycophenolate mofetil is a potent inhibitor of de novo guanine nucleotide synthesis that selectively blocks lymphocyte proliferative responses. In animal models, mycophenolate mofetil has been shown to prolong allograft survival, reverse ongoing rejection, and induce strain-specific tolerance. To assess the safety and efficacy of mycophenolate mofetil in cardiac transplantation, 30 recipients with mild rejection were enrolled in an 8-week phase I trial. Mycophenolate mofetil in doses from 500 to 3000 mg/day orally was substituted for azathioprine, while baseline cyclosporine levels and corticosteroid doses were maintained. Rejection resolved in the majority of patients, with a significant decrease in mean biopsy score. By protocol, mycophenolate mofetil was discontinued in 4 patients due to persistent mild rejection, and in 4 patients due to progression to moderate rejection. The rate of progression to moderate rejection compared favorably with that observed in patients with mild rejection maintained on azathioprine without augmentation of immunosuppression. Significant increases were observed in hematocrit, total white blood cell count, and absolute neutrophil count. Absolute lymphocyte count remained unchanged. No nephrotoxicity or hepatotoxicity was observed. Gastrointestinal side effects prompted discontinuation of mycophenolate mofetil in one patient. Two major infections occurred. Mycophenolate mofetil remained well tolerated during long-term maintenance immunosuppression, with a rate of rejection similar to that in patients receiving azathioprine. We conclude that mycophenolate mofetil is safe and well tolerated in cardiac transplant recipients, is less myelosuppressive than azathioprine, and appears to be at least equipotent to azathioprine.

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.

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.

HLA A, B and DR typing in idiopathic dilated cardiomyopathy: a search for immune response factors.

Several autoimmune diseases have been associated with increased frequencies of various histocompatibility antigen (HLA) types that may be linked to immune response genes. Idiopathic dilated cardiomyopathy (IDC) has been proposed as a disease with autoimmune features, but HLA associations have not been evaluated. We performed HLA typing in 37 consecutive patients with IDC. Patients with habitual alcoholism were excluded. Results showed that no single HLA type could account for most cases; IDC is a genetically heterogeneous disease. However, uneven distributions were noted for certain types. Haplotype frequency of B27 was 0.145 in patients vs 0.033 in 5,726 local control subjects (p less than 0.001). Other A and B frequencies (except A2) were evenly distributed. HLA DR typing also revealed differences. The DR4 locus was present in 54% (19 of 35) of patients, vs 32% (26 of 82) of blood bank control subjects (p less than 0.02). The associated relative risk of DR4 was 2.2 and the etiologic fraction 0.29. Sex, disease chronicity, functional class, ejection fraction and biopsy evidence of myocarditis did not distinguish DR4-positive from DR4-negative patients, but they were older (54 +/- 12 vs 42 +/- 14 years, p less than 0.02). Of note, 68% were positive for DR4 or B27, or both. HLA DR6Y was underrepresented; it was present in 9% (3 of 35) of patients, vs 26% (21 of 82) of control subjects (p less than 0.04). The relative risk of DR6Y was 0.27 and the preventive fraction 0.19. These associations will require independent confirmation. However, they suggest that genetically determined immune response factors associated with HLA loci may play a role in pathogenesis in certain patients with IDC.

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.

Phenotype and serine esterase production of human cardiac allograft-infiltrating lymphocytes.

Human cardiac allograft-infiltrating lymphocytes were studied by in vitro expansion in interleukin-2. Of 28 graft-infiltrating lymphocyte cultures from 17 recipients, 17 were comprised predominantly of CD4+ T cells and 10 predominantly CD8+ T cells; one culture had equal numbers of CD4+ and CD8+ cells. The mean percentages (+/- SE) of T-cell subsets for all cultures were as follows: CD4+, 49% +/- 29%; CD8+, 42% +/- 31%. No correlation was observed between the culture phenotype and histologic findings, length of time from transplantation, or number (or class) of mismatched HLA antigens. N-alpha-benzyloxycarbonyl-L-lysine thiobenzyl ester serine esterase (BLT-SE) is an enzyme associated with intracellular cytotoxic T-cell granules and with target-cell destruction. Sixteen cultures were tested for BLT-SE activity and had significantly increased enzyme activity as compared to untreated peripheral blood mononuclear cells from healthy control subjects (p < 0.002), or interleukin-2-treated control cells (p < 0.05). A low percentage (0.4% +/- 0.2%) of cells in the graft-infiltrating lymphocyte cultures expressed the phenotypic marker NKH-1, suggesting that the source of BLT-SE in these cultures was not natural killer or lymphokine-activated T cells. Elevated BLT-SE was observed in five of ten cultures containing predominantly CD4+ cells and five of six cultures containing predominantly CD8+ T cells. Mixed phenotype graft-infiltrating lymphocyte cultures depleted of either CD4+ or CD8+ T cells retained BLT-SE activity. Thus both CD4+ and CD8+ graft-derived T cells can produce this enzyme although much greater variability in enzyme production was seen for CD4+ T cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunofluorescence of endomyocardial biopsy specimens: methods and interpretation.

On the basis of our experience with prospective observation of immunofluorescence microscopy in 5775 biopsy specimens from 335 consecutively followed heart transplant cases, we describe the immunofluorescent method in detail and the potential artifacts and method of interpretation of the findings. A strategy for use of the method on the basis of the experience of the authors is proposed.

Myocardial apoptosis in a heterotopic murine heart transplantation model of chronic rejection and graft vasculopathy.

BACKGROUND: Apoptosis has been implicated in myocardial reperfusion injury and in experimental transplantation rejection. One mechanism of apoptosis is through the interaction of the cell-surface Fas receptor on target cells and the Fas ligand that is expressed on cytotoxic T cells. The purpose of this study was to look for evidence of myocardial Fas receptor, Fas ligand, and apoptosis in a murine heterotopic heart transplantation model of chronic rejection/graft vasculopathy. METHODS: Using the nick-end labeling technique, we examined a murine heterotopic heart transplantation model of chronic rejection/graft vasculopathy (strain B10.A to B10.BR) histologically for evidence of DNA fragmentation. MRNA for the Fas receptor, Fas ligand, and beta-actin was detected with reverse transcription-polymerase chain reaction. RESULTS: Hearts harvested after 30 and 60 days showed an intimal index of the allografts (0.5 +/- 0.1) (mean +/- standard error) that was at least five times more than syngeneic grafts and native (nontransplanted) hearts (p < 0.01). In situ nick end-labeling of partially degraded DNA with terminal deoxynucleotydil transferase showed an increase in apoptotic cells in allografts and syngeneic grafts compared with native hearts. Reverse transcription-polymerase chain reaction detected equal myocardial RNA signal intensity of Fas receptor and beta-actin in allografts, syngeneic grafts, and native hearts. In contrast, allografts showed a strong signal for the Fas ligand mRNA, a signal not seen in syngeneic grafts or native hearts. CONCLUSIONS: Apoptosis is occurring in both allografts and syngeneic grafts in this murine model of chronic rejection/graft vasculopathy, although distinct mechanisms may be involved.