Acute Cellular Rejection Grade Research Articles

Apollon/RNF41 Myocardial Messenger RNA Diagnoses Cardiac Allograft Apoptosis in Rejection

Endomyocardial biopsy (EMB) remains the gold standard for acute cellular rejection (ACR) diagnosis in cardiac transplantation yet is subject to interobserver variability. A method that could avoid discordant EMB analysis would be desirable. The apoptosis rate in EMB correlates with ACR severity. Apollon inhibits apoptosis, and RNF41 catalyzes its degradation. Whether tissue Apollon/RNF41 could diagnose ACR is not known. This study addressed this issue. Apollon/RNF41 messenger RNA (mRNA) was measured by real time reverse-transcriptase polymerase chain reaction and apoptosis was quantified with TUNEL assays in EMBs of 268 transplant recipients. EMBs were obtained at 1, 2, 3, 4, 7, 12, 24, and 52 posttransplant weeks. At all time points posttransplant, Apollon mRNA decreased significantly in EMBs with ACR grades 2R/3R combined (P<or=0.0010) compared with 0/1R combined, although RNF41 mRNA significantly increased in EMBs with ACR grade 1R (P<0.0001) or 2R/3R combined (P<0.0001) compared with 0. At the identified cut-off level of less than or equal to 168.2 arbitrary units, Apollon mRNA identified ACR grades 2R/3R with 100% sensitivity and 84% specificity, whereas RNF41 mRNA at the cut-off level of more than or equal to 51.8 identified ACR grades 1R-3R with 99% sensitivity and 95% specificity. Increased RNF41 (rs, 0.728; P<0.0001) and decreased Apollon (rs, -0.562; P<0.0001) expression correlated significantly with the degree of apoptosis in EMBs. Combined Apollon/RNF41 mRNA quantitatively and specifically identifies ACR associated with apoptosis in cardiac allografts and could validate ACR grading variability associated with histologic EMB analysis.

Long-Term Efficacy and Safety of Conversion to Tacrolimus in Heart Transplant Recipients with Ongoing or Recurrent Acute Cellular Rejection

Despite the widespread use of potent immunosuppressive drugs, such as cyclosporin A and mycophenolate mofetil, ongoing and recurrent cellular rejection remain a common problem after heart transplantation. We aimed to describe the long-term effects of conversion from cyclosporine A to tacrolimus in patients with ongoing and recurrent cellular rejection. This was a single-centre retrospective analysis of 17 heart transplant recipients who were switched from cyclosporine A to tacrolimus due to ongoing (5 patients) or recurrent cellular rejection (12 patients). We studied long-term efficacy and safety of this approach. 167 endomyocardial biopsies were performed during a mean follow-up of 69.1 +/- 12.7 months. Thirteen biopsies (7.8%) in eight patients (47%) revealed higher grades of acute cellular rejection (Banff 2). However, they were not hemodynamically significant and did not require intravenous antirejection therapy. The mean rejection score was reduced significantly. Conversion to tacrolimus was tolerated in 82% pts without any significant side effects during a long-term follow-up. In conclusion, the conversion to tacrolimus in heart transplant recipients with ongoing or recurrent acute cellular rejection was safe and effective also during a long-term follow-up.

International Grading Scheme for Acute Cellular Rejection in Small-Bowel Transplantation: Single-Center Experience

A standardized grading scheme for the assessment of acute cellular rejection (ACR) in small-intestine transplantation was proposed in 2003 at the Eighth International Small Intestinal Transplantation Symposium. We have implemented the current grading scheme for ACR in small-bowel transplantation since October 2003. The pathologic diagnoses of those small-intestine biopsy samples, including ACR grade and other supplementary findings were evaluated. A total of 3484 small intestine biopsy samples from 155 patients were available for evaluation in this study. Frequency of grades 0, indeterminate, 1, 2, and 3 acute cellular rejection was 33.9%, 49.1%, 12.6%, 3.7%, and 0.8%, respectively. Duration of ACR episode strongly correlated with grade of ACR episode ( P < .001). Other supplementary findings included acute vascular rejection component, 2.2%; increase in lymphoplasmacytic infiltrate in lamina propria, 15.7%; mucosal fibrosis, 0.4%; and regenerative changes, 0.3%. Our data substantiate that this grading system is reliable and useful for clinical decision making in bowel transplantation. We suggest that an assessment and quantification of supplementary findings be considered a component of the International Pathology Grading Scheme.

Tricuspid Valve Regurgitation and Endomyocardial Biopsy After Orthotopic Heart Transplantation

Objective Tricuspid valve regurgitation (TR) after heart transplantation (HTx) has been reported to be caused by endomyocardial biopsy (EMB), acute cellular rejection (ACR), or atrial anastomosis. We performed a prospective study of this problem among our HTx cohort. Methods From 1988 to 2006, we performed 274 HTx. Excluding cases within 1 year (2006), there were 178 patients in whom we had records of EMB dates, ACR grades (International Society for Heart and Lung Transplantation [ISHLT], 1990), echocardiography-measured TR, and time-to-TR. Statistical analyses were performed using nonparametric comparisons, Spearman correlation, Kaplan-Meier time to failure curves, and Cox regression model. Results All 178 patients underwent a biatrial anastomosis and underwent 2631 EMB (median, 15 times per patient; range, 0–42). The median follow-up duration was 66 months (range 2 days–194 months). Up to December 31, 2006, there were 47 patients (47/178 = 26.4%) who developed moderate-to-severe TR, which differed significantly from the prevalence rate (24/39 = 61.5%) reported by another cardiac team ( P = .001) that performed bicaval anastomoses in half of the cases (20/39 = 51%). Our 1-, 3-, and 10-year Kaplan-Meier incidence rates of remarkable TR were 14.7% (10.2%–20.8%), 19.4% (14.2%–26.2%), and 36.3% (27.2%–47.3%), respectively. A positive correlation was shown between each patient's EMB times and ACR but not TR grades, in terms of mean, maximum, or minimum over time (all P < .001 for null hypothesis of noncorrelation). Each patient's EMB times and number of definite ACRs (≥ISHLT grade II) did not differ significantly between the two groups of remarkable versus nonremarkable TR. Remarkable TR was negatively predicted by each patient's EMB times (hazard ratio = 0.93; P = .010) but not by the ACR grades or the numbers of definite ACRs. Conclusion Our cohort demonstrated that biatrial anastomosis, ACR, or EMB were not associated with the risk of remarkable TR. The protective effect of EMB on remarkable TR needs further investigation.

Effect of Ischemia–Reperfusion on the Incidence of Acute Cellular Rejection and Timing of Histologic Hepatitis C Virus Recurrence After Liver Transplantation

Background Because of a critical shortage of deceased donor (DD) livers, more extended criteria allografts are being utilized; these allografts are at increased risk for ischemia–reperfusion injury (IRI). We assessed whether, in a large cohort of patients transplanted for hepatitis C virus (HCV) either via a DD or live donor (LD), there was a relationship between the degree of IRI and the frequency and timing of acute cellular rejection (ACR) and histologic HCV recurrence. Methods During an 8-year study, patients were separated into four groups based on peak alanine aminotransferase (ALT) levels and three groups based on severity of IRI on postreperfusion liver biopsy. Results The mean follow-up time of 433 DD and 44 LD recipients was 1212 days. We noted a strong correlation in DD between peak ALT and the histologic degree of IRI ( P = .01). There was no difference in the incidence or grade of ACR among the four groups. There was no correlation between the severity of IRI and the incidence or time to histologic recurrence of HCV. Conclusions The magnitude of peak ALT correlated with the severity of IRI on postreperfusion liver biopsy. Among this large HCV cohort, there was no correlation between the severity of IRI and the incidence or timing of histologic HCV recurrence or incidence of ACR.

Characterization of C4d Immunostaining Utilizing Paraffin-Embedded Tissue of Nonpresensitized Pediatric Heart Transplant Patients

Immunoperoxidase techniques for demonstration of complement split product C4d on paraffin-embedded endomyocardial biopsy samples have been used for the evaluation of humoral rejection mainly in adult heart transplantation. Interpretation of suspected humoral rejection in children requires knowledge of the pattern and frequency of C4d deposition in nonpresensitized pediatric heart transplantation patients. Paraffin-embedded endomyocardial biopsy samples of 65 patients with no rejection, acute cellular rejection (ACR), or early ischemic/reperfusion injury were studied. Six biopsies within each grade of ACR both early (<6 months) and late (>6 months) after heart transplantation were reviewed, as were 5 biopsies of ischemic/reperfusion injury. None of the subjects was sensitized prior to transplant. All slides were blindly evaluated for histologic features traditionally associated with humoral rejection. C4d staining was quantified by counting the number of positive capillaries in 10 random high-power fields (hpf). C4d positivity (C4d+) was defined as >10 capillaries/10 hpf. C4d+ was observed in 6 (9%) endomyocardial biopsy samples; 2 in the early and 4 in the late ACR groups. Four had ACR grades 1A/B, and 2 had grade 3B/4. Thirteen (20%) endomyocardial biopsy samples had histologic features suggestive of humoral rejection. Of these, only 2 were C4d+ and both had ACR grade 3B/4. No C4d+ staining was found in the ischemic/reperfusion injury group. C4d immunostaining was negative in endomyocardial biopsy samples of the majority of our patients (91%). Endomyocardial biopsy samples with C4d+ did not show consistent ACR grade or time specificity. Contrary to previously reported data, none of the ischemic/reperfusion injury endomyocardial biopsy samples was C4d+, and histologic features alone were poor predictors of C4d+.

Correlation of Motility and Neuronal Integrity with a Focus on the Grade of Intestinal Allograft Rejection

Intestinal graft motility after small bowel transplantation (SBT) is poorly characterized. The aim of this study was to compare motor patterns with myenteric neuronal cell population as a parameter of graft viability at various degrees of acute cellular rejection (ACR). Three grades of ACR were achieved in orthotopic allografts. Syngeneic transplants and allografts with immunosuppression served as controls. Motor activities were recorded using strain gauge force transducers and analyzed visually. Quantifications of myenteric neurons in whole mounts of intestinal grafts were used to evaluate neuronal population. A typical migrating motor complex (MMC) was found in syngeneic and allogenic transplants with immunosuppression. A high prevalence of discrete clustered contractions (DCC) and nonpropagating contractions (NPC) without MMC was seen in moderately and severely rejected allografts. Neuronal cell loss in the allografts, which could be one of the causes of motor dysfunction, was noted in moderate rejection (19.3%) and progressed until severe rejection (60.1%). Monitoring motility patterns in SBT could be an effective tool for assessing intestinal rejection. Allograft dysmotility, such as absence of MMC and high prevalence of DCC or NPC, could be useful markers of progression of acute rejection and help guide treatment decisions.

Gene Expression Profiling Distinguishes a Molecular Signature for Grade 1B Mild Acute Cellular Rejection in Cardiac Allograft Recipients

Gene expression profiling distinguishes the absence or presence of moderate to severe grades of acute cellular rejection in cardiac allograft recipients using a 20-gene classifier. We explored the hypothesis that the rejection classifier also differentiates various forms of mild rejection and we performed sub-analyses based on time post-transplant and confirmatory pathology interpretations. A post hoc analysis of 265 CARGO study patients and 714 clinical encounters focused on the correlation of rejection classifier-derived gene expression (GE) scores for blood samples accompanying endomyocardial biopsies. Biopsy grades assigned by a study center pathologist (center) were re-interpreted by three pathologists (panel) in a blinded manner. Mean GE scores not only differentiated Grades >or=3A from Grade 0 (p < 0.00001, center or panel), but also from Grades 1A or 2 (p < 0.05, center or panel), based on mild rejection sub-groups defined by the ISHLT 1990 grading system. In contrast, mean GE scores for Grades 1B and >or=3A were indistinguishable, using either center or panel interpretation. Sub-group analyses of encounters from 2 to 6 months or >6 months post-transplant showed similar results for the classifier's ability to discriminate moderate to severe rejection from Grades 1A and 2 mild rejection, but indistinguishable mean GE scores for Grades >or=3A and the Grade 1B sub-group. Of the classifier's 11 informative genes, expression of MIR and WDR40 showed statistically significant increases for both Grade 1B and Grade >or=3A rejection, while expression of PDCD1 or SEMA7A showed similar directional patterns without achieving statistical significance. These data demonstrate that GE scores discriminate moderate to severe rejection from Grades 1A and 2 mild rejection. However, a sub-group of mild rejection cases, defined as Grade 1B according to the 1990 grading system, share a molecular signature more consistent with moderate to severe rejection. The clinical relevance of these data remains to be defined.

Elevated Levels of Serum Interleukin-6 are Associated With Low Grade Cellular Rejection in Patients With Heart Transplantation

Endomyocardial biopsy is the gold-standard procedure to diagnose acute cellular rejection after heart transplantation. This study assessed whether the blood levels of cytokines involved in inflammation and immune activation are useful to detect the presence of acute cellular rejection. Methods Blood specimens collected before 275 endomyocardial biopsies in 66 patients were assayed for levels of TNFα, IL6, IL1β, and IL2 receptor. The biopsies were grouped according to the presence ( n = 41) or absence ( n = 234) of acute cellular rejection grade ≥3A of the International Society for Heart and Lung Transplantation. We compared the levels of cytokines in the two groups. Results Circulating IL6 levels were significantly higher when there was a low grade (0–2) cellular rejection in the biopsy versus the group of biopsies grade ≥3A (19.8 ± 27 versus 12.9 ± 10 pg/mL; P = .001). An IL6 level higher than 30 pg/mL showed a negative predictive value of 95% for the presence of acute rejection grade ≥ 3A. Conclusion In heart transplant patients, high levels of serum IL6 were associated with low grade cellular rejection. Determination of IL6 levels may be useful to reduce the number of endomyocardial biopsies during follow-up in these patients.

Enterococcus faecium as a cause of pulmonary abscesses in kidney transplant recipient

It is well known that organ transplant recipients are susceptible to a host of atypical infections. Enterococcus faecium is an opportunistic nosocomial pathogen, spreading of which gained momentum in the last decade in hospitals worldwide. Infection by E. faecium, especially by strains resistant to vancomycin, carries a poor prognosis also after organ transplantation [1, 2]. However, pulmonary infections caused by enterococci are distinctly unusual. Herein, we report a 29-year-old male patient, who underwent his first kidney transplantation after waiting for 5 years on hemodialysis. His principal kidney disease was Goodpasture's syndrome. The surgery was uneventful and initial immunosupression consisted of tacrolimus, mycofenolate mofetil and steroids. Because of graft nonfunction, a core biopsy was performed on the fifth postoperative day showing acute cellular rejection grade IIB (Banff-’97). The patient received a course of antithymocyte globulin (Thymoglobulin, total dose of 800 mg; Sangstat-lmtix, Lyon, France). On the 22nd postoperative day, the patient developed acute abdomen, caused by chemical peritonitis due to a necrosis of the mid-portion of the ureter. After a nephrostomy drainage was placed, the kidney graft started functioning promptly, with serum creatinine dropping to 150 μmol/l. The course was further complicated by sepsis with multiple pulmonary infiltrates on the 31st postoperative day. Enterococcus faecium was found in the blood cultures. Intravenous lines were changed and chest X-ray examination was performed. A subsequent CT scan (Fig. 1) further specified the pulmonary lesions as abscesses, segmental pulmonary embolism and pericardial effusion were additional findings. Bronchoalveolar lavage revealed E. faecium as a single pathogen with the same sensitivity pattern. Our patient was initially treated by teicoplanin and after 3 days switched to linezolide according to strain sensitivity. Chest X-ray and CT scan showing multiple pulmonary infiltrates. Pericardial effusion as a side finding. The patient had rapidly recovered and received a course of linezolide for the next 3 weeks. The lung abscesses resolved within 21 days in the chest X-ray examination and control CT scan. The patient was discharged on the 52nd postoperative day with excellent kidney graft function (S-creatinine 87 μmol/l). His further course was complicated by recurrent episodes of kidney graft pyelonephritis caused by Klebsiela pneumoniae which subsided only after the implantation of the renal pelvis of his native kidney to the graft, thus restoring natural way of urinary drainage. One year later, the patient was found to be active with a stable graft function (S-creatinine 160 μmol/l). To our knowledge, this is the first report on E. faecium causing pulmonary infection (abscesses) as a single pathogen since a mixture of pulmonary pathogens including E. faecium has been documented so far [3]. Undoubtedly, both profound immunosupression with antithymocyte globulin and urinary leak contributed to this condition in our patient. This case may serve as a reminder that very unusual pathogens or unusual disease presentations can occur in organ transplant recipients under the intense immunosuppression.

Post-transplant lymphoma of the pancreatic allograft in a kidney-pancreas transplant recipient: a misleading presentation

A 37-year-old male patient underwent combined kidney–pancreas transplantation with an immunosuppressive treatment consisting of tacrolimus, mycophenolate mofetil and steroids, after induction therapy with anti-thymocyte globulin. The pancreas graft had intestinal drainage. Twenty-seven days post transplantation, the patient developed an episode of acute graft pancreatitis, which responded well to conservative measures (i.v. fluids, nil per orally, histamine-2 receptor blockade). On day 45, renal function was stable with a serum creatinine of 1.28mg/dl, corresponding to a measured creatinine clearance of 57ml/min. Because of persistent postprandial hyperglycaemia, the addition of low-dose insulin therapy was necessary for a period of 6 weeks after transplantation. Two months post transplantation, the patient was admitted because of diarrhoea and a rise in serum creatinine to 2.49mg/dl. A renal biopsy showed acute cellular rejection grade Ib according to the revised Banff 2001 criteria [1]. The patient was treated with corticosteroids, resulting in a good clinical and biochemical response. However, renewed insulin therapy was temporally required during this rejection episode and was discontinued during further follow-up. Stool cultures at that time revealed Campylobacter jejuni, for which doxycylin was administered for 10 days. Four months later, the patient presented with acute pain and swelling over the pancreatic graft. Other complaints were anorexia, weight loss and low grade fever. On clinical examination, we saw a thin patient with normal vital signs. A firm mass in the right iliac area (10 10 cm), painful on palpation, was present. Laboratory results showed haemoglobin levels of 10.0 g/dl—normochromic, normocytic—a white cell count of 9600/mm—93% neutrophils—a serum creatinine of 1.4mg/dl, urea nitrogen levels of 41mg/dl, gglutamyl transferase 68U/l, and lipase 70U/l. C-reactive protein (CRP) levels were 65.4mg/l. Amylase, triglycerides and calcium levels were normal. Tacrolimus blood levels were within therapeutic range (8–12 ng/ml). Proteinuria was 0.27 g/l without haematuria or bacteriuria. Screening for viral and fungal disease revealed the presence of Epstein–Barr virus (EBV) DNA viraemia [log 4.16 (1⁄414.510) copies/ml)]. At the time of transplantation, the patient tested positive for EBV serology (status of immunity). Abdominal ultrasound showed a heterogeneous, multinodular mass (10 10 7 cm) that was partly solid and partly heterogeneous cystic. The mass could not be distinguished from the surrounding adipose tissue. Differentiation between an inflammatory process and a tumoural mass was impossible. Magnetic resonance imaging (MRI) scan of the abdomen showed a complex heterogeneous and nodular mass (9.4 cm) with compression on the pancreatic allograft and the surrounding small intestinal loops (Figure 1). The mass was isoto hypointense on T2-weighted images and isoto slightly hyperintense on T1-weighted images. Its vascularization originated from the right iliac vessels (Figure 2). The process could not be distinguished from the head of the pancreas nor from the transplanted afferent part of the small intestine. Reaching a conclusive diagnosis was still impossible, although an inflammatory mass with necrosis or a graft pancreatitis with pseudocyst Correspondence and offprint requests to: Dirk R. J. Kuypers, MD, PhD, Department of Nephrology and Renal Transplantation, University Hospitals Leuven, Herestraat 49, B-3000 Leuven, Belgium. Email: Dirk.Kuypers@uz.kuleuven.ac.be Nephrol Dial Transplant (2006) 21: 3306–3310

Correlation between acute rejection severity and CD8-positive T cells in living related liver transplantation

The Banff schema is the most widely used standard grading system for liver allograft rejection. To investigate the relationship between the Banff rejection activity index (RAI) and the presence of lymphocyte subpopulations in allograft liver tissue, assuming these cells to probably play an important role in the mechanism of acute cellular rejection (ACR), we performed immunohistological examinations using liver tissues with various ACR severities after living related liver transplantation (LRLT). In total, 37 needle liver biopsy specimens with ACR in LRLT were examined using antibodies to CD4, CD8, and CD20. Formalin-fixed and paraffin-embedded liver tissues were used to maintain morphology. Immunohistological findings and RAI score according to the Banff schema were compared. In the results, mainly CD8-positive (CD8+), rather than CD4-positive (CD4+), cells were detected in the portal tract and were also found in bile duct epithelium and subendothelial areas of portal veins. The number of CD8+ cells increased according to ACR grade, whereas CD4+ cells tended to decrease. There were significant correlations between the presence of CD8+ cells ( p = 0.0006) and CD4+ cells ( p = 0.0003) and ACR severity. On the other hand, CD20-positive cells did not correlate with ACR severity ( p = 0.472). The results indicate that CD8+ cells play important roles in ACR severity of LRLT, suggesting the number of CD8+ cells in liver tissue to be useful as a supplementary tool, in addition to RAI of the Banff schema, for objective evaluation of ACR severity.

Does the Banff rejection activity index predict outcome in patients with early acute cellular rejection following liver transplantation?

The Banff schema incorporates a semiquantitative scoring system for grading of acute cellular rejection (ACR) of the liver allograft. The Banff rejection activity index (RAI) comprises 3 components scored from 0 to 3: venous endothelial inflammation (E); bile duct damage (B); and portal inflammation (P); the scores are combined to an overall score (the RAI). The purpose of this research was to determine the prognostic value of the Banff RAI score in predicting the response to increased immunosuppression and the long-term outcome of the graft. A retrospective study was done of patients undergoing primary liver transplantation between January 2000 and October 2004 with tacrolimus-based immunosuppression; 495 patients were included, 231 had histologically-confirmed ACR, 193 responded to 1 cycle of high-dose steroids. There was no correlation between the total RAI score and response to steroids, resistant rejection, development of chronic rejection, or graft survival. The E score was related to patient survival, a lower score being associated with a worse outcome (P = 0.048). In multivariable analysis, serum bilirubin, serum aspartate aminotransferase, and E score were significant predictors of death (P = 0.012). In univariable analysis, B score and bilirubin were significantly related to "resistant rejection" (P = 0.018 and 0.002, respectively), but only bilirubin was significant in multivariable analysis (logistic regression). In conclusion, although the Banff RAI score is a useful marker of the severity of rejection, neither the total RAI score nor any of the individual components correlated with response to steroids or graft survival.

Serum leptin concentration in patients after heart transplantation

Leptin is primarily produced by adipocytes but its receptors are expressed in a variety of tissues including the heart. Elevated plasma leptin levels predict acute myocardial infarction and it has been shown as acute phase reactant and a risk factor for coronary heart disease. The present study was undertaken to answer the question whether there exists a relationship or not, between serum leptin levels and grades of acute cellular rejection confirmed by elective endomyocardial biopsies in stable patients after orthotopic heart transplantation. We observed higher serum leptin levels compared with reference values, regardless of histopathologic biopsy findings studied. There was a positive correlation between serum leptin concentrations and body mass index, diastolic blood pressure, total cholesterol and low-density lipoprotein. The elevated leptin levels found in heart transplant recipients could be due to the result of steroid therapy. These results point the need for further studies to explain the leptin role in heart transplant recipients.

Pirfenidone inhibits inflammatory responses and ameliorates allograft injury in a rat lung transplant model

Tumor necrosis factor alpha is a proinflammatory cytokine that has been proved to play a crucial role in inducing posttransplantation lung injury. The present study was performed to determine whether pirfenidone, a new nonpeptide drug with potent anti-tumor necrosis factor alpha activity, promotes protection against acute allograft injury through inhibiting pulmonary inflammatory responses in a rat model of orthotopic lung transplantation. Three transplant groups were formed: isografts, untreated allografts, and allografts treated with pirfenidone (0.5% chow starting on day 1 after transplantation). The implants were harvested on day 21 after transplantation. Acute cellular rejection grade and degree of allograft injury were evaluated on the basis of hematoxylin-and-eosin staining. The pulmonary inflammatory response and inflammation-induced oxidative stress were assessed on the basis of neutrophil accumulation (myeloperoxidase immunoreactivity and enzymatic activity) and iron deposition (Prussian blue staining). In addition, circulating levels of tissue necrosis factor alpha in all animals were measured. The degree of allograft injury was significantly reduced in pirfenidone-treated allografts relative to untreated allografts (P < .01). The beneficial effect of pirfenidone was associated with decreased lung myeloperoxidase immunoreactivity (P < .05) and enzymatic activity (P < .01). Moreover, the untreated allografts contained a high concentration of iron, which was strikingly reduced by pirfenidone. Treatment with pirfenidone resulted in a lower level of plasma tissue necrosis factor alpha, which correlated positively with lung myeloperoxidase enzymatic activity (P < .0001). These results suggest that pirfenidone, with its anti-tissue necrosis factor alpha activity, reduced neutrophil recruitment and iron accumulation, hence limiting the acute lung allograft injury.

Diagnostic validity of hepatocyte growth factor as marker for rejection in the follow-up of patients after heart transplantation

Background Hepatocyte growth factor (HGF) is a member of growth factor with a variety of known activities, including angiogenesis promotion and antiapoptotic action. It prevents acute graft-versus-host disease after bone marrow transplantation and prolongs allogenic graft survival in rats. The aim of the study was to investigate the relationship between serum HGF concentration and the grade of acute cellular rejection of heart transplant. Methods We studied 68 male heart recipients. All of them received triple-drug immunosuppression: cyclosporine A, prednisone, and azathioprine or mycofenolate mofetil. All patients were without signs of heart failure. Blood samples were taken before elective ambulatory endomyocardial biopsy. Biopsy specimens were graded with the International Society for Heart and Lung Transplantation scale. Acute cellular rejection grade 3A and higher were considered as significant. Measurement of serum HGF was made by enzyme-linked immunosorbent assay. Results We found a positive relationship between serum HGF levels and grade of cellular rejection. As an indicator for the detection of cell rejection processes, HGF with cutoff 2000 pg/ml seems to be useful. Conclusions The results of this study demonstrate that HGF can be useful as an indicator for heart graft cell rejection.

ACCEPTANCE OF ABO-INCOMPATIBLE LIVING DONOR LIVER TRANSPLANTATIONS WITH THE USE OF ANTIGEN-SPECIFIC IMMUNOADSORPTION (GLYCOSORB® ABO), DACLIZUMAB AND MYCOPHENOLATE MOFETIL WITHOUT SPLENECTOMY.

P526 Aims: Introduction: Graft and patient survival rates in recipients of AB0 incompatible (AB0-I) liver transplantation is poor due to the greater incidence of vascular thrombosis, biliary injury and acute cellular rejection (ACR). AB0 mismatch liver transplantation is controversial especially in non-urgent situations and in living donor liver transplantation. We investigate a new protocol consisting on antigen-specific immunoadsorption (ASIAD) using Glycosorb® AB device that remove only blood group antibodies, associated with daclizumab and mycophenolate mofetil induction without splenectomy in living donor liver transplantation in adults (AALDLT). Methods: Between January and April 2003, 2 out of 10 recipients underwent AB0-I (A1>0; A2>B) AALDLT with the right lobe in our institution. Case 1 was a man 45 y.o. and case 2 was a man 52 y.o., underwent transplantation for multifocal HCC on HCV-related cirrhosis both. HSA was performed on preoperative day -2 and -1 and following AALDLT according to the anti-IgM/IgG titer increase. Immunosuppressive regimen consisted on daclizumab (1mg/Kg) induction on day -1; 1; 15; 30; 45; mycophenolate mofetil 2 × 1 g on day – 1; 0 and each postoperative day; tacrolimus 2 × 1 mg on day -1 and each postoperative day with doses adjusted to maintain serum through levels of 12-15 ng/dl. Conventional steroid tapering doses was administered. Results: After a median FU of 13 months, both patients are alive and well. In patient 1, preoperative anti-IgM/IgG titer before and after ASIAD fall from 128 to 4 and from 64 to 0 respectively. Five ASIAD sessions were required in total and no ACR episodes occurred. Anti IgG titers remained between 1 and 8. In patient 2, preoperative anti IgM/IgG titers fall from 4 to 0 and from 32 to 2. Seven ASIAD sessions were required. This patient experienced two episodes of ACR grade 2 on the third postoperative week and at 6 months. Anti IgG titers remained between 2 and 16. Both patients did not experienced major complications or infections episodes or CMV disease. After a FU of 15 and 12 months, a specific hemagglutinins level remains low. Conclusions: Antigen-specific immunoadsorption with Glycosorb AB® has showed high efficiency to decrease significantly hemagglutinins without any adverse event. The favourable outcome of these patients enrolled in this protocol using Daclizumab and Mycophenolate Mofetil could be explained by the induction of accommodation. This treatment may potentially expand the donor pool in AALDLT.

Heme-oxygenase-1 expression correlates with severity of acute cellular rejection in lung transplantation.

Heme-oxygenase-1 (HO-1) has been shown to play an important role in oxidative stress, and recent studies indicate that it is a graft survival protein in cardiac and liver transplant models. Our laboratory previously found HO-1 to be increased in human lung allografts with acute cellular rejection (ACR) and in active obliterative bronchiolitis. To better understand the role of HO-1 in ACR we studied the relationship between HO-1 expression and ACR in a rodent model of lung transplantation. Orthotopic left lung transplantation was performed from Lewis (donor) to Sprague-Dawley (recipient) rats, and ACR (Grade A0 to A4) was evaluated at days 3, 5, and 7. HO-1 expression was assessed by immunohistochemistry and Western analysis, and compared with the degree of ACR. Myeloperoxidase staining was evaluated as an indirect measure of oxidant stress. Donors and recipients were also treated with either an inhibitor of HO activity, tin protoporphyrin or an inducer, cobalt protoporphyrin, and the severity of ACR was compared with that in untreated allografts. HO-1 expression was elevated in transplanted versus native lungs or isografts, and the degree of elevation was closely correlated with ACR grade (p < 0.001). Similarly, myeloperoxidase expression increased with time and severity of ACR. Administration of the metalloporphyrins, tin protoporphyrin and cobalt protoporphyrin, produced no significant difference in the degree of ACR, but did alter the severity of ischemia-reperfusion injury. Similar to what occurs in human lung transplantation, HO-1 expression is increased in a rodent lung transplant model of ACR and correlates with the severity of rejection. Altering its expression does not appear to affect the degree of ACR.

Acute cellular rejection grading scheme for human gastric allografts

The control of acute cellular rejection (ACR) in multivisceral transplantation improves long-term survival, but monitoring this process can be challenging because different allografts can display varying forms and degrees of rejection. Criteria for ACR of small bowel and liver have been established, but a systematic analysis for ACR in stomach is lacking. For this reason we have developed a comprehensive grading scheme for the evaluation of gastric allograft rejection. The grading scheme was designed to individually grade a variety of changes in the surface epithelium, lamina propria, and glandular structures. The individual values are cumulated, and the final score determines assignment of the rejection grade. The ACR grades range from no evidence of acute cellular rejection to severe rejection. We performed a retrospective study based on 70 gastric allograft biopsies from 20 patients who received multivisceral transplantation from 1995 to 2001. We found that the scoring system showed no significant interobserver variability and allowed for an accurate designation of the ACR grade to the gastric allografts. We found with this grading system that neither clinical symptoms nor gastric endoscopic findings could serve as specific indicators of gastric ACR. Our results also showed that there were differences in the occurrence and intensity of acute rejection between the stomach and other transplanted organs, suggesting that ACR can occur independently among different allografts of the same host. In conclusion, we find that this scheme for grading ACR in gastric transplants is objective and reproducible. This grading system will likely allow for improved correlation between gastric ACR grade and clinical symptoms, as well as improve interobserver uniformity within and between institutions.

Trends in serum citrulline and acute rejection among recipients of small bowel transplants

A test for detecting acute cellular rejection (ACR) of small intestinal transplants (ITx) would be a major advance. Small preliminary studies suggest that serum citrulline levels correlate with ACR. The results for a group of 26 isolated intestinal and multivisceral transplant recipients are summarized here. Serum citrulline concentrations were determined by ion exchange chromatography and compared to biopsy-based grade of ACR. Other factors considered included patient and donor age and sex, ischemia time, and serum creatinine. Straight-line fits were employed to describe how each patient's citrulline levels changed over time. Estimated times to achieve normal citrulline (≥30 μmol/L) ranged from 1 to 730 days posttransplant for 21 patients demonstrating increasing citrulline levels over time. Using stepwise linear regression, patients' ranks for time required to achieve normal citrulline levels were the only independent predictors of both maximum ACR ( P < .0001) and average ACR ( P = .0059) after 14 days posttransplant. The rate and direction of change in citrulline over time may be an indicator of the risk of acute rejection. We plan to further examine the use of citrulline as a marker for rejection in larger prospective studies.