Marialuisa Valente

Banff 07 Classification of Renal Allograft Pathology: Updates and Future Directions

The 9th Banff Conference on Allograft Pathology was held in La Coruna, Spain on June 23–29, 2007. A total of 235 pathologists, clinicians and scientists met to address unsolved issues in transplantation and adapt the Banff schema for renal allograft rejection in response to emerging data and technologies. The outcome of the consensus discussions on renal pathology is provided in this article. Major updates from the 2007 Banff Conference were: inclusion of peritubular capillaritis grading, C4d scoring, interpretation of C4d deposition without morphological evidence of active rejection, application of the Banff criteria to zero‐time and protocol biopsies and introduction of a new scoring for total interstitial inflammation (ti‐score). In addition, emerging research data led to the establishment of collaborative working groups addressing issues like isolated ‘v’ lesion and incorporation of omics‐technologies, paving the way for future combination of graft biopsy and molecular parameters within the Banff process.

Arrhythmogenic Right Ventricular Cardiomyopathy

There have been major advances in recent years in the clinical setting of arrhythmogenic right ventricular cardiomyopathy, including new diagnostic criteria, a changing spectrum of the disease with even left dominant forms, the role of cardiac magnetic resonance and electroanatomic mapping, the expanding use of genetic screening and the existence of overlapping phenotypes. Moreover, early diagnosis at pre-participation screening with sports disqualification and risk stratification for the indication of ICD have been shown to be life-saving. In addition to traditional therapies targeting arrhythmias and congestive heart failure, an effective treatment of the disease could be based on the discovery of the molecular mechanisms involved in the pathobiology of the disease in order to block the onset and progression of cell death.ZusammenfassungDiagnostik und Management der arrhythmogenen rechtsventrikulären Kardiomyopathie (ARVC) wurden in den letzten Jahren erheblich verbessert. Dies betrifft die Präzisierung diagnostischer Kriterien, die Erweiterung des Krankheitsspektrums auf überwiegend linksventrikuläre Formen, die das klinische Bild dominieren können, den diagnostischen Beitrag der kardialen Magnetresonanztomograpie und des elektroanatomischen Mappings, die Umsetzung des genetischen Screenings in die Praxis und die Beobachtung, dass der klinische Phänotyp Überlappungen zu anderen Kardiomyopathieformen aufweisen kann. Die Früherkennung durch Screeninguntersuchung vor Wettkämpfen mit Ausschluss des Sportlers vom Wettkampfsport und ggf. prophylaktischer Defibrillatorbehandlung (ICD), je nach individueller Risikoeinschätzung, ist nachweislich lebensrettend. Neben der traditionellen Behandlung der Arrhythmie und der Herzinsuffizienz sollte eine zukünftige Behandlung auf diejenigen molekularen Mechanismen in der Pathobiologie der ARVC zielen, die die Auslösung oder Progression des myokardialen Zelltods verursachen.

Mutations in Desmoglein-2 Gene Are Associated With Arrhythmogenic Right Ventricular Cardiomyopathy

Background— Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiomyopathy characterized by progressive myocardial atrophy with fibrofatty replacement. The recent identification of causative mutations in plakoglobin, desmoplakin (DSP), and plakophilin-2 (PKP2) genes led to the hypothesis that ARVC is due to desmosomal defects. Therefore, desmoglein-2 (DSG2), the only desmoglein isoform expressed in cardiac myocytes, was screened in subjects with ARVC. Methods and Results— In a series of 80 unrelated ARVC probands, 26 carried a mutation in DSP (16%), PKP2 (14%), and transforming growth factor-&bgr;3 (2.5%) genes; the remaining 54 were screened for DSG2 mutations by denaturing high-performance liquid chromatography and direct sequencing. Nine heterozygous DSG2 mutations (5 missense, 2 insertion-deletions, 1 nonsense, and 1 splice site mutation) were detected in 8 probands (10%). All probands fulfilled task force criteria for ARVC. An endomyocardial biopsy was obtained in 5, showing extensive loss of myocytes with fibrofatty tissue replacement. In 3 patients, electron microscopy investigation was performed, showing intercalated disc paleness, decreased desmosome number, and intercellular gap widening. Conclusions— This is the first investigation demonstrating DSG2 gene mutations in a significant number of ARVC-unrelated probands. Cardiac phenotype is characterized clinically by typical ARVC features with frequent left ventricular involvement and morphologically by fibrofatty myocardial replacement and desmosomal remodeling. The presence of mutations in desmosomal encoding genes in 40% of cases confirms that many forms of ARVC are due to alterations in the desmosome complex.

COQ2 Nephropathy: A Newly Described Inherited Mitochondriopathy with Primary Renal Involvement

Primary coenzyme Q(10) (CoQ(10)) deficiency includes a group of rare autosomal recessive disorders primarily characterized by neurological and muscular symptoms. Rarely, glomerular involvement has been reported. The COQ2 gene encodes the para-hydroxybenzoate-polyprenyl-transferase enzyme of the CoQ(10) synthesis pathway. We identified two patients with early-onset glomerular lesions that harbored mutations in the COQ2 gene. The first patient presented with steroid-resistant nephrotic syndrome at the age of 18 months as a result of collapsing glomerulopathy, with no extrarenal symptoms. The second patient presented at five days of life with oliguria, had severe extracapillary proliferation on renal biopsy, rapidly developed end-stage renal disease, and died at the age of 6 months after a course complicated by progressive epileptic encephalopathy. Ultrastructural examination of renal specimens from these cases, as well as from two previously reported patients, showed an increased number of dysmorphic mitochondria in glomerular cells. Biochemical analyses demonstrated decreased activities of respiratory chain complexes [II+III] and decreased CoQ(10) concentrations in skeletal muscle and renal cortex. In conclusion, we suggest that inherited COQ2 mutations cause a primary glomerular disease with renal lesions that vary in severity and are not necessarily associated with neurological signs. COQ2 nephropathy should be suspected when electron microscopy shows an increased number of abnormal mitochondria in podocytes and other glomerular cells.

CXCR3 and Its Ligand CXCL10 Are Expressed by Inflammatory Cells Infiltrating Lung Allografts and Mediate Chemotaxis of T Cells at Sites of Rejection

The attraction of T lymphocytes into the pulmonary parenchyma represents an essential step in mechanisms ultimately leading to lung allograft rejection. In this study we evaluated whether IP-10 (CXCL10), a chemokine that is induced by interferon-gamma and stimulates the directional migration of activated T cells, plays a role in regulating the trafficking of effector T cells during lung allograft rejection episodes. Immunohistochemical examination showed that areas characterized by acute cellular rejection (grades 1 to 4) and active obliterative bronchiolitis (chronic rejection, Ca) were infiltrated by T cells expressing CXCR3, i.e., the specific receptor for CXCL10. In parallel, T cells accumulating in the bronchoalveolar lavage of lung transplant recipients with rejection episodes were CXCR3+ and exhibited a strong in vitro migratory capability in response to CXCL10. In lung biopsies, CXCL10 was abundantly expressed by graft-infiltrating macrophages and occasionally by epithelial cells. Alveolar macrophages expressed and secreted definite levels of CXCL10 capable of inducing chemotaxis of the CXCR3+ T-cell line 300-19; the secretory capability of alveolar macrophages was up-regulated by preincubation with interferon-gamma. Interestingly, striking levels of CXCR3 ligands could be demonstrated in the fluid component of the bronchoalveolar lavage in individuals with rejection episodes. These data indicate the role of the CXCR3/CXCL10 interactions in the recruitment of lymphocytes at sites of lung rejection and provide a rationale for the use of agents that block the CXCR3/CXCL10 axis in the treatment of lung allograft rejection.

Surgical Pathology of primary cardiac and pericardial tumors

OBJECTIVE Retrospective study of surgical pathology experience on cardiac and pericardial tumors at the University of Padua in the era of immunohistochemistry and endomyocardial biopsy. METHODS In the period 1970-1995, we studied 125 bioptic primary neoplasms: specimens were obtained from surgical resection in 116 cases, heart transplantation in 3, pericardiectomy in 3, endomyocardial biopsy in 2 and thoracoscopic biopsy in 1. Tumor histotype was established by light microscopy and more recently by immuno-histochemistry, using a large panel of antibodies, through avidin-biotin peroxidase method, against factor VIII-related antigen, ulex-europaeus, desmin, myoglobin, muscle-specific actin, smooth muscle-specific actin, vimentin, cytokeratins, leukocytic common antigen, neurofilaments and S100-protein. RESULTS One hundred and thirteen were benign neoplasms: myxoma was the most frequent (87 cases) followed by pericardial cyst (8), endocardial papilloma (5), fibroma (3), rhabdomyoma (3), hematic cyst (2), teratoma (2), hemangioma (1), celothelioma (1) and lipoma (1). Malignancy was diagnosed in 12 cases, and consisted of pericardial mesothelioma (3), myxosarcoma (3), angiosarcoma (2), fibrosarcoma (2) and leiomyosarcoma (2); 4 of them were intracavitary atrial masses and were supposed to be atrial myxoma on the clinical ground. Differential diagnosis included endocardial thrombosis (10), metastasis of concealed extracardiac tumors (5), echinococcosis (3), and Loefflers fibroplastic endocarditis (3). In 4 cases, cardiac mass histotype was defined without thoracotomy, through endomyocardial (3) and thoracoscopic (1) biopsy. CONCLUSIONS A large spectrum of cardiac tumors is observed in the surgical pathology practice. Although the diagnosis of cardiac masses is easily attainable by routine imaging techniques, differential diagnosis between primary and secondary tumors, malignant and benign forms, and non neoplastic masses, is achievable only by a thorough microscopic study of surgical resections. The role of the cardiac pathologist is becoming crucial as in other fields of oncology.

Calcific degeneration as the main cause of porcine bioprosthetic valve failure.

Sixty-seven glutaraldehyde-processed porcine bioprostheses (PBs), recovered at autopsy or reoperation from 65 patients, were evaluated by roentgenologic and pathologic examination. Seven patients with 8 PBs were younger than 20 years of age. The time interval of function was 2 to 138 months (average 62). Pathologically, 53 explants had signs of intrinsic dysfunction, which was ascribed to calcification in 36 (68%). By x-ray examination, calcific deposits were found in 55 of 67 PBs (82%). The mean duration of function was 70 +/- 32 months in calcified PBs vs 27 +/- 18 months in noncalcified PBs (p less than 0.001). All 26 PBs that had been in place for longer than 6 years were calcified. In 45 PBs the Ca++ deposits were considered severe (mean time of function 76 +/- 32 months) and mild in 10 (mean time of function 44 +/- 22 months) (p less than 0.005). The Ca++ deposits were located at the commissures in 54 PBs (98%), at the body of cusps in 41 (75%), at the free margin in 37 (67%) and at the aortic wall in 37 (67%). When mild, Ca++ deposits involved the commissures in 90% of cases, the body of cusps in 30% and the free margin only in 10%. Forty-seven calcified PBs were mounted on a flexible stent, and 8 had a rigid stent, with an average time of function of 63 +/- 28 and 113 +/- 18 months, respectively (p less than 0.00001). Ca++ dysfunction occurred earlier in the aortic than in the mitral position (59 +/- 19 vs 86 +/- 35 months, p less than 0.05). All the PBs explanted from young patients and 47 of 59 PBs removed from adult patients were calcified, with an average time of function of 50 +/- 21 vs 73 +/- 33 months, respectively (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Isolated Hepatocyte Transplantation for Crigler-Najjar Syndrome Type 1:

Crigler-Najjar syndrome type 1 (CN1) is an inherited disorder characterized by the absence of hepatic uridine diphosphoglucuronate glucuronosyltransferase (UDPGT), the enzyme responsible for the conjugation and excretion of bilirubin. We performed allogenic hepatocyte transplantation (AHT) in a child with CN1, aiming to improve bilirubin glucuronidation in this condition. A 9-year-old boy with CN1 was prepared with plasmapheresis and immunosuppression with prednisolone and tacrolimus. When a graft was made available, 7.5 × 109 hepatocytes were isolated and infused into the portal vein percutaneously. After 2 weeks phenobarbitone was added to promote the enzymatic activity of UDPGT of the transplanted hepatocytes. Nocturnal phototherapy was continued throughout the studied period. Total bilirubin was considered a reliable marker of allogenic cell function. There was no significant variation of vital signs nor complications during the infusion. Mean ± SD bilirubin level was 530 ± 38 μmol/L before and 359 ± 46 μmol/L after AHT (t-test, p < 0.001). However, the introduction of phenobarbitone was followed by a drop of tacrolimus level with increase of alanine aminotransferase (ALT) and increase of bilirubin. After standard treatment of cellular rejection bilirubin fell again but from then on it was maintained at a greater level. After discharge the patient experienced a further increase of bilirubin that returned to predischarge levels after readmission to the hospital. This was interpreted as poor compliance with phototherapy. Only partial correction of clinical jaundice and the poor tolerability to nocturnal phototherapy led the parents to refuse further hepatocyte infusions and request an orthotopic liver transplant. After 24 months the child is well, with good liver function on tacrolimus and prednisolone-based immunosuppression. Isolated AHT, though effective and safe, is not sufficient to correct CN1. Maintenance of adequate immunosuppression and family compliance are the main factors hampering the success of this procedure.

Immune and Nonimmune Predictors of Cardiac Allograft Vasculopathy Onset and Severity: Multivariate Risk Factor Analysis and Role of Immunosuppression

We studied 361 patients, to evaluate risk factors for cardiac allograft vasculopathy (CAV) onset and severity/diffusion in heart transplantation (HT). Rejection scores (RS) on endomyocardial biopsy were calculated (first year and whole follow‐up). CAV onset was defined as any lesion seen at yearly angiography. A CAV severity/diffusion index was calculated for each patient summing up the scores of all lesions. Cox multivariate analysis included: donor age, sex, and weight; recipient sex, age, pre‐HT diagnosis, hypertension, diabetes and hyperlipidemia post‐HT; number of treated rejections and RS; and immunosuppressive dosage at 3, 6, and 12 months. CAV frequency was 2% at 1 year, 22% at 5 and 39% at 10 years. Risk factors for CAV onset were older donor age [p < 0.0001, relative risk (RR) = 9.9], male donor (p < 0.001, RR = 3.2), high RS for severe (≥ 3A) grades (p < 0.02, RR = 2.01), high cyclosporine at 3 months (p < 0.02, RR = 1.9). Risk factors for CAV severity/diffusion were higher donor weight (p < 0.01, RR = 7.5), high prednisone dosage at 1 year (p < 0.0001, RR = 21.1), and coronary disease pre‐HT (p < 0.002, RR = 9.7). High RS was an independent predictor for CAV onset, not severity/diffusion. This suggests an immune basis for CAV onset and nonimmune modulation for progression. High RS for severe grades may provide a predictor for patients at risk.

Marked alveolar apoptosis/proliferation imbalance in end-stage emphysema.

BackgroundApoptosis has recently been proposed to contribute to the pathogenesis of emphysema.MethodsIn order to establish if cell fate plays a role even in end-stage disease we studied 16 lungs (9 smoking-associated and 7 α1antitrypsin (AAT)-deficiency emphysema) from patients who had undergone lung transplantations. Six unused donor lungs served as controls. Apoptosis was evaluated by TUNEL analysis, single-stranded DNA laddering, electron microscopy and cell proliferation by an immunohistochemical method (MIB1). The role of the transforming growth factor (TGF)-β1 pathway was also investigated and correlated with epithelial cell turnover and with the severity of inflammatory cell infiltrate.ResultsThe apoptotic index (AI) was significantly higher in emphysematous lungs compared to the control group (p ≤ 0.01), particularly if only lungs with AAT-deficiency emphysema were considered (p ≤ 0.01 vs p = 0.09). The proliferation index was similar in patients and controls (1.9 ± 2.2 vs 1.7 ± 1.1). An increased number of T lymphocytes was observed in AAT-deficiency lungs than smoking-related cases (p ≤ 0.05). TGF-β1 expression in the alveolar wall was higher in patients with smoking-associated emphysema than in cases with AAT-deficiency emphysema (p ≤ 0.05). A positive correlation between TGF-βRII and AI was observed only in the control group (p ≤ 0.005, r2 = 0.8). A negative correlation was found between the TGF-β pathway (particularly TGF-βRII) and T lymphocytes infiltrate in smoking-related cases (p ≤ 0.05, r2 = 0.99)ConclusionOur findings suggest that apoptosis of alveolar epithelial cells plays an important role even in end-stage emphysema particularly in AAT-deficiency disease. The TGFβ-1 pathway does not seem to directly influence epithelial turnover in end-stage disease. Inflammatory cytokine different from TGF-β1 may differently orchestrate cell fate in AAT and smoking-related emphysema types.