Tiberio Oggionni

Regulatory CD4+CD25+ T cells in the peripheral blood of lung transplant recipients: correlation with transplant outcome.

Background. The subset of CD4+CD25+ regulatory T cells, recently identified in humans, may play a central role in the regulation of immune tolerance to graft survival. Methods. This study assesses the frequency and functional profile of CD4+CD25+CD69− cells in the peripheral blood of lung transplant recipients (>3 years from transplantation), 10 of whom were in a stable clinical condition and 11 of whom demonstrated chronic rejection (bronchiolitis obliterans syndrome). We also studied a group of seven healthy subjects. Results. The frequency of CD4+ T cells expressing CD25 (CD4+CD25+) and the highest levels (CD25high) were lower in patients with bronchiolitis obliterans syndrome compared with healthy subjects and subjects in a stable clinical condition (P ≤0.01). Purified CD4+CD25+ cells exhibited a regulatory functional profile in vitro: they were hyporesponsive, suppressed the proliferation of CD4+CD25− cells, and produced interleukin-10. Conclusion. These results provide in vivo evidence that peripheral CD4+CD25+ T cells may represent an important regulatory subset in lung transplantation.

Impact of human metapneumovirus and human cytomegalovirus versus other respiratory viruses on the lower respiratory tract infections of lung transplant recipients

Viral respiratory tract infections in lung transplant recipients may be severe. During three consecutive winter‐spring seasons, 49 symptomatic lung transplant recipients with suspected respiratory viral infection, and 26 asymptomatic patients were investigated for presence of respiratory viruses either in 56 nasopharyngeal aspirate or 72 bronchoalveolar lavage samples taken at different times after transplantation. On the whole, 1 asymptomatic (3.4%) and 28 symptomatic (57.1%) patients were positive for human metapneumovirus (hMPV, 4 patients), influenza virus A (3 patients), and B (2 patients), respiratory syncytial virus (2 patients), human coronavirus (2 patients), human parainfluenza virus (2 patients), rhinovirus (5 patients), while 4 patients were coinfected by 2 respiratory viruses, and 5 were infected sequentially by 2 or more respiratory viruses. In bronchoalveolar lavage samples, hMPV predominated by far over the other viruses, being responsible for 60% of positive specimens, whereas other viruses were present in nasopharyngeal aspirates at a comparable rate. RT‐PCR (detecting 43 positive samples/128 examined) was largely superior to monoclonal antibodies (detecting 17 positive samples only). In addition, HCMV was detected in association with a respiratory virus in 4/18 HCMV‐positive patients, and was found at a high concentration (>105 DNA copies/ml) in 3/16 (18.7%) patients with HCMV‐positive bronchoalveolar lavage samples and pneumonia. Coinfections and sequential infections by HCMV and respiratory viruses were significantly more frequent in patients with acute rejection and steroid treatment. In conclusion: (i) about 50% of respiratory tract infections of lung transplant recipients were associated with one or more respiratory viruses; (ii) hMPV largely predominates in bronchoalveolar lavage of symptomatic lung transplant recipients, thus suggesting a causative role in lower respiratory tract infections; (iii) RT‐PCR appears to be the method of choice for detection of respiratory viruses in lung transplant recipients, (iv) a high HCMV load in bronchoalveolar lavage is a risk factor for viral pneumonia, suggesting some measure of intervention for the control of viral infection. J. Med. Virol. 78:408–416, 2006.

In situ assessment of oxidant and nitrogenic stress in bleomycin pulmonary fibrosis

Reactive oxygen species (ROS) and nitric oxide (NO) have a role in the development of pulmonary fibrosis after bleomycin administration. The ROS production induces an antioxidant response, involving superoxide dismutases (SODs), catalase, and glutathione peroxidases. We compared in situ oxidative burden and antioxidant enzyme activity in bleomycin-injured rat lungs and normal controls. ROS expression and catalase, glucose-6-phosphate-dehydrogenase (G6PHD), and NOS/NADPH-diaphorase activity were investigated by using histochemical reactions. Nitric oxide synthase (e-NOS and i-NOS) and SOD (MnSOD, Cu/ZnSOD, ECSOD) expression was investigated immunohistochemically. After treatment ROS production was enhanced in both phagocytes and in type II alveolar epithelial cells. Mn, Cu/Zn, and ECSOD were overexpressed in parenchymal cells, whereas interstitium expressed ECSOD. Catalase and G6PHD activity was moderately increased in parenchymal and inflammatory cells. NOS/NADPH-d activity and i-NOS expression increased in alveolar and bronchiolar epithelia and in inflammatory cells. It can be suggested that the concomitant activation of antioxidant enzymes is not adequate to scavenge the oxidant burden induced by bleomycin lung damage. Inflammatory cells and also epithelial cells are responsible of ROS and NO production. This oxidative and nitrosative stress may be a substantial trigger in TGF-β1 overexpression by activated type II pneumocytes, leading to fibrotic lesions.

Efficacy of tacrolimus rescue therapy in refractory acute rejection after lung transplantation

BACKGROUND Encouraging results in transplantation of other solid organs led to investigation of the use of tacrolimus in lung transplantation as a salvage immunosuppressant in persistent acute rejection. METHODS The incidence and severity of acute rejection and the number of steroid pulses were analyzed in 20 lung recipients who were converted from a cyclosporine- to a tacrolimus-based immunosuppressive regimen because of refractory biopsy-proven acute rejection. RESULTS Tacrolimus was started 12.0 +/- 13.0 months after transplantation, and the mean follow-up was 25.0 +/- 13.7 months. After shifting to tacrolimus, a significant decline was observed in both the number of acute rejections per patient (3.0 +/- 1.56 to 0.85 +/- 1.14, p < 0.0001), and the incidence of acute rejection per 100 patient-days (1.52 +/- 0.99 to 0.14 +/- 0.21, p < 0.0001). Furthermore, the average histologic grade of rejection decreased from 1.9 +/- 0.8 to 0.4 +/- 0.5 (p < 0.0001). Methylprednisolone pulses similarly decreased from 1.9 +/- 1.3/patient to 0.3 +/- 0.7/patient (p < 0.0001). During cyclosporine immunosuppression, the mean forced expiratory volume in 1 second decreased to 84.4% +/- 13.3% of individual best value. The average lung function parameters were stable 3 months after the change of medication, and then began to improve. After an average follow-up of 36.5 +/- 19.2 months, 2 patients have developed bronchiolitis obliterans syndrome (one has Stage 1 and one has Stage 3). CONCLUSION Conversion to a tacrolimus-based immunosuppressive regimen for refractory acute lung rejection is associated with reduced incidence and severity of acute rejection episodes, steroid sparing, and stabilization or improvement of pulmonary function.

Return to work after thoracic organ transplantation in a clinically-stable population

To evaluate the rate of return to work after transplantation and its determinants in a clinically‐stable population of patients transplanted and followed‐up at a single institution in Italy.

Incomplete expression of epithelial–mesenchymal transition markers in idiopathic pulmonary fibrosis

The hypothesis that epithelial-mesenchymal transition (EMT) contributes to the formation of fibroblast foci (FF), which are the histological hallmark and the site of active disease progression of Idiopathic Pulmonary Fibrosis (IPF), has not yet received a conclusive demonstration. Cells undergoing EMT lose epithelial features and acquire mesenchymal markers and morphology. Cadherin expression switch (from E to N) is one of the first events in EMT. We investigated the immunohistochemical expression of E- and N-cadherin, vimentin, fibronectin, laminin-5-γ2, α-smooth muscle actin, and fibroblast-specific protein-1 involved in EMT in 20 IPF lung biopsies, focusing on metaplastic squamous cells of bronchial basal origin, positive for laminin-5-γ2 and ΔNp63/p40, that cover FF. The results were compared with organizing pneumonia, reactive squamous cell metaplasia of bronchiolar epithelia, and squamous cell carcinoma. Bronchiolar basal metaplastic cells in IPF partially lost E-cadherin and expressed vimentin and fibronectin. Hyperplastic pneumocytes in IPF and controls coexpressed E-cadherin and N-cadherin, and were weakly positive for lam5-γ2. Reactive squamous cell metaplasia did not show any mesenchymal markers. Squamous cell carcinoma only expressed lam5-γ2. In IPF lungs, we observed two epithelial cell populations with a different expression profile of markers involved in EMT. Although neither hyperplastic pneumocytes nor bronchial basal cells showed evidence of complete EMT, only the latter seem to be specific for UIP and might have a role in its development.

Risk Factors for Chronic Renal Dysfunction in Lung Transplant Recipients

Several factors predispose to renal dysfunction (RD), a common complication of solid organ transplants. We evaluated the impact of clinical and laboratory parameters on the decline of renal function in lung and heart-lung transplant recipients. We enrolled 45 patients who survived more than 6 months after transplantation, had normal renal function and urinalysis before the surgery. The prognostic value of variables for the occurrence of RD was calculated by univariate analysis. Thirty patients developed RD, defined as doubling of serum creatinine or creatinine steadily >1.5 mg/dL after a median time of 12 months. Serum creatinine above 0.9 mg/dL during the month preceding lung transplant, systolic blood pressure above 130 mmHg, and pretransplant idiopathic pulmonary hypertension were significantly associated with the development of RD. Our findings indicate that increased systolic blood pressure, reduced glomerular filtration rate, and idiopathic pulmonary hypertension are risk factors for chronic RD in lung transplant recipients.

Time course of matrix metalloproteases and tissue inhibitors in bleomycin-induced pulmonary fibrosis

To investigate simultaneously localization and relative activity of MMPs during extracellular matrix (ECM) remodeling in bleomycin-induced pulmonary fibrosis in rat, we analyzed the time course of the expression, activity and/or concentration of gelatinases MMP-2 and MMP-9, collagenase MMP-1, matrylisin MMP-7, TIMP-1 and TIMP-2, both in alveolar space (cellular and extracellular compartments) and in lung tissue. MMP and TIMP expression was detected (immunohistochemistry) in lung tissue. MMP activity (zymography) and TIMP concentration (ELISA) were evaluated in lung tissue homogenate (LTH), BAL supernatant (BALs) and BAL cell pellet (BALp) 3, 7, 14, and 28 days after bleomycin intratracheal instillation. Immunohistochemistry showed an extensive MMP and TIMP expression from day 7 in a wide range of structural and inflammatory cells in treated rats. MMP-2 was present mainly in epithelia, MMP-9 in inflammatory cells. MMP-2 and MMP-9 activity was increased respectively in BAL fluid and BAL cells, with a peak at day 7. TIMP-1 and TIMP-2 concentration (ELISA) enhancement was delayed at day 14. In conclusion gelatinases and their inhibitors are significantly activated during bleomycin-induced pulmonary fibrosis. Marked changes in gelatinases activity are observed early in the alveolar compartment, with a prevailing extracellular activity of MMP-2 and a predominant intracellular distribution of MMP-9, while enzyme activity changes in lung parenchyma were less evident. In the repairing phase the reduction of gelatinases activity is synchronous with a peak of alveolar concentration of their inhibitors.

Post-transplant lymphoproliferative disorders and Epstein-Barr virus DNAemia in a cohort of lung transplant recipients

BackgroundPost-transplant lymphoproliferative disorders (PTLD) are serious complications in lung transplant recipients. No consensus on EBV DNAemia levels predictive of PTLD has been reached. In addition, in many instances EBV DNAemia is determined in patients with suggestive symptoms only.MethodsThe characteristics of five patients with PTLD as well as the prevalence of EBV DNAmia in a cohort of 137 consecutive patients receiving lung transplantation are described.ResultsTwenty-six out of 137 patients (18.9%) were excluded from the analysis because lost at follow-up or dead from PTLD-independent reasons within three months of transplantation. EBV DNA in peripheral blood mononuclear cells (PBMC) was determined in 83/111 patients (74.8%) because of potential PTLD-related symptoms, while 28 patients (25.2%) showed no symptoms and were not examined. EBV DNAemia was positive in 53/83 patients (63.8%), and negative in 30/83 patients (36.2%). PTLD was diagnosed in five (4.5%) patients at a median time of 270 (range 120-870) days following transplantation. All five PTLD (three large B-cell lymphomas, one Hodgkin lymphoma and one possible pre-neoplastic lesion) were potentially associated with EBV infection. However, only 3/5 patients with PTLD had detectable EBV DNAemia: < 1,000 copies EBV DNA/1 × 105 PBMC in one patient and > 1,000 copies EBV DNA/1 × 105 PBMC in two patients.ConclusionA systematic multidisciplinary (clinical, radiologic, virologic and histologic) approach is mandatory for the diagnosis and management of PTLD in lung transplant recipients, while monitoring of symptomatic patients only may provide an incomplete or late picture of the clinical problem. In addition, staining for EBV antigens and quantification of EBV DNA in biopsy specimens should always be performed to understand the role of EBV infection in the pathogenesis of PTLD.

Factors Influencing Oxidative Imbalance in Pulmonary Fibrosis: An Immunohistochemical Study

Background. Idiopathic Pulmonary Fibrosis (IPF) is a fatal lung disease of unknown etiology characterized by interstitial fibrosis determining irreversible distortion of pulmonary architecture. Reactive oxygen species (ROS) and markers of oxidative stress play a pivotal role in human IPF pathology, possibly through induction of epithelial-mesenchymal transition (EMT). Methods. We investigated by immunohistochemistry, in UIP and COP tissue samples, the expression of most relevant markers of the molecular interplay involving RAGE, oxidant/antioxidant balance regulation, tissue nitrosylation, and mediators of EMT. Results. In both UIP and COP, the degree of RAGE expression was similarly high, while SODs and i-NOS, diffusely present in COP endoalveolar plugs, were almost absent in UIP fibroblast foci. A lower degree of tissue nitrosilation was observed in UIP than in COP. Conclusions. Fibroblast lesions of UIP and of COP share a similar degree of activation of RAGE, while antioxidant enzyme expression markedly reduced in UIP.