68: Stress-induced premature senescence in airway epithelium and accelerated lung allograft aging

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Abstracts

Results: Using an A/B grading system, based on the ISHLT system for grading of pulmonary rejection, we could demonstrate that grafts from animals lacking CD26 showed a significant reduction in rejection in comparison to CD26 positive controls. Also, specific differences in leukocyte infiltration were observed. Conclusions: While an increasing rejection over time is observed in all grafts, including the CD26 negative lungs, our results suggest a significant role for CD26 in the rejection process. Thus, inhibition of CD26 may be used as a clinical tool to enhance long term survival after lung transplantation. 67 COMPARISON OF TWENTY EIGHT KEY CIRCULATING IMMUNOLOGIC MESSENGERS IN HEART TRANSPLANT RECIPIENTS TREATED WITH AZATHIOPRINE VS MYCOPHENOLATE MOFETIL N.U. Khan,1 J.E. Fildes,1 M. Al-Aloul,1 S.G. Williams,1 C.T. Leonard,1 N. Yonan,1 1The Transplant Centre, South Manchester University Hospitals NHS Trust, Wythenshawe Hospital, Manchester, United Kingdom Purpose: Azathioprine (Aza) and Mycophenolate mofetil (MMF) are major secondary immunosuppressants used in the prophylaxis and treatment of acute rejection. Several studies have identified an increased incidence of infection (including CMV activation and infection) in patients receiving MMF. The aim of this study was to compare the levels of 28 key immunologic molecules (Th1 and Th2 cytokines, chemokines and growth factors) in heart transplant recipients administered either Aza or MMF. Methods and Materials: Fourty two patients were recruited into the study. Clinical data including immunosuppressive therapy was collected from clinical notes. Peripheral blood was analysed for growth factors (EGF, FGF-b, GCSF, GM-CSF, HGF, VEGF), chemokines (eotaxin, MCP-1, MIP-1alpha, MIP-1beta, RANTES, IP-10, IL8, MIG) and cytokines (IL-1beta, IL2, IL4, IL5, IL6, IL7, IL10,, IL-12, IL-13, IL-15, IL-17, IFN-alpha, IFN-gamma, TNF alpha,) using bead based assays. Results: Thirty patients were administered MMF and 12 patients Aza. Patients in the two treatment groups did not differ in demographics, acute rejection, and CMV PCR values. We have previously identified higher expression of EGF and FGF-b in biopsy tissue of Aza treated, compared to MMF treated recipients. Using independent T test, significantly higher levels of EGF (p⫽0.001), FGF-b (p⫽0.001), GCSF (p⫽0.001), IL-12 (p⫽0.002), IL-1beta (p⫽0.001), and IL-2 (p⫽0.001) were found in the peripheral blood of Aza treated compared to MMF treated heart transplant recipients. There were no correlations between the remaining immunologic markers and secondary immunosuppression. Conclusions: The increased levels of Th1 cytokines (IL-1beta, IL-2 and IL-12) and Th1 chemokines (MCP-1, MIP-1alpha, RANTES) combined with EGF, FGF-b and GCSF in patients treated with Aza may represent the dichotomy between the incidence of infection in these two agents. On these grounds, Aza patients may be able to respond better to infection. 68 STRESS-INDUCED PREMATURE SENESCENCE IN AIRWAY EPITHELIUM AND ACCELERATED LUNG ALLOGRAFT AGING S.M. Parker,1 C. Ward,1 G. Johnson,1 F. Black,1 J.L. Lordan,1 P.A. Corris,1 G.C. Saretzki,2 A.J. Fisher,1 1Applied Immunobiology and Transplantation Research Group, University of Newcastle upon Tyne, Newcastle, United Kingdom; 2Institute for Aging and Health, University of Newcastle upon Tyne, Newcastle, United Kingdom

The Journal of Heart and Lung Transplantation February 2007

Purpose: Senescent cells lose replicative capacity yet remain biologically active with a proinflammatory phenotype. Transforming Growth Factor-beta (TGF-␤) and oxidative stress can induce senescence in cultured fibroblasts. We hypothesized that these signals may induce senescence of airway epithelial cells in the lung allograft, impair regenerative capacity, increase inflammation and thus contribute to the development of obliterative bronchiolitis (OB). Methods and Materials: Primary bronchial epithelial cells from stable lung allograft recipients (n⫽6) were treated with either hydrogen peroxide (100, 200 ␮M) for 1 hour and left to recover for up to 72 hours or TGF-␤ (10ng/ml) for 72 hours. Senescence associated beta galactosidase (SA␤Gal), p21, Ki67 and ␥H2AX immunocytochemistry were used as senescence markers. In addition, small airway biopsies from stable lung allograft recipients (n⫽10), recipients with OB (n⫽11) and normal controls (n⫽11) were stained for the cell cycle inhibitors p16 and p21. Airway epithelial staining was quantified using an image analysis system and data analysed using non parametric statistics. Results: Hydrogen peroxide caused typical senescent cell morphology changes and significant increases in SA␤Gal, p21 and ␥H2AX. Ki-67 was reduced indicating decreased cellular proliferation. TGF-␤ induced a significant increase in SA␤Gal and p21, Ki-67 was reduced. In allograft airway biopsies % epithelium positive for p16 and p21 was increased beyond that expected with normal ageing: Normal controls median p16 11%, p21 6.4%, stable recipients p16 33% p⫽0.003, p21 15% p⫽0.02, recipient with OB p16 38% p⫽0.03, p21 10.7% p⫽0.03. Conclusions: Elevated TGF-␤ and oxidative stress are associated with OB in lung allografts. We demonstrate in-vitro that these signals induce premature senescence of allograft primary airway epithelial cells. Furthermore in-vivo the lung allograft shows increased expression of p16 and p21 suggesting evidence for accelerated ageing. The relevance of this phenomenon to the development of OB requires further evaluation. 69 EXPRESSION OF GHRELIN, A NOVEL CARDIOVASCULAR HORMONE, AND ITS PEPTIDE IN THE MYOCARDIUM OF PATIENTS UNDERGOING HEART TRANSPLANTATION A. Beiras-Fernandez,1 I. Kaczmarek,1 M. Schmoeckel,1 A. Beiras,2 C. Vicol,1 B. Reichart,1 1Dept. of Cardiac Surgery, University Hospital Grosshadern; LM-University, Munich, Germany; 2Dept. of Morphological Sciences, University of Santiago de Compostela, Santiago, Spain Purpose: Ghrelin is a novel peptide hormone which acts on the pituitary and the hypothalamus to stimulate the release of growth hormone. It has been reported that administration of Ghrelin improves LV function and exercise capacity in patients with chronic heart failure. We assessed the hypothesis that the Ghrelin receptor in myocardial biopsies of patients suffering from end-stage heart failure would be up-regulated due to a decreased cellular expression of this regulatory peptide. Methods and Materials: Patients (n⫽10) undergoing heart transplantation and suffering from end-stage heart failure were included in this study. All patients gave informed consent. Biopsies (n⫽ 40) of the explanted hearts were obtained and divided according to the anatomical origin (Left and Right Ventricle; Left and Right Atrium). Expression of the peptide Ghrelin as well as its receptor was determined by means of immunohistochemistry. A control group consisting of heart muscle biopsies (n⫽8) from autopsies was designed. The histological preparations were semi-quantitative analysed by two independent investigators and image analysis software and the intensity of stain was scored from 0 to 10. The results are expressed by mean ⫾ SEM. Results: Expression of the Ghrelin receptor was significantly increased in the biopsies of patients in comparison to the control group