Stereological assessment of the blood-air barrier and the surfactant system after mesenchymal stem cell pretreatment in a porcine non-heart-beating donor model for lung transplantation.

More frequent utilization of non-heart-beating donor (NHBD) organs for lung transplantation has the potential to relieve the shortage of donor organs. In particular with respect to uncontrolled NHBD, concerns exist regarding the risk of ischaemia/reperfusion (IR) injury-related graft damage or dysfunction. Due to their immunomodulating and tissue-remodelling properties, bone-marrow-derived mesenchymal stem cells (MSCs) have been suspected of playing a beneficial role regarding short- and long-term survival and function of the allograft. Thus, MSC administration might represent a promising pretreatment strategy for NHBD organs. To study the initial effects of warm ischaemia and MSC application, a large animal lung transplantation model was generated, and the structural organ composition of the transplanted lungs was analysed stereologically with particular respect to the blood-gas barrier and the surfactant system. In this study, porcine lungs (n = 5/group) were analysed. Group 1 was the sham-operated control group. In pigs of groups 2-4, cardiac arrest was induced, followed by a period of 3 h of ventilated ischaemia at room temperature. In groups 3 and 4, 50 × 106 MSCs were administered intravascularly via the pulmonary artery and endobronchially, respectively, during the last 10 min of ischaemia. The left lungs were transplanted, followed by a reperfusion period of 4 h. Then, lungs were perfusion-fixed and processed for light and electron microscopy. Samples were analysed stereologically for IR injury-related structural parameters, including volume densities and absolute volumes of parenchyma components, alveolar septum components, intra-alveolar oedema, and the intracellular and intra-alveolar surfactant pool. Additionally, the volume-weighted mean volume of lamellar bodies (lbs) and their profile size distribution were determined. Three hours of ventilated warm ischaemia was tolerated without eliciting histological or ultrastructural signs of IR injury, as revealed by qualitative and quantitative assessment. However, warm ischaemia influenced the surfactant system. The volume-weighted mean volume of lbs was reduced significantly (P = 0.024) in groups subjected to ischaemia (group medians of groups 2-4: 0.180-0.373 µm³) compared with the sham control group (median 0.814 µm³). This was due to a lower number of large lb profiles (size classes 5-15). In contrast, the intra-alveolar surfactant system was not altered significantly. No significant differences were encountered comparing ischaemia alone (group 2) or ischaemia plus application of MSCs (groups 3 and 4) in this short-term model.

Mesenchymal stem cell pretreatment of non-heart-beating-donors in experimental lung transplantation.

BACKGROUND: Lung transplantation (LTx) is still limited by organ shortage. To expand the donor pool, lung retrieval from non-heart-beating donors (NHBD) was introduced into clinical practice recently. However, primary graft dysfunction with inactivation of endogenous surfactant due to ischemia/reperfusion-injury is a major cause of early mortality. Furthermore, donor-derived human mesenchymal stem cell (hMSC) expansion and fibrotic differentiation in the allograft results in bronchiolitis obliterans syndrome (BOS), a leading cause of post-LTx long-term mortality. Therefore, pretreatment of NHBD with recipient-specific bone-marrow-(BM)-derived hMSC might have the potential to both improve the postischemic allograft function and influence the long-term development of BOS by the numerous paracrine, immunomodulating and tissue-remodeling properties especially on type-II-pneumocytes of hMSC. METHODS: Asystolic pigs (n = 5/group) were ventilated for 3 h of warm ischemia (groups 2-4). 50x106 mesenchymal-stem-cells (MSC) were administered in the pulmonary artery (group 3) or nebulized endobronchially (group 4) before lung preservation. Following left-lung-transplantation, grafts were reperfused, pulmonary-vascular-resistance (PVR), oxygenation and dynamic-lung-compliance (DLC) were monitored and compared to control-lungs (group 2) and sham-controls (group 1). To prove and localize hMSC in the lung, cryosections were counter-stained. Intra-alveolar edema was determined stereologically. Statistics comprised ANOVA with repeated measurements. RESULTS: Oxygenation (p = 0.001) and PVR (p = 0.009) following endovascular application of hMSC were significantly inferior compared to Sham controls, whereas DLC was significantly higher in endobronchially pretreated lungs (p = 0.045) with overall sham-comparable outcome regarding oxygenation and PVR. Stereology revealed low intrapulmonary edema in all groups (p > 0.05). In cryosections of both unreperfused and reperfused grafts, hMSC were localized in vessels of alveolar septa (endovascular application) and alveolar lumen (endobronchial application), respectively. CONCLUSIONS: Preischemic deposition of hMSC in donor lungs is feasible and effective, and endobronchial application is associated with significantly better DLC as compared to sham controls. In contrast, transvascular hMSC delivery results in inferior oxygenation and PVR. In the long term perspective, due to immunomodulatory, paracrine and tissue-remodeling effects on epithelial and endothelial restitution, an endobronchial NHBD allograft-pretreatment with autologous mesenchymal-stem-cells to attenuate limiting bronchiolitis-obliterans-syndrome in the long-term perspective might be promising in clinical lung transplantation. Subsequent work with chronic experiments is initiated to further elucidate this important field.