A cross-circulatory platform for monitoring innate allo-responses in lung grafts

Abstract

Lung transplantation is the only curative option for end-stage chronic respiratory diseases. However the survival rate is only about 50% at 5 years. Although experimental evidences have shown that innate allo-responses impact on the clinical outcome, the knowledge of the involved mechanisms involved is limited. We established a cross-circulatory platform to monitor the early recruitment and activation of immune cells in an extracorporeal donor lung by coupling blood perfusion to cell mapping with a fluorescent marker in the pig, a commonly-used species for lung transplantation. The perfusing pig cells were easily detectable in lung cell suspensions, in broncho-alveolar lavages and in different areas of lung sections, indicating infiltration of the organ. Myeloid cells (granulocytes and monocytic cells) were the dominant recruited subsets. Between 6 and 10 h of perfusion, recruited monocytic cells presented a strong upregulation of MHC class II and CD80/86 expression, whereas alveolar macrophages and donor monocytic cells showed no significant modulation of expression. This cross-circulation model allowed us to monitor the initial encounter between perfusing cells and the lung graft, in an easy, rapid, and controllable manner, to generate robust information on innate response and test targeted therapies for improvement of lung transplantation outcome.

Differential early response of monocyte/macrophage subsets to intra-operative corticosteroid administration in lung transplantation

“…The lung harvest from donor pigs ( n = 10) was performed in the Animal Surgery and Medical Imaging Platform (CIMA-MIMA2-BREED-INRAE, Jouy en Josas, France) as described in a previous study ( 19 ). The ischemic durations undergone by the donor lungs are provided in Supplemental Table 1 for the UT and CST groups (warm ischemia duration: 88 ± 5.7 min for UT and 79.6 ± 11.8 min for CST group, cold ischemia duration: 107.6 ± 18.6 min for UT and 100.2 ± 12.85 min for CST group).…”

“…The overall cross-circulation procedure is depicted in Supplemental Figure 1 . Recipient pigs ( n = 10) were anesthetized as described ( 19 ). Septotryl® (0.08 ml/kg) (Vetoquinol) was injected i.m.…”

“…After a 25,000 U heparin bolus, the recipient pig was maintained on a continuous heparin infusion (100 U/Kg/h). The superior vena cava was cannulated in the recipient pig with a 20 F double lumen cannula as described ( 19 ). Carboxyfluorescein succinimidyl ester (CFSE, 25 mg, Sigma-Aldrich, Saint-Louis, MS, USA) was administered i.v.…”

“…Furthermore, rodents are not considered reliable preclinical models when examining immunomodulatory drugs in transplantation ( 18 ). We recently established a cross-circulation platform of extracorporeal donor lung coupled to cell mapping in the pig ( 19 ). We showed that this model presents several advantages over classical LT to study the cellular and molecular events occurring upon ischemia-reperfusion and the first encounter between donor and recipient cells.…”

“…Finally, an intravenous injection of CFSE that labels the whole leukocyte compartment of the perfusing pig allows to distinguish the pig donor cells from the perfusing pig cells and to take into account the regulatory systemic signaling of a complete host, which would not be the case with a classical ex vivo lung perfusion (EVLP) using whole blood. Thanks to this model, we showed that PMNs and MoCs are the dominantly recruited populations from the pig host in the first 10 h and that the recruited MoCs upregulate the MHC class II and CD80/86 antigen-presentation molecules at their cell surface ( 19 ). Here we used this cross-circulatory platform to study the effect of a CS pulse -similar to the one given upon surgery- on the innate allogeneic cell activation, and in particular on AMs and recruited classical (CD14 pos ) and non-classical/intermediate (CD16 pos ) MoCs, the latter upregulating MHC class II and CD80/86 upon reperfusion.…”