L-alanyl-L-glutamine Improves Lung Performance during Ex Vivo Lung Perfusion: From Cellular Mechanism to Porcine Donor Lungs

Abstract

Ex vivo lung perfusion (EVLP) is an established technology to assess marginal donor lungs. EVLP application has increased the use of donor lungs and is being further developed as a platform for the repair of lung injury. Steen solution is one of the most widely used EVLP perfusate solutions. Currently, clinical EVLP is generally run for 4-6 h, and most of porcine lung EVLP studies were limited to around 12 h. We hypothesize that adding nutrients to Steen solution may improve cellular function for better performance. Human pulmonary microvascular endothelial cells (HPMEC) and human lung epithelial cells (BEAS-2B) were used to determine the effects of Steen solution components on basic cellular function. Cell confluence, apoptosis and migration were visualized and quantified by IncuCyte® Live Cell Analysis System in real-time. A simulated EVLP cell culture model was established by replacing regular culture medium with cold LPD preservation solution and followed by "perfusion" with normothermic Steen solution or its modifications. Cell apoptosis was quantified. Porcine lung EVLP was performed using Steen solution with added nutrients. Cells exposed to Steen solution exhibited reduced cell confluence, increased apoptosis and suppressed cell migration compared to DMEM or DMEM+10% FBS. In an examination of adding various nutrients to Steen solution on cell function, L-alanyl-L-glutamine (gln) significantly inhibited cell apoptosis and improved cell migration (Fig A). In the simulated EVLP cell culture setting, adding gln to Steen solution significantly reduced apoptosis. In pig lung EVLP, the addition of gln to Steen solution significantly extend the period of stable lung function (Fig B). Glutamine is the most abundant free amino acid in the body with multiple cytoprotective functions. Adding L-alanyl-L-glutamine to EVLP perfusate may improve the stability and function of lungs being evaluated and treated with EVLP.