Cytoprotective and Antioxidant Effects of Steen Solution on Human Lung Spheroids and Human Endothelial Cells

Abstract

Respiratory diseases represent a major healthcare burden worldwide. Lung transplantation (LTx) is the "gold standard" for end-stage patients, strongly limited by shortage of available/suitable donor lungs. Normothermic ex vivo lung perfusion (EVLP) has significantly increased the number of lungs suitable for transplantation. Steen solution is used for EVLP, but the mechanisms involved in its beneficial properties remain to be clarified. We investigated the effects of Steen solution in an in vitro protocol of cold starvation and normothermic recovery on human lung spheroids, named pneumospheres (PSs), containing epithelial/basal cells, and on endothelial human umbilical vein endothelial cells (HUVEC). Steen solution significantly preserved the viability of PSs, reduced reactive oxygen species (ROS) release by PSs and HUVECs, decreased NADPH-oxidase (NOX) activity in PSs, and reduced inflammatory cytokines expression levels in HUVECs. Steen solution was able to specifically reduce NADPH oxidase 2 (NOX2) isoform activation, particularly in PSs, as detected by soluble-NOX2 peptide and p47-phosphorylation. Interestingly, a specific NOX2 inhibitor could partly mimic the pro-survival effect of Steen on PSs. We provide the first evidence that Steen solution can preserve lung epithelial/progenitor cells viability partially through NOX2 downregulation, and exert antioxidant effects on parenchymal cells, with consequent ROS reduction. These results suggest that NOX2 inhibition might be an additional strategy to reduce cellular damage during LTx procedures.

Impact of Different Clinical Perfusates During Normothermic Ex Situ Liver Perfusion on Pig Liver Transplant Outcomes in a DCD Model

“…21 Steen as perfusate had previously been tested in pig liver transplant studies and has been extensively used in normothermic ex vivo lung perfusion. 22,23 Steen is extensively used in clinical trials for ex vivo lung preservation, demonstrating promising results, 24–26 as well as in the first North American human trial of NEsLP. 3 Gelofusine in contrast has been used for the majority of NEsLP in European clinical studies.…”

Review 1: Lung transplant—from donor selection to graft preparation

“…Pre-clinical studies have reflected anti-oxidant properties of Steen solution in EVLP. The solution was able to particularly reduce NADPH oxidase 2 isoform activation, promoting preservation of the lung epithelial cells [145]. Furthermore, 2% sevoflurane administered intravascularly during EVLP has been shown to reduce oxidative stress and the inflammatory response [146].…”

InnovativeIn VitroModels for the Study of Lung Diseases

“…In fact, OXS strongly contributes to tissue damage during EVLP. It has been shown that inhibition of NOX2 activity during thermic stress and starvation (mimicking EVLP conditions) can reduce ROS production, thus being protective for lung epithelial cells [63,64]. Finally, specific interference of cigarette smoke with Wnt/β-catenin signaling has been described in human fibroblasts, impairing their capacity to support spheroid growth of lung epithelial cells, which can be considered in this case as a stemness assay linked to the activation of a repair mechanism [65].…”