Ischemia-reperfusion induces death receptor-independent necroptosis via calpain-STAT3 activation in a lung transplant setting

Abstract

Ischemia-reperfusion (I/R)-induced lung injury undermines lung transplantation (LTx) outcomes by predisposing lung grafts to primary graft dysfunction (PGD). Necrosis is a feature of I/R lung injury. However, regulated necrosis (RN) with specific signaling pathways has not been explored in an LTx setting. In this study, we investigated the role of RN in I/R-induced lung injury. To study I/R-induced cell death, we simulated an LTx procedure using our cell culture model with human lung epithelial (BEAS-2B) cells. After 18 h of cold ischemic time (CIT) followed by reperfusion, caspase-independent cell death, mitochondrial reactive oxygen species production, and mitochondrial membrane permeability were significantly increased. N-acetyl-Leu-Leu-norleucinal (ALLN) (calpain inhibitor) or necrostatin-1 (Nec-1) [receptor interacting serine/threonine kinase 1 (RIPK1) inhibitor] reduced these changes. ALLN altered RIPK1/RIPK3 expression and mixed lineage kinase domain-like (MLKL) phosphorylation, whereas Nec-1 did not change calpain/calpastatin expression. Furthermore, signal transducer and activator of transcription 3 (STAT3) was demonstrated to be downstream of calpain and regulate RIPK3 expression and MLKL phosphorylation during I/R. This calpain-STAT3-RIPK axis induces endoplasmic reticulum stress and mitochondrial calcium dysregulation. LTx patients' samples demonstrate that RIPK1, MLKL, and STAT3 mRNA expression increased from CIT to reperfusion. Moreover, the expressions of the key proteins are higher in PGD samples than in non-PGD samples. Cell death associated with prolonged lung preservation is mediated by the calpain-STAT3-RIPK axis. Inhibition of RIPK and/or calpain pathways could be an effective therapy in LTx.

Necroptotic Cell Death Promotes Adaptive Immunity Against Colonizing Pneumococci

“…Importantly, cytosolic contents released from necroptotic cells serve as alarmins and signal to nearby cells of imminent danger or injury and trigger inflammation (9–11). Necroptosis is known to occur following sterile injury, such as ischemia-reperfusion (12, 13), during viral infection in cells that have blocked apoptosis (14), and during infection with bacteria that produce pore-forming toxins (PFT). In the latter circumstance, PFT-induced membrane damage results in ion dysregulation and energy depletion, which activates RIPK1 in non-canonical fashion (15–19).…”

Post‐ischemic salubrinal administration reduces necroptosis in a rat model of global cerebral ischemia

“…Most necroptosis studies are focused on the TNFR1‐RIPK1‐RIPK3‐MLKL pathway, but necroptosis‐independent of this axe have been also recently described (Saveljeva et al ; Parks et al ). TNFR1‐independent pathways have been related to ER stress in the lung (Kim et al ) and heart (Zhu et al ) after I/R. Therefore, the use of an enhancer of UPR, which decreases ER stress, could provide information about a possible presence of TNFR‐independent pathways during reperfusion.…”

Review 2: Primary graft dysfunction after lung transplant—pathophysiology, clinical considerations and therapeutic targets

“…Necroptosis as a result of the calpain-STAT3-RIPK pathway activation has also been implicated in ischaemia/reperfusion pathology, and it has been suggested that blocking the said pathway may also reduce graft dysfunction [ 145 , 146 ].…”