Endoplasmic reticulum stress exacerbates ischemia-reperfusion-induced pulmonary endothelial barrier dysfunction by activating TXNDC5

INTRODUCTION: Lung ischemia-reperfusion injury (LIRI)-a fatal complication following major surgery, trauma, or shock- is characterized by pulmonary endothelial barrier (PEB) dysfunction. Although thioredoxin domain-containing protein 5 (TXNDC5) regulates endothelial homeostasis, its role in LIRI pathogenesis remains unknown. OBJECTIVES: In the present study, we investigate the role of TXNDC5 in LIRI-induced PEB disruption and explore its underlying mechanisms and therapeutic potential. METHODS: We established LIRI models in rats (left pulmonary hilum occlusion) and human umbilical vein endothelial cells (oxygen-glucose deprivation/reperfusion). TXNDC5 expression was modulated via AAV-shRNA knockdown, whereas endoplasmic reticulum (ER) stress was inhibited pharmacologically (4-PBA and Ceapin-A7). Structural and functional changes of the PEB, including HSP90/eNOS complex stability, were examined. Mechanistic studies included ChIP-seq identified ATF6 binding to the TXNDC5 promoter, followed by rescue experiments with ATF6 inhibition and TXNDC5 overexpression. Clinical validation was performed using plasma proteomics in patients with LIRI. A TXNDC5-based nomogram model for LIRI early-warning was constructed and internally validated. Incremental prediction value of TXNDC5 was evaluated using net reclassification improvement (NRI) and integrated discrimination improvement (IDI). RESULTS: Single-cell RNA sequencing revealed TXNDC5 upregulation in pulmonary endothelial cells during LIRI, correlating with PEB disruption, apoptosis, ER stress, and NF-κB activation. TXNDC5 knockdown restored HSP90/eNOS stability and improved endothelial integrity in vitro/vivo. Ceapin-A7, or 4-PBA, attenuated TXNDC5-driven PEB injury, and TXNDC5 overexpression confirmed the causal role of the ATF6-TXNDC5 axis. Mechanistically, ER stress promoted TXNDC5 transcription via ATF6 binding to its promoter. Clinically, elevated plasma TXNDC5 levels predicted LIRI and correlated with the severity of the condition. Incorporating the TXNDC5 index into the clinical predictors model enhanced the forecast of LIRI, as demonstrated by the NRI and IDI. CONCLUSION: Our findings demonstrate that ER stress-induced TXNDC5 upregulation exacerbates LIRI by impairing HSP90/eNOS-dependent endothelial function. ER stress-TXNDC5 signaling is a mechanistically distinct and potential target in LIRI.