B75-15 Indole 3 Acetic Acid Protects Against Lipopolysaccharide-induced Lung Endothelial Inflammation and Barrier Disruption

Rationale Lung microvascular endothelial cell (EC) dysfunction is the pathological hallmark of acute respiratory distress syndrome. Hsp90 plays a central role in mediating endothelial cell inflammation and barrier dysfunction in response to endotoxin exposure. Our findings demonstrate that ubiquitin-specific peptidase 40 (USP40) maintains endothelial integrity by deubiquitinating the heat shock protein HSP90β. Indole-3-acetic acid (IAA) is the most common naturally occurring plant hormone of the auxin class. Accumulating evidence suggests that IAA reduces oxidative stress and inflammation and promotes intestinal barrier function. However, little is known about the role of IAA in endothelial cells and acute lung injury. Methods Endothelial barrier function in human lung microvascular endothelial cells (HLMVECs) was assessed via ECIS and TEER measurements. HEK293/TLR4 cells pretreated with 0.2 mM IAA (3h) underwent modified IP and ubiquitin immunoblotting. USP40 DUB activity with IAA was analyzed using Ub-AMC. Lung injury was induced by intratracheal lipopolysaccharide (LPS) (2 mg/kg, 24 h). BAL fluid and lung tissue were collected for ELISA, protein assay, H&E staining, wet/dry ratio, and immunohistochemistry. Results In this study, we investigated the effect of IAA on EC barrier function and inflammation. Pretreatment with IAA attenuated LPS induced reductions in TEER and suppressed IL-6, IL8, IL1 β, and TNFα expression in HLMVECs, THP-1 cells, and 293TLR4 indicating that IAA exerts anti-inflammatory effects. Ubiquitination assays revealed that IAA decreased HSP90β ubiquitination, suggesting that IAA acts as a potent activator of USP40. The protective effects of IAA on ECs were abolished when USP40 was downregulated, confirming that USP40 mediates the IAA response. An In vivo, C57BL/6J mice were intraperitoneally injected with IAA followed by an intratracheal LPS challenge. IAA significantly reduced bronchoalveolar lavage (BAL) protein levels an indicator of alveolar-capillary leakage and lung injury in the LPS-induced acute lung injury (ALI) model. Conclusion Collectively, these findings demonstrate that IAA protects against LPS-induced endothelial dysfunction and lung injury through activation of USP40, identifying IAA as a potential anti-inflammatory modulator in vascular and pulmonary inflammation.