Revealing the Improving Effect and Molecular Mechanism of L-Clausenamide in Combating the Acute Lung Injury: Insights from Network Pharmacology, Molecular Docking, and In Vitro Validation

TL;DR

This study shows how L-Clausenamide from wampee fruit helps treat acute lung injury by reducing cell damage and targeting key proteins like AKT1 and Caspase-3.

Contribution

The novel contribution is identifying L-Clausenamide's protective effect on lung injury via AKT1 and Caspase-3, supported by network pharmacology and in vitro validation.

Findings

L-Clausenamide reduces ROS, DNA damage, and mitochondrial dysfunction in LPS-induced lung injury.

AKT1 and Caspase-3 are key targets in the protective mechanism of L-Clausenamide.

Phosphorylation of Akt increases and Caspase-3 cleavage decreases with L-Clausenamide treatment.

Abstract

Acute lung injury (ALI) is a life-threatening disease, causing intrinsic classical apoptosis. Lipopolysaccharide (LPS) is commonly used to mimic ALI in vitro. L-Clausenamide is an amide from the fruit wampee. We found it is capable of alleviating LPS-mediated reactive oxygen species (ROS) and DNA damage accumulation, ATP decline, mitochondrial depolarization, and structural and functional collapse. By prediction and verification, we found both the AKT1 protein and Caspase-3 play an important role in the alleviation effect of L-Clausenamide. This finding reveals L-Clausenamide has developmental potential for ALI treatment, and further study on this mechanism may discover the effect among L-Clausenamide, AKT1, and Caspase-3.

Acute lung injury is a severe disease with a high mortality rate, which can result in increased oxidative stress and further mitochondrial damage and cell apoptosis. L-Clausenamide is an amide from the fruit wampee. This study combined network pharmacology, molecular docking, and in vitro study to elucidate the effect of combating acute lung injury and the underlying mechanism of L-Clausenamide. Network pharmacology indicated that the 152 targets can treat acute lung injury through regulating oxidative stress. Based on PPI analysis and screening of the central target, AKT1 is the key target of the underlying mechanism. KEGG and GO enrichment analysis demonstrated that apoptosis is an important pathway for this curing effect. In the in vitro study, treatment with L-Clausenamide alleviates intracellular ROS accumulation, mitochondrial membrane potential loss, mitochondrial morphological distortion, ATP decrease, and the CASP3 activity. The SPR analysis was performed to validate the binding between AKT1 and L-Clausenamide. The Western blot result showed that L-Clausenamide increases the phosphorylation of Akt and decreases cleavage of CASP3. L-Clausenamide can alleviate lipopolysaccharide (LPS)-induced acute lung injury through targeting AKT1 and show an improvement in mitochondrial abnormality and inhibition against ROS-activated caspase-3-dependent apoptosis activation.

Figures