Protective Role of Rosmarinic Acid-Enriched Elsholtzia kachinensis Extract Against Oxidative and Inflammatory Stress in Human Trophoblasts Through the miR-138-5p/FOXC1 Pathway.

Abstract

Inflammation-induced dysregulation of microRNAs, particularly miR-138-5p, compromises trophoblast cell function and may contribute to pregnancy-related complications. While Elsholtzia species have been reported to possess therapeutic properties, the anti-inflammatory effects of Elsholtzia kachinensis Prain (EKP) in trophoblast cells have not been explored. In this study, a 70% ethanolic extract of EKP was fractionated into hexane (HEX), dichloromethane (DCM), ethyl acetate (ETAC), and water (WT) fractions. Each fraction was subjected to phytochemical profiling, antioxidant assessment, and bioactive compound identification by GC-MS and high-performance liquid chromatography (HPLC). Among them, the ETAC fraction (EKP-ETAC) showed the highest antioxidant activity, reflected by the lowest IC₅₀ values in ABTS and DPPH assays, and contained the greatest levels of phenolic and flavonoid compounds. Nontoxic concentrations were determined by an MTT assay. Pretreatment with EKP-ETAC alleviated lipopolysaccharide (LPS)-induced impairment of proliferation, migration, and invasion in HTR-8/SVneo trophoblast cells, while reducing apoptosis, intracellular ROS generation, and inflammatory cytokine expression. Mechanistic analysis revealed that EKP-ETAC suppressed LPS-induced upregulation of miR-138-5p and restored the expression of FOXC1. The dual luciferase reporter assay confirmed that FOXC1 is a direct target of miR-138-5p, mediating the effects of EKP-ETAC. LC-MS and HPLC analyses identified rosmarinic acid as the predominant bioactive component. These findings suggest that rosmarinic acid-enriched EKP-ETAC exerts protective effects on trophoblast cells by mitigating oxidative stress and inflammation through the miR-138-5p/FOXC1 axis.