Quercetin-primed BMSC-derived extracellular vesicles ameliorate chronic liver damage through miR-136-5p and GNAS/STAT3 signaling pathways

Background: Chronic liver damage (CLD) is a long-term and progressive liver condition characterized by inflammation, fibrosis, and impaired liver function, which ultimately lead to severe complications such as cirrhosis or liver Cancer. Quercetin (Que), a flavonoid in various Plants, possesses anti-inflammatory, Antiviral, anti-ischemic, and Anticancer properties. Recently, extracellular vesicles (EVs) derived from pretreated bone marrow mesenchymal stem cells (BMSCs) have shown immense potential in treating various diseases, including CLD. Thus, this study evaluated the regulatory effects of Que-preconditioned BMSC-derived EVs (Que-EVs) on LPS-stimulated RAW264.7 cells and their therapeutic effects on mice with CLD.

Methods: Que-EVs and control-EVs were harvested from the Cell Culture supernatant of BMSCs. The EVs were characterized using western blot, transmission electron microscopy, and nanoparticle tracking analysis. Further, the DIR labeling of EVs was used to detect in vitro and in vivo uptake. Next, LPS pre-stimulated RAW264.7 cells were treated with Que-EVs and control-EVs for 24 h. The relative expression of inflammatory cytokines and macrophage polarization markers genes was assessed using RT-qPCR, and western blot was conducted to evaluate the GNAS, PI3K, ERK, and STAT3 gene and protein expressions in RAW264.7 cells. Furthermore, transfection techniques were employed to induce miR-136-5p inhibition and GNAS overexpression in RAW264.7 cells to validate the role of miR-136-5p in alleviating inflammation through the GNAS/PI3K/ERK/STAT3 pathway. Subsequently, the outcomes were validated via in vitro experiments.

Results: Que enhanced miR-136-5p expression in BMSC-EVs. Furthermore, it was shown that EVs delivered miR-136-5p to macrophages, thereby attenuating M1-type macrophage polarisation through the GNAS/PI3K/ERK/STAT3 pathway, reducing liver inflammation, improving liver function and treating CLD.

BMSC-EVs; Chronic liver damage; GNAS/PI3K/ERK/STAT3; Que-EVs; Quercetin; miR-136-5p.