Spatholobus suberectus Stem-Derived Extracellular Vesicle-Like Particles Attenuate Glucocorticoid-Induced Osteoporosis via NRF2/HO-1 Signaling

Purpose: Glucocorticoid-induced osteoporosis (GIOP) is characterized by impaired osteoblast function and disrupted bone homeostasis during prolonged glucocorticoid exposure. This study investigated whether Spatholobus suberectus stem-derived extracellular vesicle-like particles (SS-EVLP) could attenuate glucocorticoid-induced osteogenic impairment and explored the potential involvement of NRF2/HO-1 signaling.

Methods: SS-EVLP were isolated by ultracentrifugation and characterized by transmission electron microscopy, nanoparticle tracking analysis, and Fourier-transform infrared spectroscopy. Cellular uptake was evaluated using PKH26 labeling. In vitro, MC3T3-E1 pre-osteoblastic cells were exposed to dexamethasone (DEX) and treated with SS-EVLP. Cell viability, Apoptosis, Alkaline Phosphatase (ALP) activity, mineralization, and protein expression were assessed by CCK-8 assay, flow cytometry, staining assays, immunofluorescence, and Western blotting. In vivo, the effects of SS-EVLP were examined in a prednisolone-induced zebrafish model by analyzing particle distribution, skeletal mineralization, osteoblast-associated signals, ALP activity, and osteogenic- and antioxidant-related gene expression.

Results: SS-EVLP exhibited vesicle-like spherical morphology and were mainly distributed between approximately 100 and 200 nm, with a major peak at 146 nm. SS-EVLP were efficiently taken up by MC3T3-E1 cells and showed low cytotoxicity. In DEX-treated cells, SS-EVLP restored cell viability, reduced Apoptosis, and partially recovered ALP staining and mineralized nodule formation. The reduced expression of RUNX2, SP7, HO-1, and NRF2 in DEX-treated cells was partially restored by SS-EVLP, accompanied by increased nuclear NRF2 signal. In zebrafish, SS-EVLP were detectable in vivo after microinjection, attenuated prednisolone-induced skeletal defects, restored ALP activity, and increased the expression of runx2a, sp7, nrf2, HO-1, gclc, gstp, and nqo1.

Conclusion: SS-EVLP attenuated glucocorticoid-induced osteogenic impairment in vitro and in vivo, at least partly through NRF2/HO-1 signaling. These findings support the potential of SS-EVLP as a plant-derived nanovesicle candidate for GIOP intervention.