Osteoclast differentiation is crucial for bone absorption, and osteoclasts are involved in bone destruction in rheumatoid arthritis (RA). Dairy Propionibacterium freudenreichii is used as a cheese starter and possesses prebiotic and postbiotic properties. It is known to stimulate the growth of bifidobacteria and produces valuable metabolites, such as vitamin B12 and propionic acid. However, limited information is available on the beneficial effects of P. freudenreichii on human disease. Herein, we aimed to investigate the inhibitory effect of P. freudenreichii MJ2 (MJ2) isolated from raw milk on osteoclast differentiation and evaluate the improvement in RA. The murine macrophage cell line, RAW 264.7, and a collagen-induced arthritis (CIA) mouse model were used to perform in vitro and in vivo studies, respectively. Heat-killed P. freudenreichii MJ2 (hkMJ2)-treated cells significantly inhibited RANKL-induced osteoclast differentiation and TRAP activity. HkMJ2-treated cells exhibited significantly decreased expression of genes and proteins related to RANKL-induced osteoclast differentiation. MJ2 administration decreased the arthritic score in the CIA mouse model. Live and dead MJ2 inhibited bone loss and afforded protection against bone erosion and joint damage in CIA mice. MJ2 decreased the levels of collagen-specific antibodies and inflammatory cytokines and the expression of osteoclast differentiation-related genes and proteins in CIA mice. Interestingly, live and dead MJ2 showed similar RA improvement effects in CIA mice. In conclusion, P. freudenreichii MJ2 inhibited osteoclast differentiation by inhibiting the NF-κB signaling pathway and ameliorated CIA.

This study investigated the effects of lemon verbena extract (LVE) on sleep regulation using both a pentobarbital-induced sleep model and an EEG-based sleep assessment model in mice. To elucidate its potential mechanisms, mice were randomly assigned to five groups: control, positive control (diazepam, 2 mg/kg b.w.), and three LVE-treated groups receiving 40, 80, or 160 mg/kg b.w. via oral administration. In the pentobarbital-induced sleep model, mice underwent a two-week oral administration of LVE, followed by intraperitoneal pentobarbital injections. The results demonstrated that LVE significantly shortened sleep latency and prolonged sleep duration compared to the control group. Notably, adenosine A1 receptor expression, both at the mRNA and protein levels, was markedly upregulated in the brains of LVE-treated mice. Furthermore, LVE’s administration led to a significant increase in the mRNA expression of gamma-aminobutyric acid type A (GABA_A) receptor subunits (α2 and β2) in brain tissue. In the electroencephalography (EEG)/electromyogram (EMG)-based sleep model, mice underwent surgical implantation of EEG and EMG electrodes, followed by one week of LVE administration. Quantitative EEG analysis revealed that LVE treatment reduced wakefulness while significantly enhancing REM and NREM sleep’s duration, indicating its potential sleep-promoting effects. These findings suggest that LVE may serve as a promising natural sleep aid, improving both the quality and duration of sleep through the modulation of adenosine and GABAergic signaling pathways.