Severe periodontitis is one of the most prevalent oral diseases globally, characterized by irreversible destruction of the periodontium resulting from destructive inflammation and immune response (GBD 2021 Oral Disorders Collaborators, 2025, Boyer et al., 2020). It is caused by microbial dysbiosis involving invasion and outgrowth of anaerobic periodontal pathogens such as Porphyromonas gingivalis (P. gingivalis; Hajishengallis & Lamont, 2021). Lipopolysaccharide (LPS), as the main virulence factor from P. gingivalis, induces and aggravates the host inflammatory response with reactive oxygen species (ROS) and cytokines affecting the alveolar bone (Akkaoui et al., 2021, Sczepanik et al., 2020).

Alveolar bone maintains a dynamic balance between bone resorption by osteoclasts and bone formation by osteoblasts. Osteoblasts express NF-κB Ligand (RANKL) and osteoprotegerin (OPG), the ratio of which is dominant in inflammatory bone destruction of periodontitis (Koide et al., 2013, Zhou et al., 2022). RANKL binds to its receptor, RANK, on osteoclast and preosteoclast cell surfaces for bone resorption, while OPG competitively binds to RANKL to inhibit the interaction (Udagawa et al., 2021, Hienz et al., 2015). The Matrix metallopeptidase-9 (MMP-9) secreted by osteoblasts is also involved in promoting the degradation of bone matrix (Li et al., 2021, Luchian et al., 2022).

Mitochondria are crucial in the inflammatory response as major sites of ROS production (Andrieux et al., 2021). Mitochondrial dysfunction has been reported to contribute to the pathogenesis of periodontitis via oxidative stress, apoptosis and inflammation (Deng et al., 2024). P. gingivalis-LPS could activate Toll-like receptor 4 (TLR4) signaling, thereby activating mitogen-activated protein kinase (MAPK) / extracellular regulated protein kinases (ERK) - dependent forkhead box O3 (FOXO3) phosphorylation and amplifying oxidative stress (Huang et al., 2025). Additionally, P. gingivalis-LPS suppresses genes involved in mitochondrial function and biogenesis, compromises the function of complexes Ⅰ, Ⅱ, and Ⅳ in the mitochondrial electron transport chain and causes abnormal oxidative phosphorylation. Consequently, adenosine triphosphate (ATP) synthesis is reduced and mitochondrial ROS (mtROS) release is further accelerated, perpetuating a vicious cycle of oxidative stress and mitochondrial impairment (Verma et al., 2023, Qiao et al., 2023). This vicious loop thus persistently exacerbates periodontal inflammation and mediates bone resorption (Jeong et al., 2023, Thouvenot et al., 2025).

In response, specific quality control mechanisms are evolved to maintain proper mitochondrial network (Eldeeb et al., 2022). The mitochondrial unfolded protein response (UPRmt) is one of the repair pathways activated by an accumulation of misfolded proteins to restore mitochondrial protein homeostasis and function under stress (Smyrnias, 2021). UPRmt is a mitochondria-to-nucleus signaling that leads to upregulation of chaperones and proteases (Zhu et al., 2021). Chaperones, including heat shock 60 kDa protein (Hsp60), heat shock 10 kDa protein (Hsp10) and mitochondrial heat shock protein 70 (mtHsp70), can facilitate the correct protein folding (Shin et al., 2021). Proteases, such as ATP-dependent Clp protease proteolytic subunit (ClpP) and Lon peptidase 1(LONP1), degrade impaired or misfolded proteins (Feng et al., 2021). In mammalian cells, UPRmt is mediated by activating transcription factor 5 (ATF5), which will be translocated to the nucleus together with C/EBP-homologous protein (CHOP), inducing the expression of chaperones and proteases. ATF5 is required by the UPRmt for protection against osteoarthritis and cardiac ischemia-reperfusion injury (Zhou et al., 2022, Wang et al., 2019).

The UPRmt is involved in immune and inflammatory responses by enhancing pathogen clearance and infection tolerance (Deng et al., 2019, Dimos et al., 2019). Presently, the UPRmt is reported to be activated and cytoprotective in cardiac stress and some neurodegenerative diseases (Wang et al., 2019, Hu et al., 2021, Lian et al., 2024). However, the role of UPRmt in periodontitis, especially the inflammatory bone destruction, remains unclear. Therefore, the purpose of this study was to explore the role of ATF5-mediated UPRmt in RANKL expression and mitochondrial function in P. gingivalis-LPS-treated osteoblasts.