Inflammatory bone loss and signaling pathways in periodontitis: mechanistic insights and emerging therapeutic strategies

Abstract

Bone resorption is a vital physiological process that enables skeletal remodeling, maintenance, and adaptation to mechanical forces throughout life. While tightly regulated under the physiological state, its dysregulation contributes to pathological conditions such as osteoporosis, rheumatoid arthritis, and periodontitis. Periodontitis is a highly prevalent chronic inflammatory disease driven by dysbiotic biofilms that disrupt the oral microbiome, leading to the progressive breakdown of the periodontal ligament, cementum, and alveolar bone and ultimately resulting in tooth loss. This review outlines the molecular and cellular mechanisms underlying periodontitis, focusing on osteoclastogenesis, the differentiation and activation of osteoclasts, the primary mediators of bone resorption. Key transcriptional regulators, including NFATc1, c-Fos, and c-Src are discussed alongside major signaling pathways such as Mitogen Activated Protein Kinase (MAPK), Janus Tyrosine Kinase/Signal Transducer and Activator of Transcription (JAK/STAT), Nuclear Factor Kappa B (NF-κB), and Phosphoinositide 3-kinase (PI3K)/Akt, to elucidate their roles in the initiation and progression of periodontal bone loss. These pathways orchestrate the inflammatory response and osteoclast activity, underscoring their relevance in periodontitis and other osteolytic conditions. Hallmark features of periodontitis, including chronic inflammation, immune dysregulation, and tissue destruction are highlighted, with emphasis on current and emerging therapeutic strategies targeting these molecular pathways. Special attention is given to small molecules, biologics, and natural compounds that have the potential to modulate key signaling pathways. Although advances in understanding these mechanisms have identified promising therapeutic targets, translation into effective clinical interventions remains challenging. Continued research into regulating bone-resorptive signaling pathways is essential for developing more effective treatments for periodontitis and related inflammatory bone diseases.

Introduction

Bone resorption is a fundamental physiological process that enables remodeling, maintaining, and continuously renewing the skeleton throughout life. However, dysregulation of this process can lead to several pathological conditions, including periodontitis.¹ Periodontitis, a non-communicable chronic and inflammatory disease linked to a dysbiotic biofilm, affects the teeth-supporting tissues due to progressive destruction of the periodontal ligament, cement, and alveolar bone.^2,3 (Fig. 1).

Fig. 1.

Schematic representation of the periodontium in healthy and diseased conditions. The periodontium’s structure differs notably between healthy and diseased conditions. In a healthy state, the gingiva is securely attached to the tooth surface, and the epithelium, periodontal ligament, cementum, gingiva, and alveolar bone remain structurally sound and undamaged. In contrast, diseased periodontium is marked by periodontal pockets, damage to the periodontal ligament, and decreased bone level due to resorption, which manifests clinically as redness, swelling, and bleeding on probing. Created with BioRender.com (License Number: VX28PGAUQ7)

This pathological process is intricately linked to the host immune response, which plays a central role in the disease initiation, progression, and exacerbation.^4–7 Severe periodontitis affects more than 700 million individuals globally, accounting for approximately 11% of the world’s population.⁸ In developed nations, prevalence rates remain high, ranging from 7.2% to 9.8%.⁹ In the United States alone, an estimated 64.7 million adults (46% of the population) are affected by periodontitis, with 8.9% experiencing severe cases.¹⁰ In Brazil, the severe forms of periodontitis account for almost 20% of the population.⁹ It is recognized as the sixth most prevalent chronic disease worldwide and remains the leading cause of tooth loss among adults.^11,12 Consequently, periodontitis represents a major public health concern, not only due to its widespread occurrence but also because of its significant impact on oral function and quality of life, including impaired mastication and aesthetic complications.¹¹ Furthermore, the financial and societal impact of dental diseases is substantial, with global treatment expenses reaching ~416 billion US dollars annually, accounting for around 5.8% of total healthcare costs.¹³ A significant portion of these expenditures, roughly 75%, is allocated to managing periodontitis. Additionally, periodontitis has been associated with a range of systemic non-communicable chronic diseases, such as rheumatoid arthritis,^14–18 diabetes mellitus,^19–22 cardiovascular disease^23–27, chronic kidney disease,²⁸ and others^29–32 (Fig. 2).

Fig. 2.

Schematic representation of the influency of periodontitis and chronic low grade inflammation in systemic conditions. The low grade chronic inflammatory condition triggered by microbial byproducts and the host immune response during periodontitis progression driven the dissemination of bacterial components or pro-inflammatory mediators through the circulation. This lead to several non-communicable chronic diseases such as cardiovascular disease, chronic obstructive pulmonary disease, rheumatoid arthritis, cancer, gastrointestinal disease, pregnancy-related complications, non alcoholic fat liver disease, diabetes/obesity, and Alzheimer’s disease. Created with BioRender.com (License Number: FJ28PGB6KS)

The periodontium is a unique and complex anatomical structure that plays a critical role in supporting the teeth and maintaining oral health. A defining feature of the periodontium is the way in which the teeth penetrate the epithelial barrier, creating a direct and constant interface between the external environment and internal tissues (Fig. 1).³³ This potentially vulnerable interface is protected by the junctional epithelium (JE), a specialized epithelial tissue that forms a tight seal around the cervical portion of the tooth.³⁴ The JE adheres firmly to the tooth surface, whether enamel or cementum, through hemidesmosomes and an internal basal lamina, creating a structurally secure attachment. Functionally, the JE serves as both a biological barrier and an immunological sentinel.³³ It restricts microbial invasion while allowing selective passage of immune cells, such as neutrophils, macrophages and lymphocytes, thus facilitating immune surveillance and maintaining tissue homeostasis. However, this protective barrier is disrupted during the onset and progression of periodontitis.³⁴ Pathogenic microorganisms in the subgingival biofilm stimulate a persistent inflammatory response that weakens the structural integrity of the JE. As a result, the JE begins to migrate apically along the tooth root, and the underlying gingival collagen fibers and connective tissue is progressively degraded mediated by matrix metalloproteinases (MMPs). This degradation marks the transition from gingivitis to periodontitis. As the JE deteriorates, the periodontium becomes increasingly exposed to bacterial pathogens and their virulence factors. Deep periodontal pockets form, providing a niche for anaerobic bacteria and further perpetuating inflammation. Concurrently, the host immune response leads to the resorption of alveolar bone.

Periodontitis begins with bacterial accumulation on the tooth surface, triggering the host’s innate immune response and recruitment of inflammatory cells. Toll-like receptors (TLRs) play a pivotal role in recognizing pathogen-associated molecular patterns (PAMPs), thereby initiating downstream signaling cascades that drive inflammation and tissue destruction in susceptible individuals.³⁵ Specifically, TLR2 and TLR4 recognize bacterial components and regulate the expression of key inflammatory mediators such as tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β) and IL-6.³⁶ This immune response activates several intracellular signaling pathways, including the Mitogen-Activated Protein Kinase (MAPK), Janus Tyrosine Kinase/Signal Transducer and Activator of Transcription (JAK/STAT), Nuclear Factor Kappa B (NF-κB), and Phosphoinositide 3-kinase (PI3K)/Akt, which together contribute to osteoclastogenesis and periodontal breakdown.^37–39 A central mechanism in bone remodeling involves the interaction between the receptor activator of nuclear factor kappa B (RANK), its ligand RANKL, and the decoy receptor osteoprotegerin (OPG), which collectively regulates the balance between bone formation and resorption.^40–42 Despite substantial advances in understanding the pathogenesis of periodontal disease,^35,43,44 many aspects remain unclear, particularly regarding the complex interplay among environmental, genetic, immunological, and epigenetic factors.

This narrative review provides a comprehensive overview of the signaling pathways implicated in the onset and progression of periodontal disease, focusing on their role in inflammatory bone loss. Additionally, we discuss therapeutic strategies aimed at modulating key molecular pathways, especially those involving NF-κB, JAK/STAT, MAPK, and PI3K/Akt. A mechanistic understanding of these pathological processes provides insight into the rationale behind current pharmacological treatments.

Pathogenesis of periodontitis

Periodontitis is a complex, multifactorial disease characterized by a disruption in the balance between host immune responses and environmental triggers.⁴⁵ Rather than resulting from a single infectious agent, periodontitis arises from a chronic inflammatory process within a susceptible host. In healthy conditions, a delicate equilibrium exists between immune surveillance and tolerance, enabling the immune system to prevent inappropriate responses while preserving tissue integrity. This immune balance plays a central role in maintaining periodontal health by preventing exaggerated inflammation.^43,46–50 When this equilibrium is disturbed, due to genetic predisposition, environmental or local factors, or immune dysregulation, a sustained inflammatory response is triggered. One proposed mechanism involves the presence of a “keystone” pathogen, such as Porphyromonas gingivalis (P. gingivalis), which can perturb host responses even in low abundance.⁵¹ This leads to an exaggerated and persistent immune activation, ultimately resulting in connective tissue degradation, loss of periodontal attachment, and radiographic evidence of alveolar bone resorption, the hallmark of periodontal disease⁵⁰ (Fig. 3).

Fig. 3.

Clinical, radiographic, microbiological, and histological aspects of periodontitis. The upper left image depicts advanced chronic periodontitis, highlighting gingival inflammation and the removal of mineralized bacterial biofilm through supragingival instrumentation. In the upper center, a clinical case of periodontitis is shown, characterized by the formation of periodontal pockets ≥4 mm in depth and alveolar bone loss due to chronic inflammation (measured using a periodontal probe). The upper right figure is a periapical radiograph demonstrating advanced bone loss, with the red line indicating the level of alveolar bone resorption (distance from the cement-enamel junction to the alveolar bone crest). The middle section contains histological images of a mouse maxilla stained with hematoxylin and eosin, comparing healthy and periodontitis-affected tissues. The right middle panel shows a detailed histological view of periodontal inflammation, highlighting infiltrated immune cells and disrupted connective tissue architecture following ligature placement. The center middle panel presents a sagittal view of a healthy tooth. Alveolar bone resorption (marked by an increased black line length) and osteoclastic activity in the periodontitis-affected sample could be observed. The progression of periodontitis results in tissue attachment loss and deepening periodontal pockets, creating an ideal environment for bacterial colonization. Among these, Porphyromonas gingivalis, a key periodontitis pathogen, is shown growing on blood agar (center left figure). At the third column, representative images from a mouse model with ligature-induced bone loss in the upper first molars are displayed. The left panel shows the clinical presentation of ligature-induced periodontitis. The middle panel provides a tomographic (micro-CT) reconstruction of the mouse maxilla, highlighting significant alveolar bone loss. The right panel displays a cross-sectional view of the micro-CT image, further illustrating the extent of bone resorption and morphological changes in the periodontal tissues. At the bottom are histological sections stained for TRAP⁺ demonstrating the osteoclasts near the alveolar bone (ab) in a mouse model of induced periodontitis

The immunological response involved in periodontal tissue breakdown is dynamic and complex, engaging both the innate and adaptive arms of immunity.^52,53 The initial response to homeostasis disruption involves rapid recruitment and activation of immune cells in the gingival connective tissue.⁵⁴ The host response is characterized by the activation of the innate immune response, providing a rapid but non-specific line of defense, and the delayed adaptive immune system, which tailors a more targeted and long-lasting response to the infection.^4,55 The interplay between these immune responses drives the pathogenesis of periodontitis, playing a role in tissue damage and resolution of inflammation (Fig. 4).^7,35

Fig. 4.

Innate and adaptive immunity drive bone resorption in periodontitis. Innate and adaptive immune responses are central drivers of the inflammatory bone resorption observed in periodontitis. The host response, initiated upon recognition of periodontal pathogenic bacteria such as Porphyromonas gingivalis, orchestrates a complex cascade of inflammatory events involving both innate and adaptive immune mechanisms. These immune responses are not only pivotal in pathogen clearance but also contribute to the chronic inflammatory microenvironment that characterizes periodontitis. Stromal cells, including gingival fibroblasts and periodontal ligament cells, interact dynamically with infiltrating immune cells to modulate the local immune milieu. They respond to microbial and inflammatory stimuli by producing chemokines, cytokines, and matrix-degrading enzymes, thereby facilitating immune cell recruitment and perpetuating tissue destruction. Meanwhile, innate immune cells such as neutrophils and macrophages initiate the early inflammatory response, whereas adaptive immune cells, particularly Th17 cells and B cells, amplify inflammation and promote osteoclastogenesis through the secretion of RANKL and other pro-osteoclastogenic factors. Together, stromal and immune cells contribute to the dysregulated immune response and bone remodeling imbalance that underlie the pathogenesis and clinical manifestations of periodontitis. Created with BioRender.com (License Number: ZP28PGCRQ5)

Host immune activation involves pattern recognition receptors (PRRs), such as TLRs expressed on the membrane of epithelial and immune cells, like macrophages and dendritic cells. These receptors sense danger-associated molecular patterns (DAMPs)³⁵ or microbial motifs and initiate signaling cascades that promote the release of pro-inflammatory mediators such as IL-1β, IL-6, and TNF-α.⁵⁶ These cytokines recruit neutrophils to the infection site. Upon sensing infection or tissue injury, neutrophils are rapidly mobilized from the bone marrow into circulation and subsequently recruited to the affected sites. There, they execute a multifaceted antimicrobial arsenal, including phagocytosis, degranulation, the production of reactive oxygen species (ROS), and the release of neutrophil extracellular traps (NETs—extracellular web-like structures composed of decondensed chromatin and antimicrobial proteins), all of which constitute the first line of defense against pathogens.^57,58 Moreover, neutrophils can contribute to an anti-inflammatory phenotype, characterized by secretion of IL-10 and CCL2, thereby favoring to immune suppression and resolution of inflammation. Conversely, in chronic inflammatory scenarios, some neutrophils exhibit hyperinflammatory profiles, producing pro-inflammatory cytokines such as IL-12 and chemokines like CCL3 and CCL4. These mediators stimulate other immune cells and contribute to the aggravation of tissue breakdown (Fig. 4).^59,60

Although neutrophils are abundant in the periodontal tissues during active periodontitis and are known to contribute to the inflammatory milieu that supports bone resorption, histopathological studies have consistently shown that these cells are infrequently found in direct proximity to bone surfaces undergoing active osteoclastic resorption.^61,62 Instead, neutrophils predominantly accumulate within the gingival crevice, junctional epithelium, and connective tissue near the microbial biofilm, where they primarily function to contain bacterial invasion through phagocytosis and the release of antimicrobial peptides and NETs.^63,64

In human periodontitis, active resorptive sites are typically occupied by osteoclasts and their precursors, which are differentiated under the influence of RANKL and pro-inflammatory cytokines secreted by immune and stromal cells, including T and B lymphocytes, macrophages, and periodontal ligament fibroblasts.⁶² While neutrophils contribute to the inflammatory environment by releasing matrix metalloproteinases (e.g., MMP-8 and MMP-9), IL-1β, TNF-α, and ROS, these factors act predominantly in a paracrine manner, influencing osteoclastogenesis indirectly rather than through direct cell-to-cell contact with osteoclast precursors at the bone surface.⁶¹

The spatial segregation between neutrophils and resorptive bone sites highlights their primary immunological function in microbial defense rather than direct participation in bone matrix degradation.⁶⁵ However, emerging data suggest that NETs and neutrophil-derived cytokines may sustain chronic inflammation and enhance the expression of RANKL by other immune cells, thereby creating a pro-resorptive environment at a distance.^66,67 Thus, while neutrophils are not in close anatomical contact with osteoclasts in vivo, their secretory profile may contribute to the amplification of osteoclastogenic signaling cascades that promote alveolar bone loss in periodontitis. Therefore, in the context of osteoclastic bone resorption in periodontitis, neutrophils should be viewed as upstream modulators of inflammation rather than direct effector cells at the bone surface. Recognizing this anatomical and functional distinction is essential to accurately interpreting the contribution of neutrophils to periodontal bone loss.^65,67

Importantly, neutrophils are now recognized as key players in trained-immunity, a form of innate immune memory characterized by long-lasting functional reprogramming following an initial challenge.^68,69 Upon exposure to certain microbial components or endogenous stimuli, neutrophils can undergo epigenetic and metabolic reprogramming that enhances their responsiveness to subsequent stimuli, even after the initial trigger has been cleared. This “training” results in augmented effector functions, such as increased production of ROS, enhanced phagocytosis, and elevated pro-inflammatory cytokine release, contributing to heightened pathogen clearance upon reinfection.^68,69 Recently, Haacke et al.⁷⁰ (2025) uncovers a novel dimension of trained immunity by demonstrating that osteoclastogenesis, the process by which osteoclasts differentiate from myeloid progenitors, is itself subject to innate immune training, thereby exacerbating inflammatory bone loss. Using β-glucan to induce trained immunity (TRIM) in mice, the authors found that this intervention led to a heightened osteoclastogenic predisposition in both bone marrow progenitors and peripheral monocytes. Upon secondary inflammatory challenge (e.g., periodontitis or arthritis), TRIM significantly increased osteoclast precursor populations, elevated expression of osteoclastogenic transcription factors, and enhanced osteoclast formation and activity in affected tissues.⁷⁰ These effects were shown to be dependent on transcriptomic rewiring of monocytes, particularly through MITF signaling, which was essential for the pro-osteoclastogenic phenotype. This study thus identifies “trained osteoclastogenesis” as a maladaptive arm of innate immune memory that may contribute to chronic inflammatory bone loss.⁷⁰

Macrophages play a more critical role in the innate immune response and tissue destruction.^71,72 Once activated, macrophages release pro-inflammatory cytokines, which amplify the inflammatory response and activate other immune cells. They further secrete catabolic enzymes such as prostaglandins and MMPs, which favor the breakdown of the extracellular matrix, leading to periodontal damage.^71,72 Additionally, dendritic cells, functioning as antigen-presenting cells, process pathogens and express bacterial antigens, after which they travel to lymph nodes to activate the adaptive immune response. This represents a critical junction between the innate and adaptive immune systems (Fig. 4).⁴³

The adaptive immune response is initiated by activating T cells, which are stimulated by antigen-presenting cells, including dendritic cells.⁷² In periodontitis, T helper 1 (Th1), Th2, and Th17 cells are key players in the immune response.⁷³ Th1 cells, driven by the transcription factor T-bet and induced by IL-12, produce interferon-gamma (IFN-γ), which activates macrophages and promotes a pro-inflammatory response. Th17 cells, under the control of the transcription factor RORγT, secrete IL-17A, a cytokine that stimulates the production of pro-inflammatory mediators such as IL-6 and TNF-α, as well as MMPs.⁷⁴ These factors contribute to tissue destruction by promoting neutrophil recruitment and activation, and by enhancing osteoclast activity, leading to bone resorption. Th17 cells are critically involved in initiating and sustaining inflammation and bone loss during the early stages of periodontal disease (Fig. 4).⁷⁵ In contrast, Th2 cells, regulated by the transcription factor GATA-3, play a more protective and regulatory role. They secrete anti-inflammatory cytokines, including IL-4, IL-5, IL-9, IL-10, and IL-13, which help modulate the immune response and inhibit osteoclastogenesis, thereby contributing to immune homeostasis and limiting tissue damage.

B cells also play a role in the adaptive immune response and disease progression. They differentiate into plasma cells, which secrete antibodies, primarily immunoglobulin G (IgG), which can specifically bind to periodontal bacteria and facilitate their clearance.⁷⁵ However, in periodontitis, the antibody response may be insufficient to clear the invaders completely, and the persistence of pathogenic insult in the periodontal tissues can result in chronic inflammation and lack of resolution. Recent studies have shown that B-cells can also function as antigen-presenting cells and release inflammatory cytokines, further exacerbating the inflammatory environment.^76,77 Over time, this leads to the stimulation of osteoclasts, leading to bone resorption and the degradation of the tooth supporting structures (Fig. 4).^35,36

The resolution of inflammation is a crucial and tightly regulated phase of the immune response that is essential for restoring tissue homeostasis following the cessation of a pro-inflammatory stimulus. Effective resolution depends on a phenotypic switch in immune cells, characterized by a coordinated transition from a pro-inflammatory to a pro-resolving state. This process involves the suppression of pro-inflammatory mediators and the activation of specialized pro-resolving pathways, including the upregulation of anti-inflammatory cytokines such as interleukin-10 and transforming growth factor-beta (TGF-β).^78,79 These cytokines are key regulators in terminating inflammation by inhibiting further immune cell activation, promoting tissue repair and regeneration, and enhancing the clearance of apoptotic cells through efferocytosis.

Specialized pro-resolving mediators (SPMs), including lipoxins, resolvins, protectins, and maresins, are biosynthesized from omega-3 and omega-6 polyunsaturated fatty acids and serve as key molecular drivers of the resolution phase.^80–83 These lipid mediators limit neutrophil recruitment, enhance macrophage-mediated efferocytosis, and promote a phenotypic switch in macrophages from a pro-inflammatory (M1) to an anti-inflammatory, tissue-reparative phenotype (M2). Additionally, regulatory T cells (Tregs) expand during this phase and secrete immunosuppressive cytokines that further promote the cessation of inflammation and support mucosal tolerance. In the context of periodontitis, however, this finely tuned resolution program is disrupted.^80–83 The persistence of microbial biofilms and inadequate plaque control leads to continuous activation of innate immune responses and sustained production of pro-inflammatory cytokines. These mediators perpetuate the inflammatory milieu and impair the production or function of SPMs, thereby blocking the transition to resolution.^84,85 Neutrophils and macrophages, which under normal circumstances would undergo apoptosis and be efficiently cleared, instead remain activated and accumulate in the periodontal tissues, contributing to extracellular matrix degradation.

Moreover, the failure to effectively clear apoptotic cells (defective efferocytosis) and to reprogram immune cells toward a reparative phenotype exacerbates tissue destruction and prevents healing. Chronic, unresolved inflammation leads to the breakdown of periodontal connective tissue and progressive alveolar bone loss. Thus, the inability to initiate or sustain the resolution phase represents a critical pathogenic mechanism in the chronicity and progression of periodontitis.

In summary, the immune response to periodontal pathogens involves a complex interplay between innate and adaptive mechanisms. While the innate system rapidly responds through PRRs and neutrophil recruitment, the adaptive response, particularly via Th1 and Th17 cells, sustains chronic inflammation. Dendritic cells, macrophages, and B cells contribute to immune activation and tissue destruction. The inability to resolve inflammation and the continued stimulation of osteoclastogenesis underlie the progressive bone loss that defines periodontitis. A detailed understanding of these cellular and molecular events in the pathogenesis of periodontitis is essential for developing effective therapeutic strategies (Fig. 5).^49,86–90

Fig. 5.

Inflammatory responses related to periodontitis. Periodontitis is characterized by the dysregulated stimulation of the innate host response and is primarily caused by an imbalance in the microbial community due to pathogenic biofilms. This imbalance activates an excessive inflammatory reaction, involving the recruitment of neutrophils, macrophages, and lymphocytes to the periodontal pocket. The condition arises from oral dysbiosis caused by biofilms composed of periodontal pathogens. Virulence factors released by pathogenic bacteria recruit neutrophils to the inflamed gingival tissue. This recruitment triggers excessive pro-inflammatory cytokine production by keratinocytes, fibroblasts, and antigen-presenting cells, leading to a hyperactive innate immune response. Macrophages contribute to this response by increasing the secretion of pro-inflammatory cytokines and MMPs, and activating B cells. The persistent elevation of TNF-α, IL-1β, INF-γ, and MMPs, coupled with the upregulation of TH1 and TH17 responses, promotes the activity of bone-resorbing cells, ultimately leading to osteoclastogenesis and bone destruction. Created with BioRender.com (License Number: GW28PGAZ0X)

Bone remodeling biology: signaling pathways and cellular interactions

Bone homeostasis is governed by two fundamental and continuous processes: remodeling and modeling.⁹¹ These processes respond to mechanical, hormonal, and nutritional stimuli to ensure bone tissue formation, maintenance, and structural adaptation throughout life.^1,92,93 Bone remodeling is a dynamic activity that constantly renews bone tissue, preserves its structural integrity, adapts to mechanical load, and maintains mineral balance.^92,94 In this context, specialized bone cells play key roles: osteoblasts synthesize the bone matrix, while osteoclasts resorb old or compromised bone.^93,95–98 The balance between synthesis and resorption ensures continuous bone renewal, preventing micro-injury accumulation that could compromise its strength and lead to fractures.⁹²

Osteoclasts, the primary bone-resorbing cell, originate from monocyte/macrophage precursors within the hematopoietic lineage in the bone marrow.^99–101 The migration of osteoclast precursors from the bone marrow and circulation to resorption sites occurs via the collagen network, guided by sphingosine-1-phosphate (S1P) signaling,¹⁰² produced by sphingosine-1 kinase (SphK) and operating in distinctive S1P receptors.^103,104 Upon reaching the bone surface, these precursors differentiate into multinucleated, mature osteoclasts with resorptive capacity, a process driven by RANKL and macrophage colony-stimulating factor (M-CSF).^105,106 These signaling molecules are produced by osteogenic cells, T cells, and vascular endothelial cells near the bone surface. Both soluble (sRANKL) and membrane-bound RANKL (mRANKL) interact with RANK receptors on osteoclast precursors, initiating intracellular signaling cascades mediated by tumor necrosis factor receptor-associated factor (TRAF) proteins, particularly TRAF2, TRAF5, and TRAF6.⁹⁹ This activation subsequently triggers NF-κB and MAPK pathways, leading to the expression of myelocytomatosis oncogene (MYC) and Fos proto-oncogene, AP-1 transcription factor subunit (FOS).⁹⁹ These factors collectively induce the nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), which plays a crucial role in osteoclast differentiation through the canonical signaling pathway.¹⁰⁷ As a key transcription factor, NFATc1 facilitates osteoclastogenesis by enhancing the expression of genes associated with bone resorption, including MMP9, cathepsin K, acid phosphatase 5 (Acp5), and tartrate-resistant acid phosphatase (TRAP).^108,109 Mature osteoclasts are multinucleated, polarized cells that adhere to the bone surface and create a resorption lacuna by secreting lysosomal proteases (Fig. 3), including TRAP, MMPs, and cathepsin K.¹¹⁰ Within this acidic microenvironment, bone minerals and organic components are degraded, endocytosed, and subsequently expelled through the secretory domain on the opposite side of the cell.¹¹¹

The resorption process and the bone remodeling occur in a four-phase cycle: i) Activation Phase: Circulating monocyte-macrophages are recruited to the affected site, where they fuse on the bone surface to form multinucleated pre-osteoclasts;^3,92,94,98 ii) Resorption Phase: The resorption process involves osteoclast attachment to the bone matrix and the formation of a ruffled border, which is regulated by integrin-mediated polarization, particularly through α5β1, α2β1, and αvβ3.^{100,112–114} Among them, αvβ3 is the most highly expressed and facilitates osteoclast adhesion to bone matrix proteins, such as osteopontin (OPN), bone sialoprotein, and fibronectin, by recognizing Arg-Gly-Asp (RGD) motifs.¹¹⁵ This binding interaction triggers structural adaptations in the osteoclast’s apical membrane and promotes actin ring formation via the phospholipase Cγ2 (Plcγ2), proline-rich tyrosine kinase 2 (Pyk2), and Src signaling pathways, which are critical for bone resorption;^50,92 iii) Reversal Phase: The formed lacunae contain mononuclear cells and osteoblast precursors, which initiate bone formation. Bone resorption activates the release of coupling factors stored within the bone matrix, including TGF-β and insulin-like growth factor-1 (IGF-1). These factors play a crucial role in recruiting osteoblast precursors to the resorption site and facilitating their differentiation, thereby linking bone resorption to bone formation;^92,116 and iv) Formation Phase: Osteoblasts, which secrete osteoid on bone surfaces, comprise approximately 4%–6% of all bone cells and are derived from mesenchymal stem cells,¹¹⁷ synthesize the organic matrix for bone mineralization, transporting phosphate and calcium to the mineralization site and degrading inhibitors to allow mineral deposition⁵⁰ (Fig. 6). Some osteoblasts become confined in the matrix and differentiate in osteocytes. Osteocytes maintain bone tissue homeostasis by coordinating the exchange of nutrients and signals through a canalicular network.^1,48,92 Moreover, osteoblasts and osteocytes can secrete RANKL, OPG, lysophosphatidic acid, and monocyte chemoattractant protein-1 to modulate osteoclast activity.¹¹⁸

Fig. 6.

Disruption in the equilibrium concerning osteoclast and osteoblast activity is a key feature of periodontitis. The downregulation of OPG enhances the communication between RANK and its ligand (RANKL), driving osteoclast differentiation and activity. Additionally, various factors, such as IL-1, IL-16, IL-17A, MMPs, and parathyroid hormone (PTH), further promote osteoclast differentiation and bone resorption. Moreover, the signaling pathways involved in bone cell communication are disrupted, leading to impaired bone homeostasis in periodontal disease, as described further below. Created with BioRender.com (License Number: FX28PGB2SL)

Osteocytes form a vast three-dimensional network by connecting through their dendritic extensions. This intricate network enables osteocytes to perceive mechanical signals by detecting fluid flow-induced shear stress along their dendrites.^119,120 In response, they modulate bone’s mechanical properties and interact with osteoblasts and osteoclasts via the RANKL/OPG pathway and the sclerostin/Dickkopf-1/WNT (SOST/Dkk1/WNT) signaling axis.⁹⁹ At the end of the remodeling cycle, ~50%–70% of osteoblasts undergo apoptosis, while the remaining osteoblasts become lining cells or osteocytes.^1,48,92 Bone lining cells regulate mineral ion transport and can redifferentiate into osteoblasts under certain stimuli, such as mechanical forces or parathyroid hormone.⁹²

Recent studies have revealed that mature osteoclasts often undergo fragmentation into smaller, non-resorptive cells known as osteomorphs rather than proceeding to apoptosis.^121,122 These osteomorphs retain the capacity to migrate across bone surfaces and can rapidly re-fuse into active osteoclasts upon subsequent RANKL stimulation. This recycling mechanism is considered more energy-efficient than de novo osteoclast formation and represents a novel regulatory feature of bone resorption dynamics.^121,122

Inflammatory bone loss and osteoclastogenesis

Pathological bone resorption arises when the homeostatic balance of bone formation and resorption is disrupted, leading to excessive osteoclast activity. This can occur through an increase in osteoclast number, function, or both, ultimately resulting in significant degradation of the bone matrix.^94,98 In periodontitis, chronic inflammation intensifies osteoclastogenesis, leading to progressive destruction of alveolar bone. Central to this process is the RANK/RANKL/OPG axis^48,98,123 which also plays a pivotal role in the pathogenesis of other skeletal disorders, including osteoporosis, primary or metastatic bone malignancy, autoimmune arthritis, and Paget’s disease.

RANKL, encoded by the TNFSF11 gene, is a member of the TNF superfamily.^{50,94,124–126} Various cell types, including osteocytes, lymphocytes, osteoblasts, stromal cells, fibroblasts, and periodontal fibroblasts, express RANKL.^{50,94,124–126} In bone, RANKL can be cleaved by MMPs to generate the soluble form sRANKL, primarily secreted by osteoblastic stromal cells.^124–126 In the periodontium, periodontal ligament cells produce RANKL in response to inflammation, promoting osteoclast differentiation by activating inflammatory signaling cascades and transcriptional regulators.^50,98 M-CSF, secreted primarily by osteoblasts and fibroblasts, supports the proliferation and survival of mononuclear osteoclast precursors and upregulates the membrane-bound RANK receptor.^{48,94,127,128} RANKL binds to RANK on these precursors, triggering signaling pathways that lead to their differentiation into mature, multinucleated osteoclasts.^124–126 This process involves activating key transcription factors such as c-Fos, NF-κB, and NFATc1, which are indispensable for osteoclast differentiation and activation.⁵⁰ OPG is a soluble glycoprotein that functions as a decoy receptor for RANKL, competitively inhibiting its binding to RANK and suppressing osteoclastogenesis. The ratio of RANKL to OPG is a critical determinant of bone resorption activity.^48,50,94 Elevated RANKL levels and reduced OPG expression, commonly observed in periodontal inflammation, create a pro-resorptive environment conducive to alveolar bone loss (Fig. 6).¹²¹ Previous studies demonstrate that RANKL-deficient (RANKL^−/−)¹²⁹ and RANK-deficient (RANK^−/−)¹³⁰ mice develop severe osteopetrosis due to impaired osteoclast formation, highlighting the essential role of the RANKL–RANK pathway in bone resorption. Conversely, osteoprotegerin-deficient (OPG^−/−) mice exhibit pronounced bone loss, underscoring OPG function as a decoy receptor that inhibits osteoclastogenesis by binding RANKL.^131,132

The resolution of inflammation plays a pivotal role in suppressing osteoclastogenesis and reestablishing bone homeostasis. This transition involves a coordinated downregulation of pro-resorptive signaling and the promotion of anti-inflammatory and anti-osteoclastogenic pathways. Key cytokines such as IL-10, IL-4, IL-13, and TGF-β act to inhibit osteoclast differentiation by reducing the expression of RANKL and enhancing OPG levels, thereby disrupting the RANK/RANKL axis essential for osteoclast maturation.^133–137 Tregs further contribute by secreting IL-10 and TGF-β, which not only suppress inflammatory cytokine production but also promote the expansion of M2 macrophages, cells associated with tissue repair and inhibition of osteoclast activity.^133–138 As levels of pro-inflammatory mediators decline, osteoclast activation is progressively limited, preventing further bone loss. This transition from inflammation to resolution enables osteoblast-mediated bone formation by activating Wnt/β-catenin signaling, bone morphogenetic protein (BMP) pathways, and the release of growth factors such as VEGF and IGF-1. Collectively, these processes restore bone homeostasis, facilitate regeneration, and ensure long-term skeletal stability.

Osteoclast transcription factors

Transcription factors are critical regulators of gene expression, and their dysregulation can contribute to the development of pathological conditions. In periodontitis, key transcription factors, such as c-Fos, NFATc1, and c-Src play pivotal roles in promoting inflammation and pathological bone resorption by enhancing osteoclastogenesis. Below, we describe the functions and implications of these transcription factors in the context of periodontal disease.

c-Fos

c-Fos is a component of the activator protein-1 (AP-1) transcription factor complex, which is activated in response to diverse extracellular stimuli, such as mechanical stress and inflammatory cytokines.¹³⁹ It plays a central role in regulating osteoclast differentiation and function.¹⁴⁰ As a member of the Fos gene family, alongside Fra-1, Fra-2, and FosB, c-Fos forms heterodimers with Jun family proteins (e.g., c-Jun, JunB, JunD) to bind AP-1 DNA motifs and regulate gene expression.¹⁴¹

The expression of c-Fos is strongly linked to RANKL signaling. It interacts with other components of the AP-1 complex, including Fosl1, to regulate genes essential for osteoclastogenesis.¹⁴² c-Fos is a crucial determinant in the lineage commitment of progenitor cells, directing them toward the osteoclast rather than macrophage lineage.¹⁴³ Mice deficient in c-Fos display impaired osteoclast formation and an expansion of bone marrow macrophages, emphasizing its role in skeletal homeostasis and inflammatory bone disease.¹⁴⁴

High levels of c-Fos are expressed in osteoclast precursors and mature osteoclasts, driving the expression of diverse osteoclast-specific genes.^145,146 In apical periodontitis¹⁴⁷ and periodontitis,¹⁴⁸ c-Fos-mediated signaling enhances osteoclastogenesis and contributes to alveolar bone resorption. Furthermore, mice lacking c-Fos exhibit elevated levels of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-1β, accompanied by reduced expression of IL-10, suggesting that c-Fos modifies the inflammatory response.¹⁴⁹ Notably, preclinical studies in rat models have demonstrated that pharmacological inhibition of c-Fos attenuates alveolar bone loss, highlighting its therapeutic potential.¹⁵⁰

NFATc1

The nuclear factor of activated T-cell cytoplasmic 1 (NFATc1) is a master transcription factor that governs osteoclast differentiation and bone-resorptive activity.^151–154 NFATc1 is activated downstream of RANKL signaling and promotes the expression of genes encoding key osteoclastic proteins, including cathepsin K, TRAP, and integrin β3.¹⁵⁵ The initial activation of NFATc1 involves cooperation with NF-κB and NFATc2, with c-Fos playing a supportive role.¹⁵⁶ The p38 MAPK pathway contributes to the induction of both c-Fos and NFATc1, facilitating sustained NFATc1 expression via an auto-amplification loop.¹⁵⁷ The crucial role of NFATc1 in osteoclast formation induced by RANKL and TNF is evident from findings showing that both cytokines can still promote osteoclastogenesis in progenitor cells lacking p50 and p52.¹⁵⁸ NF-κB is essential for heightening the transcription of c-Fos and NFATc1. However, neither c-Fos nor NF-κB appears to be required for initiating the expression of genes critical for progenitor cell differentiation and fusion, supporting the idea that NFATc1 serves as a key regulator of osteoclast formation.¹⁵⁹ Nevertheless, since osteoclastogenesis in NFATc1-overexpressing progenitors lacking p50/p52 still depended on RANKL or TNF, it appears that NFATc1 alone is insufficient to drive osteoclast differentiation. During periodontitis, increased RANKL levels lead to upregulation of NFATc1, driving osteoclast differentiation and alveolar bone resorption.^155,160

Animal studies have highlighted the importance of NFATc1 in periodontal bone loss. For example, treatment with a novel benzamide-linked molecule significantly downregulated NFATc1 and other osteoclast-related genes, reducing osteoclastogenesis and protecting against ligature-induced bone loss.¹⁶¹ Additionally, mice with myeloid-specific deletion of encoding protein tyrosine phosphatase 1B (Ptpn1) showed diminished osteoclast formation and alveolar bone loss in ligature-induced periodontitis associated with lower NFATc1 expression and reduced RANKL-mediated signaling.¹⁶² In vitro, osteoclast precursors from Ptpn1-deficient mice exhibited impaired proliferation, decreased expression of osteoclast markers, and suppressed NFATc1 activity.¹⁶² Moreover, pharmacological inhibition of NFATc1 using CsinCPI-2, a cysteine peptidase inhibitor derived from the orange tree, attenuated alveolar bone destruction in experimental periodontitis, underscoring its therapeutic value.¹⁶³ Notably, combined deficiency of NFATc1 and c-Fos drastically impairs osteoclast fusion and resorptive function, further validating their synergistic role in osteoclastogenesis.¹⁶⁴

c-Src

c-Src is a non-receptor tyrosine kinase and chromatin-remodeling factor that modulates gene accessibility and osteoclast function.¹⁵¹ Although less extensively studied in periodontitis than c-Fos and NFATc1, c-Src has emerged as a key regulator of bone resorption and inflammatory signaling.¹⁵¹ In osteoclasts, c-Src promotes actin ring formation and cytoskeletal organization, essential for bone resorptive activity.¹⁶⁵ It also modulates transcriptional programs by facilitating the binding of NFATc1 and c-Fos to their target genes. Additionally, c-Src inhibits RUNX Family Transcription Factor 2 (RUNX2), suppressing osteoblast activity and shifting the balance toward bone resorption.¹⁶⁵ Dysregulation of c-Src may enhance inflammatory responses and exacerbate osteolytic bone loss.¹⁶⁶

Preclinical studies have begun to unravel the involvement of c-Src in periodontal bone destruction. For instance, lipopolysaccharide (LPS), a component of Gram-negative bacterial cell walls, promotes c-Src expression in bone marrow-derived macrophages via TNF-α signaling. First, LPS induces TNF-α, which upregulates c-Src expression, contributing to osteoclast differentiation and bone resorption.¹⁶⁷ A recent study by Mao et al. investigated the effects of the immunomodulatory agent C-176, a stimulator of interferon genes (STING) pathway inhibitor, in a mouse model of apical periodontitis. STING-deficient mice exhibited reduced inflammatory infiltration, osteoclast activity, and bone loss compared to wild-type controls.¹⁶⁸ In vitro, LPS stimulation reduced TRAP⁺ cells and downregulated osteoclast-related genes, including c-Src and Acp5 in macrophage cultures, further implicating c-Src in inflammatory bone loss.¹⁶⁸

In summary, c-Fos, NFATc1, and c-Src are critical transcription regulators that control osteoclast differentiation, activation, and inflammatory gene expression. Their dysregulation in periodontitis contributes to excessive bone resorption and periodontal tissue destruction. A deeper understanding of their molecular interactions and regulatory networks provides promising avenues for developing targeted therapies to mitigate bone loss and preserve periodontal health.

Main signaling pathways in periodontitis

The host immune responses are orchestrated by intracellular signaling pathways that convert extracellular cues, including microbial products and cytokines, into changes in gene expression.³⁹ Ligand binding to membrane receptors, such as receptor tyrosine kinases and G protein-coupled receptors, initiates signal transduction cascades, i.e., a sequence of events in which a signal is transmitted from the cell surface to the inside of the cell, resulting in a specific cellular response.^169,170 Dysregulation of signaling cascade, through prolonged kinase activation or phosphatase inhibition, can result in chronic inflammation and tissue damage, as seen in periodontitis.^155,170 Therefore, a comprehensive understanding of how these pathways regulate immune responses is essential for identifying molecular targets to control periodontal inflammation and bone loss.^39,171,172 Key signaling pathways in periodontitis include NF-κB, JAK/STAT, MAPK, PI3K/Akt, and Wnt/β-catenin.

Nuclear factor kappa B (NF-κB) pathway

The NF-κB pathway is a central mediator of inflammation and plays a key role in the immunopathogenesis of periodontitis.³⁹ It is activated by microbial components such as LPS and by pro-inflammatory cytokines including TNF-α and IL-1β. NF-κB, a heterodimeric molecule, comprises a family of transcription factors, including c-Rel, RelB, RelA (p65), p50, and p52, which remain inactive in the cytoplasm by binding to inhibitory IκB proteins (Fig. 7).¹⁵⁷ Upon activation of TLRs, TNF receptors, or IL-1 receptors, adaptor proteins like myeloid differentiation factor 88 (MyD88) and TNF receptor-associated factor (TRAFs) are recruited, triggering the IκB kinase (IKK) complex.¹⁷³ IKK-mediated phosphorylation targets IκB for degradation, allowing NF-κB dimers (e.g., p65:p50) to translocate into the nucleus and induce transcription of genes involved in inflammation, immune cell activation, and survival.¹⁷⁴

Fig. 7.

Schematic representation of the NF-κB signaling pathway. The key components of the MAPK pathway involved in signal transduction are highlighted. When a ligand binds, most TLRs attract the connector protein MyD88, activating upstream signaling molecules, including TRAF6, IRAK, and TAK1. These molecules trigger the stimulation of MAP kinase and NF-κB signaling pathways. Additionally, pattern recognition receptors (PRRs) like Nod1 and Nod2 detect peptidoglycan remains from microbial cell walls, which can further enhance TLR-mediated stimulation of these signaling pathways. Once initiated, MAP kinases and NF-κB move into the nucleus, binding to specific promoter regions, including AP-1 and NF-κB, to initiate the transcription of early-response and inflammatory genes. This leads to mRNA production and an increase in cytokine synthesis. Furthermore, p38 MAP kinase plays a crucial role in the regulation of post-transcriptional regulation of pro-inflammatory genes, including IL-6, IL-1, and TNF, by modulating mRNA stability in the cytoplasm. Iwasaki, A. (2025). Adapted from “NF-KB Signaling Pathway.” Retrieved from https://app.biorender.com/biorender-templates. Created with BioRender.com (License Number: RK28PGBXMA)

There are two NF-κB activation pathways: (1) The canonical pathway, rapidly triggered by pro-inflammatory cytokines, primarily involves IKKβ and promotes the expression of inflammatory genes via p65:p50.¹⁷⁵ The p65/p50 heterodimer is translocated to the nucleus, where it binds to specific response elements in the promoters of various osteoclastic genes. (2) The non-canonical pathway, activated by signals such as RANKL and lymphotoxin-β, involves IKKα and activates RelB:p52 dimers, which are important for adaptive immune regulation and lymphoid organogenesis.¹⁷⁶ Tight regulation of NF-κB is essential to prevent tissue damage. Feedback mechanisms, including de novo IκB synthesis and deubiquitinating enzymes, limit pathway activation.¹⁷⁷ NF-κB also interacts with other signaling cascades, such as PI3K/Akt and MAPK, to fine-tune the inflammatory response.¹⁷⁸ Prolonged or dysregulated activation of this pathway contributes to chronic inflammatory and autoimmune conditions.¹⁷⁹

During periodontitis, NF-κB activation is a key driver of chronic inflammation and bone loss.³⁹ Bacterial challenge and immune activation stimulate NF-κB signaling in periodontal tissues, producing IL-6, IL-1β, and TNF-α cytokines.⁷ These cytokines promote the recruitment and activation of macrophages and neutrophils, perpetuating local inflammation.⁷ Importantly, NF-κB regulates osteoclastogenesis by modulating the RANK/RANKL/OPG axis.¹⁸⁰ Thus, NF-κB signaling contributes directly to alveolar bone loss in periodontitis.^180,181 Preclinical studies in rat models have shown distinct temporal patterns of NF-κB activation depending on the disease trigger. Ligature-induced periodontitis leads to rapid but transient activation of NF-κB, p38, and ERK, whereas LPS-induced models show delayed but sustained signaling.¹⁷² Therapeutically, blocking NF-κB activation, either directly or by modulating upstream signals such as TLRs or TRAFs, has been shown to reduce periodontal inflammation and bone resorption.

Janus kinase/signal transducer and activator of transcription JAK/STAT pathway

The JAK/STAT pathway is a key mediator of cytokine signaling.¹⁸² In the context of periodontitis, this pathway is activated by several cytokines and growth factors that bind to membrane receptors, stimulating JAK kinases, which phosphorylate STAT proteins.¹⁸² Once phosphorylated, STATs dimerize and translocate to the nucleus to regulate gene expression in inflammation and bone remodeling (Fig. 8).³⁹

Fig. 8.

Schematic representation of the JAK/STAT signaling pathway. The main components involved in this pathway are outlined to illustrate the signal transmission process. The initiation of this signaling pathway happens in a series of well-coordinated steps: growth factors and cytokines bind to their corresponding receptors, which leads to receptor dimerization and the recruitment of associated Janus kinases (JAKs). JAK commencement then induces tyrosine phosphorylation of the receptor, generating docking sites for signal transducer and activator of transcription (STAT) proteins. Next, JAKs phosphorylate STATs on tyrosine residues, prompting the STAT proteins to detach from the receptor. These phosphorylated STATs subsequently form either homodimers or heterodimers and move to the nucleus, where they attach to certain DNA structures to regulate the expression of target genes. Iwasaki, A. (2025). Adapted from “Cytokine Signaling through the JAK-STAT Pathway.” Retrieved from https://app.biorender.com/biorender-templates. Created with BioRender.com (License Number: GK28PGBOQ8)

During periodontitis, chronic inflammation driven by microbial infection activates cytokines such as IL-6, IL-1β, TNF-α, and IL-17A, which engage the JAK/STAT pathway and promote the expression of pro-inflammatory mediators and osteoclastogenic factors.³⁹ Among the STAT proteins, STAT3 has emerged as particularly relevant in periodontitis. IL-6-induced STAT3 activation promotes M1 macrophage polarization, a pro-inflammatory phenotype, and enhances the expression of osteoclastogenic factors, including RANKL and M-CSF.³⁹ Elevated STAT3 phosphorylation has been found in inflamed gingival tissues and is associated with greater disease severity. Experimental inhibition of STAT3 leads to reduced cytokine expression and decreased bone loss in both in vitro and in vivo models.¹⁸³ IL-17A, produced mainly by Th17 cells, also engages the STAT3 pathway and contributes to matrix degradation and osteoclast activity. Collectively, these findings position the IL-6/STAT3 and IL-17A/STAT3 axes as critical drivers of periodontal tissue destruction and potential therapeutic targets. Arce et al.¹⁸⁴ showed that STAT3 phosphorylation is elevated in gingival tissues during periodontitis, particularly in T cells and keratinocytes. Genetic and pharmacological inhibition of STAT3 reduced alveolar bone loss, reinforcing its pathogenic role.¹⁸⁴

Suppressors of cytokine signaling (SOCS) proteins, particularly SOCS1 and SOCS3, are endogenous inhibitors of the JAK/STAT pathway.¹⁸⁵ Studies using LPS-induced and ligature-induced periodontitis models have shown that SOCS3 expression correlates with the severity of inflammation and regulates STAT3 activity.¹⁸⁵ These findings suggest that modulating SOCS proteins could provide a strategy to control periodontal inflammation and tissue destruction.

Mitogen-activated protein kinase pathway

The MAPK pathway is a key signaling cascade that regulates cellular responses to stress, cytokines, and microbial stimuli.³⁹ It comprises three main subfamilies: p38 MAPKs, c-Jun N-terminal kinases (JNKs), and extracellular signal-regulated kinases (ERKs). Each of these kinases contributes to the transcriptional regulation of pro-inflammatory genes and the activation of osteoclast-related signaling in periodontal disease.³⁹ MAPK signaling is initiated through receptor engagement, such as growth factor or cytokine receptors, which activate a kinase cascade recognized as the Ras-Raf-MEK-ERK pathway, culminating in the phosphorylation of transcription factors and gene expression (Fig. 9).¹⁸⁶

Fig. 9.

Graphic illustration of the MAPK signaling pathway. The MAPK/ERK pathway components are highlighted to demonstrate signal transmission in periodontal tissues, where PRRs and innate immune signaling pathways play crucial roles. TLRs, such as TLR-2, TLR-4, and TLR-9, are prominently expressed in these tissues. After the binding of a ligand, most TLRs recruit MyD88, an adaptor protein which activates common upstream molecules, such as TAK1, TRAF6, and IRAK, subsequently triggering the MAP kinase and NF-κB pathways. Once activated, these kinases translocate to the nucleus, where they bind to specific promoter motifs, such as NF-κB and AP-1, on target genes, comprising inflammatory genes and early responses, to promote their transcription into mRNA, leading to increased cytokine production. Moreover, p38 MAP kinase contributes to the post-transcriptional regulation of pro-inflammatory genes, such as IL-6, by influencing mRNA constancy into the cytoplasm. Created with BioRender.com (License Number: RI28PGCL8E)

During periodontitis, pro-inflammatory cytokines such as IL-1β, TNF-α, and IL-17A activate the MAPK pathway, leading to increased expression of MMPs, osteoclastogenic factors, and inflammatory mediators that drive tissue breakdown and bone resorption.³⁹ Among the MAPKs, p38 MAPK plays a particularly prominent role.¹⁴⁸ Inhibition of p38 MAPK suppresses osteoclastogenesis and protects against bone loss in experimental models of periodontitis.¹⁴⁸ JNK is also involved in periodontitis by regulating the AP-1 transcription factor, composed of c-Jun and c-Fos, essential for osteoclast differentiation.¹⁸⁷ JNK signaling enhances the expression of inflammatory cytokines and MMPs, and contributes to bone resorption.¹⁷⁹ Inhibition of JNK promotes bone regeneration and periodontal repair in preclinical models.¹⁸⁸ Pro-inflammatory stimuli similarly activate ERK signaling and promote osteoclast differentiation and cytokine expression.¹⁸⁹ Cross-talk between ERK and p38 signaling pathways has been associated with enhances bone resorption in both periodontitis and arthritis.^190,191

Experimental models support the therapeutic potential of MAPK pathway inhibition.¹⁹² siRNA targeting MK2, a downstream target of p38, reduces LPS-induced inflammation and bone loss in rats.¹⁹³ Likewise, pharmacological inhibitors such as SD282 (p38 inhibitor) and Bortezomib (a proteasome inhibitor affecting MAPK-related pathways) have been shown to suppress cytokine expression, RANKL upregulation, and bone destruction in periodontitis models.¹⁹² Bortezomib also reduced the RANKL/OPG ratio and inhibited NF-κB, p38, and ERK activation in LPS-stimulated periodontal ligament cells.¹⁹⁴ Together, these findings underscore the central role of MAPK signaling, particularly the p38 MAPK and JNK branches, in periodontal inflammation and bone resorption, highlighting these kinases as promising therapeutic targets for periodontitis and related inflammatory bone diseases.

Phosphoinositide 3-kinase (PI3K)/Akt signaling pathway

The PI3K/Akt signaling is involved in a wide range of cellular processes, including cell growth, survival, metabolism, and immune regulation.¹⁹⁵ Dysregulation of the PI3K/Akt signaling pathway is increasingly recognized as a critical contributor to the pathophysiology of chronic inflammatory and bone-resorptive conditions, including periodontitis.¹⁹⁶ PI3K activation leads to the production of phosphatidylinositol (3,4,5)-trisphosphate (PIP3), which recruits Akt to the cell membrane, where it becomes phosphorylated and activated. Akt then modulates downstream targets such as mammalian target of rapamycin (mTOR), Forkhead box O1 (FOXO1), GSK3β, and NF-κB (Fig. 10).¹⁹⁶

Fig. 10.

Graphic illustration of the PI3K/Akt signaling pathway. The PI3K-Akt pathway components are highlighted to demonstrate signal transmission in periodontal tissues, where PRRs and innate immune signaling pathways play crucial roles. TLRs, such as TLR-4, are prominently expressed in these tissues. After the binding of a ligand, most TLRs recruit PI3K/Akt, which activates common molecules, such as TSC1/2, subsequently triggering the mTORC1 pathways. Once activated, these kinases translocate to the nucleus, where they bind to specific promoter motifs on target genes, comprising inflammatory genes and early responses, to promote their transcription into mRNA, leading to increased cytokine production. Team, B. (2025). Adapted from “PI3K/Akt, RAS/MAPK, JAK/STAT Signaling”. Retrieved from https://app.biorender.com/biorender-templates. Created with BioRender.com (License Number: PU28PGCWVG)

Through these downstream effectors, the PI3K/Akt pathway modulates the transcription of genes that govern osteoclast differentiation, matrix metalloproteinase expression, and inflammatory cytokine production.^127,197,198 In periodontitis, persistent activation of this pathway enhances osteoclastogenesis and promotes the survival of inflammatory cells within periodontal tissues, thereby amplifying tissue destruction and bone loss.^199–202 Additionally, impaired regulation of PI3K/Akt signaling may disrupt the balance between bone resorption and formation, contributing to the progressive and destructive nature of the disease. Given its central role in coordinating inflammatory and bone remodeling responses, targeting the PI3K/Akt pathway represents a promising therapeutic strategy for mitigating periodontal tissue degradation and preserving alveolar bone integrity.^{127,199–202}

Moreover, Akt signaling plays a pivotal role in controlling both the survival and differentiation of osteoclasts.¹²⁷ Within these cells, Akt functions as a key downstream mediator of signaling pathways initiated by three major surface receptors: colony-stimulating factor-1 receptor (c-Fms), αvβ3 integrin, and RANK. Upon stimulation with M-CSF and RANKL, Akt becomes phosphorylated and contributes significantly to osteoclastogenesis, primarily by modulating the activation of transcription factors NF-κB and NFATc1.^197,198 In a study by Moon et al. (2012), it was demonstrated that Akt facilitates osteoclast differentiation via the GSK3β/NFATc1 axis. Silencing Akt expression using small interfering RNA was shown to effectively block RANKL-induced osteoclast formation.¹⁹⁷ Furthermore, Akt appears to be particularly important during the early stages of osteoclast precursor development, as its inhibition disrupted osteoclast formation when induced by RANKL at these early differentiation stages, with less pronounced effects at later stages.¹⁹⁷

The wingless Int-1 (Wnt)/ β-catenin pathway

The Wnt/β-catenin signaling pathway is fundamental in cell fate determination, tissue homeostasis, and bone remodeling. Upon binding of Wnt ligands to Frizzled receptors, the β-catenin destruction complex is inhibited, allowing β-catenin to accumulate in the cytoplasm and translocate into the nucleus.²⁰³ There, it activates the transcription of target genes involved in cell proliferation, differentiation, and survival.²⁰³

Aberrant activation of this pathway, such as through mutations in the adenomatous polyposis coli (APC) gene, can result in excessive β-catenin accumulation and has been implicated in the pathogenesis of several cancers, including colorectal carcinoma.²⁰⁴ Although less studied in periodontitis, emerging evidence suggests that dysregulation of Wnt/β-catenin signaling may contribute to the imbalance between bone resorption and formation in periodontal disease.^205–208

Pathway interactions

Cell signaling pathways function as integrated networks, with extensive crosstalk allowing fine-tuned regulation of immune responses, tissue remodeling, and cell survival. For instance, the MAPK and NF-κB pathways can act synergistically to amplify inflammation, whereas pathways such as PI3K-Akt modulate cell survival and metabolism.²⁰⁹ On the other hand, activation of p38 MAPK initiates the senescence response in both immune and non-immune cells. Recent studies suggest that senescence, particularly stress-induced premature senescence (SIPS), plays a causal role in tooth-supporting bone loss during periodontitis.²¹⁰ Specifically, p38 MAPK: (1) stabilizes and activates p53, a central regulator of the DNA damage response; (2) induces the expression of p16Ink4a and p21Cip1, which inhibit cyclin-CDK complexes and enforce cell cycle arrest; (3) increases mitochondrial reactive oxygen species production and alters antioxidant enzyme expression, thereby disturbing redox homeostasis and promoting mitochondrial dysfunction; and (4) promotes the release of a senescence-associated secretory phenotype (SASP) enriched in RANKL and IL-17A.^211–214 Collectively, these mechanisms highlight the multifaceted role of p38 MAPK in shaping the senescent phenotype and mediating its pathological impact in periodontitis.²¹⁵ Understanding how these pathways interact is critical for identifying combinatorial therapeutic strategies that modulate inflammatory bone loss in periodontitis.

Signaling pathways as therapeutic targets

Various host-modulating strategies have been explored to complement periodontal therapy, showing promise in reducing tissue destruction and enhancing clinical outcomes when used alongside non-surgical interventions.^{11,49,216,217} These include anti-inflammatory agents, biochemical mediators, weak organic acids, antibiotics, natural compounds, and bone-protective drugs.^{11,49,216,217}

Significant progress has been made in treating osteoclast-driven diseases using agents such as bisphosphonates, denosumab, parathyroid hormone, and estrogen replacement therapy.^218–221 However, these treatments have notable limitations, including adverse effects, limited suitability for long-term use, and contraindications for oral administration. Additionally, these agents may impair immune function and exacerbate infections in immunocompromised patients or preclinical models.^11,127,217 As a result, growing attention has turned toward natural compounds for their potential to modulate biological pathways involved in inflammation and bone metabolism.²¹⁷ Active ingredients such as flavonoids, lignans, terpenoids, and limonoids have demonstrated anti-inflammatory, antioxidant, and anti-resorptive properties. These effects are largely mediated by their ability to inhibit RANKL-induced osteoclastogenesis and modulate key signaling pathways, including MAPK, NF-κB, and PI3K/Akt.¹¹

In periodontitis, therapeutic efforts increasingly target intracellular signaling cascades that drive inflammation and tissue degradation. Several animal models of induced experimental periodontitis have been widely used in research to unveil the in vivo effects of several different compounds to modulate signaling pathways and decrease inflammation and bone resorption.^222–225 Some of the most utilized animal models are described in Fig. 11.

Fig. 11.

Schematic Representation of Murine Models of Periodontitis Commonly Used to Investigate Therapeutic Strategies Targeting Signaling Pathways. The upper panel illustrates the ligature-induced bone loss model, in which a cotton, silk, or nylon thread is tied around the cervical portion of the teeth. This setup promotes bacterial accumulation, exacerbating the inflammatory process and leading to bone loss. Below, a representative model of oral inoculation with live bacteria is depicted. In this model, live bacteria are cultured under controlled conditions, mixed with carboxymethylcellulose, and applied directly into the animal’s mouth using a pipette. This method facilitates bacterial colonization in the gingival tissue surrounding the teeth, with the severity of tissue inflammation and bone loss being directly proportional to the number of applications and the duration of disease development. The third image in the panel presents a schematic of lipopolysaccharide (LPS) injections, administered directly into the palatal gingiva of the maxilla or mandible using a Hamilton syringe. Repeated LPS application amplifies the inflammatory response and promotes advanced bone resorption. Finally, in the bottom of the panel, the periapical disease model is induced by opening the crown of a tooth, with a round drill, to allow bacteria contamination inside the pulp chamber. The infection leads to the breakdown of alveolar bone surrounding the tooth apex. Created with BioRender.com (License Number: FX28PGAR2G)

Animal models have been instrumental in elucidating mechanisms underlying alveolar bone destruction in periodontitis, emphasizing the pivotal roles of osteoclasts and the innate immune response in health and in genetically modified mouse models.^226,227 Targeting key intracellular signaling molecules, including NF-κB, c-Fos, and NFATc1, have shown promise as therapeutic strategies in several animal studies.^{49,216,217,226} A review article made by Koide et al.²²⁶ elegantly examines how osteoblasts and bone marrow stromal cells control osteoclast differentiation and activity, as well as the mechanisms by which bacterial pathogens contribute to bone destruction in periodontitis. Potential treatment strategies to mitigate alveolar bone loss in periodontal disease, including inhibition of RANKL-RANK interaction, signaling molecules (NF-κB, c-Fos, and NFATc1) in osteoclast precursors, and inhibition of osteoclast function, were discussed and summarized in the next sections of this manuscript.

Therapeutic strategies targeting the NF-κB signaling pathway

Given its central role in inflammatory bone loss, the NF-κB pathway represents a promising therapeutic target in periodontitis and other osteolytic diseases such as rheumatoid arthritis and osteoporosis. Several natural compounds, most notably curcumin and resveratrol, as well as small-molecule inhibitors targeting IκB kinases (IKKs) or NF-κB itself, have shown potential in reducing inflammation and bone resorption in rodent models of experimental periodontitis.^228–230 Biological agents that neutralize TNF-α or IL-1β, key cytokines downstream of NF-κB activation, have been successfully used in treating systemic inflammatory disorders and may hold similar therapeutic value in periodontal disease and alveolar bone preservation.^231,232 Overall, a range of therapeutic approaches has been proposed to modulate NF-κB signaling or its downstream mediators to suppress chronic inflammation and mitigate excessive osteoclastic activity in periodontitis (Fig. 12).

Fig. 12.

Therapeutic Strategies Targeting the NF-κB Signaling Pathway. Various strategies have been proposed to modulate the alveolar bone resorption, including NF-κB signaling and its downstream effects, aiming to reduce inflammation and prevent excessive bone resorption, including small molecule inhibitors, natural compounds, biological agents, and strategies targeting RANK/RANKL/OPG. This figure illustrates and highlights all the beneficial effects of the aforementioned inhibitors on periodontitis. Created with BioRender.com (License Number: AL28PGAMUR)

Small molecule inhibitors

Small molecule inhibitors targeting NF-κB signaling have shown promise in managing inflammatory bone conditions such as periodontitis and rheumatoid arthritis.^233–235 These compounds primarily act by inhibiting key kinases involved in NF-κB activation, including IKKβ and NIK.

• BMS-345541, a selective IKKβ inhibitor, significantly reduces disease incidence and joint inflammation in collagen-induced arthritis models.²³³ Similarly, BMS-066, a newer IKKβ inhibitor, exhibits immunosuppressive effects at low doses by inhibiting Th17 activity, cytokine secretion, and osteoclastogenesis despite limited TNF-α inhibition.²³⁴ Although not yet studied in periodontitis, these findings support IKKβ as a viable target for inflammatory bone loss.

• Cpd33, an NF-κB-inducing kinase (NIK) inhibitor, suppresses RANKL-induced osteoclastogenesis and reduced bone resorption in both osteoporotic and periodontitis animal models, highlighting its role in modulating the non-canonical NF-κB pathway.²³⁶ Notably, local administration of Cpd33 in ligature-induced periodontitis mice reduces osteoclast numbers and the expression of inflammatory cytokines, including RANKL, in periodontal tissues, supporting its potential as a therapeutic agent for periodontitis.²³⁶

• AZD8835, a dual PI3K inhibitor, downregulates NF-κB, ERK, and AKT signaling pathways in vitro and attenuates alveolar bone loss in vivo.²³⁷ Using a rat model of periodontitis, AZD8835 effectively inhibited osteoclast differentiation and bone resorption, while suppressing the expression of key osteoclast markers, including TRAP and Cathepsin K.²³⁷

• NF-κB essential modulator (NEMO) binding domain (NBD), a cell-penetrating peptide inhibitor targeting the IκB kinase complex, effectively blocked RANKL-induced NF-κB activation and suppressed osteoclast formation in both cellular and animal models.²³⁸ These results suggest that specifically targeting NF-κB signaling could serve as a promising treatment strategy for chronic inflammatory conditions associated with excessive bone loss.²³⁸

• NAT10, an acetyltransferase involved in mRNA modification, has been shown to promotes inflammation through the NOX2–ROS–NF-κB axis.²³⁹ In an LPS-induced periodontitis mouse model, treatment with Remodelin, a selective NAT10 inhibitor, reduces macrophage infiltration, suppresses pro-inflammatory cytokine expression, and attenuates alveolar bone resorption.²³⁹

• Deferoxamine (DFO), a hypoxia mimetic, inhibits NF-κB activation and osteoclastogenesis in vitro and in vivo, enhancing HIF-1α expression.²⁴⁰ In a rat periodontitis model, DFO reduces inflammation and osteoclast formation by impairing F-actin ring assembly, downregulating osteoclast-specific markers, and attenuating alveolar bone loss.²⁴⁰

• Pirfenidone (PFD) significantly reduced alveolar bone loss and suppresses RANKL-induced TRAP⁺ osteoclast differentiation in a ligature-induced periodontitis mouse model.¹⁵⁰ Additionally, PFD inhibits the production of pro-inflammatory cytokines, including IL-1β, IL-6, and TNF-α, by blocking NF-κB signaling.¹⁵⁰

• IL-35, an anti-inflammatory cytokine, inhibits alveolar bone loss and modulates the immune response in a ligature-induced periodontitis mouse model.²⁴¹ It reduces osteoclast number and RANKL expression while increasing OPG levels in periodontal tissues. IL-35 also suppressed Th17 cells and associated cytokines (IL-6, IL-17A, IL-23) while enhancing Treg cells and the expression of IL-10, TGF-β1, and IL-35Ebi3. Notably, it decreases RANKL Th17 cells and the RANKL/OPG ratio, supporting its dual role in immune regulation and bone preservation.

• IL-37, an anti-inflammatory cytokine, reduces alveolar bone loss, osteoclast numbers, and pro-inflammatory cytokine expression in a periodontitis model. Specifically, treatment with recombinant human IL-37 (rhIL-37) significantly decreases IL-1β, IL-6, and TNF-α levels, while increasing IL-10 expression in periodontal tissues.²⁴² These effects are mediated by suppressing the NF-κB/NLRP3 signaling axis.²⁴²

• TPCA-1, an IKK inhibitor, exhibits dual effects: while promoting osteoclastogenesis in vitro under M-CSF and RANKL stimulation, it suppresses LPS-induced cytokine production and NF-κB activation, ultimately reducing alveolar bone loss in vivo.²⁴³

• A novel NIK inhibitor, 4-(3-((7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)-4-morpholinophenyl)-2-(thiazol-2-yl)-but-3-yn-2-ol, reduces osteoclastogenesis and inflammatory gene expression, and reverses alveolar bone loss in a chronic periodontitis model through inhibition of the non-canonical NF-κB pathway.²⁴⁴

These findings collectively support the therapeutic potential of small molecule inhibitors targeting canonical and non-canonical NF-κB signaling in periodontitis, offering novel avenues for inflammation and bone loss control.

Natural compounds

Several natural compounds have demonstrated the ability to inhibit NF-κB signaling, suppress inflammation, and prevent alveolar bone loss during periodontitis.

• Curcumin, a polyphenol derived from Curcuma longa, is a well-characterized NF-κB inhibitor that acts by blocking the phosphorylation of IκB proteins.²⁴⁵ In vitro and in vivo studies have shown that curcumin reduces pro-inflammatory cytokine expression, improves the OPG/RANKL ratio, and suppresses NF-κB activation.²³⁰ A chemically modified form, CMC 2.24, exhibits enhanced therapeutic efficacy in LPS-induced and diabetes-associated periodontitis rat models, significantly reducing inflammatory mediators, MMPs, and bone resorption by inhibiting p38 MAPK and NF-κB pathways.²³⁰ Notably, when CMC 2.2 is administered orally, it retains efficacy without altering systemic metabolic parameters such as blood glucose or body weight. These findings support the potential of curcumin and its derivatives as adjunctive therapies for periodontitis.

• Resveratrol, a polyphenol found in red wine, has demonstrated protective effects in periodontitis by inhibiting NF-κB signaling and reducing pro-inflammatory cytokine expression.²⁴⁶ In experimental models, it suppresses inflammation, osteoclast activity, and NLRP3 inflammasome activation.²⁴⁶ In diabetic mice with ligature-induced periodontitis, resveratrol improves glycemic control and attenuates bone loss, partly through TLR4 inhibition and downstream suppression of STAT3, p38 MAPK, and NF-κB pathways.²²⁸ Notably, it reduces the production of several cytokines, including TNF-α, IL-1β, IL-8, and IL-6.²²⁸ Encapsulation of resveratrol in nanoparticles further enhances its immunomodulatory effects in models of periodontitis and diabetes.^247,248 A chitosan-based thermosensitive injectable self-assembled hydrogel (TISH) loaded with granulocyte-macrophage colony-stimulating factor (GM-CSF) and resveratrol (TISH(GR)) to modulate immune responses and promote healing in periodontitis complicated by metabolic syndrome was evaluated.²⁴⁹ In vitro experiments revealed that TISH(GR) suppressed inflammatory signaling pathways (MAPKs and NF-κB) in dendritic cells (DCs) and promoted a tolerogenic phenotype, reducing pro-inflammatory cytokines (TNF-α, IL-6) while elevating anti-inflammatory IL-10 cytokine. In a high-fat diet (HFD)-induced periodontitis rat model, adjuvant therapy with TISH(GR) and scaling and root planing (SRP) significantly reduced inflammation and tissue destruction compared to SRP alone, as evidenced by decreased Th17 cell infiltration and enhanced periodontal healing. These findings highlight the potential of TISH(GR) as an immunomodulatory therapy to improve outcomes in periodontitis patients with metabolic comorbidities.²⁴⁹

• Tanshinone IIA, a lipophilic diterpenoid quinone derived from Salvia miltiorrhiza, a traditional Chinese medicinal herb, exerts anti-inflammatory and anti-resorptive effects by inhibiting Cathepsin K activity and suppressing NF-κB signaling.^247,248,250 In experimental periodontitis, tanshinone IIA and its sulfonate derivative significantly reduce inflammatory cell infiltration, osteoclast numbers, and the expression of IL-1β, IL-17A, and MMP-13, while improving bone density and collagen fiber integrity.²⁵¹

• Boldine, an aporphine alkaloid from the Chilean tree Peumus boldus, exerts anti-resorptive and immunomodulatory effects in ligature-induced periodontitis.²⁵² It significantly reduces alveolar bone loss and osteoclast numbers in a dose-dependent manner, downregulates RANKL, and increases OPG levels in periodontal tissues. Boldine also modulates the local immune profile by suppressing Th17 markers (IL-6, IL-17A, IL-23, RORγt) and enhancing Treg-associated cytokines and transcription factors (IL-10, IL-35, TGF-β1, Foxp3), leading to a lower RANKL/OPG ratio and restoration of Th17/Treg balance.

• Hesperetin, a flavonoid found in citrus fruits, inhibits RANKL-induced osteoclastogenesis and bone loss during osteoporosis by downregulating c-Fos, NFATc1, and other osteoclast markers via inhibition of MAPK and NF-κB pathways and activation of Nrf2/HO-1 signaling.²⁵³ Interestingly, while it prevents femoral bone loss under physiological conditions, high-dose oral administration aggravates alveolar bone loss and osteoclastogenesis in periodontitis, suggesting tissue-specific effects.²⁵⁴

• Lactobacillus rhamnosus extract, a probiotic-derived compound, inhibits osteoclast differentiation in vitro by suppressing the NF-κB/c-Fos/NFATc1 axis via TLR-2 signaling.²⁵⁵ In vivo, it reduces alveolar bone resorption in a murine periodontitis model, supporting the therapeutic potential of probiotic-based interventions.²⁵⁵

Biologic agents

Biologic therapies targeting cytokines downstream of NF-κB activation have shown efficacy in treating inflammatory bone diseases, including periodontitis.

• TNF-α inhibitors, such as adalimumab and infliximab, are widely used in inflammatory conditions. Infliximab reduces bone resorption and improves bone mineral content in rheumatoid arthritis by inhibiting TNF-α-induced NF-κB activation.²⁵⁶ In a ligature-induced periodontitis rat model, infliximab (5 mg/kg) significantly reduces alveolar bone loss, osteoclast numbers, and pro-inflammatory markers (MPO, TNF-α, IL-1β), while improving collagen fiber organization and suppressing RANK, RANKL, MMP-1, and MMP-8 expression.²⁵⁷

• Anakinra, an IL-1 receptor antagonist (IL-1Ra), inhibits NF-κB signaling and has demonstrate protective effects in inflammatory diseases.²⁵⁸ In osteoarthritis models, IL-1Ra restored autophagy and reduced extracellular matrix degradation by modulating Akt/mTOR/NF-κB and ULK1 signaling.²⁵⁹ In collagen-induced arthritis, IL-1Ra reduces inflammation, bone damage, and Th17 responses while enhancing Treg cell differentiation through STAT3/STAT5 modulation.²⁶⁰ In periodontitis-affected rats, IL-1Ra delivered via a thermosensitive hydrogel reduces TNF-α, IL-6, and IL-1β expression and alveolar bone loss in diabetic rats, highlighting its therapeutic potential.²⁶¹

• Tocilizumab, a monoclonal antibody targeting the IL-6 receptor, blocks IL-6-mediated NF-κB activation.²⁶² In ligature-induced periodontitis, tocilizumab reduces alveolar bone resorption, inflammatory infiltrates, and the expression of RANKL and Th17-related cytokines, suggesting its utility in controlling IL-6-driven periodontal inflammation.²⁶³

Targeting the RANK/RANKL/OPG axis

The RANK-RANKL-OPG signaling axis is central to osteoclast differentiation and bone resorption. Dysregulation of this pathway, particularly an increased RANKL/OPG ratio, is a hallmark of periodontitis and drives pathological bone loss. Targeting this axis has emerged as a promising therapeutic strategy for osteolytic diseases, including periodontitis.

• A previous study showed that OPG^−/− mice exhibit more severe alveolar bone resorption, particularly in cortical areas, compared to RANKL-overexpressing transgenic (Tg) mice, despite having higher circulating RANKL levels.²⁶⁴ This finding suggests that OPG deficiency exerts a more pronounced effect on bone resorption than RANKL overexpression alone. Microcomputed tomography and histological analysis revealed increased osteoclast activity and cortical porosity in OPG^−/− mice, while RANKL-Tg mice showed milder trabecular bone loss.²⁶⁴ Immunohistochemical staining identified osteocytes as a major source of OPG in cortical bone, indicating a protective role against osteoclast-mediated degradation. Additionally, systemic bone resorption markers such as serum TRAP5b activity and the RANKL/OPG ratio correlated with the severity of alveolar bone loss.²⁶⁴ Treatment of OPG^−/− mice with either anti-RANKL antibodies or risedronate significantly reduced bone loss and osteoclast numbers, with the anti-RANKL antibody showing particularly robust effects. These findings position OPG^−/− mice as a valuable model for screening anti-resorptive therapies, offering a reliable system to mimic accelerated bone destruction without the need for bacterial induction.^226,264

• Denosumab, a monoclonal antibody against RANKL, has demonstrated potent anti-resorptive effects and is widely used in osteoporosis and bone malignancies. In postmenopausal women, denosumab significantly reduced bone loss.²⁶⁵ In experimental periodontitis models, RANKL-Fc and OPG-Fc therapies also decreased osteoclast activity and alveolar bone resorption induced by P. gingivalis infection.²⁶⁶ However, denosumab use is associated with adverse effects, including osteonecrosis of the jaw and atypical femoral fractures, which limit its applicability in periodontitis.^267–270

• OPG acts as a decoy receptor for RANKL, preventing its interaction with RANK and thereby inhibiting osteoclastogenesis. Gene therapy with OPG reduces alveolar bone loss in mice by blocking RANKL–RANK binding.²⁷¹ OPG-based approaches have shown promise in mitigating bone resorption in inflammatory diseases, including periodontitis.²⁷² However, similar concerns about osteonecrosis of the jaw will likely limit OPG-centered approaches for periodontitis management.

In summary, therapeutic strategies targeting the RANK-RANKL-OPG pathway effectively control bone destruction in periodontitis. While biologics such as denosumab show strong efficacy, safety concerns highlight the need for more targeted and context-specific approaches.

Therapeutic strategies targeting the JAK/STAT pathway

Given the central role of JAK/STAT signaling in mediating inflammation and bone resorption, several therapeutic approaches have been developed to modulate this pathway in periodontitis (Fig. 13).

• JAK inhibitors (JAKi) are small molecules that block JAK activation and subsequent STAT phosphorylation. Tofacitinib, a pan-JAK inhibitor, has demonstrated efficacy in the treatment of rheumatoid arthritis and inflammatory bowel disease.²⁷³ In periodontitis patients, tofacitinib reduces serum TNF-α and IL-6 levels and improves periodontal inflammation after three months despite unchanged bacterial plaque levels.²⁷⁴ In vitro and in vivo studies further support its immunomodulatory activity via inhibiting STAT1 and downregulating inflammatory genes in macrophages.²⁷⁵ However, JAKi may also enhance osteoclastogenesis through upregulation of c-Jun and NFATc1, warranting careful evaluation of their long-term effects on bone metabolism.²⁷⁵

• STAT3 inhibitors also offer therapeutic promise. Stattic, a small-molecule inhibitor of STAT3 dimerization and DNA binding, suppresses RANKL-induced osteoclastogenesis by downregulating NFATc1 and c-Fos expression and attenuates bone loss in an ovariectomy model.²⁷⁶ Stattic also inhibited STAT3 and NF-κB signaling, with minimal impact on MAPK pathways, supporting its potential role in managing inflammatory bone diseases; however, further studies in periodontitis are needed.²⁷⁶

Fig. 13.

Therapeutic Strategies Targeting the JAK/STAT Signaling Pathway. Various strategies have been proposed to modulate bone resorption, including JAK/STAT signaling and its downstream effects, aiming to reduce inflammation and prevent excessive bone resorption, including small molecule inhibitors and STAT inhibitors. This figure illustrates and highlights all the ameliorative effects of the aforementioned inhibitors on inflammatory bone loss. Created with BioRender.com (License Number: JH28PGAI6J)

Altogether, JAK/STAT-targeting agents, including JAK inhibitors, STAT3 blockers, and cytokine modulators, show promise for controlling periodontal inflammation and bone loss. Nevertheless, their clinical application requires further investigation to ensure safety and efficacy, particularly regarding immune suppression and long-term bone health.

Therapeutic strategies targeting the MAPK pathway

Given the central role of MAPK signaling in inflammation and osteoclastogenesis in periodontitis, several therapeutic strategies have been explored (Fig. 14).

• Among them, p38 MAPK inhibitors are particularly promising, as they block the expression of MMPs and pro-inflammatory cytokines, thereby reducing osteoclast differentiation.³⁸ In response to bacterial stimuli such as LPS, p38 MAPK activation increases cytokine expression and bone resorption.³⁸ In a periodontitis model in mice, oral administration of a p38α inhibitor reduces cytokine levels and alveolar bone loss.²⁷⁷ MKP-1 overexpression further dampens p38 activity, highlighting its regulatory role in periodontitis.²⁷⁷

• Berberine, a natural alkaloid, inhibits p38 MAPK and NF-κB signaling, reducing inflammation and bone resorption in ligature-induced periodontitis.²⁷⁸ Delivery via thermo-sensitive hydrogels enhances its anti-inflammatory and osteogenic effects through PI3K/AKT modulation.²⁷⁹

• Eupatilin, a flavone from Artemisia argyi, blocks RANKL-induced differentiation of Raw264.7 cells and bone marrow-derived macrophages (BMM) into mature osteoclasts.²⁸⁰ In vivo, Eupatilin also suppresses RANKL-induced osteoclastogenesis and MAPK/NF-κB signaling, reducing bone loss.²⁸⁰

• Litcubanine A (LA), an isoquinoline alkaloid from Litsea cubeba, exhibits anti-inflammatory effects by inhibiting macrophage chemotaxis and osteoclast differentiation. In periodontitis models, LA reduces bone loss by downregulating chemokine signaling and suppressing MAPK pathway activation.²⁸¹ Similarly, Lycii Radicis Cortex (LRC), derived from Lycium chinense Mill, decreases bacterial load, oxidative stress, and inflammation while preserving connective tissue and alveolar bone via MAPK/NF-κB modulation and RANKL/OPG regulation.²⁸²

• Napyradiomycin B4 (NB4), a marine-derived compound, inhibited RANKL-induced osteoclastogenesis by suppressing NFATc1 and MEK-ERK signaling, reducing alveolar bone loss in periodontitis mice.²⁸³ Likewise, Ugonin L, from Helminthostachys zeylanica, suppresses osteoclast differentiation, promotes apoptosis of mature osteoclasts, and inhibits ERK, p38, JNK, and NF-κB activation.²⁸⁴

• An immunomodulatory hydrogel (8AGPB) releasing 8-aminoguanosine (8AG) targeted second messengers in macrophages, modulating MAPK and NF-κB pathways. In periodontitis-affected mice, 8AGPB reduces inflammatory infiltrate, osteoclastic activity, and bone resorption.²⁸⁵

• Ginsenoside Rb3, tested in LPS-induced periodontitis, significantly downregulates osteoclast markers (NFATc1, MMP9, Cathepsin K, Acp5), suppresses p38 MAPK, ERK, and NF-κB signaling, and decreases p-ERK expression in gingival tissues after Rb3 treatment.²⁸⁶

• The JNK inhibitor SP600125 reduces cytokine expression and bone resorption in arthritis models,²⁸⁷ and when delivered via carrier systems, regulates both JNK and JAK-STAT pathways, diminishing osteoclastogenesis and protecting joint integrity.²⁸⁸

• ERK inhibitors such as PD98059, which block MEK activation, reduce ERK phosphorylation and pro-inflammatory cytokine expression in models of hyperglycemia-associated periodontitis.²⁸⁹ However, further clinical validation is required, as off-target effects remain a concern.

• PF-3845, a FAAH inhibitor, suppresses MEK, ERK, and IκBα phosphorylation, reducing NFATc1 expression, osteoclast activity, and bone resorption, in a ligature-induced periodontitis model.²⁹⁰ Similarly, chitooligosaccharide (CS) inhibits MAPK activation and osteoclast-related gene expression both in vitro and in vivo, effectively preventing bone loss in ligature-induced periodontitis.¹⁴⁸

Fig. 14.

Therapeutic Strategies Targeting the MAPK Signaling Pathway. Various strategies have been proposed to modulate bone resorption, including MAPK signaling and its downstream effects, aiming to reduce inflammation and prevent excessive bone resorption, including ERK, JNK and p38 inhibitors, natural compounds, and hydrogel loaded. This figure illustrates and highlights all the beneficial effects of the aforementioned inhibitors on periodontitis. Created with BioRender.com (License Number: DV28PGACMW)

In summary, p38, JNK, and ERK MAPKs are key mediators of inflammation and bone loss in periodontitis. Targeting these pathways with inhibitors or natural compounds offers promising strategies to mitigate disease progression. However, the potential for off-target effects and safety concerns has limited clinical translation.^38,291–293 Continued investigation is required to develop more selective and effective MAPK-targeted therapies for inflammatory bone diseases.

Therapeutic strategies targeting the PI3K/Akt signaling pathway

Considering the pivotal involvement of PI3K/Akt signaling in inflammation and osteoclast differentiation associated with periodontitis, various therapeutic approaches have been investigated (Fig. 15)

• GaELNs, a garlic-derived exosome-like nanovesicle, significantly suppresses pro-inflammatory cytokine and ROS levels production in vitro. In the mouse periodontitis model, local administration of GaELNs significantly reduces gingival inflammation and alveolar bone resorption, and decreases the expression of inflammatory mediators (TNF-α, IL-12, CXCL-12, and CXCL-5) through activation of the PHGDH/PI3K/Akt signaling pathway. Its upregulation enhances PI3K/AKT phosphorylation and downstream activation of mTOR, VEGF, and Nrf2, while suppressing NF-κB activity.²⁰² These findings support the potential application of GaELNs as a safe, effective, and plant-based nanotherapy for inflammatory periodontal diseases.

• Kaempferol, a natural flavonol, restores osteogenic capacity in LPS-treated PDLSCs, enhancing ALP activity, calcium deposition, and expression of osteogenesis-related genes and proteins through activation of the PI3K/Akt and P38 MAPK pathways.²⁹⁴ In a murine ligature-induced periodontitis model, kaempferol (10 mg/kg) significantly reduces alveolar bone loss (by ~63.8%), improve bone volume/tissue volume ratio and increased expression of OPN, EphrinB2, p-PI3K, p-Akt, and p-P38 in periodontal tissues of kaempferol-treated mice. These molecular and structural effects collectively confirmed that kaempferol promotes periodontal bone regeneration by targeting EphrinB2 and activating the PI3K/Akt and P38 signaling pathways.²⁹⁴

• Kurarinone (KR), a flavonoid derived from Sophora flavescens, significantly decreases the differentiation and function of osteoclasts derived from BMM, and inhibit the PI3K/AKT/GSK-3β signaling axis.¹⁹⁹ Using a mouse model of LPS-induced bone resorption, KR treatment significantly reduces bone loss, osteoclast numbers, and bone surface erosion. KR effectively mitigates inflammatory bone loss by inhibiting osteoclast differentiation and function, suppressing oxidative stress, and downregulating inflammatory mediators.¹⁹⁹ These effects are primarily mediated through the activation of the Nrf2 antioxidant pathway and inhibition of the PI3K/AKT signaling cascade.

• PD153035, an epidermal growth factor receptor (EGFR) inhibitor, was evaluated in a periodontitis mouse model. Fibulin-3 (FBLN3), a secreted extracellular matrix (ECM) glycoprotein, was shown to bind to EGFR, leading to activation of the PI3K/AKT signaling pathway, which is essential for its pro-M1 macrophage effects. Using the PD153035 inhibitor reverses FBLN3-induced activation of PI3K/AKT and mitigate M1 polarization, significantly reduces alveolar bone loss, decreases local expression of inflammatory cytokines (IL-1β and IL-6), and shift macrophage populations toward the anti-inflammatory M2 phenotype.²⁹⁵

• Kangfuxin, a traditional Chinese medicine extract derived from Periplaneta americana, significantly attenuates key pro-inflammatory cytokines (IL-6, IL-1β, and TNF-α) at both the mRNA and protein levels in a rat periodontitis model. Histological analysis further reveals reduced leukocyte infiltration and better preservation of alveolar bone and cementum structure compared to untreated periodontitis controls. Kangfuxin inhibits phosphorylation of PI3K/Akt and mTOR, indicating suppression of the PI3K/Akt/mTOR signaling pathway. The therapeutic effects of this fraction is abolished when a PI3K activator was co-administered, confirming that modulation of this pathway is central to its activity.²⁰⁰

• Capsaicin, an active component found in chili peppers, significantly promotes the proliferation, migration, and osteogenic differentiation of PDLSCs, as evidenced by increased ALP activity, mineralization, and expression of osteogenic markers (Runx2 and ALP) induced by P. gingivalis. Capsaicin activated the PI3K/AKT/mTOR signaling pathway, enhancing the phosphorylation of key proteins (PI3K, AKT, and mTOR). This activation was linked to the upregulation of osteogenic differentiation. In a mouse periodontitis model, capsaicin pretreatment significantly reduces alveolar bone loss and increases the expression of osteogenic proteins (ALP and Runx2), demonstrating its protective role against inflammation-induced bone resorption.²⁰¹

Fig. 15.

Therapeutic Strategies Targeting the PI3K/Akt Signaling Pathway. Several therapeutic strategies have been explored to modulate bone resorption by targeting the PI3K/Akt signaling pathway and its downstream effects. These approaches aim to attenuate inflammation and prevent excessive bone loss. They include the use of PI3K/Akt pathway inhibitors, natural compounds, growth factors, and exosomes. This figure summarizes and highlights the potential benefits of these therapeutic interventions in the context of periodontitis. Created with BioRender.com (License Number: EQ28PGD0KN)

Concluding remarks and future directions

Although periodontitis is characterized by progressive alveolar bone loss, longitudinal clinical and histological studies have consistently demonstrated that this tissue destruction does not occur in a continuous linear fashion. Instead, the disease typically follows an episodic pattern marked by bursts of activity interspersed with periods of quiescence or remission.^296,297 This fluctuating course poses a critical challenge when translating therapeutic interventions from animal models to human clinical application. In contrast to human disease, experimental animal models of periodontitis, particularly those involving ligature placement, bacterial inoculation or LPS injections, tend to induce acute and synchronized osteoclastic activity over relatively short durations (e.g., 7–21 days), which does not fully recapitulate the chronic, intermittent progression observed in clinical settings.^{222,224,225,298}

The timing and delivery of therapeutic agents, especially small molecules and biologics, must therefore be strategically optimized to align with the underlying pathophysiological dynamics of human periodontitis. Current clinical tools do not adequately detect early disease activation or predict exacerbations, nor are there reliable biomarkers to distinguish between active and quiescent phases, which could further limit research models. The identification of some biomarkers, such as elevated levels of active MMP-8 or IL-1β in gingival crevicular fluid or saliva, may enable precision-targeted therapy during periods of heightened osteoclastic activity.^299,300 This targeted approach could help minimize unnecessary exposure to expensive or potentially immunosuppressive biologics, such as anti-TNF-α or anti-RANKL monoclonal antibodies, outside of periods of active bone resorption. Moreover, site-specific delivery platforms, such as injectable hydrogels, bioresorbable microspheres, or mucoadhesive gels, are emerging as promising strategies to localize drug action while reducing systemic side effects.^{249,301–303}

Given the complex etiology and chronicity of periodontitis, future research should focus on integrating molecular diagnostics, real-time disease activity monitoring, and patient stratification to guide personalized therapy. Studies are needed to (1) define molecular signatures predictive of disease activation, (2) develop controlled-release systems tailored for oral environments, and (3) evaluate the long-term safety and cost-effectiveness of these interventions in well-designed clinical trials. Moreover, leveraging multi-omics approaches, including transcriptomics, proteomics, and metabolomics, could aid in identifying patient subtypes that are more likely to benefit from biologic therapies versus adjunctive natural compounds or host-modulating agents.

The use of dietary supplements such as flavonoids, probiotics, or other herbal medicine may offer supportive benefits due to their favorable safety profile. Despite their promising biological activities, the clinical relevance of these natural compounds remains uncertain due to some limitations.^304,305 Most human studies evaluating their efficacy have been of short duration, involved small sample sizes, and often lacked rigorous design. Moreover, bioavailability is a major concern, as many of these natural compounds have poor systemic absorption, rapid metabolism, and low tissue penetration when administered orally, thereby limiting their therapeutic concentrations at periodontal sites.^306,307 While they are generally safe and well-tolerated as dietary supplements,^304–307 their effects are often modest and unlikely to produce clinically meaningful improvements in the absence of conventional periodontal therapy. Therefore, although natural compounds may serve as supportive agents for maintaining periodontal health, particularly in individuals with systemic inflammation or at risk for disease progression, their stand-alone impact on arresting or reversing periodontitis remains questionable without robust, large-scale clinical validation. The translation of findings from acute animal models to the episodic nature of human periodontitis necessitates a precision medicine framework that incorporates temporal disease patterns, biomarker surveillance, and advanced delivery technologies.

In summary, understanding the signaling pathways involved in inflammatory diseases such as periodontitis is critical for developing targeted therapies that mitigate disease progression and tissue destruction. Central to this pathological process is the activation of intracellular signaling pathways triggered by microbial and inflammatory cues. Among these, the NF-κB signaling pathway coordinates immune responses and osteoclast differentiation. Its activation in periodontal tissues in response to microbial stimuli amplifies inflammation and bone resorption. Several therapeutic approaches have been proposed to target NF-κB signaling:

• Small molecule inhibitors offer direct suppression of NF-κB activation, attenuating inflammatory cytokine production and osteoclastogenesis. While promising, their clinical translation requires further validation regarding specificity and safety.

• Natural compounds, such as curcumin, resveratrol, tanshinone, and hesperetin, exhibit anti-inflammatory and anti-resorptive properties through NF-κB inhibition, offering biocompatible alternatives or adjuncts to conventional therapy.

• Biologic agents, including IL-1, IL-6, and TNF-α inhibitors, have demonstrated remarkable efficacy in reducing systemic and local inflammation and preserving alveolar bone. However, their high cost and potential long-term effects on oral immunity warrant cautious use in periodontitis management.

• Targeting the RANK-RANKL-OPG axis, with agents such as denosumab, directly disrupts osteoclastogenesis and bone resorption. Nonetheless, these therapies must be administered with care due to risks such as medication-related osteonecrosis of the jaw.

The JAK/STAT pathway regulates key aspects of immune cell activation, survival, and cytokine signaling. Its persistent activation contributes to chronic inflammation and bone loss in periodontitis and related diseases. JAK inhibitors, STAT3 inhibitors, and cytokine antagonists have shown therapeutic potential, yet further clinical trials are essential to determine their long-term safety and efficacy in oral inflammatory contexts.

Similarly, the MAPK signaling pathway, including p38 MAPK, JNK, and ERK cascades, governs cellular responses to inflammatory stress and microbial stimuli. Its activation promotes cytokine release and osteoclast differentiation, reinforcing periodontal tissue destruction. Therapeutic agents targeting MAPKs have shown efficacy in preclinical models; however, challenges such as off-target effects and systemic toxicity have hindered clinical advancement. Precision-targeted MAPK inhibitors, alongside delivery systems that enhance local bioavailability, may overcome these limitations.

The PI3K)/Akt signaling pathway is a critical intracellular cascade involved in regulating cell survival, proliferation, metabolism, and immune responses. Upon activation by RTKs, cytokine receptors, or G protein-coupled receptors, PI3K catalyzes the conversion of PIP2 to PIP3, leading to the recruitment and activation of Akt (protein kinase B). Activated Akt phosphorylates a wide range of downstream targets to promote cell growth and survival, while inhibiting apoptotic pathways. Dysregulation of PI3K/Akt signaling is implicated in various pathological conditions, including periodontitis and its modulation have shown efficacy in inhibiting alveolar bone loss during periodontitis progression.

In conclusion, signaling pathways such as NF-κB, JAK/STAT, and MAPK represent promising therapeutic targets for controlling inflammation and bone resorption in periodontitis, at least in animal models of induced periodontitis. Advances in small molecule inhibitors, natural compounds, biologics, and pathway-specific strategies continue to expand the therapeutic landscape. Future research and clinical trials should optimize efficacy, minimize adverse effects, and integrate these approaches into personalized treatment strategies for periodontitis and other inflammatory bone disorders.

Author contributions

Conceptualization: R.S.d.M.; Methodology: R.S.d.M., R.V., G.E.O., J.P.S., E.E., L.H.T., J.J.J.P.v.d.B., and S.T.; Validation: R.S.d.M., R.V., G.E.O., J.P.S., E.E., L.H.T., J.J.J.P.v.d.B., and S.T.; Investigation: R.S.d.M., R.V., G.E.O., J.P.S., E.E., L.H.T., J.J.J.P.v.d.B., and S.T.; Writing—original draft preparation: R.S.d.M.; Writing—review and editing: R.S.d.M., RV, GEO, JPS, EE, LHT, J.J.J.P.v.d.B., and S.T.; Supervision: R.S.d.M. All authors have read and agreed to the published version of the manuscript.

Funding

Rafael Scaf de Molon is currently supported by grant provided by the Sao Paulo Research Foundation - FAPESP. Grant #2023/15750-7.

Data availability

No original data were used in this manuscript.

Competing interests

The authors have no competing interests.