To date, attempts to regenerate functional periodontal tissues (including cementum) are largely unsuccessful due to a lack of full understanding about the cellular origin (epithelial or mesenchymal cells) essential for root cementum growth. To address this issue, we first identified a rapid cementum growth window from the ages of postnatal day 28 (P28) to P56. Next, we showed that expression patterns of Axin2 and β-catenin within cementum-forming periodontal ligament (PDL) cells are negatively associated with rapid cementum growth. Furthermore, cell lineage tracing studies revealed that the Axin2⁺-mesenchymal PDL cells and their progeny rapidly expand and directly contribute to postnatal acellular and cellular cementum growth. In contrast, the number of K14⁺ epithelial cells, which were initially active at early stages of development, was reduced during rapid cementum formation from P28 to P56. The in vivo cell ablation of these Axin2⁺ cells using Axin2^CreERT2/+; R26R^DTA/+ mice led to severe cementum hypoplasia, whereas constitutive activation of β-catenin in the Axin2⁺ cells resulted in an acceleration in cellular cementogenesis plus a transition from acellular cementum to cellular cementum. Thus, we conclude that Axin2⁺-mesenchymal PDL cells, instead of K14⁺ epithelial cells, significantly contribute to rapid cementum growth.

Cementum, a specialized bony layer covering an entire molar root surface, anchors teeth into alveolar bone. Gli1, a key transcriptional activator in Hedgehog signaling, has been identified as a mesenchymal progenitor cell marker in various tissues, including the periodontal ligament (PDL). To address the mechanisms by which Gli1⁺ progenitor cells contribute to cementogenesis, we used the Gli1^lacZ/+ knock-in line to mark Gli1⁺ progenitors and the Gli1^CreERT2/+; R26R^tdTomato/+ line (named Gli1^Lin) to trace Gli1 progeny cells during cementogenesis. Our data unexpectedly displayed a biphasic feature of Gli1⁺ PDL progenitor cells and cementum growth: a negative relationship between Gli1⁺ progenitor cell number and cementogenesis but a positive correlation between Gli1-derived acellular and cellular cementoblast cell number and cementum growth. DTA-ablation of Gli1^Lin cells led to a cementum hypoplasia, including a significant reduction of both acellular and cellular cementoblast cells. Gain-of-function studies (by constitutive stabilization of β-catenin in Gli1^Lin cells) revealed a cementum hyperplasia. A loss of function (by conditional deletion of β-catenin in Gli1⁺ cells) resulted in a reduction of postnatal cementum growth. Together, our studies support a vital role of Gli1⁺ progenitor cells in contribution to both types of cementum, in which canonical Wnt/β-catenin signaling positively regulates the differentiation of Gli1⁺ progenitors to cementoblasts during cementogenesis.

Wnt–β-catenin signaling plays a key role in orthodontic tooth movement (OTM), a common clinical practice for malocclusion correction. However, its targeted periodontal ligament (PDL) progenitor cells remain largely unclear. In this study, we first showed a synchronized increase in Wnt–β-catenin levels and Axin2⁺ PDL progenitor cell numbers during OTM using immunostaining of β-catenin in wild-type mice and X-gal staining in the Axin2-LacZ knock-in line. Next, we demonstrated time-dependent increases in Axin2⁺ PDL progenitors and their progeny cell numbers within PDL and alveolar bones during OTM using a one-time tamoxifen-induced Axin2 tracing line ( Axin2^CreERT2/+; R26R^tdTomato/+). Coimmunostaining images displayed both early and late bone markers (such as RUNX2 and DMP1) in the Axin2^Lin PDL cells. Conversely, ablation of Axin2⁺ PDL cells via one-time tamoxifen-induced diphtheria toxin subunit A (DTA) led to a drastic decrease in osteogenic activity (as reflected by alkaline phosphatase) in PDL and alveolar bone. There was also a decrease in new bone mass and a significant reduction in the mineral apposition rate on both the control side (to a moderate degree) and the OTM side (to a severe degree). Thus, we conclude that the Axin2⁺ PDL cells (the Wnt-targeted key cells) are highly sensitive to orthodontic tension force and play a critical role in OTM-induced PDL expansion and alveolar bone formation. Future drug development targeting the Axin2⁺ PDL progenitor cells may accelerate alveolar bone formation during orthodontic treatment.

Notopterol, an active component isolated from the traditional Chinese medicine Notopterygium incisum Ting ex H.T. Chang, exerts anti-inflammatory activity in rheumatoid arthritis. However, its roles in suppression of inflammatory insults and halting progression of tissue destruction in periodontitis remain elusive. In this study, we reveal that notopterol can inhibit osteoclastogenesis, thereby limiting alveolar bone loss in vivo. In vitro results demonstrated that notopterol administration inhibited synthesis of inflammatory mediators such as IL-1β, IL-32, and IL-8 in LPS-stimulated human gingival fibroblasts. Mechanistically, notopterol inhibits activation of the NF-κB signaling pathway, which is considered a prototypical proinflammatory signaling pathway. RNA sequencing data revealed that notopterol activates the PI3K/protein kinase B (Akt)/NF-E2–related factor 2 (Nrf2) signaling pathway in LPS-stimulated human gingival fibroblasts, a phenomenon validated via Western blot assay. Additionally, notopterol treatment suppressed reactive oxygen species levels by upregulating the expression of antioxidant genes, including heme oxygenase 1 (HO-1), NAD(P)H quinone oxidoreductase 1 (NQO1), catalase (CAT), and glutathione reductase (GSR), indicating that notopterol confers protection against oxidative stress. Notably, inhibition of Akt activity by the potent inhibitor, MK-2206, partially attenuated both anti-inflammatory and antioxidant effects of notopterol. Collectively, these results raise the possibility that notopterol relieves periodontal inflammation by suppressing and activating the NF-κB and PI3K/AKT/Nrf2 signaling pathways in periodontal tissue, respectively, suggesting its potential as an efficacious treatment therapy for periodontitis.