Differentiation ability of Gli1⁺ cells during orthodontic tooth movement

Orthodontic tooth movement (OTM) induces bone formation on the alveolar bone of the tension side; however, the mechanism of osteoblast differentiation is not fully understood. Gli1 is an essential transcription factor for hedgehog signaling and functions in undifferentiated cells during embryogenesis. In this study, we examined the differentiation of Gli1⁺ cells in the periodontal ligament (PDL) during OTM using a lineage-tracing analysis. After the final administration of tamoxifen for 2 days to 8-week-old Gli1-Cre^ERT2/ROSA26-loxP-stop-loxP-tdTomato (iGli1/Tomato) mice, Gli1/Tomato⁺ cells were rarely observed near endomucin⁺ blood vessels in the PDL. Osteoblasts lining the alveolar bone did not exhibit Gli1/Tomato fluorescence. To move the first molar of iGli1/Tomato mice medially, nickel-titanium closed-coil springs were attached between the upper anterior alveolar bone and the first molar. Two days after OTM initiation, the number of Gli1/Tomato⁺ cells increased along with numerous PCNA⁺ cells in the PDL of the tension side. As some Gli1/Tomato⁺ cells exhibited positive expression of osterix, an osteoblast differentiation marker, Gli1⁺ cells probably differentiated into osteoblast progenitor cells. On day 10, the newly formed bone labeled by calcein administration during OTM was detected on the surface of the original alveolar bone of the tension side. Gli1/Tomato⁺ cells expressing osterix localized to the surface of the newly formed bone. In contrast, in the PDL of the compression side, Gli1/Tomato⁺ cells proliferated before day 10 and expressed type I collagen, suggesting that the Gli1⁺ cells also differentiated into fibroblasts. Collectively, these results demonstrate that Gli1⁺ cells in the PDL can differentiate into osteoblasts at the tension side and may function in bone remodeling as well as fibril formation in the PDL during OTM.