Negative feedback between PTH1R and IGF1 through the Hedgehog pathway in mediating craniofacial bone remodeling
Yi Fan, Ping Lyu, Jiahe Wang, Yali Wei, Zucen Li, Shiwen Zhang, Takehito Ouchi, Junjun Jing, Quan Yuan, Clifford J. Rosen, Chenchen Zhou
Yi Fan, Ping Lyu, Jiahe Wang, Yali Wei, Zucen Li, Shiwen Zhang, Takehito Ouchi, Junjun Jing, Quan Yuan, Clifford J. Rosen, Chenchen Zhou
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Research Article Bone biology Development

Negative feedback between PTH1R and IGF1 through the Hedgehog pathway in mediating craniofacial bone remodeling

Abstract

Regeneration of orofacial bone defects caused by inflammation-related diseases or trauma remains an unmet challenge. Parathyroid hormone 1 receptor (PTH1R) signaling is a key mediator of bone remodeling whereas the regulatory mechanisms of PTH1R signaling in oral bone under homeostatic or inflammatory conditions have not been demonstrated by direct genetic evidence. Here, we observed that deletion of PTH1R in Gli1+ progenitors led to increased osteogenesis and osteoclastogenesis. Single-cell and bulk RNA-Seq analysis revealed that PTH1R suppressed the osteogenic potential of Gli1+ progenitors during inflammation. Moreover, we identified upregulated IGF1 expression upon PTH1R deletion. Dual deletion of IGF1 and PTH1R ameliorated the bone-remodeling phenotypes in PTH1R-deficient mice. Furthermore, in vivo evidence revealed an inverse relationship between PTH1R and Hedgehog signaling, which was responsible for the upregulated IGF1 production. Our work underscored the negative feedback between PTH1R and IGF1 in craniofacial bone turnover and revealed mechanisms modulating orofacial bone remodeling.

Authors

Yi Fan, Ping Lyu, Jiahe Wang, Yali Wei, Zucen Li, Shiwen Zhang, Takehito Ouchi, Junjun Jing, Quan Yuan, Clifford J. Rosen, Chenchen Zhou

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Figure 5

IGF1 plays an important role in regulating bone turnover under both homeostasis and inflammatory microenvironment.

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IGF1 plays an important role in regulating bone turnover under both home...
(A) Schematic diagram of the experimental design. (B) Venn diagram showing coexpressed genes among Control_Sham, Control_AP, PTH1R-cKO_Sham, PTH1R-cKO_AP (FPKM > 1). Male mice were used. FPKM, fragments per kilobase million. (C) Heatmap of representative genes associated with osteogenesis, osteoclastogenesis, and IGF signaling. n = 2 for each group. (D) GO analysis of Control_Sham versus PTH1R-cKO_Sham enriched GO terms related to insulin-like growth factor, bone formation, and bone resorption. (E) RT-qPCR of Igf1 and Igf2 expression in mandibles under control and inflammation. n = 6 for control and n = 4 for PTH1R-cKO. Male mice were used. (F) Violin plot of the expression of Igf1 and Igf2 in all clusters. (G) The expression level of Igf1 in reclustered MSC population. (H) The expression level of Igf1 and Igf2 in 4 MSC subclusters presented in violin plot. (I and J) Immunofluorescence staining and quantification showed upregulated IGF1+Gli1+ cells in periapical bone of Gli1CreER PTH1Rfl/fl Rosa26Ai14 male mice under homeostasis and AP conditions. Yellow dashed lines depict the region of distal root of the mandibular first molar. Boxed areas are shown at higher magnification. n = 4. (K) HE staining of alveolar bone of healthy individuals and patients with AP. (L and N) Immunofluorescence double staining of PTH1R and IGF1 and quantification of IGF1+PTH1R+ in healthy and inflammatory alveolar bone of human samples. Boxed areas are shown at higher magnification. n = 3. (M) PTH1R and IGF1 gene expression of human healthy and inflammatory alveolar bone tissues. n = 4 in healthy individuals and n = 9 in patients with AP. Scale bar = 100 μm (J and K), 25 μm (L). Significance is determined using unpaired 2-sided Student’s t tests between 2 groups and 2-way ANOVA with Tukey’s correction for multiple comparisons. Data are mean ± SEM. *P < 0.05, **P < 0.01, ****P < 0.0001.