The Effect And Mechanism Of Primary Cilia Of Prx1~+ Cells On The Development Of Bone-tendon Interface In Mice

Background:Prx1⁺cells play an important role in the development of skeletal system.It has not been reported whether Prx1⁺participates in the development of bone-tendon Interface（BTI）.Primary cilia is an important component of progenitor cells.It has been reported that conditional knockdown of Intraflagellar transport protein 88（IFT88）on primary cilium can affect the biological behavior of cells.However,it has not been reported whether conditional knockout IFT88 of Prx1⁺cells will affect the development process of bone tendon interface and biological behavior of Prx1⁺cells.Methods:1,3,5,12,26,and 40 days old Prx1Cre ER;Rosa26td Tomato mice were used to analyze the real-time distribution of Prx1⁺cells.Primary cilium of Prx1Cre ER;Rosa26td Tomato;IFT88^fl/flmice were knocked out at 1,2,4,and 6 weeks after birth,and the specimens were harvested at 12weeks to analyze the effect of primary cilia on the fate of Prx1⁺cells.H&E staining,TB&FG staining,and microcomputed tomography were used to analyze the development of BTI.In vitro,Prx1⁺cells were extracted and obtained after sorting by flow cytometry.The expression level of IFT88 protein was knocked down by sh RNA,and it were applied to explore the effect of cilia on the proliferation and differentiation of Prx1⁺cells.The inhibitor of Smoothened（Smo）protein was used to reduce the degree of activation of hedgehog（Hh）signaling pathway,which was performed to understand the effect of Hh signaling on the differentiation capacity of Prx1⁺cells.Results:Prx1⁺cells are involved in the development of BTI,and in the early postnatal development,Prx1⁺cells are abundant in the BTI;in the late development,they are confined to the fibrocartilage layer until disappear.Locally at the BTI,knockout of cilia may attenuate the ability of Prx1⁺cells to differentiate into chondrocytes.Knocking out cilia in the early stage had a pronounced effect on BTI development,but less in the middle and late stages.In vitro,Knockdown of IFT88 by sh RNA be able to reduce the proliferation and differentiation ability of Prx1⁺cells.At the same time,inhibiting the Hh signaling pathway will also reduce the differentiation ability of Prx1⁺cells.The results suggest that primary cilia may regulate the biological function of Prx1⁺cells through the Hh signaling pathway localized in itself.Conclusion:Prx1⁺cells are deeply involved in the developmental process of BTI.In the early postnatal development,there are abundant Prx1⁺cells in BTI.Primary cilia mediate the biological function of Prx1⁺cells through the Hh signaling pathway and further affect the development process of the bone-tendon interface...