Piezo1 in Hypertrophic Chondrocytes Regulates Bone Homeostasis and Fracture Healing

Ion channels of the Piezo family are important mechanotransducers in many tissues and also influence bone formation. However, their role in different bone cell types is not fully elucidated. We recently showed that Piezo1 in osteocytes is essential for loading-induced bone formation. We further demonstrated that Piezo1 in chondrocytes is crucial for metaphyseal bone formation. During endochondral ossification in the growth plate and during bone fracture healing, chondrocytes differentiate into hypertrophic chondrocytes which then become apoptotic or transdifferentiate into osteoblasts. Here we investigated the hypothesis that Piezo1 in hypertrophic chondrocytes regulates endochondral ossification during bone development and fracture healing. To this end, we analyzed mice with Piezo1 inactivation specifically in hypertrophic chondrocytes ( Piezo1 Col10a1-Cre ). Male and female Piezo1 Col10a1-Cre mice and Cre − controls ( Piezo1 fl ) were sacrificed at the age of 12 weeks. Femora were analyzed by μCT and thereafter subjected to histology. In another experiment, 12-week-old female Piezo1 fl and Piezo1 Col10a1-Cre mice received a femur osteotomy, stabilized by an external fixator, and were euthanized 14 and 21 days post-fracture. Fracture calli of day 14 were subjected to (immuno)histology. Fractured femora of day 21 were subjected to 3-point-bending testing and thereafter analyzed by µCT and histology. Compared to Piezo1 fl mice, Piezo1 Col10a1-Cre mice displayed a significantly reduced bone volume/tissue volume (BV/TV) and trabecular number and a significantly increased trabecular separation. However, chondrocyte-to-osteoblast transdifferentiation did not seem to be disturbed, as the expression of SOX2 and RUNX2 in growth plate chondrocytes did not differ between Piezo1 fl and Piezo1 Col10a1-Cre mice. In the primary spongiosa, osteoblast numbers and surface did not differ in female mice and were rather slightly increased in male Piezo1 Col10a1-Cre vs. Piezo1 fl mice. However, osteoclast numbers and surface were significantly increased in both male and female Piezo1 Col10a1-Cre mice. Furthermore, numbers of RANKL + growth plate chondrocytes were significantly increased. 14 days post-fracture, fracture calli of Piezo1 Col10a1-Cre vs. Piezo1 fl mice also displayed significantly increased N.Oc/B.Pm and Oc.S/BS, and numbers of RANKL + hypertrophic chondrocytes were also significantly increased. After 21 days, fractured femora tended to display a reduced bending stiffness and showed a significantly reduced BV/TV in the callus, indicating disturbed fracture healing. Our results indicate an essential role of Piezo1 in hypertrophic chondrocytes for bone homeostasis and fracture healing. An increased osteoclastogenesis seems to cause the low bone mass phenotype and disturbed fracture healing in Piezo1 Col10a1-Cre mice which might be mediated by an increased RANKL expression in hypertrophic chondrocytes.