The Mdm2-p53 axis links cementocyte survival to cellular cementum volume

Cementocytes are terminally differentiated cells embedded in cellular cementum, an important hard tissue covering the apical regions of tooth roots. However, the roles of cementocytes in cellular cementum remain enigmatic. Here, we show that Murine Double Minute 2 (Mdm2), an E3 ubiquitin ligase that plays vital roles in regulating cell proliferation, apoptosis, and differentiation to influence tissue or organ development, is highly expressed in the cementocytes of mice. To investigate the role of cementocyte-expressed Mdm2, Dmp1-Cre;Mdm2flox/flox (Mdm2 cKO)mice were obtained to inactivate Mdm2 in cementocytes. The results showed that Mdm2 was successfully ablated and Mdm2 cKO mice display increased cementocyte apoptosis and reduced cellular cementum volume. p53, the canonical substrate of Mdm2, was accumulated and hyperactivated in the cementocytes of Mdm2 cKO mice and in cultured IDG-CM6 cells (a cementocyte cell line) treated with Nutlin3a, an inhibitor of Mdm2. Further experiments showed that inactivation of 1 allele of p53 significantly rescued the increased cementocyte apoptosis and the decreased cellular cementum volume in Mdm2 cKO mice. Therefore, p53 is targeted by Mdm2 for degradation and mediates the role of Mdm2 in cementocyte survival and cellular cementum volume. Notably, Mdm2 cKO mice exhibited decreased differentiation of cementoblasts (the cell type primarily responsible for cementum deposition) and reduced rate of cellular cementum deposition. Meanwhile, OCCM-30 cells (a cementoblast cell line) showed diminished migration, proliferation, differentiation, and mineralization ability after culture with conditioned medium (CM) from Nutlin3a-pretreated IDG-CM6 cells. Intriguingly, Mdm2 cKO mice displayed significantly increased osteoclast formation and cementum resorption. Meanwhile, in vitro experiments verified that CM from Nutlin3a-pretreated IDG-CM6 cells induced osteoclast differentiation of bone marrow macrophages. Collectively, these results demonstrate that Mdm2-mediated degradation of p53 promotes cementocyte survival, and that cementocytes affect the cell behaviors of cementoblasts and osteoclasts through a paracrine mode to modulate cellular cementum volume. Cementocytes are terminally differentiated cells embedded in cellular cementum and derived from surface cementoblasts. Cementoblasts are well known as primary cells responsible for secretion and formation of cellular cementum. However, the roles and underlying mechanisms of cementocytes in cellular cementum remain enigmatic. In this study, we find that Murine Double Minute 2 (Mdm2) is expressed in the cementocytes of mice. Successful inactivation of Mdm2 in cementocytes leads to cementocyte apoptosis and cellular cementum hypoplasia in Mdm2 cKO mice. Further experiments show that p53 mediates the role of Mdm2 in cementocytes. Notably, apoptotic cementocytes impede the biological activities of cementoblasts and induce the differentiation of osteoclasts through a paracrine mode, which explains the phenotype pf cellular cementum hypoplasia in Mdm2 cKO mice. Our results demonstrate that cementocytes are indeed active regulators for cellular cementum homeostasis.