title: DLX3 mutation negatively regulates odontogenic differentiation of human dental pulp cells.
abstract: OBJECTIVE
  The purpose of this study was to investigate the role of a novel mutant DLX3 on the odontogenic differentiation of human dental pulp cells (hDPCs) in tricho-dento-osseous (TDO) syndrome.
  
  
  METHODS
  hDPCs were obtained from the healthy premolars, stably-expressing wild-type DLX3 (WT), novel mutant DLX3 (Mu) and control vector (NC) cells were generated using recombinant lentiviruses. The proliferation rates of WT-hDPCs and Mu-hDPCs were measured by CCK8 assay. Odonto-differentiation of hDPCs was assessed by alkaline phosphatase (ALP) activity assay, and mineralization ability was assessed by Alizarin red staining. Odontogenic markers, including DMP-1, DSPP, Nes, ALP, and DLX5, were analyzed using real-time polymerase chain reaction (qPCR). DMP-1 and DSPP expressions were further confirmed by Western blotting.
  
  
  RESULTS
  CCK8 results showed that the novel mutant DLX3 decreased the proliferation rate of hDPCs compared with wild-type DLX3. qPCR showed that the novel mutant DLX3 weakened odontogenic differentiation by downregulating the expression of odontogenic genes. These results were further confirmed by Western blotting and ALP activity assay. Additionally, Alizarin red staining showed that the novel mutant DLX3 decreased the mineralization of hDPCs compared with wild-type DLX3.
  
  
  CONCLUSIONS
  Novel de novo mutation of DLX3 significantly decreases the proliferation rate and inhibits the odontogenic differentiation and mineralization of hDPCs, suggesting that this novel mutation of DLX3 can influence the dentinogenesis in TDO syndrome.
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology, PR China.
authors.forename: Li
authors.initials: L
authors.lastname: Zeng
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology, PR China.
authors.forename: Na
authors.initials: N
authors.lastname: Zhao
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology, PR China.
authors.forename: Dong
authors.initials: D
authors.lastname: Han
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology, PR China.
authors.forename: Haochen
authors.initials: H
authors.lastname: Liu
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology, PR China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Central Laboratory, Peking University School and Hospital of Stomatology, PR China. Electronic address: kqwangyx@bjmu.edu.cn.
authors.forename: Yixiang
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology, PR China. Electronic address: kqfenghl@bjmu.edu.cn.
authors.forename: Hailan
authors.initials: H
authors.lastname: Feng
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation / Q000235:genetics; D049109:Cell Proliferation / Q000235:genetics; D002478:Cells, Cultured; D019465:Craniofacial Abnormalities / Q000235:genetics*; D003744:Dental Enamel Hypoplasia / Q000235:genetics*; D003782:Dental Pulp / Q000166:cytology*; D015536:Down-Regulation; D006201:Hair Diseases / Q000235:genetics*; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D009154:Mutation / Q000235:genetics*; D009805:Odontogenesis / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics*
chemical_list: C088334:Distal-less homeobox proteins; D018398:Homeodomain Proteins; D014157:Transcription Factors; D000469:Alkaline Phosphatase
keywords: DLX3 mutation; Human dental pulp cells; Odontogenic differentiation; Tricho-dento-osseous syndrome