Hemicentin-1 Promotes Human Dental Pulp Cell Migration Differentiation and Mineralization
Objectives: Hemicentin-1 (HMCN-1) is a member of fibulin family of secreted extracellular matrix glycoproteins. We have previously reported that HMCN-1 is highly expressed in the odontoblasts of lingual side (root) but not on the buccal side (crown) of mouse incisors, indicating an important role in regulating root dentin formation. We hypothesized that HMCN-1 plays an important role in the differentiation and migration of dental pulp cells (DPCs) during root odontogenesis. To test this hypothesis, we knocked down Hmcn-1 in human DPSc (HDPCs) and compared their proliferation, migration, differentiation and mineralization with scramble control. Methods: Human dental pulps were extracted from intact third molars from healthy 18-20 year-old adults. HDPCs were isolated using a previously published protocol and transfected with Hmcn-1 knockdown shRNA retroviruses-GFP-PURO. Cells transfected with scrambled shRNA were used as controls. Proliferation assays were carried out with cell counting and BrdU kits at 24 hours. Scratch assay was performed at day 3 and evaluated 24 hours later to determine cellular migration rate. RT-qPCR, alkaline phosphatase (ALP) and alizarin red staining were conducted in cells treated with mineralization-induction media for 7 days. Student t test was performed to detect statistical significance (α=0.05). Results: The RT-qPCR result indicated successful knockdown of Hmcn-1 expression. The cells deficient of HMCN-1 showed no difference in proliferation but significantly (p<0.05) reduced cellular migration (open surface area) in scratch test. The expressions of odontoblast-associated genes (Runx2, Osx, Dspp) were significantly lower in Hmcn-1 KD DPSCs compared to the scramble control. ALP and alizarin red staining showed much reduced ALP activity and mineralized nodules in Hmcn-1 KD DPSC cultures. Conclusions: The results of this study indicate that HMCN-1 plays an important role in HDPC migration, differentiation and mineralization but not in proliferation. Further studies are needed to elucidate underlying regulatory pathways and potential applications in regenerative therapies.
Division:IADR/AADR/CADR General Session
Meeting:2020 IADR/AADR/CADR General Session (Washington, D.C., USA) Location:Washington, D.C., USA
Year: 2020 Final Presentation ID:0242 Abstract Category|Abstract Category(s):Pulp Biology & Regeneration Research
Authors
Li, Chuwen
( West China School of Stomatology, Sichuan University
, Chengdu
, China
; Shanghai Ninth People’s Hospital, College of Stomatology, Shanghai Jiao Tong University School of Medicine
, Shanghai
, China
; National Clinical Research Center for Oral Diseases
, Shanghai
, China
; Shanghai Key Laboratory of Stomatology and Shanghai Research Institute of Stomatology
, Shanghai
, China
)
Cui, Yujia
( West China School of Stomatology, Sichuan University
, Chengdu
, China
)
Ye, Ling
( West China School of Stomatology, Sichuan University
, Chengdu
, China
)
Zheng, Liwei
( West China School of Stomatology, Sichuan University
, Chengdu
, China
)
Zhou, Xuedong
( West China School of Stomatology, Sichuan University
, Chengdu
, China
)
Zhang, Hai
( University of Washington
, Seattle
, Washington
, United States
)
Sun, Jianxun
( West China School of Stomatology, Sichuan University
, Chengdu
, China
)
Support Funding Agency/Grant Number: Youth Program of National Natural Science Foundation of China (No.81300847)
Financial Interest Disclosure: NONE