Mechanical tensile stress effects on the expression of bone sialoprotein in bovine cementoblasts.
OBJECTIVE
To develop a new cementoblast culture method and to detect bone sialoprotein (BSP) expression in response to high and low mechanical tensile stress in cementoblast in vitro.


METHODS
Cementoblasts were collected from the roots of newborn bovine teeth and were identified with cementum-derived attachment protein (CAP) antibody 3G9. Cell proliferation was evaluated by MTT [3-(4,5-dimethylthazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay, and mineralization was confirmed by von Kossa staining. Mechanical tensile stress was applied in vitro to the cementoblast with the use of a uniaxial four-point bending system with 2000 or 4000 microstrains, at a frequency of 0.5 Hz for 3, 6, 12, 24, or 36 hours. BSP mRNA level was quantified by real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR).


RESULTS
A large amount of cementoblast was observed to be expressing CAP. Cementoblasts had a proliferation tendency similar to that of osteoblasts but different from that of periodontal ligament (PDL) cells. Cementoblasts had the ability to become mineralized between osteoblasts and PDL cells. The mechanical tensile stress significantly up-regulated BSP mRNA expression, which reached a peak at 24 hours in both 2000 and 4000 microstrain groups (P < .01) and was tenfold and sixfold higher than that of controls, respectively. BSP expression dropped toward baseline levels at 36 hours in both groups.


CONCLUSIONS
Mechanical tensile stress up-regulated the expression of BSP. Low mechanical tensile stress induced earlier and more intensive up-regulation of BSP mRNA; this might represent the optimal stimuli for cementoblast activity.
Yu
Hongyou
H
Department of Orthodontics, University of Groningen, Groningen, The Netherlands.
Ren
Yijin
Y
Sandham
Andrew
A
Ren
Aishu
A
Huang
Lan
L
Bai
Ding
D
D000818:Animals; D000831:Animals, Newborn; D000906:Antibodies; D002113:Calcification, Physiologic / Q000502:physiology; D002417:Cattle; D002448:Cell Adhesion; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D004396:Coloring Agents; D003739:Dental Cementum / Q000473:pathology*; D003799:Dental Stress Analysis; D058574:Integrin-Binding Sialoprotein; D010006:Osteoblasts / Q000473:pathology; D010513:Periodontal Ligament / Q000473:pathology; D011506:Proteins / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012795:Sialoglycoproteins / Q000032:analysis*; D013314:Stress, Mechanical; D013778:Tetrazolium Salts; D013844:Thiazoles; D013997:Time Factors; D014092:Tooth Root / Q000473:pathology; D015854:Up-Regulation
D000906:Antibodies; D004396:Coloring Agents; D058574:Integrin-Binding Sialoprotein; D011506:Proteins; D012795:Sialoglycoproteins; D013778:Tetrazolium Salts; D013844:Thiazoles; C022616:thiazolyl blue