2007 Fiscal Year Final Research Report Summary
IDENTIFICATION OF ODONTOGENIC STEM CELLAND ITS MOLECULAR MECHANISM OF PROLIFERATION AND DIFFERENTIATION.
| Project/Area Number |
17390487
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Morphological basic dentistry
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| Research Institution | Kyushu University |
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Principal Investigator |
SAKAI Hidetaka Kyushu University, Faculty of Dental Science, Professor (80136499)
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| Co-Investigator(Kenkyū-buntansha) |
KOBAYASHI Ieyoshi Kyushu University, Faculty of Dental Science, Associate Professor (40243951)
KIYOSHIMA Tamotsu Kyushu University, Faculty of Dental Science, Associate Professor (20264054)
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| Project Period (FY) |
2005 – 2007
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| Keywords | tooth germ / generation, regeneration / cDNA subtraction / in situ hybrisization / anti-sense / organ culture / thvmosin beta 4 / protoeenin |
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| Research Abstract |
We previously examined the development of the mouse mandible, and demonstrated that odontogenesis occurs between embryonic day 10.5 (E10.5) and E12. Based on the histological findings, we performed cDNA subtraction between the E10.5 and E12 mandibles to detect any differentially expressed genes which might be involved in the initiation of odontogenesis. In these differentially expressed genes, thymosin beta4 (Tb4) was strongly expressed in the developing tooth germ. Inhibition assay using anti-sense ODN method showed developmental arrest of the tooth germ and hard tissue formation, thus demonstrating the Tb4 play an important role in the morphogenesis of the tooth germ and in dentinogenesis. Furthermore, Tb4 was proven to down regulate the expression of MMP-2/-9, nucleolin
Runx2/Cbfal, DSPP, AMEL, ENML, DMP-1 and AMBN mRNAs in the odontogenic epithelial cell. We also reported Runx2/Cbfal isoform-type specific functional roles in the development of tooth. The administration of type II/II
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I Runx2/Cbfal anti-sense ODNs into the culture media resulted in an arrest of tooth germ growth at the bud-like stage in cultured mandible taken from the 11-day-old embryos, while also causing the inhibition of the differentiation of odontogenic cells into ameloblast and odontoblast in cultured tooth germs taken from the 15-day-old embryos. The expression of DSPP, AMGN, AMBN, DMP-1 was shown to be markedly suppressed in cultured tooth germ by the semi-quantitative RT PCR method. These data indicate that the type II/III Runx2/Cbfal isoform is closely related to the development and differentiation of tooth germ. inhibition of translation by anti-sense ODN.
An inhibition of nucleolin by anti-sense ODN in cultured mouse mandibles at embryonic day 11.0 showed a marked growth inhibition of tooth germ. Meanwhile, no developmental arrest was found in the cultured tooth germ at E15.0 treated with nucleolin antisense AS S-ODN. A significant reduction in the midkine (MK)mRNA expression was observed in the mouse mandible after being treated with nucleolin anti-sense ODN. This inhibition assay indicated that nucleolin could thus be involved in the early stage of tooth germ initiation and morphologenesis, possibly by regulating the MK expression.
Protogenin (Prtg) has been identified as a previously unknown gene that we have found in prior study to be one of the highly expressed genes in the mouse mandible at embryonic day 10.5(E10.5).Both Prtg mRNA and protein expressions are widely demonstrated in the mesenchymal cells in the mandible at E10.5. Oral epithelial cells are also positive for in situ signal of Prtg. knockdown of Prtg function is likely to be responsible for inhibition of the tooth germ development. Less
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