| Budget Amount *help |
¥3,890,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥390,000)
Fiscal Year 2007: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
Fiscal Year 2006: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
To identify the unknown periodontal ligament-related molecules, we performed the following studies.
1. The changes of the tendon/ligament markers expression by induction of chondrogenic differentiation and TGF-β3 treatment in human periodontal ligament cells (HPLC)
Many tendon/ligament marker molecules may play a role in healing and maintenance of constancy of periodontal ligament as well as tendon/ligament. Further, transforming growth factor-β(TGF-β) stimulates tendon healing. In the present study, we assessed the changes of the tendon/ligament markers expression by induction of chondrogenic differentiation and TGF-β3 treatment in HPLC. When HPLC were aggregated into micropellets and incubated with chondrogenic differentiatin medium with TGF-β3, chondrocyte-like lacunae formation was not detected in HPLC pellets. However, the expression levels of periostin, periodontal ligament associated protein-1 (PLAP-1), biglycan and decorin genes were higher, but growth and differentiation factor-
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5 (GDF-5) and cartilage oligomeric protein (COMP) gene expression were lower levels, as compared with these pellets cultured with control medium. Further, scleraxis gene expression did not change throughout the culture periods. In monolayer cultures of HPLC, TGF-β3 stimulated expression of periostin, PLAP-1, biglycan, scleraxis and COMP, but suppressed GDF-5 expression, while, this cytokine had no effect on decorin expression. These results demonstrate that the expression of these tendon/ligament marker molecules in HPLC is modulated under the conditions. which induce chondrogenesis. as well as by the TGF-β3 stimulation.
2. Characteristics of side population cells existing in HPLC.
The various types of stem cells exist in side population (SP) effused strongly Hoechst 33342 dye. The SP cells in bone marrow express various mesenchymal stem cell (MSC) markers and are highly enriched for MSC activity. In the present study, we determined the characteristics of SP cells isolated from HPLC. SP cells and non-SP (main population : MP) cells were isolated from HPLC stained with Hoechst 33342 using flowcytometry. The frequency of SP cells existed in HPLC was mean 0.07% of total cells. Then, the gene expression profile, PDL-marker gene expression, activity of cell proliferation and multi-lineage differentiation were determined in SP cells and MP cells. Expression of CD73, a MSC marker gene, in SP cells was higher levels than that in MP cells, while MP cells expressed higher levels of extracellular molecule genes characterized in HPL (type I/III collagen, PLAP-1, periostin, decorin). Further, SP cells had higher cell proliferation activity than MP cells. However, activities of differentiation into osteoblasts and adipocytes were not different between the 2 cell types. Therefore, MSC-like cells may not be enriched in the SP fraction of HPLC. Less
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