2003 Fiscal Year Final Research Report Summary
Basic research on decelerating disc degeneration -Activation of nucleus pulposus cells-
| Project/Area Number |
14571406
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Orthopaedic surgery
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| Research Institution | Tokai University |
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Principal Investigator |
MOCHIDA Joji Tokai University, School of Medicine, Professor, 医学部, 教授 (50174347)
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| Co-Investigator(Kenkyū-buntansha) |
NOMURA Takeshi Tokai University, School of Medicine, Assistant Professor, 医学部, 講師 (60246121)
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| Project Period (FY) |
2002 – 2003
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| Keywords | intervertebral disc / nucleus pulposus cells / mesenchymal stem cells / coculture |
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| Research Abstract |
NP cells were cocultured with mesenchymal stem cells (MSCs) in order to evaluate the effect of mesenchymal stem cells on activation of NP cells in vitro and in vivo. Methods : NP cells were harvested from rabbit discs. A 12-well multiple culture plates were used for coculture. A membrane culture insert was used. NP cells were seeded into each well at 1.0 X 10^4 cells / well, with or without 1.0 X 10^4 MSCs inside a membrane culture insert. On day 7 of coculture, they were evaluated for cell proliferation by using WST-8, DNA synthesis by measuring the uptake of [3H]-thymidine and proteoglycan (PG) synthesis by measuring the uptake of [35S]-sulphate. Statistical analysis by Wilcoxon's signed-ranks test was used with significance set at p<0.01. Cocultured NP cells were then reinserted into disc degeneration models in rabbits. Disc degeneration was evaluated at 2, 4, 8 and 16 weeks post reinsertion. Results : In cell proliferation assay, NP cells seeded with MSCs inside the insert (group M) was 2.97 X 10^5 cells, where as NP cells cultured monolayer (group N) only counted 4.02 X 10^4 cells / well (p<0.01). The average disintegration per second (DPM) measured for [3H]-thymidine indicating DNA synthesis was 188.56±5.20 DPM / cell in group M and 9.54±0.65 DPM / cell in group N (p<0.01). Results for [35S]-sulphate demonstrated 64.16±1.66 DPM / cell in group M and 6.42±0.15 DPM / cell in group N (p<0.01). These results correlated well with the results of in vivo studies. Conclusions : Coculture of nucleus pulposus cells by mesenchymal stem cells resulted in an improvement of cell proliferation and matrix synthesis over monolayer culture and reinsertion of nucleus pulposus cells activated by mesenchymal stem cells using coculture method was more efficient than our previous studies in deceleration of intervertebral disc degeneration.
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