2001 Fiscal Year Final Research Report Summary
Experimental studies on allogeneic nerve graft using specific tolerance and cryopreservation
| Project/Area Number |
09671518
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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 | Keio University |
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Principal Investigator |
IKEGAMI Hiroyasu Keio University, Orthopedics, instructor, 医学部, 助手 (00193186)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAYAMA Shinichiro Keio University, Orthopedics, assistant professor, 医学部, 講師 (40138045)
ISHII Seika Keio University, Orthopedics, instructor, 医学部, 助手 (60255487)
NAKAO Yasushi Keio University, Orthopedics, instructor, 医学部, 助手 (30188883)
HORIUCHI Yukio Keio University, Orthopedics, assistant professor, 医学部, 講師 (10138125)
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| Project Period (FY) |
1997 – 2000
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| Keywords | Immunosuppression / Nerve graft / Monoclonal antibody |
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| Research Abstract |
Current studies have shown that monoclonal antibody (mAb) to cell surface adhesion molecules can prolong allogeneic graft survival. We examined the immunosuppressive effect of mAb therapy on peripheral nerve allografts in mice. Additionally, a method for long-term preservation of rat nerve tissue with viable Schwann cells was examined using a cryopreservation techniques.
After engraftment, recipients were treated with both anti-intercellular adhesion molecule-1 ICAM-1) and anti-lymphocyte function-associated antigen-1 (LFA-1) mAbs for 12 days. Serum mAb levels were assessed with flow cytometry. Rejection response and nerve regeneration were evaluated historically at 8 weeks. Subsequent skin graft at 9 weeks and cytotoxic T lymphocyte (CTL) assay at 12 weeks assessed recipient immune system. In the preservation study, nerve segments were treated with cooling down phase to -40℃ at the rate of -1℃ /minute in program freezer, suspending phase at -80℃ for 30 minutes and preservation phase in
… More
liquid nitrogen at -196℃ for 1 week. In the rapid-freeze group, nerve segments were directly placed into liquid nitrogen.
Although circulating mAbs were undetectable 3 weeks after final treatment, the allografts from mAb-treated recipients showed no rejection pattern and excellent nve regeneration, similar to the isografts, was observed. Conversely, the allograft from untreated animals showed poor nerve regeneration and severe rejection appearance. In the mAb-treated recipients, the mean survival time of nerve-donor skin grafts was significantly prolonged and CTL activity was markedly suppressed against nerve-donor cells. The programd-freeze group showed well-preserved stratiform construction of perineurium and numerous myelinated axons. In contrast, the rapid-freeze group showed complete destruction of the nerve architecture. In cell culture examinations, a large number of surviving spindle-shaped cells were observed in the programd-freeze group and non-freeze group, but no viable cell in the rapid-freeze group.
A short course of mAb therapy against ICAM-1 and LFA-1 induced specific tolerance and permitted excellent nerve regeneration in nerve allograft model. The viability of Schwann cells can be maintained if the peripheral nerve is preserved by the programd freezing method. Less
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