2001 Fiscal Year Final Research Report Summary
Development of a novel bone formation therapy by inhibition of the calponin gene expression
| Project/Area Number |
12557128
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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 | 展開研究 |
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| Research Field |
Orthopaedic surgery
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| Research Institution | Osaka Medical Center for Cancer and Cardiovascular Diseases |
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Principal Investigator |
TAKAHASHI Katsuhito Osaka Medical Center, Molecular Medicine, Chief, 病態生理学部門, 部長 (40211338)
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| Co-Investigator(Kenkyū-buntansha) |
YOSHIKAWA Hideki Osaka University, Orthopedic Surgery, Professor, 大学院・医学研究科, 教授 (60191558)
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| Project Period (FY) |
2000 – 2001
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| Keywords | Calponin / Bone Formation / Bone Fracture Repair / Antisense oligoDNA / Adenovirus |
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| Research Abstract |
Myoblast differentiation into osteoblast induced by recombinant human BMP-2 (40 ng/ml) was enhanced by addition of anti-sense oligoDNA of mouse calponin h1 into the culture medium. Myoblast was prepared from the thigh muscle by trypsin digestion. Phosphothioate-modified anti-sense oligoDNA (18 mers) to the mouse calponin h1 sequence including the initiation codon was designed and synthesized. Treatment with the anti-sense oligoDNA but not with control oligoDNA at the concentrations of 80 μg/ml for 48 hr reduced the calponin h1 mRNA expression and 32.4±3.9% of the cells were differentiated into the osteogenic-lineage cells as demonstrated by the expression of alkaline phosphatase. We also demonstrated the accelerated healing of experimental bone fracture by local delivery of the anti-sense oligoDNA with the pluronic F127 gel at 48 h after bone fracture. The rate of union three weeks after fracture, as assessed by radiographic and histological examination, was significantly increased in the presence of the anti-sense oligoDNA (50.9%, n=28/55) as compared with control groups (sense oligoDNA ; 31.3%, n=16/51, mutated anti-sense oligoDNA ; 28.6%, n=4/14, pluronic gel only ; 37.5%, n=6/16). We further constructed adenovirus vector harboring anti-sense cDNA of human calponin h1 for inhibition of calponin mRNA expression in human mesenchymal cells.
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