Development of a novel bone formation therapy by inhibition of the calponin gene expression

通过抑制钙调蛋白基因表达开发新型骨形成疗法

• 批准号: 12557128
• 负责人: TAKAHASHI Katsuhito
• 金额: $ 8.19万
• 依托单位: Osaka Medical Center for Cancer and Cardiovascular Diseases
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12557128/
• 关键词: Calponin; Bone Formation; Bone Fracture Repair; Antisense oligoDNA; Adenovirus; 骨芽細胞; 骨折治療; 組織修復

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.

肌细胞区分诱导的BMP-2（40 ng/ml）通过将小鼠calponin 1的抗固定寡素添加到培养基中。小鼠Calponin H1序列设计并合成Ligodna，但在40μg/ml的浓度下，ligodna均未证明少神经的浓度为80μg/ml在骨折后48小时，抗敏感性的抗氧化寡素体加速了实验性骨折的愈合。 28/55）与对照组（感官寡虫； 31.3％，n = 16/51，静音抗sense-oligodna; 28.6％，n = 4/14，仅pluronic凝胶； 37.5％，n = 6/16）在人间充质细胞中含有抗calponin h1的抗calponin H1的抗粘蛋白H1的抗浓度cDNA的腺病毒载体。

项目成果

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Takahashi K.: "Regualation of shortening velocity by calponin in intact contracting smooth muscles."Biochem.Biophys.Res.Commun.. 279. 150-157 (2000)

Takahashi K.：“完整收缩平滑肌中钙调素对缩短速度的调节。”Biochem.Biophys.Res.Commun.. 279. 150-157 (2000)

Youichi Sugenoya: "Smooth-muscle calponin in mesangial cells : Regulation of expression and a role in suppressing glomerulonephritis"J.Am.Soc.Nephrol.. 13. 322-331 (2002)

Youichi Sugenoya：“系膜细胞中的平滑肌钙调蛋白：表达调节和抑制肾小球肾炎的作用”J.Am.Soc.Nephrol.. 13. 322-331 (2002)

Hisako Yamamura: "Identification of the transcriptional regulatory sequences of human calponin promoter and their use in targeting of a conditionally replicating herpes vector to"Cancer Res.. 61. 3969-3977 (2001)

Hisako Yamamura：“人钙调蛋白启动子转录调控序列的鉴定及其在条件复制疱疹载体靶向中的应用”Cancer Res.. 61. 3969-3977 (2001)

Sugimoto,T.: "A malignant rhabdoido-tumor cell line showing neural and smooth-muacle-cell pheotypes."Intern.J.Cancer. 82. 678-686 (2000)

Sugimoto,T.：“一种恶性横纹肌瘤细胞系，显示神经细胞和平滑粘液细胞表型。”Intern.J.Cancer。