Experimental Studies on the effects of BMP-2 Expressing Adenoviral Vector to the Distraction Osteogenesis

表达BMP-2的腺病毒载体对牵张成骨作用的实验研究

• 批准号: 14571924
• 负责人: FUJIMURA Kazuma
• 金额: $ 2.18万
• 依托单位: KANAZAWA MEDICAL UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14571924/
• 关键词: Adenoviral vector; Distraction; Osteogenesis; Bone Morphogenetic Protein; rhBMP-2; アデノウイルスベクター; 骨延長術

1.To examine the effectiveness of a gene transfer procedure for the delivery of BMP-2 to the distraction osteogenesis, we constructed a human BMP-2--expressing replication-deficient adenoviral vector, AxCAOBMP-2, and evaluated its osteoinductive activity in vitro and in vivo.2.C2C12 myoblasts were infected in vitro with this viral vector or an Escherichia coil LacZ gene-expressing control adenovirus vector (AxCALacZ). Twenty-four hours after the infection, indirect immunofluorescence was performed. On day 5 after the infection, alkaline phosphatase (ALP) in the cells and osteocalcin in the culture medium were measured. Furthermore, to examine the effectiveness of gene transfer of BMP-2 in vivo, we evaluated osteoinduction by AxCAOBMP-2, under transient immunosuppression with cyclophosphamide, given at a dose of 125 mg/kg intraperitoneally the day before injection of the adenoviral vector. Twenty-five microliters of AxCAOBMP-2 (8.75 x 10^8 plaque-forming units [pfu],'Group I) and AxCALa … More cZ (1.75 x 10^8 pfu, control group) and 5 microl of AxCAOBMP-2 (1.75x 10^8 pfu, Group II) were injected into a right calf muscle of Wistar rats. On day 21, bone formation in each group was investigated radiologically and histologically.3.Abundant BMP-2 expression in C2C12 cells infected with this viral vector was confirmed by immunofluorescence. C2C12 cellstransferred with the BMP-2 gene by this vector produced ALP in the cells and also produced and secreted osteocalcin in the culture medium. Osteoinduction was found only in the AxCAOBMP-2 treated groups with immunosuppression. Osteoinduction activity was higher in Group I than in Group II.3.Twelve animals, performed distraction, were treated with AxCAOBMP-2 (8.75 x 10^8 pfu) 25μg in each, during distraction osteogenesis at 7 days and 14 days. In results, Bone formation and ALP activity were increased significantly in comparison with non-treatment groups. It was suggested that Gene transfer procedure for the delivery of BMP-2 effected to the distraction osteogenesis and had a potential shorting distraction period. Less

1.构建了表达BMP-2的复制缺陷型腺病毒载体AxCAOBMP-2，并对其在体内外的成骨诱导活性进行了研究。2.将该病毒载体和表达LacZ基因的对照腺病毒载体（AxCALacZ）分别感染C2 C12成肌细胞。感染后24小时，进行间接免疫荧光。在感染后第5天，测量细胞中的碱性磷酸酶（ALP）和培养基中的骨钙素。此外，为了检查BMP-2在体内的基因转移的有效性，我们评估了AxCAOBMP-2的骨诱导作用，在注射腺病毒载体的前一天，在环磷酰胺的瞬时免疫抑制下，以125 mg/kg的剂量腹膜内给药。25微升AxCAOBMP-2（8.75 × 10^8空斑形成单位[pfu]，I组）和AxCALa ...更多信息 将cZ（1.75 x 10^8 pfu，对照组）和5 μ l AxCAOBMP-2（1.75 x 10^8 pfu，组II）注射到Wistar大鼠的右小腿肌肉中。第21天，通过放射学和组织学观察各组的骨形成情况。3.免疫荧光法证实，BMP-2在感染的C2 C12细胞中有大量表达。用该载体转染BMP-2基因的C2 C12细胞在细胞内产生ALP，并在培养液中产生和分泌骨钙素。骨诱导作用仅在免疫抑制的AxCAOBMP-2处理组中发现。第一组的成骨活性高于第二组。3. 12只动物进行牵张成骨，在第7天和第14天牵张成骨过程中，每只动物给予AxCAOBMP-2（8.75 × 10^8 pfu）25μg。结果表明，与非治疗组相比，骨形成和ALP活性显著增加。结果表明，BMP-2基因转染法对牵张成骨有促进作用，并有缩短牵张成骨周期的潜力。少

项目成果

Fujimura K, Bessho K, Okubo Y, Kusumoto K, Segami N, Iizuka. T: "The effect of fibroblast growth factor-2 to osteoinducing activity by recombinant human bone morphogenetic protein-2 in rat muscle."Arch Oral Biol.. 47(8). 577-584 (2002)

藤村 K、别所 K、大久保 Y、楠本 K、濑上 N、饭冢。

Fujimura K, Bessho K, et al.: "The effect of fibroblast growth factor-2 to osteoinducing activity by recombinant human bone morphogenetic protein-2 in rat muscle."Arch Oral Biol.. 48. 577-584 (2002)

Fujimura K、Bessho K 等人：“成纤维细胞生长因子 2 对大鼠肌肉中重组人骨形态发生蛋白 2 的骨诱导活性的影响。”Arch Oral Biol.. 48. 577-584 (2002)

(Y.Okubo), K.Fujimura, et al.: "Expression of bone morphogenetic protein-2 in the course of osteoinduction by recombinant human bone morphogenetic protein-2."Clin Oral Impl Res.. 13. 80-85 (2002)

(Y.Okubo)、K.Fujimura 等：“重组人骨形态发生蛋白 2 在骨诱导过程中骨形态发生蛋白 2 的表达。”Clin Oral Impl Res.. 13. 80-85 (2002)

Kusumoto K, Bessho K, Fujimura K, Akioka J, Okubo Y, Wang Y, Iizuka T, Ogawa Y.: "Osteoinduction by recombinant human bone morphogenetic protein-2 in muscles of non-human primates."J Int Med Res.. 30(3). 251-259 (2002)

Kusumoto K、Bessho K、Fujimura K、Akioka J、Okubo Y、Wang Y、Iizuka T、Okawa Y.：“非人类灵长类动物肌肉中重组人骨形态发生蛋白 2 的骨诱导。”J Int Med Res..

Fujimura K, Bessho K, Okubo Y, et al.: "A bioactive bone cement containing Bis-GMA resin and A-W Glass-ceramic as an augmentation graft material on mandibular bone."Clin Oral Impl Res.. 14. 659-667 (2003)

Fujimura K、Bessho K、Okubo Y 等人：“一种含有 Bis-GMA 树脂和 A-W 玻璃陶瓷的生物活性骨水泥，作为下颌骨的增强移植材料。”Clin Oral Impl Res.. 14. 659-667（