中国修复重建外科杂志

中国修复重建外科杂志

局部注射促血管生成素 2 调控自噬促进体内组织工程人工骨早期血管化和骨缺损修复的研究

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目的探讨兔桡骨骨缺损处局部注射促血管生成素 2(angiopoietin 2,Ang-2),通过调控自噬促进组织工程人工骨早期血管化和骨缺损修复的机制。方法取 48 只新西兰大白兔,构建单侧 1.5 cm 长桡骨缺损模型,于骨缺损处植入羟基磷灰石/胶原支架后,随机分为两组,对照组(A 组)和 Ang-2 组(B 组)分别于术后 2 周内每日于骨缺损处注射 1 mL 生理盐水和 1 mL 溶于生理盐水的 400 ng/mL Ang-2。采用 Western blot 检测骨痂区自噬相关蛋白微管相关蛋白 1 轻链 3(microtubule associated protein 1 lightchain 3,LC3)和 Beclin-1,血管生成相关蛋白 VEGF 及自噬降解底物蛋白 SQSTMl/p62 的表达。术后 4、8、12 周行 X 线片检查,采用 Lane-Sandhu X 线评分法对骨缺损修复情况进行评分;术后 12 周处死动物取标本行大体观察,采用 HE 染色观察骨缺损区血管生成情况。结果Western blot 检测示,B 组 LC3-Ⅱ/LC3-Ⅰ、Beclin-1 及 VEGF 相对表达量高于 A 组,SQSTMl/p62 相对表达量低于 A 组,差异均有统计学意义(P<0.05)。影像学及标本大体观察示 A 组仅有少量骨痂生成,但骨缺损未修复;B 组新生骨痂较多,骨缺损处完全修复。术后 4、8、12 周 B 组 Lane-Sandhu 评分均显著高于 A 组(P<0.05)。HE 染色示 B 组哈佛管排列整齐,新生血管显著多于 A 组,差异有统计学意义(t=–11.879,P=0.000)。结论局部注射适宜浓度 Ang-2 可能通过增强自噬促进兔体内组织工程人工骨早期血管化和骨缺损修复。

ObjectiveTo investigate the mechanism of early vascularization of the tissue engineered bone in the treatment of rabbit radial bone defect by local injection of angiopoietin 2 (Ang-2).MethodsForty-eight New Zealand white rabbits were established unilateral 1.5 cm long radius defect models. After implantation of hydroxyapatite/collagen scaffolds in bone defects, the rabbits were randomly divided into 2 groups: control group (group A) and Ang-2 group (group B) were daily injected with 1 mL normal saline and 1 mL saline-soluble 400 ng/mL Ang-2 at the bone defect within 2 weeks after operation, respectively. Western blot was used to detect the expressions of autophagy related protein [microtubule associated protein 1 light chain 3 (LC3), Beclin-1], angiogenesis related protein [vascular endothelial growth factor (VEGF)], and autophagy degradable substrate protein (SQSTMl/p62) in callus. X-ray films examination and Lane-Sandhu X-ray scoring were performed to evaluate the bone defect repair at 4, 8, and 12 weeks after operation. The rabbits were sacrificed at 12 weeks after operation for gross observation, and the angiogenesis of bone defect area was observed by HE staining.ResultsWestern blot assay showed that the relative expressions of LC3-Ⅱ/LC3-Ⅰ, Beclin-1, and VEGF in group B were significantly higher than those in group A, and the relative expression of SQSTMl/p62 was significantly lower than that in group A (P<0.05). Radiographic and gross observation of specimens showed that only a few callus were formed in group A, the bone defect was not repaired; more callus were formed and complete repair of bone defect was observed in group B. The Lane-Sandhu scores in group B were significantly higher than those in group A at 4, 8, and 12 weeks after operation (P<0.05). HE staining showed that the Harvard tubes in group B were well arranged and the number of new vessels was significantly higher than that in group A (t=–11.879, P=0.000).ConclusionLocal injection of appropriate concentration of Ang-2 may promote early vascularization and bone defect repair of tissue engineered bone in rabbits by enhancing autophagy.

关键词: 促血管生成素 2; 自噬; 血管生成; 组织工程人工骨; 骨缺损;

Key words: Angiopoietin 2; autophagy; angiogenesis; tissue engineered bone; bone defect; rabbit

引用本文: 殷建, 王斌, 朱超, 孙超, 刘新晖. 局部注射促血管生成素 2 调控自噬促进体内组织工程人工骨早期血管化和骨缺损修复的研究. 中国修复重建外科杂志, 2018, 32(9): 1150-1156. doi: 10.7507/1002-1892.201804105 复制

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