Sonic hedgehog gene-enhanced tissue engineering for bone regeneration
Sonic hedgehog gene-enhanced tissue engineering for bone regeneration
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DOI：
10.1038/sj.gt.3302386
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发表时间：
2005-01-01
期刊：
影响因子：
5.1
通讯作者：
Mason, JM
中科院分区：
文献类型：
--
作者：
Edwards, PC;Ruggiero, S;Mason, JM
在继发于肿瘤切除、先天性畸形和修复牙齿重建之前的显著骨缺损患者的颅面康复中，需要改进的骨再生方法。在这项研究中，使用基因增强的组织工程方法来评估Sonic hedgehog（Shh）转导的牙龈成纤维细胞、间充质干细胞和脂肪来源的细胞在藻酸盐/胶原蛋白基质中递送到兔颅骨缺损的骨再生能力。将从胎儿肺组织分离的人Shh cDNA克隆到复制缺陷型逆转录病毒表达载体LNCX中，其中鼠白血病病毒逆转录病毒LTR驱动新霉素抗性基因的表达。大鼠β-肌动蛋白增强子/启动子复合物被工程化以驱动Shh的表达。逆转录-聚合酶链反应分析表明，转导的原代兔细胞群表达Shh RNA。通过酶联免疫吸附测定（ELISA）确认Shh蛋白分泌。将含有2 × 10（6）个Shh转导细胞的藻酸盐/I型胶原构建体引入雄性新西兰白色兔颅骨缺损（8 mm）中。总共检查了八组（N = 6）：未恢复的空缺陷、单独的基质、基质加上用对照和Shh表达载体两者转导的三个细胞群。在植入后6周和12周，对Shh基因增强的细胞的骨再生能力进行大体、X线和组织学评估。6周后，在Shh转导组中，观察到直接从藻酸盐/胶原基质发出的新的全厚度骨。组织切片的定量二维数字分析证实，与对照组相比，所有三个Shh增强组的骨再生量具有统计学显著性（P < 0.05）。尸检未能证明任何治疗相关副作用的证据。这是第一项证明Shh递送到骨缺损的研究，在这种情况下，通过一种新的基因增强的组织工程方法，导致显着的骨再生。这鼓励了Shh基因增强的骨再生组织工程方法的进一步发展。
Improved methods of bone regeneration are needed in the craniofacial rehabilitation of patients with significant bone deficits secondary to tumor resection, congenital deformities, and prior to prosthetic dental reconstruction. In this study, a gene-enhanced tissue-engineering approach was used to assess bone regenerative capacity of Sonic hedgehog (Shh)-transduced gingival fibroblasts, mesenchymal stem cells, and fat-derived cells delivered to rabbit cranial bone defects in an alginate/collagen matrix. Human Shh cDNA isolated from fetal lung tissue was cloned into the replication-incompetent retroviral expression vector LNCX, in which the murine leukemia virus retroviral LTR drives expression of the neomycin-resistance gene. The rat beta-actin enhancer/promoter complex was engineered to drive expression of Shh. Reverse transcriptase-polymerase chain reaction analysis demonstrated that the transduced primary rabbit cell populations expressed Shh RNA. Shh protein secretion was confirmed by enzyme-linked immunosorbent assay (ELISA). Alginate/type I collagen constructs containing 2 x 10(6) Shh-transduced cells were introduced into male New Zealand White rabbit calvarial defects (8 mm). A total of eight groups (N = 6) were examined: unrestored empty defects, matrix alone, matrix plus the three cell populations transduced with both control and Shh expression vectors. The bone regenerative capacity of Shh gene enhanced cells was assessed grossly, radiographically and histologically at 6 and 12 weeks postimplantation. After 6 weeks, new full thickness bone was seen emanating directly from the alginate/collagen matrix in the Shh-transduced groups. Quantitative two-dimensional digital analysis of histological sections confirmed statistically significant (P < 0.05) amounts of bone regeneration in all three Shh-enhanced groups compared to controls. Necropsy failed to demonstrate any evidence of treatment-related side effects. This is the first study to demonstrate that Shh delivery to bone defects, in this case through a novel gene-enhanced tissue-engineering approach, results in significant bone regeneration. This encourages further development of the Shh gene-enhanced tissue-engineering approach for bone regeneration.