The effect of regional gene therapy with bone morphogenetic protein-2-producing bone-marrow cells on the repair of segmental femoral defects in rats
The effect of regional gene therapy with bone morphogenetic protein-2-producing bone-marrow cells on the repair of segmental femoral defects in rats
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DOI：
10.2106/00004623-199907000-00002
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发表时间：
1999-07-01
影响因子：
5.3
通讯作者：
Witte, ON
中科院分区：
文献类型：
--
作者：
Lieberman, JR;Daluiski, A;Witte, ON
背景资料：重组人骨形态发生蛋白（rhBMPs）具有诱导成骨的作用，但由于缺乏理想的载体，限制了其临床应用。我们使用离体腺病毒基因转移来创建BMP-2产生骨髓细胞，其允许将BMP-2递送到特定解剖部位。方法：将5 × 10（6）BMP-2产生的骨髓细胞通过腺病毒基因转移的方法移植到5组大鼠的股骨缺损中（24个股骨，组I）; 20 μ g rhBMP-2（16个股骨，第II组）; 5 × 10（6）个产生β-半乳糖苷酶的大鼠骨髓细胞，通过腺病毒基因转移lacZ基因产生（12个股骨，组III）; 5 x 10（6）个未感染的大鼠骨髓细胞（10个股骨，组IV）;或仅盐酸胍提取的脱矿骨基质（10个股骨，组V）。盐酸胍提取的脱钙骨基质作为所有实验组的底物。术后两个月取出的标本进行组织学和组织形态学分析以及生物力学测试。I组24个缺损中的22个术后两个月，第二组（rhBMP-2）和第二组（rhBMP-2）的所有16个缺损在放射学上都已愈合，而三个对照组的32个缺损中只有一个缺损愈合（产生β-半乳糖苷酶的大鼠骨髓细胞、未感染的大鼠骨髓细胞和单独的盐酸胍提取的脱矿骨基质）。样本的组织学分析显示，接受了产生BMP-2的骨髓细胞的缺损（组I）在术后两个月时被粗骨小梁填充，而在接受rhBMP-2的那些（组II）中，骨是薄的和花边状的。用产生β-半乳糖苷酶的骨髓细胞治疗的缺损（III组），未感染的骨髓细胞（第四组），组织形态计量学分析显示，BMP-2产生骨髓细胞处理的缺损处的骨形成总面积明显大于rhBMP-2处理的缺损处（p = 0.036）。生物力学测试表明没有显着差异，可用的数字，愈合的股骨已收到BMP-2生产骨髓细胞和未处理的（对照）股骨方面的最终扭矩失败或能量failure.Conclusions：这项研究表明，BMP-2生产骨髓细胞通过腺病毒基因转移产生足够的蛋白质来愈合节段性股骨缺损。我们还建立了离体基因转移的可行性与使用生物急性自体短期文化的骨髓cells.Clinical Relevance：区域基因治疗是一种新的方法来治疗骨缺损。与第一代腺病毒载体相关的转基因表达的有限持续时间对于该临床应用是有利的。这种离体基因转移和随后用含有BMP-2 cDNA的腺病毒感染骨髓细胞的系统可用于增强人体骨形成。
Background: Recombinant human bone morphogenetic proteins (rhBMPs) can induce bone formation, but the inability to identify an ideal delivery system limits their clinical application. We used ex vivo adenoviral gene transfer to create BMP-2-producing bone-marrow cells, which allow delivery of the BMP-2 to a specific anatomical site. The autologous BMP-2-producing bone-marrow cells then were used to heal a critical-sized femoral segmental defect in syngeneic rats.Methods: Femoral defects in five groups of rats were filled with 5 x 10(6) BMP-2-producing bone-marrow cells, created through adenoviral gene transfer (twenty-four femora, Group I); twenty micrograms of rhBMP-2 (sixteen femora, Group II); 5 x 10(6) beta-galactosidase-producing rat-bone-marrow cells, created through adenoviral gene transfer of the lacZ gene (twelve femora, Group III); 5 x 10(6) unfected rat-bone-marrow cells (ten femora, Group IV); or guanidine hydrochloride-extracted demineralized bone matrix only (ten femora, Group V). Guanidine hydrochloride-extracted demineralized bone matrix served as a substrate in all experimental groups. Specimens that were removed two months postoperatively underwent histological and histomorphometric analysis as well as biomechanical testing.Results: Twenty-two of the twenty-four defects in Group I (BMP-2-producing bone-marrow cells) and all sixteen defects in Group II (rhBMP-2) had healed radiographically at two months postoperatively compared with only one of the thirty-two defects in the three control groups (beta-galactosidase-producing rat-bone-marrow cells, uninfected rat-bone-marrow cells, and guanidine hydrochloride-extracted demineralized bone matrix alone).Histological analysis of the specimens revealed that defects that had received BMP-2-producing bone-marrow cells (Group I) were filled with coarse trabecular bone at two months postoperatively, whereas in those that had received rhBMP-2 (Group II) the bone was thin and lace-like. Defects that had been treated with bone-marrow cells producing beta-galactosidase (Group III), uninfected bone-marrow cells (Group IV), or guanidine hydrochloride-extracted demineralized bone matrix only (Group V) demonstrated little or no bone formation.Histomorphometric analysis revealed a significantly greater total area of bone formation in the defects treated with the BMP-2-producing bone-marrow cells than in those treated with the rhBMP-2 (p = 0.036). Biomechanical testing demonstrated no significant differences, with the numbers available, between the healed femora that had received BMP-2-producing bone-marrow cells and the untreated (control) femora with respect to ultimate torque to failure or energy to failure.Conclusions: This study demonstrated that BMP-2-producing bone-marrow cells created by means of adenoviral gene transfer produce sufficient protein to heal a segmental femoral defect. We also established the feasibility of ex vivo gene transfer with the use of biologically acute autologous short-term cultures of bone-marrow cells.Clinical Relevance: Regional gene therapy is a novel approach to the treatment of bone defects. The limited duration of transgenic expression associated with first-generation adenoviral vectors is advantageous for this clinical application. This system of ex vivo gene transfer and the subsequent infection of bone-marrow cells with an adenovirus containing the BMP-2 cDNA could be adapted to enhance bone formation in humans.