Recombinant adeno-associated virus vectors efficiently and persistently transduce chondrocytes in normal and osteoarthritic human articular cartilage
Recombinant adeno-associated virus vectors efficiently and persistently transduce chondrocytes in normal and osteoarthritic human articular cartilage
复制标题
DOI：
10.1089/104303403321208998
复制
发表时间：
2003-03-01
影响因子：
4.2
通讯作者：
Trippel, SB
中科院分区：
文献类型：
--
作者：
Madry, H;Cucchiarini, M;Trippel, SB
成功地将基因转移到关节软骨中是关节疾病基因治疗的先决条件。在本研究中，我们测试了重组腺相关病毒（rAAV）载体能够在体外分离的关节软骨细胞、体外关节软骨组织和体内关节损伤部位中影响基因转移的假设。使用在巨细胞病毒（CMV）立即早期启动子/增强子（rAAV-lacZ）控制下携带大肠杆菌β-半乳糖苷酶基因（lacZ）的rAAV载体，对于分离的正常成人关节软骨细胞和骨关节炎人关节软骨细胞，转导效率超过70％。这些与用新生牛关节软骨细胞获得的转导效率相当。关节软骨外植体培养物的转导导致报告基因在所有三种软骨类型的组织内表达，深度超过 450 毫米，并一直存在到 150 天。当rAAV-lacZ载体应用于大鼠膝关节模型体内股骨软骨缺损和骨软骨缺损时，转导后至少10天实现报告基因表达。这些数据表明，基于AAV的载体可以在关节软骨细胞（包括正常和骨关节炎人类关节软骨的软骨细胞）中有效转导并稳定表达外源基因。数据进一步表明，相同的rAAV载体能够在其天然基质内原位转导软骨细胞，转导深度足以具有潜在的临床意义。最后，数据表明这些rAAV载体能够有效地将重组基因递送至体内软骨和骨软骨缺损处。
Successful gene transfer into articular cartilage is a prerequisite for gene therapy of articular joint disorders. In the present study we tested the hypothesis that recombinant adeno-associated virus (rAAV) vectors are capable of effecting gene transfer in isolated articular chondrocytes in vitro, articular cartilage tissue in vitro, and sites of articular damage in vivo. Using an rAAV vector carrying the Escherichia coli beta-galactosidase gene (lacZ) under the control of the cytomegalovirus (CMV) immediate-early promoter/enhancer (rAAV-lacZ), transduction efficiency exceeded 70% for isolated normal human adult articular chondrocytes, and osteoarthritic human articular chondrocytes. These were comparable to the transduction efficiency obtained with neonatal bovine articular chondrocytes. Transduction of explant cultures of articular cartilage resulted in reporter gene expression within the tissue of all three cartilage types to a depth exceeding 450 mm, which remained present until 150 days. When rAAV-lacZ vectors were applied to femoral chondral defects and osteochondral defects in vivo in a rat knee model, reporter gene expression was achieved for at least 10 days after transduction. These data suggest that AAV-based vectors can efficiently transduce and stably express foreign genes in articular chondrocytes, including chondrocytes of normal and osteoarthritic human articular cartilage. The data further suggest that the same rAAV vectors are capable of transducing chondrocytes in situ within their native matrix to a depth sufficient to be of potential clinical significance. Finally, the data demonstrate that these rAAV vectors are capable of effectively delivering recombinant genes to chondral and osteochondral defects in vivo.