Basic Research of Gene Therapy for Joint Diseases

为了探讨基因治疗人类关节疾病的可行性，我们将携带β-半乳糖苷酶（beta-gal）或转化生长因子（transforming growth factor，TGF）-β 1基因的腺病毒载体直接注射到Hartley豚鼠的关节内。通过5-溴-4-氯-3-吲哚-β-D-吡喃半乳糖苷（X-gal）染色和逆转录-聚合酶链反应（RT-PCR）检测转导的β-gal基因的表达。通过酶联免疫吸附试验（ELISA）评估传递到关节然后转移到关节液的TGF-β 1的水平。LacZ的表达在几乎整个滑膜组织和软骨细胞表面的退化软骨。直接注射载体对其他器官的影响也是出乎意料的，没有观察到递送基因的表达。因此，关节内直接注射将实现限于关节腔的基因递送，可能消除不必要的全身效应。关节液中TGF-β 1的水平在2周内明显高于对照组，腺病毒载体体内基因转染方法可直接将基因导入关节腔，具有临床应用价值。

We evaluated the in vivo applicability of adenovirus-mediated gene delivery in order to consider the feasibility of gene therapy for human joint diseases.We directly injected vectors harbouring beta-galactosidase (beta-gal) or transforming growth factor (TGF)-beta1 gene into the joints of Hartley guinea pigs. Expressions of transduced beta-gal genes were examined by 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) staining and reverse transcription- polymerase chain reaction (RT-PCR). The levels of TGF-beta1 which was delivered to the joint and then transferred to the joint fluid, were assessed by enzyme-linked immunosorbent assay (ELISA). LacZ expression was observed in almost the entire synovial tissues and in chondrocytes on the surface of degenerated cartilage. Un expected effects of the direct vector inj ection on the other organs were also examined, no expression of delivered gene was observed. Therefore, intraarticular direct injection would achieve gene delivery limited to the joint cavity, possibly eliminating unecessary systemic effects. TGF-beta1 levels in the joint fluid following the gene delivery had been significantly higher than the levels in the controls for 2 weeks.Direct gene delivery into the joint cavity is realistic with the in vivo gene delivery method using adenovirus vector, and it would be clinically applicable.