Institute for Biomedical Engineering, University and ETH Zürich, Zürich, Switzerland.
J Tissue Eng Regen Med. 2010 Jul;4(5):387-94. doi: 10.1002/term.251.
Anti-inflammatory gene therapy can inhibit inflammation driven by TNFalpha in experimental models of rheumatoid arthritis. However, assessment of the therapeutic effect on cartilage and bone quality is either missing or unsatisfactory. A multimodal imaging approach, using confocal laser scanning microscopy (CLSM) and micro-computed tomography (microCT), was used for gathering 3D quantitative image data on diseased and treated murine joints. As proof of concept, the efficacy of anti-TNF-based gene therapy was assessed, comparing imaging techniques with classical investigations. SCID mice knees were injected with human synoviocytes overexpressing TNFalpha. Two days later, electric pulse-mediated DNA transfer was performed after injection of the pGTRTT-plasmid containing a dimeric soluble-TNF receptor (dsTNFR) under the control of a doxycycline-inducible promoter. After 21 days the mice were sacrificed, TNFalpha levels were measured and the joints assessed for cartilage and bone degradation, using CLSM, microCT and histology. TNFalpha levels were decreased in the joints of mice treated with the plasmid in the presence of doxycycline. Concomitantly, histological analysis showed an increase in cartilage thickness and a decrease in specific synovial hyperplasia and cartilage erosion. Bone morphometry revealed that groups with the plasmid in the presence of doxycycline displayed a higher cortical thickness and decreased porosity. Using an anti-TNF gene therapy approach, known to reduce inflammation, as proof of concept, 3D imaging allowed quantitative evaluation of its benefits to joint architecture. It showed that local delivery of a regulated anti-TNF vector allowed decreasing arthritis severity through TNFalpha inhibition. These tools are valuable for understanding the efficacy of gene therapy on whole-joint morphometry.
抗炎基因治疗可以抑制 TNFalpha 驱动的实验性类风湿关节炎模型中的炎症。然而，对软骨和骨质量的治疗效果的评估要么缺失，要么不尽如人意。采用共聚焦激光扫描显微镜（CLSM）和微计算机断层扫描（microCT）的多模态成像方法，对患病和治疗的鼠关节进行了 3D 定量图像数据的采集。作为概念验证，通过与经典研究的比较，评估了基于抗 TNF 的基因治疗的疗效。在注射过表达 TNFalpha 的人滑膜细胞后，将 SCID 小鼠的膝盖注射到 SCID 小鼠的膝盖中。两天后，在注射含有双体可溶性 TNF 受体（dsTNFR）的 pGTRTT 质粒后，通过电脉冲介导的 DNA 转移，该质粒受强力霉素诱导型启动子的控制。21 天后，处死小鼠，测量 TNFalpha 水平，并使用 CLSM、microCT 和组织学评估关节软骨和骨降解。在强力霉素存在的情况下，用质粒处理的小鼠关节中的 TNFalpha 水平降低。同时，组织学分析显示软骨厚度增加，滑膜增生和软骨侵蚀特异性减少。骨形态计量学显示，在强力霉素存在的情况下带有质粒的组显示出更高的皮质厚度和降低的孔隙率。使用已知可减少炎症的抗 TNF 基因治疗方法作为概念验证，3D 成像允许对其对关节结构的益处进行定量评估。它表明，局部递送电调控的抗 TNF 载体可以通过抑制 TNFalpha 来减轻关节炎的严重程度。这些工具对于了解基因治疗对整个关节形态计量学的疗效非常有价值。
