Protocol approval and animal treatments

The present study was approved by the Research and Animal Ethics Association of Zhongshan Hospital affiliated to Fudan University under a license number No. GDR452, in compliance with the institutional guidelines for the care and use of animals. A protocol was prepared before study without registration. A total of 40 1-year-old female Sprague-Dawley rats (140–160 g) were purchased from the Animal Laboratory of the Academy of Medical Sciences (Beijing, China) and used in strict accordance with the guidelines for the Care and Use of Laboratory Animals. For each experiment, power calculations (P<0.05) were used to include an adequate number of animals for the observed effects to be legitimate. An allocation concealment method was used for randomization in allocating experimental units to the control and treatment groups. The potential confounders were minimized. No criteria were used for excluding animals (or experimental units) during the experiment, and no data were excluded during the analysis. The study did not have humane endpoints. The rats were kept in separate cages with free access to food and water, and in a 12/12-hour light/dark cycle (temperature, 24 °C; humidity, 50%). Rats were randomly divided into 4 groups of 10. The sham group received only a skin incision, which was then sutured to close the wound. The rats in the other 3 groups received removal of the sacrospinal muscles, spinous processes, supraspinous ligaments, interspinous ligaments, and posterolateral halves of the bilateral zygapophysial joints of the lumbar spine (a model for LDD). Afterwards, the rats in the LDD group received an orthotopic injection of 150 µL normal saline, rats in the LDD + AAV-scr group received an orthotopic injection of 10¹¹ AAV-scr in a 150 µL volume, and rats in the LDD + AAV-IGF1/VEGF group received an orthotopic injection of 10¹¹ AAV-IGF1/VEGF in a 150 µL volume. Rats were kept for 8 weeks before analysis.