AAV6 as an effective gene delivery vector for prolonged transgene expression in intervertebral disc cells in vivo

Intervertebral disc degeneration is the main contributor to low back pain, now the leading cause of disability worldwide. Gene transfer, either in a therapeutic attempt or in basic research to understand the mechanisms of disc degeneration, is a fascinating and promising tool to manipulate the complex physiology of the disc. Viral vectors based on the adenoassociated virus (AAV) have emerged as powerful transgene delivery vehicles yet a systematic investigation into their respective tropism, transduction efficiency, and relative toxicity have not yet been performed in the disc in vivo. Herein, we used in vivo bioluminescence imaging to systematically compare multiple AAV serotypes, injection volumes, titers, promoters, and luciferase reporters to determine which result in high transduction efficiency of murine nucleus pulposus (NP) cells in vivo. We find that AAV6 using a CAG promoter to drive transgene expression, delivered into the NP of murine caudal discs at a titer of 1011 GC/mL, provides excellent transduction efficiency/kinetics and low toxicity in vivo. We also show, for the first time, that the transduction of NP cells can be significantly boosted in vivo by the use of small cell permeabilization peptides. Finally, to our knowledge, we are the first to demonstrate the use of optical tissue clearing and three-dimensional lightsheet microscopy in the disc, which was used to visualize fine details of tissue and cell architecture in whole intact discs following AAV6 delivery. Taken together, these data will contribute to the success of using AAV-mediated gene delivery for basic and translational studies of the IVD. Copyright (c) 2020, Chongqing Medical University. Production and hosting by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/ by-nc-nd/4.0/).

Automated summary

Intervertebral disc degeneration is a major cause of low back pain worldwide. In this study, the researchers investigated the use of adenoassociated virus (AAV) as a tool for gene transfer in the intervertebral disc. They compared different AAV serotypes and parameters, and found that AAV6 with a CAG promoter at a titer of 1011 GC/mL achieved high transduction efficiency and low toxicity in murine nucleus pulposus cells. They also demonstrated the use of small cell permeabilization peptides to boost transduction efficiency, and utilized optical tissue clearing and three-dimensional lightsheet microscopy to visualize tissue and cell architecture after AAV6 delivery.