Yanxia Gao, Kailun Fang, Zixiang Yan, Haiwei Zhang, Guannan Geng, Weiwei Wu, Ding Xu, Heng Zhang, Na Zhong, Qifang Wang, Minqing Cai, Erwei Zuo, and Hui Yang
Abstract
Reprogramming Müller glia (MG) into functional cells is considered a promising therapeutic strategy to treat ocular diseases and vision loss. However, current AAV-based system for MG-tracing was reported to have high leakage in recent studies. Here, we focused on reducing the leakage of AAV-based labeling systems and found that different AAV serotypes showed a range of efficiency and specificity in labeling MG, leading us to optimize a human GFAP-Cre reporter system packaged in the AAV9 serotype with the woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) removed. The leakage ratio of the AAV9-hGFAP-Cre-ΔWPRE decreased by an approximate 40-fold compared with the AAV9-hGFAP-Cre-WPRE labeling system. In addition, we validated the specificity of the AAV-ΔWPRE system for tracing MG reprogramming under Ptbp1-suppression and observed strict non-MG-conversion, similar to previous studies using genetic lineage tracking mouse models. Thus, the AAV9-hGFAP-Cre-ΔWPRE system showed high efficiency and specificity for MG labeling, providing a promising tool for tracing cell fate in vivo.
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