Extracellular Vesicle-Mediated Delivery of Circular RNA SCMH1 Promotes Functional Recovery in Rodent and Nonhuman Primate Ischemic Stroke Models

Li Yang; Bing Han; Zhiting Zhang; Shuguo Wang; Ying Bai; Yuan Zhang; Ying Tang; Lingli Du; Ling Xu; Fangfang Wu; Lei Zuo; Xufeng Chen; Yu Lin; Kezhong Liu; Qingqing Ye; Biling Chen; Bin Li; Tianci Tang; Yu Wang; Ling Shen; Guangtian Wang; Minzi Ju; Mengqin Yuan; Wei Jiang; John H Zhang; Gang Hu; Jianhong Wang; Honghong Yao

doi:10.1161/CIRCULATIONAHA.120.045765

Extracellular Vesicle-Mediated Delivery of Circular RNA SCMH1 Promotes Functional Recovery in Rodent and Nonhuman Primate Ischemic Stroke Models

Circulation. 2020 Aug 11;142(6):556-574. doi: 10.1161/CIRCULATIONAHA.120.045765. Epub 2020 May 22.

Authors

Li Yang^#¹, Bing Han^#¹, Zhiting Zhang^{2

3

4}, Shuguo Wang⁵, Ying Bai¹, Yuan Zhang¹, Ying Tang¹, Lingli Du², Ling Xu², Fangfang Wu¹, Lei Zuo⁶, Xufeng Chen⁷, Yu Lin², Kezhong Liu², Qingqing Ye¹, Biling Chen¹, Bin Li¹, Tianci Tang¹, Yu Wang¹, Ling Shen¹, Guangtian Wang¹, Minzi Ju¹, Mengqin Yuan⁸, Wei Jiang⁸, John H Zhang^{3

9}, Gang Hu¹⁰, Jianhong Wang^{2

11

12

13}, Honghong Yao^{1

14

15}

Affiliations

¹ Department of Pharmacology, School of Medicine (L.Y., B.H., Y.B., Y.Z., Y.T., F.W., Q.Y., B.C., B.L., T.T., Y.W., L.S., G.W., M.J., H.Y.), Southeast University, Nanjing, China.
² National Research Facility for Phenotypic & Genetic Analysis of Model Animals (Primate Facility) (Z.Z., L.D., L.X., Y.L., K.L., J.W.), Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China.
³ Institutes of Physical Science and Information Technology, Anhui University, Hefei, China (Z.Z., K.L.).
⁴ State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica & Neuroscience Center, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China (Z.Z.).
⁵ Department of Neurosurgery, First Affiliation Hospital of Kunming Medical University, Kunming, China (S.W.).
⁶ Department of Neurology of Affiliated ZhongDa Hospital, Institute of Neuropsychiatry of Southeast University (L.Z.), Southeast University, Nanjing, China.
⁷ Emergency Department, Jiangsu Province Hospital and Nanjing Medical University First Affiliated Hospital, Nanjing, China (X.C.).
⁸ College of Automation Engineering, Nanjing University of Aeronautics and Astronautics, Nanjing, China (M.Y., W.J.).
⁹ Department of Physiology, School of Medicine, Loma Linda University, Loma Linda, CA (J.H.Z.).
¹⁰ Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, Nanjing, China (G.H.).
¹¹ National Resource Center for Non-Human Primates (Kunming Primate Research Center) (J.W.), Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China.
¹² Key Laboratory of Animal Models and Human Disease Mechanisms of the Chinese Academy of Science & Yunnan Province, (J.W.), Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China.
¹³ KIZ-CUHK Joint Laboratory of Bioresources and Molecular Research in Common Diseases (J.W.), Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China.
¹⁴ Institute of Life Sciences, Key Laboratory of Developmental Genes and Human Disease (H.Y.), Southeast University, Nanjing, China.
¹⁵ Co-innovation Center of Neuroregeneration, Nantong University, Nantong, China (H.Y.).

^# Contributed equally.

PMID: 32441115
DOI: 10.1161/CIRCULATIONAHA.120.045765

Abstract

Background: Stroke is a leading cause of adult disability that can severely compromise the quality of life of patients, yet no effective medication currently exists to accelerate rehabilitation. A variety of circular RNA (circRNA) molecules are known to function in ischemic brain injury. Lentivirus-based expression systems have been widely used in basic studies of circRNAs, but safety issues with such delivery systems have limited exploration of the potential therapeutic roles for circRNAs.

Methods: Circular RNA SCMH1 (circSCMH1) was screened from the plasma of patients with acute ischemic stroke by using circRNA microarrays. Engineered rabies virus glycoprotein-circSCMH1-extracellular vesicles were generated to selectively deliver circSCMH1 to the brain. Nissl staining was used to examine infarct size. Behavioral tasks were performed to evaluate motor functions in both rodent and nonhuman primate ischemic stroke models. Golgi staining and immunostaining were used to examine neuroplasticity and glial activation. Proteomic assays and RNA-sequencing data combined with transcriptional profiling were used to identify downstream targets of circSCMH1.

Results: CircSCMH1 levels were significantly decreased in the plasma of patients with acute ischemic stroke, offering significant power in predicting stroke outcomes. The decreased levels of circSCMH1 were further confirmed in the plasma and peri-infarct cortex of photothrombotic stroke mice. Beyond demonstrating proof-of-concept for an RNA drug delivery technology, we observed that circSCMH1 treatment improved functional recovery after stroke in both mice and monkeys, and we discovered that circSCMH1 enhanced the neuronal plasticity and inhibited glial activation and peripheral immune cell infiltration. CircSCMH1 binds mechanistically to the transcription factor MeCP2 (methyl-CpG binding protein 2), thereby releasing repression of MeCP2 target gene transcription.

Conclusions: Rabies virus glycoprotein-circSCMH1-extracellular vesicles afford protection by promoting functional recovery in the rodent and the nonhuman primate ischemic stroke models. Our study presents a potentially widely applicable nucleotide drug delivery technology and demonstrates the basic mechanism of how circRNAs can be therapeutically exploited to improve poststroke outcomes.

Keywords: MeCP2 protein; circular RNA; extracellular vesicles; neuronal plasticity; primate; stroke.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Brain / pathology*
Disease Models, Animal
Drug Delivery Systems
Extracellular Vesicles
Gene Expression Regulation
Genetic Vectors
Humans
Ischemic Stroke / genetics
Ischemic Stroke / rehabilitation*
Lentivirus / genetics*
Methyl-CpG-Binding Protein 2 / genetics
Methyl-CpG-Binding Protein 2 / metabolism
Mice
RNA, Circular / genetics*
Rabies / immunology*
Rabies Vaccines / immunology*
Rabies virus / physiology*
Rats
Recovery of Function

Substances

Mecp2 protein, mouse
Methyl-CpG-Binding Protein 2
RNA, Circular
Rabies Vaccines