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Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway

Rational: Ischemic injury of the skeletal muscle remains a serious clinical problem and currently there is no effective therapy. The aim of the present study is to determine whether human umbilical cord mesenchymal stem cells- derived exosomes (UMSC-Exo) could repair ischemic injury by releasing cir...

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Autores principales: Yan, Bing, Zhang, Yu, Liang, Chun, Liu, Bin, Ding, Fengzhi, Wang, Yanli, Zhu, Bao, Zhao, Ranzun, Yu, Xi-Yong, Li, Yangxin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Ivyspring International Publisher 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7295049/
https://www.ncbi.nlm.nih.gov/pubmed/32550900
http://dx.doi.org/10.7150/thno.42259
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author Yan, Bing
Zhang, Yu
Liang, Chun
Liu, Bin
Ding, Fengzhi
Wang, Yanli
Zhu, Bao
Zhao, Ranzun
Yu, Xi-Yong
Li, Yangxin
author_facet Yan, Bing
Zhang, Yu
Liang, Chun
Liu, Bin
Ding, Fengzhi
Wang, Yanli
Zhu, Bao
Zhao, Ranzun
Yu, Xi-Yong
Li, Yangxin
author_sort Yan, Bing
collection PubMed
description Rational: Ischemic injury of the skeletal muscle remains a serious clinical problem and currently there is no effective therapy. The aim of the present study is to determine whether human umbilical cord mesenchymal stem cells- derived exosomes (UMSC-Exo) could repair ischemic injury by releasing circular RNA. Methods and Results: To create hindlimb ischemia, we surgically ligated the left femoral artery in C57BL/6 mice. Using circRNA-seq analyses of total RNA from ischemic and control muscles, we found reduced expression of circHIPK3 in the ischemic muscle. To explore the role of circHIPK3 in ischemic injury, the mice were randomly assigned into three groups after surgery: 1) vehicle; 2) UMSC-Exo; 3) UMSC-Exo and siRNA targeting circHIPK3 (UMSC-Exo /si-circHIPK3). UMSC-Exo treatment significantly increased expression of circHIPK3 and improved blood perfusion, running distance and muscle force, which were reversed by injection of UMSC-Exo /si-circHIPK3, suggesting that UMSC-Exo improve muscle function by releasing circHIPK3. UMSC-Exo treatment also inhibited ischemia induced pyroptosis - cell death caused by inflammasome as evidenced by activation of NLRP3, cleaved caspase-1, and subsequent increase of IL-1β and IL-18, and the effects were reversed by injection UMSC-Exo /si-circHIPK3. Bioinformatic analysis identified miR-421/FOXO3a as a potential target for circHIPK3, which was confirmed by luciferase reporter assay. Knockdown of circHIPK3 in C2C12 cells resulted in increased expression of miR-421. We established an in vitro model of pyroptosis by stimulating C2C12 cells with LPS and ATP. LPS and ATP treatment resulted in reduced expression of circHIPK3 and increased expression of miR-421, which was prevented by UMSC-Exo. Western blot analysis showed reduced levels of NLRP3 and cleaved caspase-1 when cells were treated by UMSC-Exo. The expression of FOXO3a in C2C12 cells was increased in the presence of miR-421 inhibitor, and the expression was reduced when cells were treated by LPS and ATP. Importantly, the expression of FOXO3a was upregulated by UMSC-Exo but was reduced when si-circHIPK3 was present. Conclusions: Using loss/gain-of function method, we demonstrated that miR-421/FOXO3a is the direct target of circHIPK3, and UMSC-Exo prevent ischemic injury by releasing circHIPK3, which in turn down regulate miR-421, resulting in increased expression of FOXO3a, leading to inhibition of pyroptosis and release of IL-1β and IL-18.
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spelling pubmed-72950492020-06-17 Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway Yan, Bing Zhang, Yu Liang, Chun Liu, Bin Ding, Fengzhi Wang, Yanli Zhu, Bao Zhao, Ranzun Yu, Xi-Yong Li, Yangxin Theranostics Research Paper Rational: Ischemic injury of the skeletal muscle remains a serious clinical problem and currently there is no effective therapy. The aim of the present study is to determine whether human umbilical cord mesenchymal stem cells- derived exosomes (UMSC-Exo) could repair ischemic injury by releasing circular RNA. Methods and Results: To create hindlimb ischemia, we surgically ligated the left femoral artery in C57BL/6 mice. Using circRNA-seq analyses of total RNA from ischemic and control muscles, we found reduced expression of circHIPK3 in the ischemic muscle. To explore the role of circHIPK3 in ischemic injury, the mice were randomly assigned into three groups after surgery: 1) vehicle; 2) UMSC-Exo; 3) UMSC-Exo and siRNA targeting circHIPK3 (UMSC-Exo /si-circHIPK3). UMSC-Exo treatment significantly increased expression of circHIPK3 and improved blood perfusion, running distance and muscle force, which were reversed by injection of UMSC-Exo /si-circHIPK3, suggesting that UMSC-Exo improve muscle function by releasing circHIPK3. UMSC-Exo treatment also inhibited ischemia induced pyroptosis - cell death caused by inflammasome as evidenced by activation of NLRP3, cleaved caspase-1, and subsequent increase of IL-1β and IL-18, and the effects were reversed by injection UMSC-Exo /si-circHIPK3. Bioinformatic analysis identified miR-421/FOXO3a as a potential target for circHIPK3, which was confirmed by luciferase reporter assay. Knockdown of circHIPK3 in C2C12 cells resulted in increased expression of miR-421. We established an in vitro model of pyroptosis by stimulating C2C12 cells with LPS and ATP. LPS and ATP treatment resulted in reduced expression of circHIPK3 and increased expression of miR-421, which was prevented by UMSC-Exo. Western blot analysis showed reduced levels of NLRP3 and cleaved caspase-1 when cells were treated by UMSC-Exo. The expression of FOXO3a in C2C12 cells was increased in the presence of miR-421 inhibitor, and the expression was reduced when cells were treated by LPS and ATP. Importantly, the expression of FOXO3a was upregulated by UMSC-Exo but was reduced when si-circHIPK3 was present. Conclusions: Using loss/gain-of function method, we demonstrated that miR-421/FOXO3a is the direct target of circHIPK3, and UMSC-Exo prevent ischemic injury by releasing circHIPK3, which in turn down regulate miR-421, resulting in increased expression of FOXO3a, leading to inhibition of pyroptosis and release of IL-1β and IL-18. Ivyspring International Publisher 2020-05-18 /pmc/articles/PMC7295049/ /pubmed/32550900 http://dx.doi.org/10.7150/thno.42259 Text en © The author(s) This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.
spellingShingle Research Paper
Yan, Bing
Zhang, Yu
Liang, Chun
Liu, Bin
Ding, Fengzhi
Wang, Yanli
Zhu, Bao
Zhao, Ranzun
Yu, Xi-Yong
Li, Yangxin
Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway
title Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway
title_full Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway
title_fullStr Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway
title_full_unstemmed Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway
title_short Stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circHIPK3/ FOXO3a pathway
title_sort stem cell-derived exosomes prevent pyroptosis and repair ischemic muscle injury through a novel exosome/circhipk3/ foxo3a pathway
topic Research Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7295049/
https://www.ncbi.nlm.nih.gov/pubmed/32550900
http://dx.doi.org/10.7150/thno.42259
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