{"title":"Mesenchymal Stem Cell-Derived Extracellular Vesicles Protect Rat Nucleus Pulposus Cells from Oxidative Stress.","authors":"Sobia Ekram, Shumaila Khalid, Faiza Ramzan, Asmat Salim, Imtiaz Bashir, Marie Christine Durrieu, Irfan Khan","doi":"10.1177/19476035231172154","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Oxidative stress (OS) is mainly associated with the pathogenesis of intervertebral disc (IVD) degeneration; it causes nucleus pulposus cells (NPCs) to undergo senescence and triggers autophagy and apoptosis. This study aims to evaluate the regeneration potential of extracellular vesicles (EVs) derived from human umbilical cord-mesenchymal stem cells (hUC-MSCs) in an <i>in vitro</i> rat NPC-induced OS model.</p><p><strong>Design: </strong>NPCs were isolated from rat coccygeal discs, propagated, and characterized. OS was induced by hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>), which is confirmed by 2,7-dichlorofluorescein diacetate (H<sub>2</sub>DCFDA) assay. EVs were isolated from hUC-MSCs and characterized by analyzing the vesicles using fluorescence microscope, scanning electron microscope (SEM), atomic force microscope (AFM), dynamic light scattering (DLS), and Western blot (WB). The <i>in vitro</i> effects of EVs on migration, uptake, and survival of NPCs were determined.</p><p><strong>Results: </strong>SEM and AFM topographic images revealed the size distribution of EVs. The phenotypes of isolated EVs showed that the size of EVs was 403.3 ± 85.94 nm, and the zeta potential was -0.270 ± 4.02 mV. Protein expression analysis showed that EVs were positive for CD81 and annexin V. Treatment of NPCs with EVs reduced H<sub>2</sub>O<sub>2</sub>-induced OS as evidenced by a decrease in reactive oxygen species (ROS) levels. Co-culture of NPCs with DiI-labeled EVs showed the cellular internalization of EVs. In the scratch assay, EVs significantly increased NPC proliferation and migration toward the scratched area. Quantitative polymerase chain reaction analysis showed that EVs significantly reduced the expression of OS genes.</p><p><strong>Conclusion: </strong>EVs protected NPCs from H<sub>2</sub>O<sub>2</sub>-induced OS by reducing intracellular ROS generation and improved NPC proliferation and migration.</p>","PeriodicalId":9626,"journal":{"name":"CARTILAGE","volume":null,"pages":null},"PeriodicalIF":2.7000,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11418459/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"CARTILAGE","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1177/19476035231172154","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2023/5/4 0:00:00","PubModel":"Epub","JCR":"Q1","JCRName":"ORTHOPEDICS","Score":null,"Total":0}
引用次数: 0
Abstract
Background: Oxidative stress (OS) is mainly associated with the pathogenesis of intervertebral disc (IVD) degeneration; it causes nucleus pulposus cells (NPCs) to undergo senescence and triggers autophagy and apoptosis. This study aims to evaluate the regeneration potential of extracellular vesicles (EVs) derived from human umbilical cord-mesenchymal stem cells (hUC-MSCs) in an in vitro rat NPC-induced OS model.
Design: NPCs were isolated from rat coccygeal discs, propagated, and characterized. OS was induced by hydrogen peroxide (H2O2), which is confirmed by 2,7-dichlorofluorescein diacetate (H2DCFDA) assay. EVs were isolated from hUC-MSCs and characterized by analyzing the vesicles using fluorescence microscope, scanning electron microscope (SEM), atomic force microscope (AFM), dynamic light scattering (DLS), and Western blot (WB). The in vitro effects of EVs on migration, uptake, and survival of NPCs were determined.
Results: SEM and AFM topographic images revealed the size distribution of EVs. The phenotypes of isolated EVs showed that the size of EVs was 403.3 ± 85.94 nm, and the zeta potential was -0.270 ± 4.02 mV. Protein expression analysis showed that EVs were positive for CD81 and annexin V. Treatment of NPCs with EVs reduced H2O2-induced OS as evidenced by a decrease in reactive oxygen species (ROS) levels. Co-culture of NPCs with DiI-labeled EVs showed the cellular internalization of EVs. In the scratch assay, EVs significantly increased NPC proliferation and migration toward the scratched area. Quantitative polymerase chain reaction analysis showed that EVs significantly reduced the expression of OS genes.
Conclusion: EVs protected NPCs from H2O2-induced OS by reducing intracellular ROS generation and improved NPC proliferation and migration.
期刊介绍：
CARTILAGE publishes articles related to the musculoskeletal system with particular attention to cartilage repair, development, function, degeneration, transplantation, and rehabilitation. The journal is a forum for the exchange of ideas for the many types of researchers and clinicians involved in cartilage biology and repair. A primary objective of CARTILAGE is to foster the cross-fertilization of the findings between clinical and basic sciences throughout the various disciplines involved in cartilage repair.
The journal publishes full length original manuscripts on all types of cartilage including articular, nasal, auricular, tracheal/bronchial, and intervertebral disc fibrocartilage. Manuscripts on clinical and laboratory research are welcome. Review articles, editorials, and letters are also encouraged. The ICRS envisages CARTILAGE as a forum for the exchange of knowledge among clinicians, scientists, patients, and researchers.
The International Cartilage Repair Society (ICRS) is dedicated to promotion, encouragement, and distribution of fundamental and applied research of cartilage in order to permit a better knowledge of function and dysfunction of articular cartilage and its repair.
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