Chitosan oligosaccharides packaged into rat adipose mesenchymal stem cells-derived extracellular vesicles facilitating cartilage injury repair and alleviating osteoarthritis

(2021) 19:343 Li et al. J Nanobiotechnol https://doi.org/10.1186/s12951-021-01086-x Journal of Nanobiotechnology Open Access RESEARCH Chitosan oligosaccharides packaged into rat adipose mesenchymal stem cells‑derived extracellular vesicles facilitating cartilage injury repair and alleviating osteoarthritis Shenglong Li1,2† , Jie Liu3†, Siyu Liu1, Weijie Jiao1 and Xiaohong Wang1,4* Abstract Objectives: This study aimed to investigate the roles of adipose mesenchymal stem cell (AMSC)-derived extracellular vesicles (EVs) binding with chitosan oligosaccharides (COS) in cartilage injury, as well as the related mechanisms. Results: IL-1β treatment significantly inhibited the viability and migration of chondrocytes and enhanced cell apoptosis (P < 0.05), while chitosan oligosaccharides and extracellular vesicles-chitosan oligosaccharide conjugates (EVsCOS/EVs-COS conjugates) reversed the changes induced by IL-1β (P < 0.05), and the effects of extracellular vesicleschitosan oligosaccharide conjugates were better than those of chitosan oligosaccharides (P < 0.05). After cartilage damage, IL-1β, OPN, and p53 were significantly upregulated, COL1A1, COL2A1, OCN, RUNX2, p-Akt/Akt, PI3K, c-Myc, and Bcl2 were markedly downregulated, and extracellular vesicles-chitosan oligosaccharide conjugates reversed the expression induced by cartilage injury. Through sequencing, 760 differentially expressed genes (DEGs) clustered into four expression patterns were associated with negative regulation of the canonical Wnt, PI3K-Akt, AMPK, and MAPK signaling pathways. Conclusion: Extracellular vesicles-chitosan oligosaccharide conjugates may serve as a new cell-free biomaterial to facilitate cartilage injury repair and improve osteoarthritis. Keywords: EVs, Chitosan oligosaccharides, EVs-COS conjugates, Cartilage injury repair, Osteoarthritis *Correspondence: ; wangxiaohong@mail. tsinghua.edu.cn † Shenglong Li and Jie Liu contributed equally to this work 1 Department of Tissue Engineering, Center of 3D Printing & Organ Manufacturing, School of Intelligent Medicine, China Medical University (CMU), No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China Full list of author information is available at the end of the article © The Author(s) 2021. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativeco mmons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Li et al. J Nanobiotechnol (2021) 19:343 Page 2 of 19 Graphical Abstract Background Articular cartilage, a non-self-repairing tissue, is mainly composed of water, proteoglycan, and collagen, which together determine the functional characteristics of cartilage tissues [1]. Cartilage injury usually marks the occurrence of tissue degeneration, progressive deterioration, subchondral osteosclerosis, and osteoarthritis (OA) [2]. OA clinically manifests as slow progression of joint pain, tenderness, stiffness, joint swelling, limited movement, and joint deformity [3], and is a major cause of disability, affecting approximately 240 million people globally [4]. Currently, drugs used to alleviate the symptoms of OA include steroid injections, non-steroidal anti-inflammatory drugs (NSAIDs), and opioids [5, 6]. However, the long-term use of these drugs may result in side effects, such as gastrointestinal, renal, and cardiovascular diseases [7]. Nanocomposites, including organic–inorganic, inorganic-inorganic, and bioinorganic nanomaterials, have been reported in bone tissue regeneration engineering, such as hydroxyapatite (HA) with chitosan, polycaprolactone/bioglass, and HA-gelatin nanocomposites [8]. By combining nanotechnology-based drug delivery systems, the bioavailability, pharmacokinetics and pharmacodynamics of drugs in bone tissue can be improved, thus improving therapeutic efficacy while reducing side effects. Owing to the immunogenicity of the receptor cells, rapid blood clearance, cytotoxicity, and poor biological distribution of these nanocomposites, their use has been limited [9]. Therefore, more therapeutic strategies are urgently needed to improve cartilage injury repair and manage OA. Mesenchymal stem cells (MSCs), which can be isolated from many adult organs, are self-renewing multipotent progenitors, and can differentiate into a variety of cell lineages, such as adipocytes, osteoblasts, and chondrocytes [10]. Increasing evidence has shown that MSC transplantation promotes tissue regeneration, including fracture, wound healing, and cartilage repair [11]. A previous study indicated that bone marrow MSCs could enhance articular cartilage repair and regeneration, as well as improve the quality of life of knee OA [12]. In addition, owing to their relatively easy isolation, high yield, and strong potential for proliferation and differentiation [13], adipose MSCs (AMSCs) have been widely used in various biomedical applications. A previous double-blinded clinical trial showed that intraarticular injection of AMSCs could improve cartilage defects and relieve pain in knee OA patients, without causing adverse events at 6 months’ follow-up [14]. Another study reported that AMSCs with BMP9 overexpression promoted cartilage repair and differentiation through the Notch1/Jagged1 signaling pathway [15]. These findings suggest that AMSCs can promote cartilage injury repair and improve OA. Nevertheless, the clinical effects of traditional AMSC transplantation methods have been greatly limited by their stability, safety, and immune-mediated rejection. Extracellular vesicles (EVs) are released by a variety of cells [16], and can serve as a tool for cell-to-cell communication. They can selectively encapsulate protein Li et al. J Nanobiotechnol (2021) 19:343 molecules, genes (RNA and DNA), cytokines, and other functional bioactive substances derived from the cells and deliver them to the extracellular environment or other target cells [17]. EVs derived from AMSCs have been considered an important part of cell-free regenerative medicine because they carry special bioactive substances and possess the distincti (...truncated)