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Engineering exosomes derived from subcutaneous fat MSCs specially promote cartilage repair as miR-199a-3p delivery vehicles in Osteoarthritis.

ABSTRACT: Osteoarthritis (OA) is a degenerative joint disease involving cartilage. Exosomes derived from Mesenchymal stem cells (MSCs) therapy improves articular cartilage repair, but subcutaneous fat (SC) stromal cells derived exosomes (MSCs^SC-Exos), especially engineering MSCs^SC-Exos for drug delivery have been rarely reported in OA therapy. This objective of this study was to clarify the underlying mechanism of MSCs^SC-Exos on cartilage repair and therapy of engineering MSCs^SC-Exos for drug delivery in OA. MSCs^SC-Exos could ameliorate the pathological severity degree of cartilage via miR-199a-3p, a novel molecular highly enriched in MSCs^SC-Exos, which could mediate the mTOR-autophagy pathway in OA rat model. Intra-articular injection of antagomiR-199a-3p dramatically attenuated the protective effect of MSCs^SC-Exos-mediated on articular cartilage in vivo. Furthermore, to achieve the superior therapeutic effects of MSCs^SC-Exos on injured cartilage, engineering exosomes derived from MSCs^SC as the chondrocyte-targeting miR-199a-3p delivery vehicles were investigated in vitro and in vivo. The chondrocyte-binding peptide (CAP) binding MSCs^SC-Exos could particularly deliver miR-199a-3p into the chondrocytes in vitro and into deep articular tissues in vivo, then exert the excellent protective effect on injured cartilage in DMM-induced OA mice. As it is feasible to obtain human subcutaneous fat from healthy donors by liposuction operation in clinic, meanwhile engineering MSCs^SC-Exos to realize targeted delivery of miR-199a-3p into chondrocytes exerted excellent therapeutic effects in OA animal model in vivo. Through combining MSCs^SC-Exos therapy and miRNA therapy via an engineering approach, we develop an efficient MSCs^SC-Exos-based strategy for OA therapy and promote the application of targeted-MSCs^SC-Exos for drug delivery in the future.

SUBMITTER: Zhao S

PROVIDER: S-EPMC10515007 | biostudies-literature | 2023 Sep

REPOSITORIES: biostudies-literature

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Engineering exosomes derived from subcutaneous fat MSCs specially promote cartilage repair as miR-199a-3p delivery vehicles in Osteoarthritis.

Zhao Shu S Xiu Guanghui G Wang Jian J Wen Yi Y Lu Jinyuan J Wu Baitong B Wang Guangming G Yang Danjing D Ling Bin B Du Dajiang D Xu Jun J

Journal of nanobiotechnology 20230922 1

Osteoarthritis (OA) is a degenerative joint disease involving cartilage. Exosomes derived from Mesenchymal stem cells (MSCs) therapy improves articular cartilage repair, but subcutaneous fat (SC) stromal cells derived exosomes (MSCsSC-Exos), especially engineering MSCsSC-Exos for drug delivery have been rarely reported in OA therapy. This objective of this study was to clarify the underlying mechanism of MSCsSC-Exos on cartilage repair and therapy of engineering ...[more]

PMID: 37736726

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Project description:BackgroundOsteoarthritis (OA) is the most common joint disorder, and places a heavy burden on individuals and society. As conventional therapies, such as surgery, rarely cure the disorder, targeted therapies represent a promising alternative. This research sought to explore the potential effect of miR-199a-5p on the development of OA.MethodsBased on the OA rat model, the serum was collected at 6 and 12 weeks, and microRNA (miRNA) sequencing was performed. A bioinformatics analysis was conducted to examine the differentially expressed micro ribonucleic acids, and qRT-PCR (real-time quantitative PCR) was conducted to determine their expression in the joint tissues of rats with OA. Rats articular chondrocytes were collected and treated with a miR-199a-5p antagomir or agomir. Afterwards, cell viability, autophagy was determinated. Dual luciferase was used to verify that miR-199a-5p targets the regulation of mitogen-stimulated protein kinase 4 (MAPK4). Subsequently, in chondrocytes, MAPK was knockdown to rescue the effect of miR-199a-5p inhibition, and cell viability and autophagy were examined. Finally, the OA model was treated with miR-199a-5p antagomir to detect joint pathology, cartilage tissue and inflammatory factor and autophagy was measured.ResultsMiR-199a-5p was greatly upregulated in OA, and miRNA was found to be differentially expressed in OA tissues. MAPK4 was identified to be a target gene of miR-199-5p. Inhibiting miR-199a-5p not only decreased the survival of chondrocytes and induced apoptosis, but also relieved inflammation and decreased the content of pro-inflammatory cytokines. Further, the silencing of miR-199a-5p protected the articular cartilage and improved gait abnormalities, but this effect was abrogated by the silencing of MAPK4.ConclusionsThe silencing of miR-199a-5p appears to improve gait abnormalities, promote the survival of chondrocytes, and improve the condition of OA. Our findings may lead to the development of miR-199a-5p-based targeted therapy for OA.

Project description:BackgroundsAbnormal cartilage calcification is one of the pathological changes of temporomandibular joint (TMJ) osteoarthritis (OA). Recent studies have reported that exosomes can regulate the formation of abnormal calcified nodules in diseases including atherosclerosis and chronic kidney disease. However, the influences of chondrocyte-derived exosomes on abnormal cartilage calcification in TMJ OA are still unclear.MethodsTMJ OA was induced by unilateral anterior crossbite (UAC) for 4, 8, or 12 weeks in rats to observe abnormal calcification in TMJ condylar cartilage and exosome formation. Concomitantly, GW4869, the inhibitor of exosome formation, was locally injected to the TMJ of rats under stimulation of UAC, while the exosomes extracted from primary condylar chondrocytes stimulated with fluid flow shear stress (FFSS) were locally injected to rats TMJ.ResultsAbnormal calcification was enhanced in the degenerative cartilage of TMJ OA in UAC rats, and a large number of exosome-like structures with diameters of 50-150 nm were found in the calcified cartilage together with decreased expression of matrix Gla protein (MGP) and increased expression of CD63, tissue-nonspecific alkaline phosphatase (TNAP) and nucleotide pyrophosphatase/phosphodiesterase-1 (NPP1). After FFSS stimulation, the number of exosomes secreted by chondrocytes and the numbers of calcified nodules were increased in cultured cells, and the protein levels of MGP, TNAP, and NPP1 in exosomes were changed. Inhibition of exosome formation, TNAP, and NPP1 or supplementation with exogenous MGP effectively alleviated FFSS-induced chondrocyte calcification. Local injection of GW4869, the exosome inhibitor, alleviated TMJ OA-related cartilage degeneration and calcification in UAC rats. Local injection of exosomes obtained from chondrocytes stimulated by FFSS to the TMJs of normal rats induced cartilage degeneration and calcification similar to that in TMJ OA.ConclusionsAbnormal biomechanical loading leads to enhanced formation of chondrocyte-derived exosomes, in which promoters of calcification increased and inhibitors decreased, resulting in accelerating abnormal cartilage calcification in TMJ OA. The inhibition of degenerative chondrocyte-derived exosomes is expected to be a new way to prevent and treat TMJ OA.

Project description:ObjectiveOur previous studies revealed that normal synovial exosomes promoted chondrogenesis, and microRNA (miR)-19b-3p independently related to osteoarthritis (OA) risk. Subsequently, this study intended to further explore the effect of OA fibroblast-like synoviocyte (OA-FLS) exosomal miR-19b-3p on OA ferroptosis and its potential mechanisms.MethodsInterleukin (IL)-1β-stimulated chondrocytes and medial meniscus surgery were used to construct the OA cellular model and the OA rat model, respectively. OA-FLS exosomes with/without miR-19b-3p modification were added to the IL-1β-stimulated chondrocytes and OA rat models, followed by direct miR-19b-3p mimic/inhibitor transfection with/without SLC7A11 overexpression plasmids. miR-19b-3p, ferroptosis-related markers (malondialdehyde (MDA), glutathione (GSH)/oxidized glutathione (GSSG), ferrous ion (Fe2+), glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), and acyl-CoA synthetase long-chain family member 4 (ACSL4)), mitochondrial membrane potential (MMP), and reactive oxygen species (ROS) levels were detected.ResultsEnhanced ferroptosis reflected by dysregulated ferroptosis-related markers, a reduced MMP, and an increased ROS was observed in cartilage tissues from OA patients vs. controls, IL-1β-stimulated chondrocytes vs. normal ones, and OA rat models vs. sham, so did miR-19b-3p. OA-FLS exosomes promoted MDA, Fe2+, ACSL4, and ROS but reduced cell viability, GSH/GSSG, GPX4, SLC7A11, and MMP in IL-1β-stimulated chondrocytes, whose effect was enhanced by miR-19b-3p mimics and attenuated by miR-19b-3p inhibitors. miR-19b-3p negatively regulated SLC7A11 and directly bound to SLC7A11 via luciferase reporter gene assay. Furthermore, SLC7A11 overexpression weakened miR-19b-3p mimics' effect on ferroptosis-related markers, MMP, or ROS in IL-1β-stimulated chondrocytes. OA-FLS exosomes also induced cartilage damage and ferroptosis in OA rats whose influence was tempered by miR-19b-3p inhibitors.ConclusionOA-FLS exosomal miR-19b-3p enhances cartilage ferroptosis and damage by sponging SLC7A11 in OA, indicating a potential linkage among synovium, cartilage, and ferroptosis during the OA process.

Project description:Exosomes (Exos) secreted by adipose-derived stem cells (ADSCs) have shown potential in alleviating osteoarthritis (OA). Previous studies indicated that infrapatellar fat pad (IPFP) derived stem cells (IPFSCs) may be more suitable for the treatment of OA than subcutaneous adipose tissue (ScAT) derived stem cells (ScASCs). However, it remains unclear which type of Exos offers superior therapeutic benefit for OA. This study first compared the differences between Exos derived from IPFP stem cells (ExosIPFP) and ScAT stem cells (ExosScAT) in OA treatment. Results suggested that ExosIPFP significantly inhibit the degradation of cartilage extracellular matrix (ECM) than ExosScAT, following this, the differences in microRNA (miRNA) expression between the two types of Exos using small RNA sequencing were performed. Subsequently, miR-99 b-3p was chosen and over-expressed in ExosScAT (ExosScAT-99b-3p), both in vivo and in vitro experiments demonstrated its efficacy in inhibiting the expression of ADAMTS4, promoting the repair of the ECM in OA. Finally, microfluidic technology was performed to fabricate a hyaluronan-based hydrogel microparticles (HMPs) for encapsulating Exos (HMPs@exos), the injectability, sustained release of Exos and long-term therapeutic effect on OA were validated. In summary, these results suggest miR-99 b-3p regulates the degradation of cartilage ECM by targeting ADAMTS4, the upregulation of miR-99 b-3p in ExosScAT would enable them to exhibit comparable or even superior effectiveness to ExosIPFP for OA treatment, making it a promising approach for OA treatment. Considering the abundant resources of ScAT and the limited availability of IPFP, ScAT harvested through liposuction could be genetically engineered to yield Exos for OA treatment. Furthermore, the encapsulation of Exos in HMPs provides an injectable sustained local drug release system, which could potentially enhance the efficacy of Exos and hold potential as future therapeutic strategies.

Project description:Osteoarthritis (OA) is a most common form of arthritis worldwide leading to significant disability. MicroRNAs (miRNAs) are non-coding RNAs involved in various aspects of cartilage development, homoeostasis and pathology. Several miRNAs have been identified which have shown to regulate expression of target genes relevant to OA pathogenesis such as matrix metalloproteinase (MMP)-13, cyclooxygenase (COX)-2, etc. Epigallocatechin-3-O-gallate (EGCG), the most abundant and active polyphenol in green tea, has been reported to have anti-arthritic effects, however, the role of EGCG in the regulation of miRNAs has not been investigated in OA. Here, we showed that EGCG inhibits COX-2 mRNA/protein expression or prostaglandin E2 (PGE2 ) production via up-regulating microRNA hsa-miR-199a-3p expression in interleukin (IL)-1β-stimulated human OA chondrocytes. This negative co-regulation of hsa-miR-199a-3p and COX-2 by EGCG was confirmed by transfection of OA chondrocytes with anti-miR-199a-3p. Transfection of OA chondrocytes with anti-miR-199a-3p significantly enhanced COX-2 expression and PGE2 production (P < 0.001), while EGCG treatment significantly inhibited anti-miR-199a-3p transfection-induced COX-2 expression or PGE2 production in a dose-dependent manner. These results were further re-validated by co-treatment of these transfection OA chondrocytes with IL-1β and EGCG. EGCG treatment consistently up-regulated the IL-1β-decreased hsa-miR-199a-3p expression (P < 0.05) and significantly inhibited the IL-1β-induced COX-2 expression/PGE2 production (P < 0.05) in OA chondrocytes transfected with anti-hsa-miR-199a-3p. Taken together, these results clearly indicate that EGCG inhibits COX-2 expression/PGE2 production via up-regulation of hsa-miR-199a-3p expression. These novel pharmacological actions of EGCG on IL-1β-stimulated human OA chondrocytes provide new suggestions that EGCG or EGCG-derived compounds inhibit cartilage breakdown or pain by up-regulating the expression of microRNAs in human chondrocytes.

Dataset Information

Engineering exosomes derived from subcutaneous fat MSCs specially promote cartilage repair as miR-199a-3p delivery vehicles in Osteoarthritis.

Publications

Engineering exosomes derived from subcutaneous fat MSCs specially promote cartilage repair as miR-199a-3p delivery vehicles in Osteoarthritis.

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