Synovial Mesenchymal Stem Cells Research Articles

Small extracellular vesicles in combination with sleep-related circRNA3503: A targeted therapeutic agent with injectable thermosensitive hydrogel to prevent osteoarthritis

Osteoarthritis (OA), characterized by chondrocyte apoptosis and disturbance of the balance between catabolism and anabolism of the extracellular matrix (ECM), is the most common age-related degenerative joint disease worldwide. As sleep has been found to be beneficial for cartilage repair, and circular RNAs (circRNAs) have been demonstrated to be involved in the pathogenesis of OA, we performed RNA sequencing (RNA-seq), and found circRNA3503 was significantly increased after melatonin (MT)-induced cell sleep. Upregulation of circRNA3503 expression completely rescued the effects of interleukin-1β (IL-1β), which was used to simulate OA, on apoptosis, ECM degradation- and synthesis-related genes. Mechanistically, circRNA3503 acted as a sponge of hsa-miR-181c-3p and hsa-let-7b-3p. Moreover, as we previously showed that small extracellular vesicles (sEVs) derived from synovium mesenchymal stem cells (SMSCs) can not only successfully deliver nucleic acids to chondrocytes, but also effectively promote chondrocyte proliferation and migration, we assessed the feasibility of sEVs in combination with sleep-related circRNA3503 as an OA therapy. We successfully produced and isolated circRNA3503-loaded sEVs (circRNA3503-OE-sEVs) from SMSCs. Then, poly(D,l-lactide)-b-poly(ethylene glycol)-b-poly(D,l-lactide) (PDLLA-PEG-PDLLA, PLEL) triblock copolymer gels were used as carriers of sEVs. Through in vivo and in vitro experiments, PLEL@circRNA3503-OE-sEVs were shown to be a highly-effective therapeutic strategy to prevent OA progression. Through multiple pathways, circRNA3503-OE-sEVs alleviated inflammation-induced apoptosis and the imbalance between ECM synthesis and ECM degradation by acting as a sponge of hsa-miR-181c-3p and hsa-let-7b-3p. In addition, circRNA3503-OE-sEVs promoted chondrocyte renewal to alleviate the progressive loss of chondrocytes. Our results highlight the potential of PLEL@circRNA3503-OE-sEVs for preventing OA progression.

Thawed cryopreserved synovial mesenchymal stem cells show comparable effects to cultured cells in the inhibition of osteoarthritis progression in rats

Intra-articular injections of mesenchymal stem cells (MSCs) can inhibit the progression of osteoarthritis (OA). Previous reports have used cultured MSCs, but the ability to use thawed cryopreserved MSC stocks would be highly advantageous. Our purpose was to elucidate whether thawed cryopreserved MSCs show comparable inhibitory effects on OA progression in rats to those obtained with cultured MSCs. Cultured rat synovial MSCs or thawed MSCs were compared for in vitro viability and properties. The inhibitory effect of thawed MSCs on OA progression was evaluated by injecting cryopreservation fluid and thawed MSCs in meniscectomized rats. Cartilage degeneration was assessed using gross finding and histological scores. Cultured MSCs were then injected into one knee and thawed MSCs into the contralateral knee of the same individual to compare their effects. Cultured MSCs and MSCs thawed after cryopreservation had comparable in vitro colony formation and chondrogenic potentials. In the rat OA model, the gross finding and histological scores were significantly lower in the thawed MSC group than in the cryopreservation fluid group at 8 weeks. Finally, cartilage degeneration did not differ significantly after injection of cultured and thawed MSCs. In conclusion, thawed MSCs showed comparable inhibitory effects on OA progression to cultured MSCs.

Intraarticular injections of mesenchymal stem cells in knee osteoarthritis

Purpose: Several studies have reported improvement in knee pain following mesenchymal stem cell (MSC) injections for knee osteoarthritis (OA). We developed a novel 3D magnetic resonance imaging (MRI) analysis software program that provides “projected cartilage area ratios” for automatic detection of changes in cartilage amounts. The primary objective of this prospective interventional study was to compare alterations in the projected cartilage area ratio (thickness ≥ 1.5mm) at the femoral posteromedial region between 30 weeks before and 30 weeks after synovial MSC injections.

Intra‐articular Injection of PDGF-BB Explored in a Novel in Vitro Model Mobilizes Mesenchymal Stem Cells From the Synovium Into Synovial Fluid in Rats

Drugs that can induce mesenchymal stem cell (MSC) mobilization from synovium into synovial fluid will enable regenerative medicine in joints without use of exogenous MSCs. An in vitro synovial MSC migration model had previously been developed for screening but had problems in practical application. We herein developed a novel in vitro model, explored cytokines for synovial MSC mobilization with this model, and verified whether MSCs in synovial fluid increase following intra-articular injection of the cytokine. Human synovial MSCs embedded in a mixture of Matrigel and type 1 collagen hydrogel were placed on a culture insert and then put in medium containing migration factor. Of the six cytokines, we identified the one that mobilizes the highest number of MSCs. PDGF-BB or PBS was injected into rat knees, and 48h later, synovial fluid was collected and cultured. The cells were examined for MSC properties. PDGF-BB was the most effective for synovial MSC mobilization among six cytokines. The effect of PDGF-BB was inhibited by a PRGFR inhibitor. Injection of PDGF-BB into rat knees increased colony-forming cells in the synovial fluid. These cells had surface epitopes and multipotency comparable to MSCs and a higher capacity for proliferation, colony formation, and chondrogenesis. This novel in vitro model recapitulated the migration of MSCs from synovium into synovial fluid. Our exploration of cytokines revealed that PDGF-BB strongly induced in vitro synovial MSC migration, while intra-articular injection of PDGF-BB increased in vivo MSC numbers in synovial fluid in rats.

Fibrin Glue-Kartogenin Complex Promotes the Regeneration of the Tendon-Bone Interface in Rotator Cuff Injury.

Objective Rotator cuff injury healing is problematic because the tendon-bone junction often forms cicatricial tissues, rather than fibrocartilage, which leads to mechanical impairment and is prone to redamage. Kartogenin (KGN) is a newly discovered small molecule compound which can induce cartilage formation through chondrogenesis of endogenous mesenchymal stem cells. Methods In this study, we used KGN with fibrin glue (FG) to repair the rotator cuff injury by promoting the formation of fibrocartilage at the tendon to bone interface. Firstly, we assessed the release rate of KGN from the FG-KGN complex and then created a rabbit rotator cuff tendon graft-bone tunnel model. The rabbits received saline, FG-KGN, or FG injections onto the tendon to bone interface after injury. Shoulder tissues were harvested at 6 and 12 weeks, and the sections were stained with HE and Safranin O/Fast green. The samples were assessed by histologic evaluation and biomechanical testing. Synovial mesenchymal stem cells derived from the synovial tissue around the rotator cuff were harvested for western blotting and qRT-PCR analysis. Results KGN was released rapidly from the FG-KGN complex during first 4 hrs and followed by a slow release until 7 days. The tendon graft-bone interface in the control (saline) group and the FG group was filled with scar tissue, rather than cartilage-like tissue, and only a small number of chondrocytes were found at the adjacent bone surface. In the FG-KGN group, the tendon to bone interface was fully integrated and populated by chondrocytes with proteoglycan deposition, indicating the formation of fibrocartilage-like tissues. At 12 weeks, the maximum tensile strength of the FG-KGN group was significantly higher than that of the FG and control groups (P < 0.01). The RNA expression levels of tendinous genes such as Tenascin C and the chondrogenic gene Sox-9 were substantially elevated in SMSCs treated with the FG-KGN complex compared to the other two groups. Conclusion These results indicated that fibrin glue is an effective carrier for KGN, allowing for the sustained release of KGN. The FG-KGN complex could effectively promote the regeneration and formation of fibrocartilage tissue of the tendon-bone interface in the rabbit rotator cuff tendon graft-bone tunnel model.

Characteristics of MSCs in Synovial Fluid and Mode of Action of Intra-Articular Injections of Synovial MSCs in Knee Osteoarthritis.

We have been studying mesenchymal stem cells (MSCs) in synovial fluid and the intra-articular injection of synovial MSCs in osteoarthritis (OA) knees. Here, mainly based on our own findings, we overview the characteristics of endogenous MSCs in the synovial fluid of OA knees and their mode of action when injected exogenously into OA knees. Many MSCs similar to synovial MSCs were detected in the synovial fluid of human OA knees, and their number correlated with the radiological OA grade. Our suspended synovium culture model demonstrated the release of MSCs from the synovium through a medium into a non-contacting culture dish. In OA knees, endogenous MSCs possibly mobilize in a similar manner from the synovium through the synovial fluid and act protectively. However, the number of mobilized MSCs is limited; therefore, OA progresses in its natural course. Synovial MSC injections inhibited the progression of cartilage degeneration in a rat OA model. Injected synovial MSCs migrated into the synovium, maintained their MSC properties, and increased the gene expressions of TSG-6, PRG-4, and BMP-2. Exogenous synovial MSCs can promote anti-inflammation, lubrication, and cartilage matrix synthesis in OA knees. Based on our findings, we have initiated a human clinical study of synovial MSC injections in OA knees.

ADULT MESENCHYMAL STEM CELL-BASED APPROACHES FOR OSTEOARTHRITIS: CURRENT PERSPECTIVES AND CHALLENGES

Osteoarthritis (OA) is a degenerative joint disease that may cause joint inflammation, stiffness, and pain. Current therapy for OA involves symptomatic treatment, mainly pain management. Therefore, it does not repair degenerated cartilage or attenuate joint inflammation. Because articular cartilage cannot heal or regenerate these tissues, tissue regeneration remains one of the most important objectives of new and potential OA therapeutics. The main features of adult mesenchymal stem cells (MSCs) are simple acquisition from adult tissues, rapid proliferation in vitro, immunomodulation in vivo, and lasting existence in the host, which are beneficial for OA treatment. In the past 15 years, adult MSCs, including bone marrow-, adipose-, and synovial membrane-derived MSCs, and their secretome have been successfully used in different animal (preclinical) models or in genetic manipulation for regenerating degenerated cartilage, reducing inflammation, or relieving pain. Furthermore, the implantation of adult MSCs showed pain reduction, anti-inflammation, and cartilage protection or healing in early-phase clinical trials. Adult MSCs are the most extensively explored as potential regenerative medicine for OA because of their efficacy in chondrocyte differentiation and their immunomodulatory properties. In this review paper, we highlighted current knowledge and future perspectives regarding preclinical tests, clinical application, and MSC-based/related products for curing OA.

Biofunctional Extracellular Matrix-Polycaprolactone-Hydroxyapatite Scaffold and Synovium Mesenchymal Stem Cells/Chondrocytes for Repairing Cartilage Defects.

Articular cartilage defects and degeneration can be caused by multiple factors, and the current clinical treatment schemes for pathological changes are relatively limited. Engineered cartilage tissue represents an alternative therapy for repairing cartilage defects in regenerative medicine. The scaffold material is considered the framework of tissue engineering; thus, scaffold material selection plays a crucial role in the therapy outcome. Polycaprolactone (PCL)-hydroxyapatite (HA) has been applied as a scaffold material for bone and cartilage tissue engineering with nontoxic, harmless metabolites and proper physical properties. The extracellular matrix (ECM) is mainly composed of collagen and proteoglycan, as well as a large number of growth factors and cytokines, which provide a tissue-specific microenvironment for host cells. Adipose-derived stem cells are pluripotent stem cells, and transforming growth factor-β3 (TGF-β3) enables mesenchymal stem cells to promote ECM production. This study, via in vitro and in vivo experiments, elucidated that the synovium mesenchymal stem cells (SMSCs) + chondrocytes + ECM-PCL-HA repair system, which is constructed upon the ECM-PCL-HA scaffold material, exhibits an adequate chondrogenic ability and reparatory effect. Overall, ECM-PCL-HA can be defined as a biofunctional scaffold material. The SMSCs + chondrocytes + ECM-PCL-HA repair system showed good confluency between the new cartilage and the surface, as well as the interface of the adjacent host cartilage. Furthermore, the structure of new cartilage tissue is consistent with adjacency. Thus, it can be used as a preferred plan for articular cartilage defect repair. Impact statement Studies investigating the chondrogenic ability and reparatory effect of the synovium mesenchymal stem cells (SMSCs) + chondrocytes + extracellular matrix (ECM)-polycaprolactone (PCL)-hydroxyapatite (HA) repair system provided a theoretical and practical basis for choosing ECM-PCL-HA as the scaffold material for cartilage tissue engineering. In this study, the transforming growth factor-β3 (TGF-β3) gene was introduced into adipose-derived stem cells (ADSCs) using a lentiviral vector to enhance ECM production. The decellularized ADSCs-ECM-PCL-HA acted as a biofunctional scaffold material with suitable physicochemical properties, which was advantageous for SMSC and chondrocyte adhesion and growth. Lastly, the SMSCs + chondrocytes + ECM-PCL-HA repair system showed excellent capability in the flush fusion state of the prosthetic surface and interface.

Histological Analysis of Cartilage Defects Repaired with an Autologous Human Stem Cell Construct 48 Weeks Postimplantation Reveals Structural Details Not Detected by T2-Mapping MRI

The aim of this study was to elucidate the efficacy of T2-mapping MRI and correlation with histology for the evaluation of tissue repair quality following the first-in-human implantation of an autologous tissue engineered construct. We directly compared the results of T2-mapping MRI of cartilage repair tissue with the histology of a biopsy specimen from the corresponding area at 48 weeks postoperatively in 5 patients who underwent the implantation of a scaffold-free tissue-engineered construct generated from autologous synovial mesenchymal stem cells to repair an isolated cartilage lesion. T2 values and histological scores were compared at each of 2 layers of equally divided halves of the repair tissue (upper and lower zones). Histology showed that the repair tissue in the upper zone was dominated by fibrous tissue and the ratio of hyaline-like matrix increased with the depth of the repair tissue. There were significant differences between upper and lower zones in histological scores. Conversely, there were no detectable statistically significant differences in T2 value detected among zones of the repair tissue, but zonal differences were detected in corresponding healthy cartilage. Accordingly, there were no correlations detected between histological scores and T2 values for each repair cartilage zone. Discrepancies in the findings between T2 mapping and histology suggest that T2 mapping was limited in ability to detect details in the architecture and composition of the repair cartilage.

MiR-129-5p shuttled by human synovial mesenchymal stem cell-derived exosomes relieves IL-1β induced osteoarthritis via targeting HMGB1

AimsTo explore the therapeutic effect of miR-129-5p carried by exosomes from Human Synovial Mesenchymal Stem Cell (HS-MSC) on osteoarthritis(OA). Materials and methodsThe levels of miR-129-5p and high mobility group protein -1 (HMGB1) and interleukin-1β (IL-1β) in the joint fluid of OA patients were respectively detected via real-time quantitative reverse transcription-PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). IL-1β was taken to act on chondrocytes for the establishment of OA model in vitro. Ultracentrifugation was conducted to isolate HS-MSC exosomes (HS-MSC-Exo) from the supernatant. Western blot and ELISA were carried out to measure the expression of iNOS, COX2, MMP13, Collagen 2, TLR4, NF-κB, Caspase3, Bcl-2, HMGB1 in chondrocytes. Flow cytometry was conducted to detect the apoptosis of chondrocytes. Besides, bioinformatics was employed to predict the targeted relationship between miR-129-5p and HMGB1, which was further verified via dual luciferase activity experiments. Key findingsThe results illustrated that miR-129-5p was decreased in OA patients and IL-1β-induced chondrocytes, while HMGB1 was notably upregulated. HS-MSC-Exo rich in miR-129-5p remarkably declined the inflammatory response and apoptosis of chondrocytes, while HS-MSC-Exo deficient in miR-129-5p increased the IL-1β-mediated inflammatory response and apoptosis of chondrocytes. In terms of mechanism, miR-129-5p targets the 3’UTR end of HMGB1 and inhibits IL-1β-mediated upregulation of HMGB1. SignificanceIn a word, this paper proved that miR-129-5p, existing in HS-MSC-Exo, can suppress the IL-1β-mediated OA by inhibiting HMGB1 release.

Therapeutic Potential of Exosomal circRNA Derived from Synovial Mesenchymal Cell via Targeting circEDIL3/miR-485-3p/PIAS3/STAT3/VEGF Functional Module in Rheumatoid Arthritis

Background: Synovial inflammation and its associated activation of angiogenesis play critical roles in rheumatoid arthritis (RA). Exosomes, as carriers of genetic information including circular RNAs (circRNAs), have been studied as delivery vehicles for therapeutic molecules. However, effects and mechanisms of synovial mesenchymal stem cells (SMSCs)-derived exosomal circRNAs in RA progression remain unclear. Methods: SMSCs derived exsomes (SMSCs-Exo) were administered to a RA fibroblast-like synoviocytes (RA-FLS) and human dermal microvascular endothelial cells (HDMECs) co-culture in vitro as well as collagen-induced arthritis (CIA) mice model in vivo . Therapeutic effect and angiogenic activity plus VEGF expression were evaluated. Exosomes from circEDIL-overexpressed SMSCs (Ad-circEDIL3-SMSCs-Exo) were used for further determination of circEDIL’s role in SMSCs-Exo-based therapy. Findings: VEGF induced by IL-6/sIL-6R complex in supernatants of RA-FLS and HDMECs co-culture as well as cocultured RA-FLS were significantly downregulated by SMSCs-Exo and Ad-circEDIL3-SMSCs-Exo, and the decreased extent was more significant in the latter group. Subsequent angiogenic activity were significantly downregulated by SMSCs-Exo and Ad-circEDIL3-SMSCs-Exo via targeting VEGF. CircEDIL3 was further determined to be a sponge for the PIAS3-targeting miR-485-3p , and STAT3 activity was then suppressed, with reducement of downstream VEGF. Synovial VEGF and consequently arthritis severity were verified to be reduced in CIA mice model under injection of SMSCs-Exo or Ad-circEDIL3-SMSCs-Exo. Interpretation: The intracellular transfer of circEDIL3 by SMSCs-Exo suggests a potential novel therapeutic strategy for RA. Funding: National Natural Science Found (NSFC 81901638), Postdoctoral Science Foundation of Chinese (2019M651309) and Heilongjiang (LBH-Z18226), Innovation Foundation of First (2019B18) and Second Affiliated Hospital (KYCX2019-10) of Harbin Medical University. Declaration of Interest: None to declare. Ethical Approval: The study was approved by the Ethics Committee of the First Affiliated Hospital of Harbin Medical University.

Promoting Effect of Basic Fibroblast Growth Factor in Synovial Mesenchymal Stem Cell-Based Cartilage Regeneration.

Synovial mesenchymal stem cell (SMSC) is the promising cell source of cartilage regeneration but has several issues to overcome such as limited cell proliferation and heterogeneity of cartilage regeneration ability. Previous reports demonstrated that basic fibroblast growth factor (bFGF) can promote proliferation and cartilage differentiation potential of MSCs in vitro, although no reports show its beneficial effect in vivo. The purpose of this study is to investigate the promoting effect of bFGF on cartilage regeneration using human SMSC in vivo. SMSCs were cultured with or without bFGF in a growth medium, and 2 × 105 cells were aggregated to form a synovial pellet. Synovial pellets were implanted into osteochondral defects induced in the femoral trochlea of severe combined immunodeficient mice, and histological evaluation was performed after eight weeks. The presence of implanted SMSCs was confirmed by the observation of human vimentin immunostaining-positive cells. Interestingly, broad lacunae structures and cartilage substrate stained by Safranin-O were observed only in the bFGF (+) group. The bFGF (+) group had significantly higher O’Driscoll scores in the cartilage repair than the bFGF (−) group. The addition of bFGF to SMSC growth culture may be a useful treatment option to promote cartilage regeneration in vivo.

Morphological and Molecular Features of Porcine Mesenchymal Stem Cells Derived From Different Types of Synovial Membrane, and Genetic Background of Cell Donors.

Synovial mesenchymal stem cells (SMSCs) have become a great cell source for musculoskeletal stem cell research, especially related to cartilage and bone tissue regeneration, due to their superior cell proliferation properties and multidifferentiation potential into various cell lineages. This study revealed isolation methods, culture conditions, and morphological and molecular characterization of SMSCs derived fibrous synovium (FS) and adipose synovium (FP) of two pig breeds differing in growth performance [German Landrace (DL), and fat deposition (Angeln Saddleback (AS)]. Herein, FS possessed nucleated cell numbers nearly twice as high as those of FP at Passage 0. SMSCs derived from different types of synovial membrane and genetic background show similar cell morphologies and immunophenotypes, which were assessed by cell surface epitopes and multilineage differentiation potential, but differ significantly in their molecular characteristics. In addition, transcripts of SMSCs from AS were more enriched in IGF-1 signaling and VEGF ligand receptor, while SMSCs from DL were more enriched in growth hormone signaling and bone metabolism. The results indicate that genetics and tissues play significant roles for SMSC characteristics so that SMSCs can be traced back to the original cell donor and be used for fine turning in applications of medical research and therapies.

Exosomes derived from miR-155-5p-overexpressing synovial mesenchymal stem cells prevent osteoarthritis via enhancing proliferation and migration, attenuating apoptosis, and modulating extracellular matrix secretion in chondrocytes.

Synovial mesenchymal stem cells (SMSCs) have the potential to attenuate osteoarthritis (OA)-induced injury. The role and mechanism of SMSC-derived exosomes (SMSC-Exos), pivotal paracrine factors of stem cells, in OA-associated injury remain unclear. We aimed to confirm the effect of SMSC-Exos with specific modifications on OA-induced damage and to investigate the potential molecular mechanisms. Exosomes derived from miR-155-5p-overexpressing SMSCs (SMSC-155-5p-Exos) and SMSCs (SMSC-Exos) were isolated and characterized. CCK-8, Transwell, and Western blot analyses were used to detect proliferation, migration, extracellular matrix (ECM) secretion, and apoptosis of osteoarthritic chondrocytes. The therapeutic effect of exosomes in a mouse model of OA was examined using immunohistochemical staining and OARSI scores. SPSS 17.0 and GraphPad software were used for all statistical analyses in this study. The SMSC-Exos enhanced the proliferation and migration and inhibited the apoptosis of osteoarthritic chondrocytes but had no effect on ECM secretion. The miR-155-5p-overexpressing exosomes showed common characteristics of exosomes in vitro and further promoted ECM secretion by targeting Runx2. Thus, the SMSC-155-5p-Exos promoted proliferation and migration, suppressed apoptosis and enhanced ECM secretion of osteoarthritic chondrocytes, and effectively prevented OA in a mouse model. In addition, overexpression of Runx2 partially reversed the effect of the SMSC-155-5p-Exos on osteoarthritic chondrocytes. Given the insufficient effect of the SMSC-Exos on the ECM secretion of osteoarthritic chondrocytes, we modified the SMSM-Exos and demonstrated that the SMSC-155-5p-Exos could prevent OA. Exosomes derived from modified SMSCs may be a new treatment strategy to prevent OA. Graphical abstract.

THE EFFECT OF SINGLE VERSUS MULTIPLE INTRA-ARTICULAR INJECTION OF SYNOVIAL FLUID MESENCHYMAL STEM CELLS ON RAT TEMPOROMANDIBULAR JOINT WITH INDUCED ARTHRITIS. BIOCHEMICAL AND HISTOLOGIC ANALYSIS

Introduction: Osteoarthritis (OA) is the most common chronic musculoskeletal disorder. Nowadays, the role of synovial fluid mesenchymal stem cells (SF MSCs) is increasingly attributed to the treatment of Temporomandibular joint osteoarthritis. This study aimed to compare single versus multiple intra-articular injection of SFMSCs in terms of macroscopic, biochemical, and histological analysis. Material & methods: 18 young adult male 200–300 grams albino rats were used in the current study divided randomly into 3 equal groups. Group (A) comprised control in which OA was induced with no treatment, group (B) comprised MSCs single intra-articular injection group, and group (C) comprised MSCs multiple intra-articular injection group. Biochemical and histologic analysis were performed. Results: histological findings showed that both groups B, C showed regeneration of meniscus and retrodiscal tissues, but group C was superior in regeneration compared to group B. Biochemical analysis showed that TNF-α and IL-1B had the highest value in the control group, followed by the single injection group, while the lowest value was found in multiple injections group respectively. Conclusion: Multiple Intra-articular SF MSCs is a potential disease-modifying therapy for patients suffering from TMJ-OA.

Mesenchymal Stem Cells in Synovial Fluid Increase in Knees with Degenerative Meniscus Injury after Arthroscopic Procedures through the Endogenous Effects of CGRP and HGF.

Mesenchymal stem cells (MSCs) in synovial fluid increase after traumatic meniscus injuries. However, MSC kinetics in synovial fluid may differ for knees with degenerative meniscus injuries. Furthermore, the combination of surgical repair and synovial MSC transplantation has been found to improve clinical symptoms in patients with degenerative meniscus injury, and in this treatment, only the operation procedure without MSC transplantation might increase MSCs in synovial fluid; if so, soluble factors in synovial fluid will be involved. The purpose is this study was to examine whether MSCs exist in synovial fluid of knees with degenerative meniscus injury, to investigate whether MSCs in synovial fluid increase after harvest of synovium and meniscus repair, and to explore what soluble factors in synovial fluids affect the number of MSCs in synovial fluid. Subjects were 7 patients with degenerative meniscus injury who underwent meniscal repair and synovial MSC transplantation. Synovial fluid (Pre) was aspirated from knees before harvest of synovium and meniscus repair. After 2weeks, synovial fluid (Post) was aspirated again before transplantation of synovial MSCs. A half volume of the synovial fluid was plated and cultured for 2weeks to count the colony formation. The other half was used for antibody array analysis, and the correlation coefficients between the signal intensity and colony number were measured in 503 factors. Factors with high correlation coefficients were verified by migration assay. While cell colonies derived from synovial fluid (Pre) were hardly observed, greater numbers of colonies from synovial fluid (Post) were demonstrated. Of the 503 factors, calcitonin gene-related peptide (CGRP) and hepatocyte growth factor (HGF) had high correlation coefficients between colony number and expression level. Both CGRP and HGF promoted migration of synovial fluid MSCs. MSCs in synovial fluid were hardly seen in knees with degenerated meniscus injury. They significantly increased 2weeks after harvest of synovium and meniscus repair. Both CGRP and HGF in synovial fluid can possibly induce MSCs from synovium into synovial fluid. Graphical abstract.

Synovial Mesenchymal Stem Cell-Derived EV-Packaged miR-31 Downregulates Histone Demethylase KDM2A to Prevent Knee Osteoarthritis

Extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) have emerged as important mediators of intercellular communication in response to cartilage damage. In this study, we sought to characterize the inhibitory role of microRNA (miR)-31 encapsulated in synovial MSC (SMSC)-derived EVs in knee osteoarthritis (OA). The expression of miR-31, lysine demethylase 2A (KDM2A), E2F transcription factor 1 (E2F1), and pituitary tumor transforming gene 1 (PTTG1) was validated in cartilage tissues of knee OA patients. Following SMSC-EV extraction and identification, chondrocytes with the miR-31 inhibitor were added with SMSC-EVs, whereupon the effects of miR-31 on proliferation and migration of chondrocytes were assessed. The interaction among miR-31, KDM2A, E2F1, and PTTG1 in chondrocyte activities was probed in vitro, along with an in vivo mouse knee OA model. We identified downregulated miR-31, E2F1, and PTTG1 and upregulated KDM2A in cartilage tissues of knee OA patients. SMSC-EV-packaged miR-31 potentiated chondrocyte proliferation and migration as well as cartilage formation by targeting KDM2A. Mechanistically, KDM2A bound to the transcription factor E2F1 and inhibited its transcriptional activity. Enrichment of E2F1 in the PTTG1 promoter region activated PTTG1 transcription, accelerating chondrocyte proliferation and migration. SMSC-EVs and EVs from miR-31-overexpressed SMSCs alleviated cartilage damage and inflammation in knee joints in vivo. SMSC-EV-encapsulated miR-31 ameliorates knee OA via the KDM2A/E2F1/PTTG1 axis.

Morphological changes in synovial mesenchymal stem cells during their adhesion to the meniscus

Synovial mesenchymal stem cells (MSCs) are an attractive cell source for transplantation because of their high chondrogenic potential, especially in areas like the meniscus of the knee. A synovial MSC suspension placed onto the meniscus for 10 min promoted healing of repaired meniscal tears that generally do not heal. Here, we quantified the proportion of human synovial MSCs that adhered to a porcine abraded meniscus, clarified their morphological changes, and revealed the mechanism by which the synovial MSCs adhered to the meniscus. The numbers of adhering cells at immediately after 10, 60 min and 6, 24 h after suspension placement were calculated. The meniscus surface was examined by scanning electron microscopy, and 50 cells were randomly selected at each time period, classified, and quantified for each of the six donors. Approximately 28% of the synovial MSCs immediately adhered to the meniscus after placement and the proportion of adhered cells increased further with time. All cells maintained a round shape for 60 min, and then transformed to a mixture of round and semi-flattened cells. By 24 h, flattened cells covered the meniscus. Microspikes were observed in 36% of the floating synovial MSCs and in 76% of the cells on the meniscus shortly after placement on the meniscus, then the proportion of cells with pseudopodia increased. The bleb-dominant cell proportion significantly decreased, and the smooth-dominant cell proportion increased within 60 min. Microspikes or the bodies of synovial MSCs were trapped by meniscal fibers immediately after placement. The proportion of adhered cells increased with time, and the cell morphology changed dynamically for 24 h as the synovial MSCs adhered to the meniscus. The MSCs in the round morphological state had a heterogeneous morphology. The microspikes, and the subsequent development of pseudopodia, may play an important role in adhesion onto the meniscus.

Evaluation of the Effect of Low Level Laser Therapy and Synovial Fluid-Mesenchymal Stem Cells in Cases of Temporomandibular Joint Disorders

Purpose: Temporomandibular joint disorders (TMD) might be the main cause of the non-dental orofacial pain. Varioustreatment modalities were introduced to treat that condition. Low level laser therapy (LLLT) had an anti-inflammatory andanalgesic effect by enhancing the circulation and the cellular respiration. Synovial fluid-mesenchymal stem cells (SF-MSCs)had a regenerative and immunoregulatory effect. Both methods were applied as a treatment for the condition; but with nocomparative studies in the literature. So, the current study aimed at evaluating their therapeutic effects.Material and methods: A total of 27 patients with TMD in 40 TMJs were randomly divided into two equal groups. In group I,LLLT was applied at four successive weeks. For group II, SF-MSCs were isolated, proliferated and pooled at Passage 4 to beinjected into the joint space at four successive weeks.Results: SF-MSCs therapy was superior to the LLLT with regards to the enhancement of the maximum mouth opening and thereduction of the inflammatory cytokines IL-1β and TNFα. With regards to interleukin-6, there was no statistically significantdifference. However, for pain reduction, LLLT was better than SF-MSCs; but not after the third week.Conclusion: Both treatment methods are accepted modalities; but SF-MSCs were more superior and had more sustainedresults.

Isolation, Characterization, Proliferation and Differentiation of Synovial Membrane-derived Mesenchymal Stem Cells (SM-MSCs) from Osteoarthritis Patients

Background: Mesenchymal stem cells (MSCs) are the cells which has high renewal capacity and and are capable for differentiating into some types of cells. MSCs can be obtained from several tissues including bone marrow, synovial membrane, blood, adipose tissue and periosteum. The proliferation and self-repair ability of MSCs are the advantages to use as stem cells-based therapy of various diseases. The aim of this study was to determine the differentiation, characterization and priliferation of synovial membrane-derived MSCs (SM-MSCs).Materials and Methods: The cells proliferation capacity was determined by cell counting using trypan blue, characterization of MSCs (cluster of differentiation (CD)90, CD11b, CD73, CD34, CD19, CD45, CD105 and human leukocyte antigen-DR isotype (HLA-DR)) using flow cytometry analysis, and differentiation capability into three lineage cells was determined with red alcian blue, oil red O and alizarin staining.Results: The type culture of SM-MSCs was adherent and showed positive CD44, CD105, CD73, CD90 and negative of CD19, HLA-DR, CD11b, CD45, CD34 surface marker. Based on the result, SM-MSCs P3 showed differentiation potency into adipogenic, chondrogenic, and osteogenic lineage cells. The population doubling time of SM-MSCs has increased from P3 to P8. The population doubling time of SM-MSCs P3 was 1.69 days and SM-MSCs P8 was 3.64 days.Conclusion: The results indicated that SM-MCSCs from osteoarthritis patients are able to differentiate into osteocytes, chondrocytes, adipocytes and highly express of CD105, CD73, CD90, CD44 and negative for CD34, CD45, CD14, CD19.Keywords: synovial membrane, mesenchymal stromal cells, adipocyte, chondrocyte, osteocyte