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Background WNT5A is known to be involved in the pathogenesis of osteoarthritis. This study investigated the molecular mechanism of exosomal miR-92a-3p and WNT5A in chondrogenesis and cartilage degeneration. Methods Exosomal miR-92a-3p expression was assessed in vitro in a human mesenchymal stem cell (MSC) model of chondrogenesis and in normal and OA primary human chondrocytes (PHCs). MSCs and PHCs were treated with exosomes derived from MSC-miR-92a-3p (MSC-miR-92a-3p-Exos) or its antisense inhibitor (MSC-anti-miR-92a-3p-Exos), respectively. Small interfering RNAs (siRNAs) and luciferase reporter assay were used to reveal the molecular role of exosomal miR-92a-3p and WNT5A in chondrogenesis. The protective effect of exosomes in vivo was measured using Safranin-O and Fast Green staining and immunohistochemical staining. Results Exosomal miR-92a-3p expression was elevated in the MSC chondrogenic exosome, while it was significantly reduced in the OA chondrocyte-secreted exosome compared with normal cartilage. Treatment with MSC-miR-92a-3p-Exos promoted cartilage proliferation and matrix genes expression in MSCs and PHCs, respectively. In contrast, treatment with MSC-anti-miR-92a-3p-Exos repressed chondrogenic differentiation and reduced cartilage matrix synthesis by enhancing the expression of WNT5A. Luciferase reporter assay demonstrated that miR-92a-3p suppressed the activity of a reporter construct containing the 3’-UTR and inhibited WNT5A expression in both MSCs and PHCs. MSC-miR-92a-3p-Exos inhibit cartilage degradation in the OA mice model. Conclusions Our results suggest that exosomal miR-92a-3p regulates cartilage development and homeostasis by directly targeting WNT5A. This indicates that exosomal miR-92a-3p may act as a Wnt inhibitor and exhibits potential as a disease-modifying osteoarthritis drug.

Objectives Aberrations in exosomal circular RNA (circRNA) expression have been identified in various human diseases. In this study, we investigated whether exosomal circRNAs could act as competing endogenous RNAs (ceRNAs) to regulate the pathological process of osteoarthritis (OA). This study aimed to elucidate the specific MSC-derived exosomal circRNAs responsible for MSC-mediated chondrogenic differentiation using human bone marrow-derived MSCs (hMSCs) and a destabilization of the medial meniscus (DMM) mouse model of OA. Methods Exosomal circRNA deep sequencing was performed to evaluate the expression of circRNAs in human bone marrow-derived MSCs (hMSCs) induced to undergo chondrogenesis from day 0 to day 21. The regulatory and functional roles of exosomal circRNA_0001236 were examined on day 21 after inducing chondrogenesis in hMSCs and were validated in vitro and in vivo. The downstream target of circRNA_0001236 was also explored in vitro and in vivo using bioinformatics analyses. A luciferase reporter assay was used to evaluate the interaction between circRNA_0001236 and miR-3677-3p as well as the target gene sex-determining region Y-box 9 ( Sox9 ). The function and mechanism of exosomal circRNA_0001236 in OA were explored in the DMM mouse model. Results Upregulation of exosomal circRNA_0001236 enhanced the expression of Col2a1 and Sox9 but inhibited that of MMP13 in hMSCs induced to undergo chondrogenesis. Moreover, circRNA_0001236 acted as an miR-3677-3p sponge and functioned in human chondrocytes via targeting miR-3677-3p and Sox9 . Intra-articular injection of exosomal circRNA_0001236 attenuated OA in the DMM mouse model. Conclusions Our results reveal an important role for a novel exosomal circRNA_0001236 in chondrogenic differentiation. Overexpression of exosomal circRNA_0001236 promoted cartilage-specific gene and protein expression through the miR-3677-3p/Sox9 axis. Thus, circRNA_0001236-overexpressing exosomes may alleviate cartilage degradation, suppressing OA progression and enhancing cartilage repair. Our findings provide a potentially effective therapeutic strategy for treating OA.

MicroRNAs play critical roles in the pathogenesis of osteoarthritis, the most common chronic degenerative joint disease. Exosomes derived from miR‐95‐5p‐overexpressing primary chondrocytes (AC‐miR‐95‐5p) may be effective in treating osteoarthritis. Increased expression of HDAC2/8 occurs in the tissues and chondrocyte‐secreted exosomes of patients with osteoarthritis and mediates cartilage‐specific gene expression in chondrocytes. We have been suggested that exosomes derived from AC‐miR‐95‐5p (AC‐miR‐95‐5p‐Exos) would enhance chondrogenesis and prevent the development of osteoarthritis by directly targeting HDAC2/8. Our in vitro experiments showed that miR‐95‐5p expression was significantly lower in osteoarthritic chondrocyte‐secreted exosomes than in normal cartilage. Treatment with AC‐miR‐95‐5p‐Exos promoted cartilage development and cartilage matrix expression in mesenchymal stem cells induced to undergo chondrogenesis and chondrocytes, respectively. In contrast, co‐culture with exosomes derived from chondrocytes transfected with an antisense inhibitor of miR‐95‐5p (AC‐anti‐miR‐95‐5p‐Exos) prevented chondrogenic differentiation and reduced cartilage matrix synthesis by enhancing the expression of HDAC2/8. MiR‐95‐5p suppressed the activity of reporter constructs containing the 3ʹ‐untranslated region of HDAC2/8, inhibited HDAC2/8 expression and promoted cartilage matrix expression. Our results suggest that AC‐miR‐95‐5p‐Exos regulate cartilage development and homoeostasis by directly targeting HDAC2/8. Thus, AC‐miR‐95‐5p‐Exos may act as an HDAC2/8 inhibitor and exhibit potential as a disease‐modifying osteoarthritis drug.

Objective The association between Metabolic Syndrome (MetS), its components, and the risk of osteoarthritis (OA) has been a topic of conflicting evidence in different studies. The aim of this present study is to investigate the association between MetS, its components, and the risk of OA using data from the UK Biobank. Methods A prospective cohort study was conducted in the UK Biobank to assess the risk of osteoarthritis (OA) related to MetS. MetS was defined according to the criteria set by the International Diabetes Federation (IDF). Additionally, lifestyle factors, medications, and the inflammatory marker C-reactive protein (CRP) were included in the model. Cox proportional hazards regression was used to calculate hazard ratios (HR) and 95% confidence intervals (CI). The cumulative risk of OA was analyzed using Kaplan–Meier curves and log-rank tests. To explore potential nonlinear associations between MetS components and OA risk, a restricted cubic splines (RCS) model was employed. In addition, the polygenic risk score (PRS) of OA was calculated to characterize individual genetic risk. Results A total of 45,581 cases of OA were identified among 370,311 participants, with a median follow-up time of 12.48 years. The study found that individuals with MetS had a 15% higher risk of developing OA (HR = 1.15, 95%CI:1.12–1.19). Additionally, central obesity was associated with a 58% increased risk of OA (HR = 1.58, 95%CI:1.5–1.66), while hyperglycemia was linked to a 13% higher risk (HR = 1.13, 95%CI:1.1–1.15). Dyslipidemia, specifically in triglycerides (HR = 1.07, 95%CI:1.05–1.09) and high-density lipoprotein (HR = 1.05, 95%CI:1.02–1.07), was also found to be slightly associated with OA risk. When stratified by PRS, those in the high PRS group had a significantly higher risk of OA compared to those with a low PRS, whereas no interaction was found between MetS and PRS on OA risks. Furthermore, the presence of MetS significantly increased the risk of OA by up to 35% in individuals with elevated CRP levels (HR = 1.35, 95% CI:1.3–1.4). Conclusion MetS and its components have been found to be associated with an increased risk of OA, particularly in individuals with elevated levels of CRP. These findings highlight the significance of managing MetS as a preventive and intervention measure for OA.

Background Bone marrow mesenchymal stem cells (BMMSCs) are commonly used for cell transplantation to treat refractory diseases. However, the presence of inflammatory factors, such as tumour necrosis factor-alpha (TNF-α), at the transplantation site severely compromises the stemness of BMMSCs, thereby reducing the therapeutic effect of cell transplantation. Aspirin (AS) is a drug that has been in use for over a century and has a wide range of effects, including the regulation of cell proliferation, multidirectional differentiation, and immunomodulatory properties of stem cells. However, it is still unclear whether AS can delay the damaging effects of TNF-α on BMMSC stemness. Methods This study investigated the effects of AS and TNF-α on BMMSC stemness and the molecular mechanisms using colony formation assay, western blot, qRT-PCR, and overexpression or knockdown of YAP and SMAD7. Results The results demonstrated that TNF-α inhibited cell proliferation, the expression of stemness, osteogenic and chondrogenic differentiation markers of BMMSCs. Treatment with AS was shown to mitigate the TNF-α-induced damage to BMMSC stemness. Mechanistic studies revealed that AS may reverse the damage caused by TNF-α on BMMSC stemness by upregulating YAP and inhibiting the expression of SMAD7. Conclusion AS can attenuate the damaging effects of TNF-α on BMMSC stemness by regulating the YAP-SMAD7 axis. These findings are expected to promote the application of AS to improve the efficacy of stem cell therapy.

Background Osteoarthritis (OA) is a highly prevalent human degenerative joint disorder that has long plagued patients. Glucocorticoid injection into the intra-articular (IA) cavity provides potential short-term analgesia and anti-inflammatory effects, but long-term IA injections cause loss of cartilage. Synovial mesenchymal stem cells (MSCs) reportedly promote cartilage proliferation and increase cartilage content. Methods CD90 + MCS-derived micro-vesicle (CD90@MV)-coated nanoparticle (CD90@NP) was developed. CD90 + MCSs were extracted from human synovial tissue. Cytochalasin B (CB) relaxed the interaction between the cytoskeleton and the cell membranes of the CD90 + MCSs, stimulating CD90@MV secretion. Poly (lactic-co-glycolic acid) (PLGA) nanoparticle was coated with CD90@MV, and a model glucocorticoid, triamcinolone acetonide (TA), was encapsulated in the CD90@NP (T-CD90@NP). The chondroprotective effect of T-CD90@NP was validated in rabbit and rat OA models. Results The CD90@MV membrane proteins were similar to that of CD90 + MCSs, indicating that CD90@MV bio-activity was similar to the cartilage proliferation-inducing CD90 + MCSs. CD90@NP binding to injured primary cartilage cells was significantly stronger than to erythrocyte membrane-coated nanoparticles (RNP). In the rabbit OA model, the long-term IA treatment with T-CD90@NP showed significantly enhanced repair of damaged cartilage compared to TA and CD90 + MCS treatments. In the rat OA model, the short-term IA treatment with T-CD90@NP showed effective anti-inflammatory ability similar to that of TA treatment. Moreover, the long-term IA treatment with T-CD90@NP induced cartilage to restart the cell cycle and reduced cartilage apoptosis. T-CD90@NP promoted the regeneration of chondrocytes, reduced apoptosis via the FOXO pathway, and influenced type 2 macrophage polarization to regulate inflammation through IL-10. Conclusion This study confirmed that T-CD90@NP promoted chondrocyte proliferation and anti-inflammation, improving the effects of a clinical glucocorticoid treatment plan. Graphical Abstract Highlights Long-term injection of glucocorticoids in the knee joint cavity promotes loss of cartilage content. CD90-positive stem cell vesicles encapsulated with Triamcinolone acetonide-loaded nanoparticles have good materials. T-CD90@NPs drive anti-inflammatory properties and promote cartilage regeneration after osteoarthritis. T-CD90@NPs regulate the polarization of type 2 macrophages to resist inflammation. T-CD90@NPs promote chondrocyte regeneration through the FOXO signaling pathway.

MicroRNAs (miRNAs, miR) play a key role in the pathogenesis of osteoarthritis (OA). Few studies have examined the regulatory role of P21-activated kinases (PAKs), a family of serine/threonine kinases, in OA. The aim of this study was to determine whether miR-455-3p can regulate cartilage degeneration in OA by targeting PAK2. MiR-455-3p knockout mice showed significant degeneration of the knee cartilage. MiR-455-3p expression increased and PAK2 expression decreased in the late stage of human adipose-derived stem cell (hADSC) chondrogenesis and in chondrocytes affected by OA. Furthermore, in both miR-455-3p-overexpressing chondrocytes and PAK2-suppressing chondrocytes, cartilage-specific genes were upregulated, and hypertrophy-related genes were downregulated. A luciferase reporter assay confirmed that miR-455-3p regulates PAK2 expression by directly targeting the 3′-untranslated regions (3′UTRs) of PAK2 mRNA. IPA-3, a PAK inhibitor, inhibited cartilage degeneration due to OA. Moreover, suppressing PAK2 promoted R-Smad activation in the TGF/Smad signaling pathway in chondrocytes. Altogether, our results suggest that miR-455-3p promotes TGF-β/Smad signaling in chondrocytes and inhibits cartilage degeneration by directly suppressing PAK2. These results thus indicate that miR-455-3p and PAK2 are novel potential therapeutic agents and targets, respectively, for the treatment of OA. Arthritis: A molecular mechanism for cartilage maintenance Functional insights into a short RNA strand that prevents cartilage degeneration could lead to new therapeutic strategies for treating osteoarthritis. The microRNA miR-455-3p regulates genes in tissues throughout the body, but Weiming Liao and Zhiqi Zhang of the First Affiliated Hospital of Sun Yat-sen University in Guangzhou, China have observed that it appears to play a particularly prominent role in cartilage-producing chondrocyte cells. Liao, Zhang and colleagues have now explored this mechanism in detail, and determined that miR-455-3p selectively blocks the effects of a protein called PAK2 in chondrocytes. PAK2 normally inhibits an important signaling pathway underlying cartilage generation, and such inhibition has previously been observed in MDCK epithelial cells. The authors conclude that this microRNA or other drugs that replicate its PAK2-inhibiting effects could prevent or slow the joint damage associated with this degenerative disorder.

Mitochondrial dysfunction is related to the pathogenesis of osteoarthritis (OA); however, there are no effective drugs to treat OA for maintaining mitochondrial homeostasis. Studies have shown that mitochonic acid-5 (MA-5) has a protective effect against mitochondrial damage and plays a role in mitophagy. However, it is not clear whether MA-5 has a beneficial effect on inflammatory articular cartilage. Here, human OA cartilage was obtained from patients undergoing total joint replacement. Interleukin-1β (IL-1β) was used to stimulate chondrocytes and induce inflammatory injury. Cell Counting Kit-8, TUNEL, and flow cytometry assays were used to assess apoptosis. Gene expression was examined using quantitative reverse transcription-polymerase chain reaction. Mitochondrial function was evaluated using immunoblotting, mitochondrial membrane potential assay, JC-1 staining, and immunofluorescence analysis. Mitophagy was detected using immunoblotting and immunofluorescence. 3-(1H-1,2,3-triazol-4-yl) pyridine (3-TYP), a specific inhibitor of Sirtuin 3 (SIRT3), was used to block the SIRT3/Parkin pathway. Mitophagy in the cartilage sections was evaluated via immunohistochemistry. IL-1β was found to induce chondrocyte apoptosis by inhibiting SIRT3 expression and mitophagy. In addition, inflammatory damage reduced the mitochondrial membrane potential and promoted the production of intracellular reactive oxygen species (ROS), leading to increased mitochondrial division, mitochondrial fusion inhibition, and the consequent mitochondrial damage. In contrast, the MA-5 treatment inhibited excessive ROS production by upregulating mitophagy, maintaining the mitochondrial membrane potential, and reducing mitochondrial apoptosis. After chemically blocking SIRT3 with 3-TYP, Parkin-related mitophagy was also inhibited, an effect that was prevented by pretreatment of the chondrocytes with MA-5, thereby suggesting that SIRT3 is upstream of Parkin. Overall, MA-5 was found to enhance the activity of SIRT3, promote Parkin-dependent mitophagy, eliminate depolarized/damaged mitochondria in chondrocytes, and protect cartilage cells. In conclusion, MA-5 inhibits IL-1β-induced oxidative stress and protects chondrocytes by upregulating the SIRT3/Parkin-related autophagy signaling pathway.

The aim of this work was to determine whether miR-455-3p regulates DNA methylation during chondrogenic differentiation of hMSCs. The expression of miR-455-3p and de novo methyltransferase DNMT3A was assessed in micromass culture of hBMSCs, which induced chondrogenic differentiation in vitro, and in E16.5 mice in vivo. A luciferase reporter assay was used to confirm whether miR-455-3p directly targets DNMT3A by interaction with the 3′-UTR. Using an Illumina Infinium Methylation EPIC microarray, genome-wide DNA methylation of hBMSCs with or without overexpressed miR-455-3p was examined for 28 days during induced chondrogenic differentiation. Here, we showed that miR-455-3p was more expressed during the middle stage of hBMSC chondrogenic differentiation, and less expressed in the late stage. DNMT3A was less expressed in the middle stage and more expressed in the late stage, and was also more expressed in the palms of miR-455-3p deletion mice compared to those of wild-type mice. The luciferase reporter assay demonstrated that miR-455-3p directly targets DNMT3A 3′-UTR. miR-455-3p overexpression inhibits the degenerate process during chondrogenic differentiation, while deletion of miR-455-3p in mice accelerated cartilage degeneration. Genome-wide DNA methylation analysis showed miR-455-3p overexpression regulates DNA methylation of cartilage-specific genes. GO analysis revealed PI3K-Akt signaling pathway was most hypomethylated. Our data show that miR-455-3p can regulate hMSC chondrogenic differentiation by affecting DNA methylation. Overexpression of miR-455-3p and DNA methylation inhibitors can thus potentially be utilized to optimize chondrogenic differentiation.

Platelet adhesion to blood vessel walls in shear flow is essential to initiating the blood coagulation cascade and prompting clot formation in vascular disease processes and prosthetic cardiovascular devices. Validation of predictive adhesion kinematics models at the single platelet level is difficult due to gaps in high resolution, dynamic morphological data or a mismatch between simulation and experimental parameters. Gel-filtered platelets were perfused at 30 dyne/cm2 in von Willebrand Factor (vWF)-coated microchannels, with flipping platelets imaged at high spatial and temporal resolution. A semi-unsupervised learning system (SULS), consisting of a series of convolutional neural networks, was used to segment platelet geometry, which was compared with expert-analyzed images. Resulting time-dependent rotational angles were smoothed with wavelet-denoising and shifting techniques to characterize the rotational period and quantify flipping kinematics. We observed that flipping platelets do not follow the previously-modeled modified Jefferey orbit, but are characterized by a longer lift-off and shorter reattachment period. At the juncture of the two periods, rotational velocity approached 257.48 ± 13.31 rad/s. Our SULS approach accurately segmented large numbers of moving platelet images to identify distinct adhesive kinematic characteristics which may further validate the physical accuracy of individual platelet motion in multiscale models of shear-mediated thrombosis.