Disease-modifying potential of diphenyl diselenide in an experimental osteoarthritis model.

Oxidative stress is a key factor in the disruption of cartilage homeostasis during the development of osteoarthritis (OA). Organic selenium (Se)-containing compounds such as diselenides have excellent antioxidant activity and may prevent related diseases. We aimed to examine the benefits of the synthetic small molecule diphenyl diselenide (DPDSe) in OA models in vitro and in vivo. Our findings showed that DPDSe could maintain extracellular matrix (ECM) homeostasis and inhibit reactive oxygen species (ROS) production in IL-1ß-treated chondrocytes. In a destabilization of the medial meniscus (DMM)-induced OA mouse model, intra-articular administration of DPDSe alleviated joint degeneration, as evidenced by a decrease in the OARSI score and the restoration of collagen II (COL2) and MMP-13 expression in cartilage tissues. We confirmed that DDS activated the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in IL-1ß-treated chondrocytes, and its chondroprotective effects were significantly counteracted when Nrf2 signaling was blocked by the inhibitor ML385 or by siRNA-mediated Nrf2 knockdown. The relatively strong performance of DPDSe makes it an ideal candidate for further trials as a disease-modifying OA drug (DMOAD).

Urolithin B reduces cartilage degeneration and alleviates osteoarthritis by inhibiting inflammation.

Osteoarthritis is the most prevalent degenerative joint disease reported worldwide. Conventional treatment strategies mainly focus on medication and involve surgical joint replacement. The use of these therapies is limited by gastrointestinal complications and the lifespan of joint prostheses. Hence, safe and efficacious drugs are urgently needed to impede the osteoarthritis progression. Urolithin B, a metabolite of ellagic acid in the gut, exhibits anti-inflammatory and antioxidant properties; however, its role in osteoarthritis remains unclear. In this study, we demonstrated that urolithin B efficiently inhibits the inflammatory factor-induced production of matrix metalloproteinases (MMP3 and MMP13) in vitro and upregulates the expression of type II collagen and aggrecan. Urolithin B alleviates cartilage erosion and osteophyte formation induced by anterior cruciate ligament transections. Moreover, urolithin B inhibits the activation of the NF-κB pathway by reducing the phosphorylation of Iκb-α and the nuclear translocation of P65. In summary, urolithin B significantly inhibits inflammation and alleviates osteoarthritis. Hence, urolithin B can be considered a potential agent suitable for the effective treatment of osteoarthritis in the future.

Restrained Mitf-associated autophagy by Mulberroside A ameliorates osteoclastogenesis and counteracts OVX-Induced osteoporosis in mice.

Bone and mineral metabolism homeostasis accounts for the maintenance of normal skeletal remodeling. However, with aging and changes in hormone levels, over-activated osteoclasts disrupt homeostasis, induce osteoporosis, and even cause osteoporotic fractures, leading to an enormous economic burden. Despite the rapid development of pharmacological therapy for osteoporosis, safer and more effective treatments remain to be explored. Here, we demonstrate that Mulberroside A (Mul-A), a natural component extracted from mulberry bark and branches, effectively suppresses osteoclastogenesis in vitro and counteracts bone loss caused by ovariectomy (OVX). The mechanism underlying this effect involves the repression of autophagic flux during osteoclastogenesis by Mul-A, which can be attributed to the restrained expression of microphthalmia-related transcription factor (Mitf) and its nuclear translocation. Importantly, Mitf overexpression partially reverses the inhibitory effects of Mul-A on autophagy and osteoclastogenesis. Moreover, applying two autophagy agonizts, rapamycin and Torin 1, attenuates the osteoclastogenic regulatory role of Mul-A. Collectively, our study demonstrates that Mul-A damages osteoclast differentiation and ameliorates osteoporosis caused by estrogen deficiency by modulation of Mitf-associated autophagy, indicating its therapeutic potential against osteoporosis.

[Arthroscopic assistance of latissimus dorsi tendon transposition for the treatment of unrepairable rotator cuff tear].

OBJECTIVE: To explore clinical effect of arthroscopy-assisted rotator cuff tendon transfer in treating irreparable rotator cuff tears (IRCT). METHODS: From May 2015 to May 2018, 23 patients with unrepairable rotator cuff tears were treated with arthroscopy-assisted rotator cuff tendon transfer, and 21 patients were followed up finally, including 8 males and 13 females, aged from 48 to 82 years old with an average of(64.3±9.1) years old;the courses of disease ranged from 6 to 36 months with an average of (14.0±6.4) months. American Rotator and Elbow Surgeons Score(ASES) and Constant-Murley score were used to evaluate clinical efficacy before surgery and at the latest follow-up. RESULTS: All 21 patients were followed up for 36 to 54 months with an average of (39.4±4.4) months. Axillary incision of 1 patient was redness, swelling and exudation after surgery, which healed after 3 weeks of dressing change, and exudate culture was negative. At the latest follow-up, MRI showed partial tearing of the metastatic tendon in 2 patients, but pain and movement of the affected shoulder were still better than before surgery. ASES increased from preoperative (41.0±9.6) scores to the latest follow-up (75.6±14.0) scores, and had statistical difference (t=10.50, P<0.01). Constant-Murley score increased from (49.8±7.1) scores before operation to (67.5±11.6) scores at the latest follow-up (t=11.27, P<0.01). CONCLUSION: Arthroscopic assisted latissimus dorsalis tendon transposition restores physiological and anatomical structure of glenohumeral joint by reconstructing balance of horizontal and vertical couples of shoulder joint, thus achieving the stability of the shoulder joint, relieving shoulder pain and improving shoulder joint function.

Treatment of porcine ovarian follicles with tert-butyl hydroperoxide as an ovarian senescence model in vitro.

Ovarian aging is the main reason of female reproductive problems. Excessive oxidative stress can induce ovarian senescence and follicular atresia, thereby reducing the reproductive performance. Follicles were divided into five groups for in vitro culture based on the duration of stimulation with tert-butyl hydroperoxide (t-BHP)-control group and groups 1 h, 2 h, 6 h, and 12 h. The results revealed that the ratio of progesterone (P4) to estradiol (E2) was increased after 24 and 36 h of follicle culture, shifting follicles toward atresia (P < 0.05). Stimulated by 200 µM t-BHP, follicles showed progressive aging phenotype. Senescence-associated ß-galactosidase staining (SA-ß-Gal) showed a significant increase in the number of positive cells (P < 0.05). Reactive oxygen species were also significantly upregulated (P < 0.05). t-BHP treatment for 6 h induced significant increases in Caspase 3, P53, and Foxo1 mRNA and protein levels (P < 0.05) and significant decreases in SOD mRNA and protein levels (P < 0.05). Transcriptome sequencing analysis of the follicles showed that the aged and treatment groups were clustered together in hierarchical clustering. Correlation analysis indicated significant changes at the transcriptome level in the treatment groups versus the control group. The common differentially expressed genes in the treatment groups were enriched in three growth-factor signaling pathways associated with cell proliferation and apoptosis (P53, mTOR, and MAPK). In conclusion, induction of follicular senescence by treatment with 200 µM t-BHP for 6 h is an effective in vitro model to simulate ovarian senescence in sows.

Effect of high-fat diet on the lipid profile of ovarian granulosa cells and female reproduction in mice.

Currently, comorbidities of obesity are becoming increasingly frequent. For example, obese women are more susceptible to reproductive diseases; however, the underlying mechanism remains poorly understood. The present study aimed to explore the effect of obesity on female reproduction and discuss changes of the lipid profile in ovarian granulosa cells. Fifty female mice were randomly divided into two groups, one group was fed high-fat diet, the other group was fed standard control diet, food and water freely. After 12 weeks of feeding, the average body weight of the high-fat diet mice (19.027g) was significantly higher than that of the standard control diet mice (36.877g) (P < 0.05). The tissue sections were stained with oil red O, and the online software mage Pro plus 6.0 analyzed the staining results, the lipids in the ovaries and endometria were found to be different between the two groups. Liquid chromatography-electrospray ionization with tandem mass spectrometry (LC-ESI-MS/MS) analysis of ovarian granulosa cells (GCs) was performed, with a total of 228 different lipids being identified, the abundant of 147 were increased and 81 were decreased in the high-fat diet group. Among them, PI (18:1/20:1) was the most different lipid, and high-fat feeding was 85 times higher than standard control group. Among these different lipids, 44% in phospholipid metabolism, 30% in glycerolipid metabolism, and 30% in fat digestion and absorption. The results of this study laid a theoretical foundation of the effects of diet-induced obesity on female reproduction.

CircFNDC3B regulates osteoarthritis and oxidative stress by targeting miR-525-5p/HO-1 axis.

Osteoarthritis (OA) is a common chronic degenerative joint disease associated with a variety of risk factors including aging, genetics, obesity, and mechanical disturbance. This study aimed to elucidate the function of a newly discovered circular RNA (circRNA), circFNDC3B, in OA progression and its relationship with the NF-κB signaling pathway and oxidative stress. The circFNDC3B/miR-525-5p/HO-1 axis and its relationship with the NF-κB signaling pathway and oxidative stress were investigated and validated using fluorescence in situ hybridization, real-time PCR, western blotting, immunofluorescence analysis, luciferase reporter assays, pull-down assays, and reactive oxygen species analyses. The functions of circFNDC3B in OA was investigated in vitro and in vivo. These evaluations demonstrated that circFNDC3B promotes chondrocyte proliferation and protects the extracellular matrix (ECM) from degradation. We also revealed that circFNDC3B defends against oxidative stress in OA by regulating the circFNDC3B/miR-525-5p/HO-1 axis and the NF-κB signaling pathway. Further, we found that overexpression of circFNDC3B alleviated OA in a rabbit model. In summary, we identified a new circFNDC3B/miR-525-5p/HO-1 signaling pathway that may act to relieve OA by alleviating oxidative stress and regulating the NF-κB pathway, resulting in the protection of the ECM in human chondrocytes, highlighting it as a potential therapeutic target for the treatment of OA.

ERK1 loss accelerates the progression of osteoarthritis in aged mice via NRF2/BACH1 signaling.

Osteoarthritis (OA) is now a common degenerative joint related disease. However, the clinical efficacy of drugs associated with cartilage regeneration remains limited. In our study, we firstly explored the role of ERK1 in the progression of OA. We clarified that ERK1-deficient mice were susceptible to age-related OA. The higher OARSI scores and more severe cartilage degeneration was observed in the ERK1-deficient mice. ERK1 deficiency decreased the nuclear transportation of Nrf2 in the chondrocytes and accelerated chondrocyte aging in vitro. Moreover, chondrocytes with ERK1 deficiency elevated the nuclear expression of BACH1, resulting in lowered expression of antioxidant enzymes in ERK1-deficient chondrocytes. The Nrf2 activator dimethyl fumarate (DMF) was used. Our experiments demonstrated the protective function of DMF against OA in ERK1 knockout mice. Above all, we confirmed the effects of ERK1 on the progression of OA and clarified the mechanisms underlying these effects. DMF might has significant use in the development of novel drugs for the therapy of OA in the future.

USP1 inhibition suppresses the progression of osteosarcoma via destabilizing TAZ.

Mutations and altered expression of deubiquitinating enzymes (DUBs) profoundly influence tumor progression. Ubiquitin-specific protease 1 (USP1) is a well-characterized human DUB reportedly overexpressed in and associated with maintaining the mesenchymal stem cell status of osteosarcoma (OS); however, the potential mechanisms of USP1 in OS remain poorly understood. In this study, we identified that USP1 directly interacts with Transcriptional Co-Activator With PDZ-Binding Motif (TAZ) in OS cell lines, and with mechanistic analysis indicating that the anti-OS effects of USP1 inhibition could be partially attributed to TAZ instability, with its reduced nuclear accumulation responsible for a subsequent decrease in the expression of downstream genes associated with the Hippo signaling pathway. Moreover, pharmacological inhibition USP1 by ML323 presented the similar effects on Hippo signaling pathway and suppressed OS growth and metastasis both in vitro and in vivo. Taken together, our results revealed a novel molecular mechanism underlying the function of USP1 in OS and a potential role of ML323 as a therapeutic strategy for the clinical treatment of OS.

Progress on the study of nucleosome reorganization during cellular reprogramming.

Cellular reprogramming is the process during which epigenetic markers of nuclear genome are deleted and remodeled during sperm-egg binding or nuclear transplantation, thereby rendering differentiated cells totipotent. The main cellular reprogramming methods are cell fusion, somatic cell nuclear transplantation, and induced pluripotent stem cells. Nucleosomes are the basic structural and functional units of chromatin, and nucleosome localization has an important role in regulating gene expression and the state of the cell. The occupancy and location of nucleosomes also change dramatically during cellular reprogramming, while the occupancy of nucleosomes around the transcriptional start site also decreases to promote the expression of pluripotency genes. In this review, we summarize the role of nucleosome localization in gene activation and repression, chromatin remodeling, and transcription factor recognition, with the aim of providing an important basis for an in-depth analysis of cellular reprogramming mechanisms.

DUSP6 expression is associated with osteoporosis through the regulation of osteoclast differentiation via ERK2/Smad2 signaling.

Osteoporosis-related fractures, such as femoral neck and vertebral fractures, are common in aged people, resulting in increased disability rate and health-care costs. Thus, it is of great importance to clarify the mechanism of osteoclast-related osteoporosis and find effective ways to avoid its complication. In this study, gene expression profile analysis and real-time polymerase chain reaction revealed that DUSP6 expression was suppressed in human and mice osteoporosis cases. In vitro experiments confirmed that DUSP6 overexpression prevented osteoclastogenesis, whereas inhibition of DUSP6 by small interference RNA or with a chemical inhibitor, (E/Z)-BCI, had the opposite effect. (E/Z)-BCl significantly accelerated the bone loss process in vivo by enhancing osteoclastogenesis. Bioinformatics analyses and in vitro experiments indicated that miR-181a was an upstream regulator of DUSP6. Moreover, miR-181a positively induced the differentiation and negatively regulated the apoptosis of osteoclasts via DUSP6. Furthermore, downstream signals by ERK2 and SMAD2 were also found to be involved in this process. Evaluation of ERK2-deficiency bone marrow-derived macrophages confirmed the role of ERK2 signaling in the DUSP6-mediated osteoclastogenesis. Additionally, immunoprecipitation assays confirmed that DUSP6 directly modified the phosphorylation status of SMAD2 and the subsequent nuclear transportation of NFATC1 to regulate osteoclast differentiation. Altogether, this study demonstrated for the first time the role of miRNA-181a/DUSP6 in the progression of osteoporosis via the ERK2 and SMAD2 signaling pathway. Hence, DUSP6 may represent a novel target for the treatment of osteoclast-related diseases in the future.

circPDE4B prevents articular cartilage degeneration and promotes repair by acting as a scaffold for RIC8A and MID1.

OBJECTIVES: Circular RNAs (circRNAs) have emerged as significant biological regulators. Herein, we aimed to elucidate the role of an unidentified circRNA (circPDE4B) that is reportedly downregulated in osteoarthritis (OA) tissues. METHODS: The effects of circPDE4B were explored in human and mouse chondrocytes in vitro. Specifically, RNA pull-down (RPD)-mass spectrometry analysis (MS), immunoprecipitation, glutathione-S-transferase (GST) pull-down, RNA immunoprecipitation and RPD assays were performed to verify the interactions between circPDE4B and the RIC8 guanine nucleotide exchange factor A (RIC8A)/midline 1 (MID1) complex. A mouse model of OA was also employed to confirm the role of circPDE4B in OA pathogenesis in vivo. RESULTS: circPDE4B regulates chondrocyte cell viability and extracellular matrix metabolism. Mechanistically, FUS RNA binding protein (FUS) was found to promote the splicing of circPDE4B, while downregulation of circPDE4B in OA is partially caused by upstream inhibition of FUS. Moreover, circPDE4B facilitates the association between RIC8A and MID1 by acting as a scaffold to promote RIC8A degradation through proteasomal degradation. Furthermore, ubiquitination of RIC8A at K415 abrogates RIC8A degradation. The circPDE4B-RIC8A axis was observed to play an important role in regulating downstream p38 mitogen-activated protein kinase (MAPK) signalling. Furthermore, delivery of a circPDE4B adeno-associated virus (AAV) abrogates the breakdown of cartilage matrix by medial meniscus destabilisation in mice, whereas a RIC8A AAV induces the opposite effect. CONCLUSION: This work highlights the function of the circPDE4B-RIC8A axis in OA joints, as well as its regulation of MAPK-p38, suggesting this axis as a potential therapeutic target for OA.

ML264 inhibits osteosarcoma growth and metastasis via inhibition of JAK2/STAT3 and WNT/ß-catenin signalling pathways.

Osteosarcoma, the most common bone malignancy, has a high morbidity rate and poor prognosis. Krüppel-like factor 5 (KLF5) is a key transcriptional regulator of cellular proliferation whose overexpression is observed in osteosarcoma cell lines (U2OS, 143B, MG63 and SAOS2). ML264, a small-molecule inhibitor of KLF5, exerts antiproliferative effects in colorectal cancer; however, its function in osteosarcoma remains unknown. Here, we explored the possible antitumour effects of ML264 on 143B and U2OS cell lines and murine tumour xenograft model. ML264 suppressed proliferation and clonogenic ability of osteosarcoma cells in a dose-dependent manner. Moreover, ML264 induced G0/G1 cell cycle arrest, with no influence on apoptosis, and inhibited the migratory and invasive abilities of osteosarcoma cells, as demonstrated by wound-healing and Transwell assays. Exposure to ML264 reduced the mRNA and protein levels of molecules associated with epithelial-mesenchymal transition phenotype, including N-cadherin, vimentin, Snail, matrix metalloproteinase (MMP) 9 and MMP13. Inhibition of signal transducer and activator of transcription (STAT) 3 phosphorylation and Wnt signalling was also observed. In the murine model of osteosarcoma, tumour growth was efficiently suppressed following a 10-day treatment with ML264. Collectively, our findings demonstrate the potential value of ML264 as a novel anticancer drug for osteosarcoma.

The Nrf2 activator RTA-408 attenuates osteoclastogenesis by inhibiting STING dependent NF-κb signaling.

The dysregulation of ROS production and osteoclastogenesis is involved in the progress of osteoporosis. To identify novel and effective targets to treat this disease, it is important to explore the underlying mechanisms. In our study, we firstly tested the effect of the Nrf2 activator RTA-408, a novel synthetic triterpenoid under clinical investigation for many diseases, on osteoclastogenesis. We found that it could inhibit osteoclast differentiation and bone resorption in a time- and dose-dependent manner. Further, RTA-408 enhanced the expression and activity of Nrf2 and significantly suppressed RANKL-induced reactive oxygen species (ROS) production. Nrf2 regulates the STING expression and STING induces the production of IFN-ß. Here, we found that RTA-408 could suppress STING expression, but that it does not affect Ifnb1 expression. RANKL-induced degradation of IκBα and the nuclear translocation of P65 was suppressed by RTA-408. Although this compound was not found to influence STING-IFN-ß signaling, it suppressed the RANKL-induced K63-ubiquitination of STING via inhibiting the interaction between STING and the E3 ubiquitin ligase TRAF6. Further, adenovirus-mediated STING overexpression rescued the suppressive effect of RTA-408 on NF-κB signaling and osteoclastogenesis. In vivo experiments showed that this compound could effectively attenuate ovariectomy (OVX)-induced bone loss in C57BL/6 mice by inhibiting osteoclastogenesis. Collectively, we show that RTA-408 inhibits NF-κB signaling by suppressing the recruitment of TRAF6 to STING, in addition to attenuating osteoclastogenesis and OVX-induced bone loss in vivo, suggesting that it could be a promising candidate for treating osteoporosis in the future.

Octyl itaconate inhibits osteoclastogenesis by suppressing Hrd1 and activating Nrf2 signaling.

The endogenous metabolite itaconate has emerged as a regulator of macrophage function that limits inflammation. However, its effect on cell differentiation and osteoclast-related diseases is unclear. Here, for the first time, we explored the effect of itaconate and its cell-permeable itaconate derivative, 4-octyl itaconate (OI) on osteoclast differentiation in vitro and in vivo. Firstly, we demonstrated that itaconate concentration was lower in estrogen-deficient mice. OI released itaconate and induced the expression of nuclear factor-erythroid 2-related factor 2 (Nrf2) in bone marrow-derived macrophages during osteoclastogenesis. Furthermore, OI significantly suppressed the early, middle, and late stages of osteoclastogenesis induced by receptor activator of NF-κB ligand in vitro, as confirmed by tartrate-resistant acid phosphatase staining. Moreover, it significantly inhibited fibrous actin ring formation and bone resorption in vitro. Mechanistically, we observed that OI enhanced Nrf2 expression by suppressing its association with ubiquitin via inhibition of the E3 ubiquitin ligase (Hrd1). OI also inhibited LPS-induced the reactive oxygen species and inflammatory responses via Hrd1. An estrogen deficiency (via ovariectomy)-induced osteoporosis model was also established. Here, on micro-computed tomography and histologic analysis showed that OI effectively suppressed ovariectomy-induced bone loss. In summary, OI, an itaconate derivative, can inhibit osteoclastogenesis in vitro and in vivo, indicating that OI could be a potential drug to treat osteoclast-related diseases; our results also link itaconate to the development of osteoporosis.-Sun, X., Zhang, B., Pan, X., Huang, H., Xie, Z., Ma, Y., Hu, B., Wang, J., Chen, Z., Shi, P. Octyl itaconate inhibits osteoclastogenesis by suppressing Hrd1 and activating Nrf2 signaling.

Activation of the P38/CREB/MMP13 axis is associated with osteoarthritis.

PURPOSES: Osteoarthritis (OA) is a common joint disease characterized by the degradation of articular cartilage and joint inflammation. Interleukin-1ß induces P38/cAMP response element binding protein (CREB) pathway activation, resulting in increased expression of matrix metallopeptidase-13 (MMP13) in chondrocytes. However, the role of the P38/CREB/MMP13 axis is unclear in the progression of OA. In this study, we aimed to answer the following questions: (1) how does the P38/CREB/MMP13 axis in cartilage from patients with OA compare with control specimens? (2) Can the P38 agonist anisomycin (ANS) induce mouse OA? MATERIALS AND METHODS: Surgical specimens of human cartilage were divided into OA and control groups. Surgical specimens of mouse cartilage were divided into control and ANS-induced groups. Safranin O staining of the cartilage tissues was performed to evaluate the extracellular matrix. Reverse transcription-polymerase chain reaction was performed using these tissues to investigate messenger RNA expressions of type II collagen, aggrecan, MMP13, and ADAM metallopeptidase with thrombospondin type 1 motif 5. Phosphorylated (p)-P38, p-CREB, and MMP13 were evaluated by Western blot analysis. Anisomycin was used to activate P38, and p-P38, p-CREB, and MMP13 were evaluated by immunofluorescence and Western blot analysis. RESULTS: Safranin O staining showed that the extracellular matrix degraded in humans with OA and ANS-induced mouse cartilage samples. The expressions of p-P38, p-CREB, and MMP13 were all upregulated in osteoarthritic cartilage or anisomycin-induced chondrocytes, suggesting that the P38/CREB/MMP13 axis may play a role in the progression of OA. CONCLUSIONS: The P38/CREB/MMP13 axis is active in osteoarthritic chondrocytes and may cause the degeneration of cartilage. Effective new therapy directed against this pathway could be developed.

EGR1 promotes the cartilage degeneration and hypertrophy by activating the Krüppel-like factor 5 and ß-catenin signaling.

Osteoarthritis is one of the most common orthopedic diseases in elderly people who have lost their mobility. In this study，we observed abnormally high EGR1 expression in the articular cartilage of patients with osteoarthritis. We also found significantly high EGR1 expression in the articular cartilage of mice with destabilized medial meniscus (DMM)-induced osteoarthritis and 20-month-old mice. In vitro experiments indicated that IL-1ß could significantly enhance EGR1 expression in primary mouse chondrocytes. EGR1 over-expression in chondrocytes using adenovirus could inhibit COl2A1 expression and enhance MMP9 and MMP13 expression. And silencing EGR1, using RNAi, had the opposite effects. Moreover, EGR1 over-expression accelerated chondrocyte hypertrophy in vitro, and EGR1 knockdown reversed this effect. We then explored the underlying mechanism. EGR1 over-expression increased Kruppel-Like Factor 5 (KLF5) protein level without influencing its synthesis. Enhanced EGR1 expression induced its integration with KLF5, leading to suppressed ubiquitination of KLF5. Moreover, EGR1 prompted ß-catenin nuclear transportation to control chondrocyte hypertrophy. Ectopic expression of EGR1 in articular cartilage aggravated the degradation of the cartilage matrix in vivo. The EGR1 inhibitor, ML264, protected chondrocytes from IL-1ß-mediated cartilage matrix degradation in vitro and DMM-induced osteoarthritis in vivo. Above all, we demonstrate the effect and mechanisms of EGR1 on osteoarthritis and provide evidence that the ML264 might be a potential drug for treating osteoarthritis in the future.

CircSERPINE2 protects against osteoarthritis by targeting miR-1271 and ETS-related gene.

OBJECTIVES: Circular RNAs (circRNA) expression aberration has been identified in various human diseases. In this study, we investigated whether circRNAs could act as competing endogenous RNAs to regulate the pathological process of osteoarthritis (OA). METHODS: CircRNA deep sequencing was performed to the expression of circRNAs between OA and control cartilage tissues. The regulatory and functional role of CircSERPINE2 upregulation was examined in OA and was validated in vitro and in vivo, downstream target of CircSERPINE2 was explored. RNA pull down, a luciferase reporter assay, biotin-coupled microRNA capture and fluorescence in situ hybridisation were used to evaluate the interaction between CircSERPINE2 and miR-1271-5 p, as well as the target mRNA, E26 transformation-specific-related gene (ERG). The role and mechanism of CircSERPINE2 in OA was also explored in rabbit models. RESULTS: The decreased expression of CircSERPINE2 in the OA cartilage tissues was directly associated with excessive apoptosis and imbalance between anabolic and catabolic factors of extracellular matrix (ECM). Mechanistically, CircSERPINE2 acted as a sponge of miR-1271-5 p and functioned in human chondrocytes (HCs) through targeting miR-1271-5 p and ERG. Intra-articular injection of adeno-associated virus-CircSERPINE2-wt alleviated OA in the rabbit model. CONCLUSIONS: Our results reveal an important role for a novel circRNA-CircSERPINE2 in OA progression. CircSERPINE2 overexpression could alleviate HCs apoptosis and promote anabolism of ECM through miR-1271-ERG pathway. It provides a potentially effective therapeutic strategy for OA progression.

TGF-ß inhibits osteogenesis by upregulating the expression of ubiquitin ligase SMURF1 via MAPK-ERK signaling.

High incidence of osteoporotic fractures emphasizes the necessity of developing effective measures to promote osteogenesis. In our study, we investigated a possible role of MAPK-ERK signaling in the TGF-ß-mediated osteoblastic differentiation. Our results indicated that TGF-ß activated the MAPK-ERK pathway and inhibited osteogenesis in mesenchymal pluripotent cell line, C3H10T1/2, and preosteoblastic cell line, MC3T3 cells. And the downregulation of MAPK-ERK signaling using pharmacological inhibitor U0126 and RNA interference rescued osteoblast differentiation suppressed by TGF-ß, which was confirmed by Alkaline phosphatase (ALP) staining and alizarrn red staining, and the enhanced expression of osteogenesic markers. Western blotting analysis indicated that TGF-ß induced protein expression of E3 ubiquitin-protein ligase SMURF1, which contributed to the degradation of RUNX2 and SMAD1 as evidenced by SMURF1 inhibition using RNA interference and proteasome inhibitor MG132. Moreover, we observed that the expression of SMURF1 was decreased, while that of SMAD1 and RUNX2 increased by MAPK-ERK inhibitor U0126 in TGF-ß-treated differentiating preosteoblasts, suggesting that MAPK-ERK regulated the transcription of osteogenesis-related genes. Furthermore, a synergistic effect between U0126 and bone morphogenic protein (BMP)-2 on osteoblast differentiation and bone formation was observed both in cell cultures and experimental animals. In conclusion, our results revealed that TGF-ß inhibited osteoblastic differentiation by inducing the MAPK-ERK pathway which upregulated the expression of ubiquitin ligase SMURF1 and resulted in reduced presence of osteogenic proteins. In addition, the potentiation of BMP-2 on osteogenic activity by ERK1/2 inhibitor U0126 suggests that it may have potential clinical utility for promoting osteogenesis in bone fracture repair.