Chitosan oligosaccharide suppresses osteosarcoma malignancy by inhibiting CEMIP via the PI3K/AKT/mTOR pathway.

Osteosarcoma is a malignant bone tumor that is prone to metastasize early and primarily affects children and adolescents. Cell migration-inducing protein (CEMIP) plays a crucial role in the progression and malignancy of various tumor diseases, including osteosarcoma. Chitosan oligosaccharide (COS), an oligomer isolated from chitin, has been found to have significant anti-tumor activity in various cancers. This study investigates the effects of COS on CEMIP expression in osteosarcoma and explores the underlying mechanism. In present study, in vitro experiments were conducted to confirm the inhibitory activity of COS on human osteosarcoma cells. Our results demonstrate that COS possesses inhibitory effects against human osteosarcoma cells and significantly suppresses CEMIP expression in vitro. Next, we studied the inhibition of the expression of CEMIP by COS and then performed bioinformatics analysis to explore the potential inhibitory mechanism of COS against signaling pathways involved in regulating CEMIP expression. Bioinformatics analysis predicted a close association between the PI3K signaling pathway and CEMIP expression and that the inhibitory effect of COS on CEMIP expression may be related to PI3K signaling pathway regulation. The results of this study show that COS treatment significantly inhibits CEMIP expression and the PI3K/AKT/mTOR signaling pathway, as observed both in vitro and in vivo. This study demonstrates that COS could inhibit the expression of CEMIP, which is closely related to osteosarcoma malignancy. This inhibitory effect may be attributed to the inhibition of the PI3K/AKT/mTOR signaling pathway in vitro and in vivo.

Nesfatin-1 suppresses autophagy of chondrocytes in osteoarthritis via remodeling of cytoskeleton and inhibiting RhoA/ROCK signal pathway.

Autophagy and cytoskeleton integrity of chondrocytes are a considered as major factors in the progression of osteoarthritis (OA) involving excessive chondrocyte apoptosis and senescence. Nesfatin-1, an adipokine, has been reported to be closely related to cell autophagy and cytoskeleton malfunction. Our previous study found that nesfatin-1 was highly correlated with OA progress in OA patient, and the expression of nesfatin-1 rises in knee articular tissue, serum and chondrocytes. In current study, we aimed to explore the therapeutic effect of nesfatin-1 on OA and its molecular mechanism related to chondrocyte autophagy and cytoskeleton malfunction. We firstly demonstrated that nesfatin-1 effectively suppressed excessive autophagy of OA chondrocytes at both gene and protein levels. Meanwhile, we also found that nesfatin-1 significantly improved cytoskeleton integrity by showing higher F-actin/G-actin ratio, as well as more organized actin fiber structure. Mechanistically, utility of RhoA activator and inhibitor revealed that regulation of autophagy and cytoskeleton integrity via nesfatin-1 was realized via RhoA/ROCK pathway. We also confirmed that nesfatin-1 significantly ameliorated IL-1ß induced cartilage degeneration via destabilization of the medial meniscus (DMM) model. Overall, our study indicates that nesfatin-1 might be a promising therapeutic molecule for OA intervention.

Nitidine Chloride Alleviates Inflammation and Cellular Senescence in Murine Osteoarthritis Through Scavenging ROS.

Osteoarthritis (OA) is one of the most common chronic musculoskeletal disorder worldwide, representing a major source of disability, pain and socioeconomic burden. Yet the effective pharmaceutical treatments applied in the clinical works are merely symptomatic management with uncertainty around their long-term safety and efficacy, namely no drugs currently are capable of modulating the biological progression of OA. Here, we identified the potent anti-inflammatory as well as anti-oxidative properties of Nitidine Chloride (NitC), a bioactive phytochemical alkaloid extracted from natural herbs, in IL-1ß-treated rat articular chondrocytes (RACs), LPS-stimulated RAW 264.7 and rat osteoarthritic models in vivo. We demonstrated NitC remarkably inhibited the production of inflammatory mediators including COX2 and iNOS, suppressed the activation of MAPK and NF-κB cell signaling pathway and reduced the expression of extracellular matrix (ECM) degrading enzymes including MMP3, MMP9 and MMP13 in IL-1ß-treated RACs. Several emerging bioinformatics tools were performed to predict the underlying mechanism, the result of which indicated the potential reactive oxygen species (ROS) clearance potential of NitC. Further, NitC exhibited its anti-oxidative potential through ameliorating cellular senescence in IL-1ß-treated RACs and decreasing NLRP3 inflammasomes activation in LPS-stimulated RAW 264.7 via scavenging ROS. Additionally, X-ray, micro-CT and other experiments in vivo demonstrated that intra-articular injection of NitC significantly alleviated the cartilage erosion, ECM degradation and subchondral alterations in OA progression. In conclusion, the present study reported the potent anti-inflammatory and anti-oxidative potential of NitC in OA biological process, providing a promising therapeutic agent for OA management.

The effect of mitochondrial fusion on chondrogenic differentiation of cartilage progenitor/stem cells via Notch2 signal pathway.

BACKGROUND: Osteoarthritis (OA) is a debilitating disease that inflicts intractable pain, a major problem that humanity faces, especially in aging populations. Stem cells have been used in the treatment of many chronic diseases, including OA. Cartilage progenitor/stem cells (CPSCs) are a type of stem cells with the ability to self- renew and differentiate. They hold a promising future for the understanding of the progression of OA and for its treatment. Previous studies have reported the relationship between mitochondrial dynamics and mesenchymal stem cell (MSC) proliferation, differentiation and aging. Mitochondrial dynamic and morphology change during stem cell differentiation. METHODS: This study was performed to access the relationship between mitochondrial dynamics and chondrogenic differentiation of CPSCs. Mitochondrial fusion and fission levels were measured during the chondrogenic differentiation process of CPSCs. After that, we used mitochondrial fusion promoter to induce fusion in CPSCs and then the chondrogenic markers were measured. Transmission electron microscopy (TEM) and confocal microscopy were used to capture the mass and fusion status of mitochondria. Lentiviruses were used to detect the role of mitofusin 2 (Mfn2) in CPSC chondrogenic differentiation. In vivo, Mfn2 was over-expressed in sheets of rat CPSCs, which were then injected intra-articularly into the knees of rats. RESULTS: Mitochondrial fusion markers were upregulated during the chondrogenic induction process of CPSCs. The mass of mitochondria was higher in differentiated CPSC, and the fusion status was obvious relative to un-differentiated CPSC. Chondrogenesis of CPSCs was upregulated with the induction by mitochondrial fusion promoter. Mfn2 over-expression significantly increased chondrocyte-specific gene expression and reversed OA through NOTCH2 signal pathway. CONCLUSIONS: Our study demonstrated that the mitochondrial fusion promotes chondrogenesis differentiation of CPSCs. Mfn2 accelerates the chondrogenesis differentiation of CPSCs through Notch2. In vivo, Mfn2-OE in sheets of rCPSCs ameliorated OA in the rat model.

circFAM160A2 Promotes Mitochondrial Stabilization and Apoptosis Reduction in Osteoarthritis Chondrocytes by Targeting miR-505-3p and SIRT3.

Competitive endogenous RNAs (ceRNAs), as a newly identified regulating mechanism, have been demonstrated to play a crucial role in various human diseases. An increasing number of recent studies have revealed that circular RNAs (circRNAs) can function as ceRNAs. However, little is known about the role of circFAM160A2 in the pathological process of osteoarthritis (OA). This study is the first to examine the crucial role of the circFAM160A2-miR-505-3p-SIRT3 axis in osteoarthritis progression. miR-505-3p was selected from the interaction of a microRNA (miRNA) microarray comparing chondrocytes in OA and normal conditions and prediction results from TargetScan. RT-qPCR was performed to assess the expression of circFAM160A2, miR-505-3p, and SIRT3. A dual luciferase assay was used to validate the binding of circFAM160A2, miR-505-3p, and SIRT3. We used lentivirus and adeno-associated virus to establish in vitro and in vivo overexpression models. Western blotting, apoptosis assay, ROS detection assay, Safranin O staining, and CCK-8 assay were employed to assess the role of circFAM160A2, miR-505-3p, and SIRT3. We found that miR-505-3p was upregulated and circFAM160A2 was downregulated in OA. While overexpression of circFAM160A2 decreased the production of extracellular matrix (ECM) degrading enzymes and ameliorated chondrocyte apoptosis and mitochondrial dysfunction, inhibition of miR-505-3p could reverse the protective effect of circFAM160A2 on the OA phenotype both in vitro and in vivo. In conclusion, circFAM160A2 can promote mitochondrial stabilization and apoptosis reduction in OA chondrocytes by targeting miR-505-3p and SIRT3, which might be a potential therapeutic target for OA therapy.

Spironolactone Ameliorates Senescence and Calcification by Modulating Autophagy in Rat Tendon-Derived Stem Cells via the NF-κB/MAPK Pathway.

Tendinopathy is a disabling musculoskeletal disease, the pathological process of which is tightly associated with inflammation. Spironolactone (SP) has been widely used as a diuretic in clinical practice. Recently, SP has shown anti-inflammatory features in several diseases. Tendon-derived stem cells (TDSCs), a subset cell type from tendon tissue possessing clonogenic capacity, play a vital role in the pathological process of tendinopathy. In the present study, the protective effect of SP on TDSCs was demonstrated under simulated tendinopathy conditions both in vitro and in vivo. SP contributed to the maintenance of TDSC-specific genes or proteins, while suppressing the interleukin- (IL-) 1ß-induced overexpression of inflammation-mediated factors. Additionally, IL-1ß-induced cellular senescence in TDSCs was inhibited, while autophagy was enhanced, as determined via ß-galactosidase activity, western blot (WB), and quantitative real-time polymerase chain reaction analysis. With the aid of several emerging bioinformatics tools, the mitogen-activated protein kinase (MAPK) pathway likely participated in the effect of SP, which was further validated through WB analysis and the use of MAPK agonist. Immunofluorescence analysis and an NF-κB agonist were used to confirm the inhibitory effect of SP on IL-1ß-induced activation of the NF-κB pathway. X-ray, immunofluorescence, immunohistochemistry, hematoxylin and eosin staining, histological grades, and Masson staining showed that SP ameliorated tendinopathy in an Achilles tenotomy (AT) rat model in vivo. This work elucidates the protective role of SP on the pathological process of tendinopathy both in vitro and in vivo, indicating a potential therapeutic strategy for tendinopathy treatment.

SIRT3 ameliorates osteoarthritis via regulating chondrocyte autophagy and apoptosis through the PI3K/Akt/mTOR pathway.

Osteoarthritis (OA) is the most common form of joint disease. The aim of this study was to explore the functions of SIRT3 on OA pathophysiology and the mechanism involved. Rat chondrocytes and destabilized medial meniscus (DMM) rat OA model were used as in vitro and in vivo models. In addition, lentivirus and plasmid were used to overexpress SIRT3, while siRNA was applied to establish SIRT3 knockdown. IL-1ß induced inflammation, apoptosis, mitochondrial dysfunction, and chondrocyte degeneration were inhibited by SIRT3 overexpression, which were enhanced in SIRT3-knockdown rat chondrocytes. Furthermore, overexpression of SIRT3 could restore IL-1ß-induced autophagy inhibition. We also found that IL-1ß-induced PI3K/Akt/mTOR signaling pathway activation was inhibited by SIRT3 overexpression, which was enhanced by SIRT3 knockdown. Last, intra-articular SIRT3 overexpression alleviated the severity of OA-induced rat joint damage. Our results demonstrated that SIRT3 is an important protective agent against OA pathophysiology via inhibiting PI3K/Akt/mTOR signaling.

Nesfatin-1 Promotes the Osteogenic Differentiation of Tendon-Derived Stem Cells and the Pathogenesis of Heterotopic Ossification in Rat Tendons via the mTOR Pathway.

Heterotopic ossification (HO) is a pathological condition involved in tendinopathy. Adipokines are known to play a key role in HO of tendinopathy. Nesfatin-1, an 82-amino acid adipokine is closely reportedly associated with diabetes mellitus (DM), which, in turn, is closely related to tendinopathy. In the present study, we aimed to investigate the effects of nesfatin-1 on the osteogenic differentiation of tendon-derived stem cells (TDSCs) and the pathogenesis of tendinopathy in rats. In vitro, TDSCs were incubated in osteogenic induction medium for 14 days with different nesfatin-1 concentration. In vivo, Sprague Dawley rats underwent Achilles tenotomy to evaluate the effect of nesfatin-1 on tendinopathy. Our results showed that the expression of nesfatin-1 expression in tendinopathy patients was significantly higher than that in healthy subjects. Nesfatin-1 affected the cytoskeleton and reduced the migration ability of TDSCs in vitro. Furthermore, nesfatin-1 inhibited the expression of Scx, Mkx, and Tnmd and promoted the expression of osteogenic genes, such as COL1a1, ALP, and RUNX2; these results suggested that nesfatin-1 inhibits cell migration, adversely impacts tendon phenotype, promotes osteogenic differentiation of TDSCs and the pathogenesis of HO in rat tendons. Moreover, we observed that nesfatin-1 suppressed autophagy and activated the mammalian target of rapamycin (mTOR) pathway both in vitro and in vivo. The suppression of the mTOR pathway alleviated nesfatin-1-induced HO development in rat tendons. Thus, nesfatin-1 promotes the osteogenic differentiation of TDSC and the pathogenesis of HO in rat tendons via the mTOR pathway; these findings highlight a new potential therapeutic target for tendinopathy.

DUSP5 suppresses interleukin-1ß-induced chondrocyte inflammation and ameliorates osteoarthritis in rats.

Osteoarthritis (OA) is a chronic degenerative joint disease characterized by deterioration of articular cartilage. Dual specificity phosphatase 5 (DUSP5), a member of the DUSP subfamily, is known to regulate cellular inflammation. Here, we studied the relationship between DUSP5 and OA by knockdown and overexpression DUSP5, respectively. Results from in vitro experiments demonstrated that the knockdown of DUSP5 increased interleukin-1ß (IL-1ß)-induced expression of inflammatory genes, such as inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2), and matrix metalloproteinases (MMPs) in chondrocytes, whereas it decreased the expression of anti-inflammatory genes, such as tissue inhibitor of metalloproteinase 3 (TIMP3) and IL-10. Conversely, the overexpression of DUSP5 suppressed the IL-1ß-induced expression of iNOS, COX-2, and MMPs, and upregulated the expression of TIMP3 and IL-10. Moreover, knockdown of DUSP5 enhanced the IL-1ß-induced activation of NF-κB and ERK pathways, whereas its overexpression inhibited these pathways. DUSP5 overexpression prevented cartilage degeneration in a rat OA model, while its knockdown reversed that effect. Our findings reveal that DUSP5 suppresses IL-1ß-induced chondrocyte inflammation by inhibiting the NF-κB and ERK signaling pathways and ameliorates OA.

Tectorigenin Alleviates Inflammation, Apoptosis, and Ossification in Rat Tendon-Derived Stem Cells via Modulating NF-Kappa B and MAPK Pathways.

Tendinopathy is a common musculoskeletal disorder that mainly affects athletes and people of older age. Tumor necrosis factor-α (TNF-α) plays an important role in initiating tendinopathy. Tectorigenin, an extract component of Belam-canda Chinesis, possesses anti-inflammatory and anti-apoptosis activity. The present study was established to investigate the role of tectorigenin against the pathogenetic effects of TNF-α on tendon-derived stem cells (TDSCs) in vivo and in vitro. The findings indicated that TNF-α is able to induce TDSC inflammation, apoptosis, and ossification, as well as activate nuclear factor-kappa B and mitogen-activated protein kinase (MAPK). Furthermore, the results confirmed that tectorigenin is able to inhibit the TNF-α-induced inflammation, apoptosis, and ossification. Tectorigenin treatment decreases activation of NF-kappa B and MAPK signaling in TDSCs. Tectorigenin ameliorates tendinopathy in the in vivo rat model. Thus, these data reveal that tectorigenin can serve as a potential treatment for tendinopathy.

Rat Chondrocyte Inflammation and Osteoarthritis Are Ameliorated by Madecassoside.

As a joint disease, osteoarthritis (OA) is caused by the breakdown of subchondral bone and cartilage damage. Inflammatory factors, such as interleukin- (IL-) 1ß, mediate the progression of OA. Madecassoside (MA), a triterpenoid component derived from the gotu kola herb (Centella asiatica), exhibits various pharmacological effects, including antioxidative and anti-inflammatory properties. In the present study, the protective effects and possible mechanism of MA on the treatment of OA were investigated. MA was demonstrated to significantly suppress the IL-1ß-induced overexpression of matrix metalloproteinase- (MMP-) 3, MMP-13, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) and to decrease the IL-1ß-induced degradation of type II collagen and sox9. Additionally, MA was able to reduce the IL-1ß-induced phosphorylation of p65 in osteoarthritic chondrocytes. Furthermore, in a rat OA model, MA prevented cartilage degeneration and reduced the OARSI score in the MA-treated group compared with the OA group. The present study showed that MA suppresses the nuclear factor-κB signaling pathway, reducing IL-1ß-induced chondrocyte inflammation, which indicates the therapeutic potential of MA in patients with OA.

Laser Acupuncture for Patients with Knee Osteoarthritis: A Systematic Review and Meta-Analysis of Randomized Placebo-Controlled Trials.

OBJECTIVES: To provide updated evidence from randomized controlled trials (RCTs) on the effectiveness of laser acupuncture for patients with knee osteoarthritis (KOA). METHODS: A literature search in 9 databases was conducted from their inception through February 2019. Randomized controlled trials (RCTs) written in English that compared active laser acupuncture with placebo in KOA patients were included. Two authors independently extracted data from these trials. Meta-analysis software was used to analyze the data. Included studies were assessed in terms of the follow-up period, the methodological quality, and appropriateness of their technical features. RESULTS: Of 357 studies, seven RCTs (totaling 395 patients) met the inclusion criteria. The short-term outcomes showed that laser acupuncture offered significant pain relief over placebo when assessed by the 100 mm visual analog scale (VAS) pain score (p = 0.02), while there was no significant difference between laser acupuncture and placebo based on Western Ontario and McMaster Universities Arthritis Index (WOMAC) pain score (p = 0.25). For subgroup analysis, laser acupuncture had superiority over placebo in terms of both VAS and WOMAC pain scores in the appropriate technical features subgroup and the excellent methodological quality subgroup. But the effect of laser acupuncture on pain relief was not maintained in terms of either VAS (p = 0.19) or WOMAC pain score (p = 0.60). The pooled effect showed no significant difference between laser acupuncture and placebo at either time point according to WOMAC function scale, WOMAC stiffness scale, and quality of life outcome. CONCLUSIONS: Our findings indicate that laser acupuncture can effectively reduce knee pain for patients with KOA at short term when appropriate technical features are applied, but the effect likely fades away during the subsequent follow-up period.

Costunolide inhibits matrix metalloproteinases expression and osteoarthritis via the NF­κB and Wnt/ß­catenin signaling pathways.

Osteoarthritis (OA) is a chronic joint disease involving cartilage erosion and matrix degradation. Costunolide is a sesquiterpene lactone that has been demonstrated to exert anti­inflammatory activities in a wide variety of cells. The aim of the present study was to investigate the effect of costunolide in OA treatment, using rat chondrocytes and an OA rat model, in which animals were subjected to destabilization of the medial meniscus. The results revealed that costunolide (2­6 µM) had no effect on chondrocyte viability or phenotype maintenance. Costunolide decreased the interleukin (IL)­1ß­induced upregulation of matrix metalloproteinases (MMPs), inducible nitric oxide synthase, cyclooxygenase­2 and IL­6, and increased the expression of collagen II and transcription factor SOX­9, which were inhibited by IL­1ß. Costunolide significantly decreased p65 phosphorylation induced by IL­1ß and the translocation of p65 into the nucleus of rat chondrocytes, as observed by western blot analysis and immunofluorescence staining. In addition, activation of the Wnt/ß­catenin signaling pathway was inhibited by costunolide, as demonstrated by the level of activation of ß­catenin and the transfer of ß­catenin into the nucleus induced by IL­1ß. In vivo, cartilage treated with costunolide exhibited attenuated degeneration and lower Mankin scores compared with the OA group. The present study investigated the anti­osteoarthritic effects of costunolide, which exerted anti­inflammatory activities and inhibited MMPs expression. Taken together, these results indicate that costunolide may have a potential value in the treatment of OA.

Schisandrin B ameliorated chondrocytes inflammation and osteoarthritis via suppression of NF-κB and MAPK signal pathways.

INTRODUCTION: Osteoarthritis (OA) is the most prevalent joint disorder in the elderly population, and inflammatory mediators like IL-1ß were thought to play central roles in its development. Schisandrin B, the main active component derived from Schisandra chinensis, exhibited anti-oxidative and antiinflammatory properties. METHODS: In the present study, the protective effect and the underlying mechanism of Schisan-drin B on OA was investigated in vivo and in vitro. RESULTS: The results showed that Schisandrin B decreased IL-1ß-induced upregulation of matrix metalloproteinase 3 (MMP3), MMP13, IL-6, and inducible nitric oxide synthase (iNOS) and increased IL-1ß-induced downregulation of collagen II, aggrecan, and sox9 as well. Schisandrin B significantly decreased IL-1ß-induced p65 phosphorylation and nuclear translocation of p65 in rat chondrocytes. Mitogen-activated protein kinase (MAPK) activation was also inhibited by Schisandrin B, as evidenced by the reduction of p38, extracellular signal-regulated kinase (Erk), and c-Jun amino-terminal kinase (Jnk) phosphorylation. In addition, Schisandrin B prevented cartilage degeneration in rat OA model with significantly lower Mankin's score than the control group. CONCLUSION: Our study demonstrated that Schisandrin B ameliorated chondrocytes inflammation and OA via suppression of nuclear factor-κB (NF-κB) and MAPK signal pathways, indicating a therapeutic potential in OA treatment.