MiR-106b inhibition suppresses inflammatory bone destruction of wear debris-induced periprosthetic osteolysis in rats.

Aseptic loosening caused by periprosthetic osteolysis (PPO) is the main reason for the primary artificial joint replacement. Inhibition of inflammatory osteolysis has become the main target of drug therapy for prosthesis loosening. MiR-106b is a newly discovered miRNA that plays an important role in tumour biology, inflammation and the regulation of bone mass. In this study, we analysed the in vivo effect of miR-106b on wear debris-induced PPO. A rat implant loosening model was established. The rats were then administrated a lentivirus-mediated miR-106b inhibitor, miR-106b mimics or an equivalent volume of PBS by tail vein injection. The expression levels of miR-106b were analysed by real-time PCR. Morphological changes in the distal femurs were assessed via micro-CT and histopathological analysis, and cytokine expression levels were examined via immunohistochemical staining and ELISA. The results showed that treatment with the miR-106b inhibitor markedly suppressed the expression of miR-106b in distal femur and alleviated titanium particle-induced osteolysis and bone loss. Moreover, the miR-106b inhibitor decreased TRAP-positive cell numbers and suppressed osteoclast formation, in addition to promoting the activity of osteoblasts and increasing bone formation. MiR-106b inhibition also significantly regulated macrophage polarization and decreased the inflammatory response as compared to the control group. Furthermore, miR-106b inhibition blocked the activation of the PTEN/PI3K/AKT and NF-κB signalling pathways. Our findings indicated that miR-106b inhibition suppresses wear particles-induced osteolysis and bone destruction and thus may serve as a potential therapy for PPO and aseptic loosening.

Aspirin inhibits osteoclast formation and wear-debris-induced bone destruction by suppressing mitogen-activated protein kinases.

Excessive osteoclast recruitment and activation is the chief cause of periprosthetic osteolysis and subsequent aseptic loosening, so blocking osteolysis may be useful for protecting against osteoclastic bone resorption. We studied the effect of aspirin on titanium (Ti)-particle-induced osteolysis in vivo and in vitro using male C57BL/6J mice randomized to sham (sham surgery), Ti (Ti particles), low-dose aspirin (Ti/5 mg·kg-1 ·d-1 aspirin), and high-dose aspirin (Ti/30 mg·kg-1 ·d-1 aspirin). After 2 weeks, a three-dimensional reconstruction evaluation using micro-computed tomography and histomorphology assessment were performed on murine calvariae. Murine hematopoietic macrophages and RAW264.7 lineage cells were studied to investigate osteoclast formation and function. Aspirin attenuated Ti-particle-induced bone erosion and reduced osteoclasts. In vitro, aspirin suppressed osteoclast formation, osteoclastic-related gene expression, and osteoclastic bone erosion in a dose-dependent manner. Mechanically, aspirin reduced osteoclast formation by suppressing receptor activator of nuclear factor kappa-B ligand-induced activation of extracellular signal-related kinase, p-38 mitogen-activated protein kinase, and c-Jun N-terminal kinase. Thus, aspirin may be a promising option for preventing and curing osteoclastic bone destruction, including peri-implant osteolysis.

Aspirin-Mediated Attenuation of Intervertebral Disc Degeneration by Ameliorating Reactive Oxygen Species In Vivo and In Vitro.

Intervertebral disc (IVD) degeneration (IDD) is a major cause of low back pain. The pathogenesis of IDD is associated with the disturbance of reactive oxygen species (ROS) equilibrium, inflammation, and matrix loss. Aspirin is a nonsteroidal anti-inflammatory drug that effectively inhibits inflammation and oxidative stress and has been widely used for the treatment of back pain. Therefore, we hypothesize that aspirin reverses the IDD process via antioxidative and anti-inflammatory effects on the AMPK signaling pathway. In vitro, aspirin diminished cellular oxygen free radicals (ROS, nitric oxide (NO)) and inflammatory cytokines (interleukin- (IL-) 1ß and IL-6 and tumor necrosis factor alpha (TNF-α)) induced by lipopolysaccharides (LPS) in nucleus pulposus cells (NPCs). We found that aspirin preserved the extracellular matrix (ECM) content of collagen type II (COL2) and aggrecan while inhibiting the expression of matrix-degenerating enzymes, including matrix metalloproteinase 3 and 13 (MMP-3 and MMP-13) and A disintegrin and metalloproteinase with thrombospondin motifs 4 and 5 (ADAMTS-4, ADAMTS-5). Aspirin significantly promoted the ratios of p-AMPK to AMPK and p-ACC to ACC expression in NPCs. Furthermore, pretreatment with the AMPK inhibitor compound C abrogated the antioxidant effects of aspirin. In vivo, an IDD model was established in Sprague-Dawley rats via percutaneous disc puncture with the 20-gauge needle on levels 8-9 and 9-10 of the coccygeal vertebrae. Imaging assessment showed that after aspirin treatment, improvements in disc height index (DHI) ranged from 1.22-fold to 1.54-fold and nucleus pulposus signal strength improved from 1.26-fold to 1.33-fold. Histological analysis showed that aspirin treatment prevented the loss of COL2 and decreased MMP-3 and MMP-13, inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), IL-1ß, and TNF-α expression in the IVD tissues. These results suggest that treatment with aspirin could reverse the IDD process via the AMPK signaling pathway, which provides new insights into the potential clinical applications of aspirin, particularly for IDD treatment.

Melatonin Increases Bone Mass around the Prostheses of OVX Rats by Ameliorating Mitochondrial Oxidative Stress via the SIRT3/SOD2 Signaling Pathway.

Bone mass loss around prostheses is a major cause of implant failure, especially in postmenopausal osteoporosis patients. In osteoporosis, excess oxidative stress largely contributed abnormal bone remodeling. Melatonin, which is synthesized from the pineal gland, promotes osteoblast differentiation and bone formation and has effectively been used to combat oxidative stress. Thus, we determined if melatonin can inhibit oxidative stress to promote osteogenesis and improve bone mass around prostheses in osteoporosis. In this study, we observed that received melatonin at 50 mg/kg body weight significantly increased periprosthetic bone mass as well as implant fixation intensity in ovariectomized (OVX) rats. Meanwhile, it decreased the expression of oxidative stress markers (NAPDH oxidase 2 and cytochrome c) and enhanced expressing level of the formation markers of bones (alkaline phosphatase, osteocalcin, and osterix) around prostheses compared to that in the control group. Additionally, melatonin decreased hydrogen peroxide- (H2O2-) induced oxidative stress and restored the osteogenesis potential of MC3T3-E1 cells. Mechanistically, melatonin clearly increased mitochondrial sirtuin 3 (SIRT3) expression and decreased the ratio of acetylated superoxide dismutase 2 (AC-SOD2)/SOD2 compared to the H2O2 group. SIRT3 inhibition counteracted the protective effects of melatonin on oxidative stress and bone formation. Together, the results showed that melatonin ameliorated oxidative stress in mitochondrial via the SIRT3/SOD2 signaling pathway, thereby promoting osteogenesis, improving bone mass around the prostheses, and increasing initial stability. Thus, melatonin might be a suitable candidate to decrease the rate of implant failure and lengthen the lifespan of prostheses after total joint arthroplasty.

Exenatide ameliorates inflammatory response in human rheumatoid arthritis fibroblast-like synoviocytes.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease of unknown etiology characterized by degradation of cartilage and bone, accompanied by unimpeded proliferation of synoviocytes of altered phenotype. In the present study, we investigated the involvement of the glucagon-like peptide 1 (GLP-1) receptor on human fibroblast-like synoviocytes (FLS) in the pathogenesis of RA using the selective GLP-1 agonist exenatide, a licensed drug used for the treatment of type 2 diabetes. Our results indicate that exenatide may play a role in regulating tumor necrosis factor-α-induced mitochondrial dysfunction by increasing mitochondrial membrane potential, oxidative stress by reducing the production of reactive oxygen species, the expression of NADPH oxidase 4, expression of matrix metalloproteinase (MMP)-3 and MMP-13, release of proinflammatory cytokines including interleukin-1ß (IL-1ß), IL-6, monocyte chemoattractant protein-1, and high-mobility group protein 1, as well as activation of the p38/nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor, α/nuclear factor κB signaling pathway in primary human RA FLS. These positive results indicate that exenatide may have potential as a therapeutic agent for the treatment and prevention of RA. © 2019 IUBMB Life, 9999(9999):1-9, 2019.

Tenocyte-derived exosomes induce the tenogenic differentiation of mesenchymal stem cells through TGF-ß.

Mesenchymal stem cells (MSCs) hold great potential to treat tissue damage based on their multipotent property, and are also considered as suitable cell resources to create tissue-engineered grafts for tendon repair. However, the clinical application of MSCs is still limited by the lack of efficient methods to induce tenogenic differentiation. In this study, by performing the experiments in transwell system, we found that paracrine factors from tenocytes could induce MSCs to undergo the tenogenic differentiation. We further verified that tenocytes could secrete exosomes and these tenocyte-derived exosomes efficiently initiated the tenogenic differentiation of MSCs. Finally, we revealed that the TGF-ß existing in tenocyte-derived exosomes mediated the process, as the inhibition of TGF-ß signaling abolished the effects of tenocyte-derived exosomes on MSCs. By investigating the effects of tenocytes on MSCs, we found that tenocytes-derived exosomes can induce tenogenic differentiation of MSCs in a TGF-ß dependent manner. These studies provided critical information about the multipotency of MSCs and suggested potential strategies for clinical translation.

Efficient Inhibition of Wear-Debris-Induced Osteolysis by Surface Biomimetic Engineering of Titanium Implant with a Mussel-Derived Integrin-Targeting Peptide.

Wear-debris-induced osteolysis and subsequent aseptic loosening of the prosthesis are the main reasons for failed arthroplasty. Inhibition of wear-debris-induced osteoclastogenesis is thus a promising method to prolong the lifetime of prostheses. In this work, a mussel-derived peptide, capped with an integrin-targeting RGD tripeptide and a titanium (Ti)-affinity tetrapeptide with catechol groups at each end is biomimetically designed. The RGD peptide can interfere with integrin αv ß3 to affect the cytoskeletal organization and functions of osteoclasts and subsequently inhibit osteoclast hyperactivation and osteoclastogenesis in vitro and in vivo, while the catechol groups could easily adhere to the TiO2 layer of the Ti implant via Ti-catechol coordination. This design thus enables the stable immobilization of the RGD peptide on Ti implants to inhibit osteoclast formation and reduce osteolysis in vivo, even with the existence of Ti wear particles. Further study reveals that the suppression of osteoclastogenesis and osteoclast polarization on the peptide coating is regulated by blocking the PI3K/AKT and NF-κB signal pathways. Considering the simplicity in the surface engineering, the highly biomimetic nature of the bioactive peptide, and efficient inhibition of wear-debris-caused osteolysis, this work thus presents a simple and efficient strategy to improve clinical outcome of prosthesis implantation in challenging conditions.

KLF2 regulates osteoblast differentiation by targeting of Runx2.

Osteoblast differentiation plays a critical role in bone formation and maintaining balance in bone remodeling. Runt-related transcription factor 2 (Runx2) is a central transcription factor regulating osteoblast differentiation and promoting bone mineralization. Until now, the molecular regulatory basis and especially the gene regulatory network of osteogenic differentiation have been unclear. Krüppel-like factor 2 (KLF2) is a zinc finger structure and DNA-binding transcription factor. The current study aimed to investigate the physiological function of KLF2 in osteoblast differentiation. Our results indicate that KLF2 is expressed in pre-osteoblast MC3T3-E1 cells and primary osteoblasts. Interestingly, KLF2 expression is increased in osteoblasts during the osteoblastic differentiation process. Overexpression of KLF2 in MC3T3-E1 cells promoted the expression of the osteoblastic differentiation marker genes Alp, Osx, and Ocn, and stimulated mineralization by increasing Runx2 expression at both the mRNA and protein levels. In contrast, knockdown of KLF2 produced the opposite effects. Importantly, we found that KLF2 could physically interact with Runx2. KLF2 promoted osteoblast differentiation by regulating Runx2 and physically interacting with Runx2. Taken together, the findings of this study identify KLF2 as a novel regulator of osteoblast differentiation. Our findings suggest that KLF2 might be a new therapeutic target for bone disease.

Shikimic acid prevents cartilage matrix destruction in human chondrocytes.

Abnormal reduction of extracellular matrix (ECM), including type II collagen and aggrecan, caused by tumor necrosis factor-α (TNF-α) is an important pathological feature of osteoarthritis (OA). Shikimic acid (SA), derived from natural plants, has displayed effective pharmacological properties in diverse diseases. The biological roles of SA in OA have not been reported before. Here, we found that treatment with SA (1 mM, 10 mM) prevented TNF-α-induced degradation of type II collagen and aggrecan ECM in human primary chondrocytes culture in vitro. Importantly, we also reported that SA treatment reduced TNF-α-induced expression of matrix metalloproteinase­1, ­3, and ­13 (MMP­1, ­3, and ­13) and increased expression of tissue inhibitor of metalloproteinase­1 and ­2 (TIMP­1, ­2). Additionally, SA treatment attenuated TNF-α-induced expression of a disintegrin and metalloprotease­4 and ­5 (ADAMTS­4, ­5). Mechanistically, we found that SA prevented activation of the nuclear factor-κB (NF­κB) pathway. Our findings suggest that SA might act as an important therapeutic agent in the treatment of OA.