Chemical Inhibitors Targeting the Oncogenic m6A Modifying Proteins.

Epigenetics is brought to RNA, introducing a new dimension to gene expression regulation. Among numerous RNA modifications, N6-methyladenosine (m6A) is an abundant internal modification on eukaryote mRNA first identified in the 1970s. However, the significance of m6A modification in mRNA had been long neglected until the fat mass and obesity-associated (FTO) enzyme was identified as the first m6A demethylase almost 40 years later. The m6A modification influences nearly every step of RNA metabolism and thus broadly affects gene expression at multiple levels, playing a critical role in many biological processes, including cancer progression, metastasis, and immune evasion. The m6A level is dynamically regulated by RNA epigenetic machinery comprising methyltransferases such as methyltransferase-like protein 3 (METTL3), demethylases FTO and AlkB human homologue 5 (ALKBH5), and multiple reader proteins. The understanding of the biology of RNA epigenetics and its translational drug discovery is still in its infancy. It is essential to further develop chemical probes and lead compounds for an in-depth investigation into m6A biology and the translational discovery of anticancer drugs targeting m6A modifying oncogenic proteins.In this Account, we present our work on the development of chemical inhibitors to regulate m6A in mRNA by targeting the FTO demethylase, and the elucidation of their mode of action. We reported rhein to be the first substrate competitive FTO inhibitor. Due to rhein's poor selectivity, we identified meclofenamic acid (MA) that selectively inhibits FTO compared with ALKBH5. Based on the structural complex of MA bound with FTO, we designed MA analogs FB23-2 and Dac51, which exhibit significantly improved activities compared with MA. For example, FB23-2 is specific to FTO inhibition in vitro among over 400 other oncogenic proteins, including kinases, proteases, and DNA and histone epigenetic proteins. Mimicking FTO depletion, FB23-2 promotes the differentiation/apoptosis of human acute myeloid leukemia (AML) cells and inhibits the progression of primary cells in xenotransplanted mice. Dac51 treatment impairs the glycolytic activity of tumor cells and restores the function of CD8+ T cells, thereby inhibiting the growth of solid tumors in vivo. These FTO inhibitors were and will continue to be used as probes to promote biological studies of m6A modification and as lead compounds to target FTO in anticancer drug discovery.Toward the end, we also include a brief review of ALKBH5 demethylase inhibitors and METTL3 methyltransferase modulators. Collectively, these small-molecule modulators that selectively target RNA epigenetic proteins will promote in-depth studies on the regulation of gene expression and potentially accelerate anticancer target discovery.

Exploring m6A-linked aging genes in osteoarthritis and broad cancer spectrum: Prospects for diagnostic and therapeutic advancements.

Osteoarthritis (OA) is a prevalent degenerative joint disease that significantly impacts individuals and healthcare systems worldwide. However, the exploration of N6-methyladenosine (m6A)-related aging genes in OA pathogenesis remains largely underexplored. This study aimed to elucidate the role of m6A-related aging genes in OA and to develop a robust diagnostic model based on their expression profiles. Leveraging publicly available gene expression datasets, we conducted consensus clustering to categorize OA into distinct subtypes, guided by the expression patterns of m6A-related aging genes. Utilizing XGBoost, a cutting-edge machine learning approach, we identified key diagnostic genes and constructed a predictive model. Our investigation extended to the immune functions of these genes, shedding light on potential therapeutic targets and underlying regulatory mechanisms. Our analysis unveiled specific OA subtypes, each marked by unique expression profiles of m6A-related aging genes. We pinpointed a set of pivotal diagnostic genes, offering potential therapeutic avenues. The developed diagnostic model exhibited exceptional capability in distinguishing OA patients from healthy controls. To corroborate our computational findings, we performed quantitative real-time polymerase chain reaction analyses on two cell lines: HC-OA (representing adult osteoarthritis cells) and C-28/I2 (representative of normal human chondrocytes). The gene expression patterns observed were consistent with our bioinformatics predictions, further validating our initial results. In conclusion, this study underscores the significance of m6A-related aging genes as promising biomarkers for diagnosis and prognosis, as well as potential therapeutic targets in OA. Although these findings are encouraging, further validation and functional analyses are crucial for their clinical application.

Merlin functions as a critical regulator in Staphylococcus aureus-induced osteomyelitis.

Merlin is known as a tumor suppressor, while its role in osteomyelitis remains unclear. This study aimed to investigate the role of Merlin in Staphylococcus aureus-induced osteomyelitis and its underlying mechanisms. S. aureus-induced osteomyelitis mouse model was established in Merlinfl/fl Lyz2cre/+ and Merlinfl/fl Lyz2+/+ mice. Bone marrow-derived macrophages (BMDMs) were isolated and stimulated by lipopolysaccharide (LPS). Bioassays, including quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blot analysis, and enzyme-linked immunosorbent assays, were conducted to determine the levels of target genes or proteins. Immunoprecipitation was applied to determine the interactions between proteins. DCAF1fl/fl mice were further crossed with Lyz2-Cre mice to establish myeloid cell conditional knockout mice (DCAF1fl/fl Lyz2cre/+ ). It was found that the level of Merlin was elevated in patients with osteomyelitis and S. aureus-infected BMDMs. Merlin deficiency in macrophages suppressed the production of inflammatory cytokines and ameliorated the symptoms of osteomyelitis induced by S. aureus. Merlin deficiency in macrophages also suppressed the production of proinflammatory cytokines in BMDMs induced by LPS. The inhibitory effects of Merlin deficiency on the inflammatory response were associated with DDB1-Cul4-associated factor 1 (DCAF1). In summary, Merlin deficiency ameliorates S. aureus-induced osteomyelitis through the regulation of DCAF1.

Involvement of mechanosensitive ion channels in the effects of mechanical stretch induces osteogenic differentiation in mouse bone marrow mesenchymal stem cells.

Bone marrow mesenchymal stem cells (BMSCs) can be induced to process osteogenic differentiation with appropriate mechanical and/or chemical stimuli. The present study described the successful culture of murine BMSCs under mechanical strain. BMSCs were subjected to 0%, 3%, 8%, 13%, and 18% cyclic tensile strain at 0.5 Hz for 8 hr/day for 3 days. The expression of osteogenic markers and mechanosensitive ion channels was evaluated with real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blot. The expression of alkaline phosphatase (ALP) and matrix mineralization were evaluated with histochemical staining. To investigate the effects of mechanosensitive ion channel expression on cyclic tensile strain-induced osteogenic differentiation, the expression of osteogenic markers was evaluated with real-time RT-PCR in the cells without mechanosensitive ion channel expression. This study revealed a significant augment in osteogenic marker in BMSC strained at 8% compared to other treatments; therefore, an 8% strain was used for further investigations. The ALP expression and matrix mineralization were enhanced in osteogenic induced BMSCs subjected to 8% strain after 7 and 14 days, respectively. Under the same conditions, the osteogenic marker and mechanosensitive ion channel expression were significantly promoted. However, the loss function of mechanosensitive ion channels resulted in the inhibition of osteogenic marker expression. This study demonstrated that strain alone can successfully induce osteogenic differentiation in BMSCs and the expression of mechanosensitive ion channels was involved in the process. The current findings suggest that mechanical stretch could function as efficient stimuli to induce the osteogenic differentiation of BMSCs via the activation of mechanosensitive ion channels.

Inhibition of microRNA-222 up-regulates TIMP3 to promotes osteogenic differentiation of MSCs from fracture rats with type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM) is the most common diabetes and has numerous complications. Recent studies demonstrated that T2DM compromises bone fracture healing in which miR-222 might be involved. Furthermore, tissue inhibitor of metalloproteinase 3 (TIMP-3) that is the target of miR-222 accelerates fracture healing. Therefore, we assume that miR-222 could inhibit TIMP-3 expression. Eight-week-old rats were operated femoral fracture or sham, following the injection of streptozotocin (STZ) to induce diabetes one week later in fractured rats, and then, new generated tissues were collected for measuring the expression of miR-222 and TIMP-3. Rat mesenchymal stem cells (MSCs) were isolated and treated with miR-222 mimic or inhibitor to analyse osteogenic differentiation. MiR-222 was increased in fractured rats and further induced in diabetic rats. In contrast, TIMP-3 was reduced in fractured and further down-regulated in diabetic rats. Luciferase report assay indicated miR-222 directly binds and mediated TIMP-3. Furthermore, osteogenic differentiation was suppressed by miR-222 mimic and promoted by miR-222 inhibitor. miR-222 is a key regulator that is promoted in STZ-induced diabetic rats, and it binds to TIMP3 to reduce TIMP-3 expression and suppressed MSCs' differentiation.

The promotion of bone regeneration through CS/GP-CTH/antagomir-133a/b sustained release system.

Few studies reported the application of miRNA in bone regeneration. In this study, the expression of miR133a and miR133b in murine BMSCs was inhibited via antagomiR-133a/b and the osteogenic differentiation in murine BMSCs was evaluated. The RT-PCR, flow cytometry, cell counting kit-8, and annexin V-FITC/PI double staining assays were performed. Double knockdown miR133a and miR133b can promote BMSC osteogenic differentiation. At optimum N/P ration (15:1), the loading efficiency can reach over 90%. CTH-antagomiR-133a/b showed no cytotoxicity to BMSCs and diminished miR133a and miR133b expression in BMSCs. Furthermore, chitosan-based sustained delivery system can facilitate continuous dosing of antagomiR-133a/b, which enhanced calcium deposition and osteogenic specific gene expression in vitro. The new bone formation was enhanced after the sustained delivery system containing CTH-antagomiR-133a/b nanoparticles was used in mouse calvarial bone defect model. Our results demonstrate that CTH nanoparticles could facilitate continuous dosing of antagomiR133a/b, which can promote osteogenic differentiation.

Ubiquitin-Specific Peptidase 8 Is Critical for the Onset of Infectious Osteomyelitis by Targeting RIPK2 Ubiquitination.

Receptor-interacting serine/threonine kinase (RIPK) is associated with cellular inflammation and immune regulation. The current study explored the role of RIPK2 in osteomyelitis and the potential upstream targets of RIPK2. A Staphylococcus aureus-induced osteomyelitis mouse model was established using wild-type (WT) and ubiquitin-specific peptidase 8 (USP8)-deficient (USP-/-) mice, and the osteomyelitis-related symptoms were evaluated. Bone marrow-derived macrophages (BMDMs) were isolated from the WT and USP-/- mice. Enzyme-linked immunosorbent assays, quantitative polymerase chain reaction, and immunoblot analysis were used to determine the levels of target biomarkers, which were induced by lipopolysaccharide (LPS), CpG, or PAM3CSK4. USP8 promoted RIPK2-mediated NF-κB activation. USP8 is indispensable for RIPK2-mediated LPS-induced NF-κB activation in BMDMs. USP8 is required for the production of inflammatory cytokines induced by LPS, CpG, or PAM3CSK4 in BMDMs. In addition, USP-/- mice exhibited ameliorated symptoms, including less body weight and cortical bone loss, and reduced bacterial load and reactive bone formation in the S. aureus-induced osteomyelitis mouse model. USP8 is critical in the S. aureus-induced osteomyelitis mouse model by targeting RIPK2 ubiquitination.

The Ubiquitination of RIPK2 is Mediated by Peli3 and Negatively Regulates the Onset of Infectious Osteomyelitis.

Osteomyelitis is the infection and destruction of the bone. To date, there is no universal protocol for its treatment. Receptor-interacting serine/threonine-protein kinase 2 (RIPK2) has been implicated in osteomyelitis development. However, the detailed mechanism remains unknown. Here, 6-8w wild-type or Pellino E3 Ubiquitin Protein Ligase Family Member 3 (Peli3)-deficient mice were injected with Staphylococcus aureus to induce osteomyelitis. RAW264.7 cells or bone marrow-derived macrophages isolated from mice were treated with lipopolysaccharide (LPS). Knocking down Peli3 in RAW264.7 cells increased the expression of inflammatory cytokines (interleukin-1ß, interleukin-6, and tumor necrosis factor-α) after LPS stimulation. Inflammation was also activated in S. aureus-induced Peli3-deficient mice. Moreover, S. aureus-infected Peli3-deficient mice also displayed more severe symptoms of osteomyelitis than S. aureus-infected wild-type mice. Moreover, Peli3 targets and degrades RIPK2 through K48-linked ubiquitination, and negatively modulates osteomyelitis by degrading RIPK2. Our data further expands the current understanding of RIPK2 in osteomyelitis, and suggests that RIPK2 might serve as a novel therapeutic target for treating osteomyelitis.

Effects of astragaloside IV on glucocorticoid-induced avascular necrosis of the femoral head via regulating Akt-related pathways.

We investigated the role of astragaloside IV (AS-IV) in preventing glucocorticoid-induced avascular necrosis of the femoral head (ANFH) and the underlying molecular mechanisms. Network pharmacology was used to predict the molecular targets of AS-IV. Molecular dynamic simulations were performed to explore the binding mechanism and interaction mode between AS-IV and Akt. Rat models of glucocorticoid-induced ANFH with AS-IV intervention were established, and osteogenesis, angiogenesis, apoptosis and oxidative stress were evaluated before and after blocking the PI3K/Akt pathway with LY294002. The effects of glucocorticoid and AS-IV on bone marrow mesenchymal stem cells and human umbilical vein endothelial cells incubated with and without LY294002 were determined. Downregulated p-Akt expression could be detected in the femoral heads of glucocorticoid-induced ANFH patients and rats. AS-IV increased trabecular bone integrity and vessel density of the femoral head in the model rats. AS-IV increased Akt phosphorylation and upregulated osteogenesis-, angiogenesis-, apoptosis- and oxidative stress-related proteins and mRNA and downregulated Bax, cleaved caspase-3 and cytochrome c levels. AS-IV promoted human umbilical vein endothelial cell migration, proliferation and tube formation ability; bone marrow mesenchymal stem cell proliferation; and osteogenic differentiation under glucocorticoid influence. AS-IV inhibited apoptosis. LY294002 inhibited these effects. AS-IV prevented glucocorticoid-induced ANFH by promoting osteogenesis and angiogenesis via the Akt/Runx2 and Akt/HIF-1α/VEGF pathways, respectively, and suppressing apoptosis and oxidative stress via the Akt/Bad/Bcl-2 and Akt/Nrf2/HO-1 pathways, respectively.

miR-146a Protects against Staphylococcus aureus-Induced Osteomyelitis by Regulating Inflammation and Osteogenesis.

Osteomyelitis is a Staphylococcus aureus-caused bone infection. In this study, the effects of miR-146a on osteomyelitis were evaluated. Using the osteoblast cell model and S. aureus-induced osteomyelitis mice model, we monitored the miR-146 expression and explored the effects of miR-146a on cell proliferation of osteoblasts, bone remodeling, osteoclastogenesis, inflammatory cytokine production, and bacterial burden. Upregulated miR-146a was found in mice with S. aureus-induced osteomyelitis. miR-146a attenuated S. aureus-induced cell loss of osteoblasts, rescued the expression of osteogenic markers, altered the bone remodeling, and inhibited inflammatory cytokine production and osteoclastogenesis. miR-146a knockout mice had higher S. aureus burden. In conclusion, miR-146a protects against S. aureus-induced osteomyelitis by regulating inflammation and osteogenesis.

Staphylococcus aureus Infection Initiates Hypoxia-Mediated Transforming Growth Factor-ß1 Upregulation to Trigger Osteomyelitis.

Little is unknown about the regulatory mechanisms underlying the pathogenesis of osteomyelitis induced by Staphylococcus aureus. Hypoxia-inducible factor-1α (HIF-1α) and transforming growth factor ß1 (TGF-ß1) were both upregulated in S. aureus-infected MC3T3-E1 cells and osteomyelitis patients. HIF-1α directly targets the hypoxia-responsive elements (HREs) of TGF-ß1 mRNA to induce its expression. Silencing HIF-1α and TGF-ß1, as well as treatment of hypoxia inhibitor IDF-11774, consistently elevated OPN and RUNX2 expression and alizarin Red S (ARS) and alkaline phosphatase (ALP) staining levels in MC3T3-E1 cells with S. aureus infection. S. aureus infection increased HIF-1α expression and serum TGF-ß1 concentration in a mouse model of osteomyelitis. Hypoxia inhibitor IDF-11774 treatment reduced serum levels of interleukin (IL)-6, IL-1ß, and C-reactive protein. Upon S. aureus infection, hypoxia was activated to trigger TGF-ß1 upregulation through direct targeting of HRE on TGF-ß1 mRNA by HIF-1α, eventually leading to osteomyelitis symptoms in terms of osteogenesis and mineralization deficiencies as well as elevated inflammation. This study hereby suggests a novel signaling cascade involving hypoxia/HIF-1α/TGF-ß1 in osteomyelitis pathogenesis, which could potentially serve as a target for therapeutic measures. IMPORTANCE The pathogenesis of osteomyelitis induced by Staphylococcus aureus remains unclear. To develop therapeutic approaches for osteomyelitis, it is important to understand the molecular mechanisms of its pathogenesis. Our results suggests that hypoxia/HIF-1α/TGF-ß1 signaling is involved in osteomyelitis pathogenesis. Thus, these findings highlight the potential of this signaling components as therapeutic targets for the treatment of osteomyelitis.

Gallium Porphyrin and Gallium Nitrate Reduce the High Vancomycin Tolerance of MRSA Biofilms by Promoting Extracellular DNA-Dependent Biofilm Dispersion.

Biofilms, structured communities of bacterial cells embedded in a self-produced extracellular matrix (ECM) which consists of proteins, polysaccharide intercellular adhesins (PIAs), and extracellular DNA (eDNA), play a key role in clinical infections and are associated with an increased morbidity and mortality by protecting the embedded bacteria against drug and immune response. The high levels of antibiotic tolerance render classical antibiotic therapies impractical for biofilm-related infections. Thus, novel drugs and strategies are required to reduce biofilm tolerance and eliminate biofilm-protected bacteria. Here, we showed that gallium, an iron mimetic metal, can lead to nutritional iron starvation and act as dispersal agent triggering the reconstruction and dispersion of mature methicillin-resistant Staphylococcus aureus (MRSA) biofilms in an eDNA-dependent manner. The extracellular matrix, along with the integral bacteria themselves, establishes the integrated three-dimensional structure of the mature biofilm. The structures and compositions of gallium-treated mature biofilms differed from those of natural or antibiotic-survived mature biofilms but were similar to those of immature biofilms. Similar to immature biofilms, gallium-treated biofilms had lower levels of antibiotic tolerance, and our in vitro tests showed that treatment with gallium agents reduced the antibiotic tolerance of mature MRSA biofilms. Thus, the sequential administration of gallium agents (gallium porphyrin and gallium nitrate) and relatively low concentrations of vancomycin (16 mg/L) effectively eliminated mature MRSA biofilms and eradicated biofilm-enclosed bacteria within 1 week. Our results suggested that gallium agents may represent a potential treatment for refractory biofilm-related infections, such as prosthetic joint infections (PJI) and osteomyelitis, and provide a novel basis for future biofilm treatments based on the disruption of normal biofilm-development processes.

IKKe in osteoclast inhibits the progression of methylprednisolone-induced osteonecrosis.

Previous studies have described that NF-κB signaling mediated by NFκB-inducing kinase (NIK) plays a critical role of the differentiation of osteoclasts. We aim to explore the role of IKKe in methylprednisolone -induced osteonecrosis of the femoral head (ONFH). Methylprednisolone-induced ONFH mice model was successfully established, and subjected to micro computed tomography to detect the femoral head image of the mice. Bone marrow cells from experimental mice were collected and cultured. qPCR and immunoblot were performed to examine the possible signal pathways of IKKe involvement, and osteoclast-related gene expressions in IKKe+/+ and IKKe-/- cells in vitro and in vivo were examined. It was found that the levels of IKKe decreased in ONFH patients, and IKKe interacted with NIK in the NF-κB signal pathway to suppress osteoclasts via inhibiting the transcription of NIK. Furthermore, IKKe knockout promoted the osteoclastogenesis in mice model. Finally, IKKe knockout suppressed methylprednisolone-induced ONFH and pro-inflammatory responses in mice model. Our findings show a mechanism of IKKe inhibition of the progression of methylprednisolone-induced ONFH via the NIK/NF-κB pathway.

Bone morphogenetic protein 2 controls steroid-induced osteonecrosis of the femoral head via directly inhibiting interleukin-34 expression.

Increased inflammatory response is one of the major characteristics of osteonecrosis of the femoral head (ONFH). We aimed to investigate the function of bone morphogenetic protein 2 (BMP-2)/interleukin (IL)-34 axis in the inflammatory responses of ONFH. The systemic and local expression of BMPs in ONFH patients was detected by qRT-PCR and ELISA. In vitro osteoclast differentiation and ONFH mouse models, induced by 20 mg/kg methylprednisolone through i.m. injection, were established using WT and BMP-2-/- mice to explore the regulatory role of BMP-2 in pro-inflammatory responses and bone defects of ONFH. IL-34 expression and function were examined in vitro and in vivo through qRT-PCR, tartrate-resistant acid phosphatase (TRAP) staining, and gene knockout. The systemic and local expression of BMPs was elevated in ONFH patients. BMP-2 reduced the production of pro-inflammatory cytokines and inhibited the differentiation of osteoclasts. Mechanistically, BMP-2 inhibited osteoclasts formation through suppressing IL-34 expression and then promoted bone repair and alleviated ONFH. In conclusion, our study reveals that BMP-2 inhibits inflammatory responses and osteoclast formation through downregulating IL-34.

Astragaloside IV ameliorates steroid-induced osteonecrosis of the femoral head by repolarizing the phenotype of pro-inflammatory macrophages.

Osteonecrosis of the femoral head (ON-FH) is a common complication of steroid use. Pro-inflammatory macrophages play a crucial role in the apoptosis of osteocytes. The objective of the study was to evaluate a plant extract astragaloside IV (AS-IV) in treating ON-FN. Bone-marrow-derived macrophages (BMDMs) were treated with lipopolysaccharides (LPS), IFN-γ or IL-4 to induce M1 and M2-like phenotypes. Quantitative real-time PCR and Western blot were used to examine M1 and M2 phenotypic markers. Flow cytometry was used to analyze MHC II, CD206, F4/80, and CD11b levels and cell apoptosis. Glucocorticoid was used to induce ON-FN in mice. TNF-α and IL-1ß levels in femoral head were determined using enzyme-linked immunosorbent assay. AS-IV repolarized macrophages from M1 to M2 phenotypes. Culture medium from AS-IV treated M1 macrophages induced less cell apoptosis osteocytes compared to that from untreated M1 macrophages. In ON-FH mice, the ratio of M1 macrophages was decreased in the femoral head by AS-IV, concomitant with a decrease in TNF-α and IL-1ß levels. AS-IV is effective in alleviating ON-FH through its effects in repolarizing macrophages from M1-like phenotype to M2-like phenotype, promoting survival of osteocytes, reducing arthritic symptoms, and decreasing inflammatory cytokines.

Tumor necrosis factor-α promotes Staphylococcus aureus-induced osteomyelitis through downregulating endothelial nitric oxide synthase.

BACKGROUND: Infections of Staphylococcus aureus (S. aureus) often result in osteomyelitis, which is the acute or chronic infections of the bone marrow or bones. TNF-α is long recognized as a key factor contributing to the pathogenesis of osteomyelitis, but little is known about the underlying molecular mechanism. METHODS: Expression levels of TNF-α, and several candidate genes, including endothelial nitric oxide synthase (eNOS), known to be downregulated by TNF-α were analysed in MC3T3-E1 cells with S. aureus infection and osteomyelitis patient blood. MicroRNA(miR)-129-5p was predicted and experimentally verified to target eNOS. Alizarin red sulfate (ARS) and alkaline phosphatase (ALP) staining assays were conducted on MC3T3-E1 cells with S. aureus infection to assess the role of TNF-α/miR-129-5p/eNOS on mineralization defect. RESULTS: TNF-α and miR-129-5p were upregulated while eNOS was downregulated in MC3T3-E1 cells with S. aureus infection and osteomyelitis patients, showing inversely correlated expression profiles. MiR-129-5p directly binds to the 3'-UTR of eNOS mRNA to suppress eNOS expression in MC3T3-E1 cells. TNF-α blocker inhibited miR-129-5p and elevated eNOS expression, likely contributing to rescued mineralization defect in S. aureus-infected MC3T3-E1 cells. During S. aureus infection, upregulated TNF-α increases endogenous miR-129-5p expression, which in turn inhibits eNOS, contributing to osteomyelitis. CONCLUSION: Our study thereby proposes a novel signalling cascade involving TNF-α/miR-129-5p/eNOS in the pathogenesis of osteomyelitis, which may also serve as therapeutic targets.

MicroRNA-23b-3p participates in steroid-induced osteonecrosis of the femoral head by suppressing ZNF667 expression.

BACKGROUND: Clinical treatment with high-dose of steroid hormone causes steroid-induced osteonecrosis of the femoral head (SONFH), whereas the internal regulation mechanism remains elusive. Numerous studies have reported that microRNAs participated in the development of SONFH through modulating gene expression. The aim of the current study was to clarify the function of microRNA-23b-3p (miR-23b-3p) and ZNF667 in SONFH. EXPERIMENTAL DESIGN: Bioinformatics prediction and luciferase reporter system were utilized to confirm the target relation between miR-23b-3p and ZNF667. To examine the function of miR-23b-3p in vivo, rat SONFH models were established by specific inducers. The morphological changes, plasma viscosity, blood lipid, and inflammatory cytokines were measure by corresponding experiments. RESULTS: MiR-23b-3p and ZNF667 was negatively correlated in SONFH patient tissues, miR-23b-3p was down-regulated, while ZNF667 was up-regulated. MiR-23b-3p targeted ZNF667, the expression level of ZNF667 was suppressed by miR-23b-3p activation whereas strengthened by miR-23b-3p inhibition. SONHF rats with overexpressed miR-23b-3p displayed alleviated symptoms, including reduced plasma viscosity, declined blood lipids, decreased levels of pro-inflammatory cytokines and improved bone integrality. Moreover, elevation of ZNF667 reversed the repression of SONFH induced by miR-23b-3p overexpression. CONCLUSIONS: We found that miR-23b-3p played a protective role in SONFH by targeting ZNF667, which provided a novel reference for SONFH prevention and therapy.

miR-26a Attenuated Bone-Specific Insulin Resistance and Bone Quality in Diabetic Mice.

Diabetes mellitus is a prevalent disease result in several complications, including bone problems. Previous studies have shown that microRNA (miR)-26a regulates glucose metabolism and plays a protective role in diabetes. However, whether miR-26a also affects bone quality in diabetes remains unknown. In the present study, we evaluated the potential effects of miR-26a on bone in diabetic mice. We administrated miR-26a in streptozotocin-induced diabetic mice. The metabolic parameters, bone quality, osteoblast and osteoclast markers, and insulin signaling activation were measured. miR-26a ameliorated insulin resistance and glucose tolerance, improved bone microarchitecture and quality, increased osteoblasts and bone formation, decreased osteoclasts, and promoted the insulin signaling pathway in diabetic mice. These effects were abolished in insulin receptor-compromised Col1a1-Insr+/- mice. In conclusion, miR-26a could ameliorate bone-specific insulin resistance and bone quality in diabetic mice, which depended on the insulin receptors on osteoblasts. Our findings highlight the potential of miR-26a as a therapeutic target for diabetes mellitus-related bone metabolism and diseases.

20(S)-Protopanaxadiol Inhibits Titanium Particle-Induced Inflammatory Osteolysis and RANKL-Mediated Osteoclastogenesis via MAPK and NF-κB Signaling Pathways.

Osteolysis is a principal reason for arthroplasty failure like aseptic loosening induced by Titanium (Ti) particle. It is a challenge for orthopedic surgeons. Recent researches show that 20(S)-protopanaxadiol can inhibit inflammatory cytokine release in vitro. This study aims to assess the effect of 20(S)-protopanaxadiol on Ti particle-induced osteolysis and RANKL-mediated osteoclastogenesis. Micro-CT and histological analysis in vivo indicated the inhibitory effects of 20(S)-protopanaxadiol on osteoclastogenesis and the excretion of inflammatory cytokines. Next, we demonstrated that 20(S)-protopanaxadiol inhibited osteoclast differentiation, bone resorption area, and F-actin ring formation in a dose-dependent manner. Moreover, mechanistic studies suggested that the suppression of MAPK and NF-κB signaling pathways were found to mediate the inhibitory effects of 20(S)-protopanaxadiol. In conclusion, 20(S)-protopanaxadiol may suppress osteoclastogenesis in a dose- dependent manner and it could be a potential treatment of Ti particle-induced osteolysis.