Double-network hydrogel enhanced by SS31-loaded mesoporous polydopamine nanoparticles: Symphonic collaboration of near-infrared photothermal antibacterial effect and mitochondrial maintenance for full-thickness wound healing in diabetes mellitus.

Diabetic wound healing has become a serious healthcare challenge. The high-glucose environment leads to persistent bacterial infection and mitochondrial dysfunction, resulting in chronic inflammation, abnormal vascular function, and tissue necrosis. To solve these issues, we developed a double-network hydrogel, constructed with pluronic F127 diacrylate (F127DA) and hyaluronic acid methacrylate (HAMA), and enhanced by SS31-loaded mesoporous polydopamine nanoparticles (MPDA NPs). As components, SS31, a mitochondria-targeted peptide, maintains mitochondrial function, reduces mitochondrial reactive oxygen species (ROS) and thus regulates macrophage polarization, as well as promoting cell proliferation and migration, while MPDA NPs not only scavenge ROS and exert an anti-bacterial effect by photothermal treatment under near-infrared light irradiation, but also control release of SS31 in response to ROS. This F127DA/HAMA-MPDA@SS31 (FH-M@S) hydrogel has characteristics of adhesion, superior biocompatibility and mechanical properties which can adapt to irregular wounds at different body sites and provide sustained release of MPDA@SS31 (M@S) NPs. In addition, in a diabetic rat full thickness skin defect model, the FH-M@S hydrogel promoted macrophage M2 polarization, collagen deposition, neovascularization and wound healing. Therefore, the FH-M@S hydrogel exhibits promising therapeutic potential for skin regeneration.

Identifying therapeutic biomarkers of zoledronic acid by metabolomics.

Zoledronic acid (ZOL) is a potent antiresorptive agent that increases bone mineral density (BMD) and reduces fracture risk in postmenopausal osteoporosis (PMOP). The anti-osteoporotic effect of ZOL is determined by annual BMD measurement. In most cases, bone turnover markers function as early indicators of therapeutic response, but they fail to reflect long-term effects. We used untargeted metabolomics to characterize time-dependent metabolic shifts in response to ZOL and to screen potential therapeutic markers. In addition, bone marrow RNA-seq was performed to support plasma metabolic profiling. Sixty rats were assigned to sham-operated group (SHAM, n = 21) and ovariectomy group (OVX, n = 39) and received sham operation or bilateral ovariectomy, respectively. After modeling and verification, rats in the OVX group were further divided into normal saline group (NS, n = 15) and ZOL group (ZA, n = 18). Three doses of 100 µg/kg ZOL were administrated to the ZA group every 2 weeks to simulate 3-year ZOL therapy in PMOP. An equal volume of saline was administered to the SHAM and NS groups. Plasma samples were collected at five time points for metabolic profiling. At the end of the study, selected rats were euthanatized for bone marrow RNA-seq. A total number of 163 compound were identified as differential metabolites between the ZA and NS groups, including mevalonate, a critical molecule in target pathway of ZOL. In addition, prolyl hydroxyproline (PHP), leucyl hydroxyproline (LHP), 4-vinylphenol sulfate (4-VPS) were identified as differential metabolites throughout the study. Moreover, 4-VPS negatively correlated with increased vertebral BMD after ZOL administration as time-series analysis revealed. Bone marrow RNA-seq showed that the PI3K-AKT signaling pathway was significantly associated with ZOL-mediated changes in expression (adjusted-p = 0.018). In conclusion, mevalonate, PHP, LHP, and 4-VPS are candidate therapeutic markers of ZOL. The pharmacological effect of ZOL likely occurs through inhibition of the PI3K-AKT signaling pathway.

Polymeric coating on ß-TCP scaffolds provides immobilization of small extracellular vesicles with surface-functionalization and ZEB1-Loading for bone defect repair in diabetes mellitus.

Repair of critical-size bone defects in patients with diabetes mellitus (DM) has always been a challenge in clinical treatment. The process of bone defect regeneration can be impaired by underlying diseases including DM, but the mechanism remains unclear. In bone tissue engineering, the integration of bionic coatings and bioactive components into basic scaffolds are common function-enhancing strategies. Small extracellular vesicles (sEVs) have been applied for cell-free tissue regeneration in the last few years. We previously reported that sEVs have flexible and easily-extensible potential, through modular design and engineering modification. The impairment of CD31hiendomucinhi endothelial cells (ECs) whose function is coupling of osteogenesis and angiogenesis, is considered an important contributor to diabetic bone osteopathy, and ZEB1, which is highly expressed in CD31hiendomucinhi ECs, promotes angiogenesis-dependent bone formation. Thus we believe these ECs hold much promise for use in bone regeneration. In addition, c(RGDfC) has been reported to be a highly-effective peptide targeting αvß3, which is highly expressed in the bone microenvironment. In this study, we developed a hyaluronic acid (HA)/poly-L-lysine (PLL) layer-by-layer (LbL) self-assembly coating on ß-TCP (ß-tricalcium phosphate) scaffolds providing immobilization of modularized engineered sEVs (with c(RGDfC) surface functionalization and ZEB1 loading) to facilitate bone defect regeneration under DM conditions. RNA-seq was used to explore possible molecular mechanisms, and the therapeutic effects of bone regeneration were systematically evaluated in vitro and in vivo. Our data demonstrated that this strategy could be very effective in promoting the repair of diabetic bone defects, by enhancing angiogenesis, promoting osteogenesis and inhibiting osteoclast formation.

Small extracellular vesicles in combination with sleep-related circRNA3503: A targeted therapeutic agent with injectable thermosensitive hydrogel to prevent osteoarthritis.

Osteoarthritis (OA), characterized by chondrocyte apoptosis and disturbance of the balance between catabolism and anabolism of the extracellular matrix (ECM), is the most common age-related degenerative joint disease worldwide. As sleep has been found to be beneficial for cartilage repair, and circular RNAs (circRNAs) have been demonstrated to be involved in the pathogenesis of OA, we performed RNA sequencing (RNA-seq), and found circRNA3503 was significantly increased after melatonin (MT)-induced cell sleep. Upregulation of circRNA3503 expression completely rescued the effects of interleukin-1ß (IL-1ß), which was used to simulate OA, on apoptosis, ECM degradation- and synthesis-related genes. Mechanistically, circRNA3503 acted as a sponge of hsa-miR-181c-3p and hsa-let-7b-3p. Moreover, as we previously showed that small extracellular vesicles (sEVs) derived from synovium mesenchymal stem cells (SMSCs) can not only successfully deliver nucleic acids to chondrocytes, but also effectively promote chondrocyte proliferation and migration, we assessed the feasibility of sEVs in combination with sleep-related circRNA3503 as an OA therapy. We successfully produced and isolated circRNA3503-loaded sEVs (circRNA3503-OE-sEVs) from SMSCs. Then, poly(D,l-lactide)-b-poly(ethylene glycol)-b-poly(D,l-lactide) (PDLLA-PEG-PDLLA, PLEL) triblock copolymer gels were used as carriers of sEVs. Through in vivo and in vitro experiments, PLEL@circRNA3503-OE-sEVs were shown to be a highly-effective therapeutic strategy to prevent OA progression. Through multiple pathways, circRNA3503-OE-sEVs alleviated inflammation-induced apoptosis and the imbalance between ECM synthesis and ECM degradation by acting as a sponge of hsa-miR-181c-3p and hsa-let-7b-3p. In addition, circRNA3503-OE-sEVs promoted chondrocyte renewal to alleviate the progressive loss of chondrocytes. Our results highlight the potential of PLEL@circRNA3503-OE-sEVs for preventing OA progression.

EWSAT1 Acts in Concert with Exosomes in Osteosarcoma Progression and Tumor-Induced Angiogenesis: The "Double Stacking Effect".

The prognosis for osteosarcoma (OS) continues to be unsatisfactory due to tumor recurrence as a result of metastasis and drug resistance. Several studies have shown that Ewing sarcoma associated transcript 1 (EWSAT1) plays an important role in the progression of OS. Exosomes (Exos) act as important carriers in intercellular communication and play an important role in the tumor microenvironment, especially in tumor-induced angiogenesis. Nonetheless, the specific mechanism via which EWSAT1 and Exos regulate OS progression is unknown, and whether they can be effective therapeutic targets also requires verification. Hence, in this study, it is aimed to investigate the mechanisms of action of EWSAT1 and Exos. EWSAT1 significantly promotes proliferation, migration, colony formation, and survival of OS. EWSAT1 regulates OS-induced angiogenesis via two mechanisms, called the "double stacking effect," which is a combination of the increase in sensitivity/reactivity of vascular endothelial cells triggered by Exos-carrying EWSAT1, and the EWSAT1-induced increase in angiogenic factor secretion. In vivo experiments further validates the "double stacking effect" and shows that EWSAT1-KD effectively inhibits tumor growth in OS. The above observations indicate that EWSAT1 can be used as not only a potential diagnostic and prognostic marker, but also as a precise therapeutic target for OS.

Valproic acid prevents glucocorticoid­induced osteonecrosis of the femoral head of rats.

Glucocorticoids (GCs) are the most common cause of atraumatic osteonecrosis of the femoral head (ONFH) because their effect compromises the osteogenic capability of bone marrow­derived mesenchymal stem cells (BMSCs). Valproic acid (VPA) is a widely used anti­epileptic and anti­convulsant drug. Previous studies have reported that VPA promotes osteogenic differentiation of MSCs in vitro and osteogenesis in vivo as a histone deacetylase (HDAC) inhibitor. The purpose of the present study was to investigate the efficacy of VPA as a precautionary treatment of ONFH after GC treatment in rats. In vitro, the effect of VPA, dexamethasone or a combination treatment of the two on the proliferation and osteogenic differentiation of human BMSCs was assessed using a Cell Counting Kit­8 and apoptosis assays, and by measuring the expression of proteins associated with osteogenesis. In vivo, a GC­induced ONFH model was established in rats and VPA was added during GC treatment to investigate the preventive effect of VPA against ONFH. Rat BMSCs were also extracted to investigate the osteogenic capacity. The results of micro­computed tomography scanning, angiography of the femoral head and histological and immunohistochemical analyses indicated that 11 of 15 rats induced with methylprednisolone (MP) presented with ONFH, while only 2 of 15 rats treated with a combination of MP and VPA developed ONFH. VPA produced beneficial effects on subchondral bone trabeculae in the femoral head with significant preservation of bone volume and blood supply, as well as improved osteogenic capability of BMSCs compared with those in rats treated with GC alone. In conclusion, VPA attenuated the inhibitory effect of GC on BMSC proliferation and osteogenesis by inhibiting apoptosis and elevating the expression of proteins associated with osteogenesis, which may contribute to the prevention of GC­induced ONFH in rats.

Association between SNPs and haplotypes in the METTL21C gene and peak bone mineral density and body composition in Chinese male nuclear families.

The methyltransferase-like 21C gene (METTL21C), which is mainly expressed in muscle, can promote the differentiation of myoblasts to myotubes and reduce glucocorticoid-induced apoptosis of osteocytes. The purpose of this study was to explore the association between single nucleotide polymorphisms of METTL21C and peak bone mineral density (BMD), body mass index, total fat mass (TFM), and total lean mass (TLM) in Chinese young men. Fifteen tagging single nucleotide polymorphisms were genotyped, and haplotype blocks were derived in 400 Chinese male nuclear families. The peak BMD of the lumbar and hip, TFM, and TLM were measured by dual-energy X-ray absorptiometry. The association analyses were performed by a quantitative transmission disequilibrium test. Both TLM and TFM had a significant positive effect on peak BMD, but the positive regulation of TLM was stronger than that of TFM. After 1000 permutations, significant within-family associations were found between rs9585961 and lumbar spine BMD and femoral neck BMD, rs9518810 and femoral neck BMD, and rs599976 and body mass index, TFM, and percentage fat mass (all P < 0.05). The association analyses with haplotypes showed that haplotype AG in block 1 was significantly associated with TFM (P = 0.031) and haplotype CAG in block 2 was significantly associated with lumbar spine BMD (P = 0.020). Our study, for the first time, demonstrates that the polymorphisms and haplotypes of METTL21C contribute to the peak BMD and TFM in Chinese males, which suggests that as a quantitative trait locus with potential pleiotropy it may have an influence on osteoporosis and obesity.

Sirt1 is involved in decreased bone formation in aged apolipoprotein E-deficient mice.

AIM: Apolipoprotein E (ApoE) plays an important role in the transport and metabolism of lipids. Recent studies show that bone mass is increased in young apoE(-/-) mice. In this study we investigated the bone phenotype and metabolism in aged apoE(-/-) mice. METHODS: Femurs and tibias were collected from 18- and 72-week-old apoE(-/-) mice and their age-matched wild-type (WT) littermates, and examined using micro-CT and histological analysis. Serum levels of total cholesterol, oxidized low-density lipoprotein (ox-LDL) and bone turnover markers were measured. Cultured bone mesenchymal stem cells (BMSCs) from tibias and femurs of 18-week-old apoE(-/-) mice were used in experiments in vitro. The expression levels of Sirt1 and Runx2 in bone tissue and BMSCs were measured using RT-PCR and Western blot analysis. RESULTS: Compared with age-matched WT littermates, young apoE(-/-) mice exhibited high bone mass with increased bone formation, accompanied by higher serum levels of bone turnover markers OCN and TRAP5b, and higher expression levels of Sirt1, Runx2, ALP and OCN in bone tissue. In contrast, aged apoE(-/-) mice showed reduced bone formation and lower bone mass relative to age-matched WT mice, accompanied by lower serum OCN levels, and markedly reduced expression levels of Sirt1, Runx2, ALP and OCN in bone tissue. After BMSCs were exposed to ox-LDL (20 µg/mL), the expression of Sirt1 and Runx2 proteins was significantly increased at 12 h, and then decreased at 72 h. Treatment with the Sirt1 inhibitor EX527 (10 µmol/L) suppressed the expression of Runx2, ALP and OCN in BMSCs. CONCLUSION: In contrast to young apoE(-/-) mice, aged apoE(-/-) mice showe lower bone mass than age-matched WT mice. Long-lasting exposure to ox-LDL decreases the expression of Sirt1 and Runx2 in BMSCs, which may explain the decreased bone formation in aged apoE(-/-) mice.