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ABSTRACTWhether long non-coding RNA Mir-99a-Let-7c Cluster Host Gene (LncRNA MIR99AHG) is involved in osteoporosis (OP) remains vague, so we hereby center on its implication. Old C57BL/6J mice were injected with the silencing lentivirus of MIR99AHG and subjected to microCT analysis and immunohistochemistry on osteogenic cells. The osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) with or without transfection was determined by alkaline phosphatase (ALP) and Alizarin Red S staining. Total N(6)-methyladenosine (m6A) on the bone marrow mesenchymal stem cells (BMSCs) was quantified. The potential methylation site and the complementary binding sites with candidate microRNA (miR) were predicted via bioinformatic analyses, with the latter being confirmed via dual-luciferase reporter, RNA immunoprecipitation and RNA pull-down assays. Quantitative real-time PCR and Western blot were used for quantification assays. MIR99AHG was decreased during the osteogenic differentiation of BMSCs, where increased Osterix (OSX), Collagen, Type I, Alpha 1 (Col1A1), Osteocalcin (OCN) and RUNX Family Transcription Factor 2 (RUNX2) as well as more color-stained areas were found. Also, silencing MIR99AHG relieved the OP in mice and reduced the loss of osteogenic cells. M6A methylation in undifferentiated BMSCs was low and MIR99AHG overexpression abolished the effects of overexpressed METTL3 on promoting osteogenic differentiation. MiR-4660, which was downregulated in BMSCs without differentiation but increased during osteogenic differentiation, could bind with MIR99AHG. Furthermore, miR-4660 promoted osteogenic differentiation and reversed the effects of overexpressed MIR99AHG. The present study demonstrated that METTL3-mediated LncRNA MIR99AHG methylation enhanced the osteogenic differentiation of BMSCs via targeting miR-4660.

As the demand for green energy with high efficiency and low carbon dioxide (CO2) emissions has increased, solid oxide fuel cells (SOFCs) have been intensively developed in recent years. Integrated gasification fuel cells (IGFCs) in particular show potential for large-scale power generation to further increase system efficiency. Thus, for commercial application of IGFCs, it is important to design reliable multi-stacks for large systems that show long-term stability and practical fuel gas for application to industrial equipment. In this work, a test rig (of a 5 kW SOFC system, with syngas from industrial gasifiers as fuel) was fabricated and subjected to long-term tests under high fuel utilization to investigate its performance. The maximum steady output power of the system was 5700 W using hydrogen and 5660 W using syngas and the maximum steady electrical efficiency was 61.24% while the fuel utilization efficiency was 89.25%. The test lasted for more than 500 h as the fuel utilization efficiency was larger than 83%. The performances of each stack tower were almost identical at both the initial stage and after long-term operation. After 500 h operation, the performances of the stack towers decreased only slightly under lower current and showed almost no change under high current. These results demonstrate the reliability of the multi-stack design and the prospect of this SOFC power-generation system for further enlarging its application in a MWthdemonstration.

ABSTRACTPrevious research has identified N-myc downstream-regulated gene 2 (NDRG2) as one of the differentially expressed genes common to rat models of osteoarthritis (OA) and human OA. The purpose of this study was to investigate the role of NDRG2 in OA. In this study, an in vitro OA model was constructed by challenging ATDC5 chondrocytes with 10 ng/ml IL-1β. After transfection of pcDNA3.1(+)/NDRG2, qPCR and western blot were performed to assay NDRG2 expression. The analyses of cell viability, apoptosis and inflammatory molecule expression were employed respectively by CCK-8, TUNEL and ELISA. The protein expression related to apoptosis, inflammation or extracellular matrix (ECM) degradation was detected by western blot. The binding of Krüppel-like factor 5 (KLF5) to NDRG2 promoter was verified by means of dual-luciferase reporter assay. After overexpression of both NDRG2 and KLF5 in IL-1β-stimulated ATDC5 chondrocytes, corresponding assays were performed to examine cell viability, apoptosis, inflammatory response and ECM degradation. In ATDC5 chondrocytes challenged with IL-1β, NDRG2 expression was much lower than that in the control group, whereas it’s overexpression helped restored cell viability and reduce cell apoptosis, inflammatory response and ECM degradation. It was also observed that KLF5 expression was decreased in IL-1β-stimulated ATDC5 chondrocytes, and that KLF5 bound to the NDRG2 promoter. Importantly, overexpressing KLF5 could reverse the protective effect of NDRG2 overexpression on IL-1β-stimulated ATDC5. Overall, NDRG2 could be transcriptionally regulated by transcription factor KLF5 and may play a protective role against chondrocyte the inflammatory response and ECM degradation in OA.

Surface modification of medical implants is considered as an effective method to improve cellular behaviors and the integration of tissues with materials. Titanium (Ti)-based materials with four different micro/nano-structures and compositions were prepared by acid etching, electrochemical anodization and alkali-heat treatment. The surface morphologies and compositions of the different surface-modified Ti materials were characterized by field-emission scanning electron microscopy (FE-SEM), atomic force microscopy (AFM) and X-ray diffraction (XRD). The effects of the micro/nano structured and compositions of the surfaces on cellular responses were investigated in vitroby observing the morphology, adhesion, proliferation and osteogenic differentiation of osteoblasts. To further investigate the underlying mechanisms, an RT-PCR assay was performed to analyze the expression levels of cell adhesion-related genes. Our results indicated that the nanosized structure and anatase composition could promote the adhesion and proliferation of MC3T3-E1 pre-osteoblast, as well as alkaline phosphatase activity and extracellular matrix mineralization via the integrin-FAK signaling pathway. Taken together, our innovation presented in this work demonstrated that the surface nano-structure design and composition of biomedical implants can be modified of for future orthopaedic applications.

Computationally-efficient optimal energy-management strategies are required for the optimal design of hybrid electric vehicles with respect to fuel economy and other criteria. In this paper, the novel Selective Hamiltonian Minimization (SHM) technique is introduced. Based on Pontryagin’s Minimum Principle, the SHM is developed by explicitly calculating possible optimal control modes, and then selecting the one that minimizes the Hamiltonian function. Parametric analytical models of powertrain components are developed to compute Hamiltonians in closed form. Engine and battery models are further expressed in terms of their main design parameters and technologies thanks to a statistical analysis over several sample components. An example of design optimization is presented to illustrate the effectiveness of the proposed approach.

Wear particle-induced osteolysis is the leading cause of aseptic loosening, which is the most common reason for THA (total hip arthroplasty) failure and revision surgery. Although existing studies suggest that osteoblast apoptosis induced by wear debris is involved in aseptic loosening, the underlying mechanism linking wear particles to osteoblast apoptosis remains almost totally unknown. In the present study, we investigated the effect of autophagy on osteoblast apoptosis induced by CoCrMo metal particles (CoPs) in vitro and in a calvarial resorption animal model. Our study demonstrated that CoPs stimulated autophagy in osteoblasts and PIO (particle-induced osteolysis) animal models. Both autophagy inhibitor 3-MA (3-methyladenine) and siRNAof Atg5could dramatically reduce CoPs-induced apoptosis in osteoblasts. Further, inhibition of autophagy with 3-MA ameliorated the severity of osteolysis in PIO animal models. Moreover, 3-MA also prevented osteoblast apoptosis in an antiautophagic way when tested in PIO model. Collectively, these results suggest that autophagy plays a key role in CoPs-induced osteolysis and that targeting autophagy-related pathways may represent a potential therapeutic approach for treating particle-induced peri-implant osteolysis.

Osteosarcoma is a primary malignant bone tumour with high morbidity that occurs mainly in children and adolescents. While the molecular basis of osteosarcoma has received considerable attention, the cellular and molecular mechanisms underlying pre- and postoperative metastasis formation and the development of chemoresistance remain unclear. MicroRNAs (miRNAs), a class of 22-nucleotide noncoding RNAs, have emerged as critical components of gene-regulatory networks controlling numerous important pathophysiological processes, including the initiation and progression of cancers. Studies on miRNAs have opened new avenues for both the diagnosis and treatment of cancer. This review discusses the roles of miRNAs in osteosarcoma and their potential applications for the diagnosis, prognosis and treatment of this malignancy. As a rapidly evolving field of basic and biomedical science, miRNA research will have a revolutionary impact on the management of osteosarcoma.

Bone tissue engineering has generated promising results in bone defect repair, but is limited by the inherently poor nutrient supply to nonvascularized tissue-engineered bone grafts. In this study, we investigated the delivery of vascular endothelial growth factor (VEGF) and the effect on in vitrocultured human umbilical vein endothelial cells (HUVECs), in an attempt to provide experimental basis for promoting the vascularization of tissue-engineered bone grafts. A mechanical stimulator was developed to generate a periodic compressive load analogous to goat locomotor characteristics, simulating the mechanical stimulation applied on the fracture ends of the load-bearing bone. Poly-l-lysine–coated VEGF/alginate microspheres were combined with demineralized bone matrices into composites, and the in vitrorelease of VEGF from these composites was evaluated under periodic compression. The effect of the release media on HUVECs was also investigated. Compression slightly accelerated VEGF release at the early stage (<11 days) compared with noncompressed composites, although the release profiles of the two composite groups were generally similar. The released VEGF promoted HUVEC proliferation. In addition, the periodic compression applied on composites containing both HUVECs and VEGF/alginate microspheres promoted the expression of matrix metalloproteinases-2/9 in HUVECs. This study provides a model for investigating VEGF release under simulated in vivobiomechanical conditions and without the disadvantage of the rapid degradation of VEGF in in vivoinvestigation of VEGF release. The results also provide important guidelines for future in vivoexperiments.

Recently, bone marrow–derived mesenchymal stem cells (MSCs) have been paid more attention for cartilage regeneration. This study evaluated the potential of using MSCs seeded in plasmid transforming growth factor β1 (pTGF-β1)–activated three-dimensional chitosan/gelatin scaffolds for improving cartilage repair in vivo. Significant cell proliferation and transforming growth factor β1 protein expression were observed in vitroin pTGF-β1-activated scaffolds. Transforming growth factor β1–activated scaffolds showed high collagen type II and aggrecan expression and low collagen type I expression during in vitrocultivation. MSC-based pTGF-β1-activated scaffolds also exhibited cartilage histology with high secretion of collagen type II in vitrounder the stimulation of pTGF-β1. In rabbits with full-thickness cartilage defects, the implantation of MSC-based pTGF-β1-activated scaffolds not only significantly promoted chondrogenic differentiation of MSCs and hyalin-like cartilage matrix synthesis, but also remarkably improved the overall repair of rabbit cartilage defects and exhibited favorable tissue integrity at 10 weeks postsurgery. These results suggest that MSC-based localized pTGF-β1-activated scaffolds have potential applications for in vivocartilage repair.

Biotin is one of the water-soluble B-complex group of vitamins. Recent studies have found that the relative protein expression of BMP2, BSP and OPG in MC3T3-E1 cells is prominent after 14 days of co-culture with biotin film, especially for BMP2. It is also found that the rapid degradation of biotin film in vivo limits its application value. In this work, magnesium-doped hydroxyapatite (MgHA) film can form a porous network structure as a biological sustained-release film. Therefore, the multilayer (MgHA|biotin|MgHA|biotin) film was prepared by pulsed laser assisted electron beam deposition technique. The morphology, structure and properties of biotin film and multilayer film were analyzed and characterized. Also, the osteogenic effect of biotin film and multilayer film was evaluated after implantation into the femoral bone marrow cavity of SD rats. The results of micro-CT scan and 3D reconstruction showed that there were a large number of trabecular bones around the multilayer film, which was superior to biotin film in osteogenesis. Hematoxylin-eosin staining showed cancellous bone structure and intact bone marrow structure around the multilayer film, and the newly formed bone became lamellar. Masson-trichromatic staining revealed abundant osteoid and braided bone formation around the multilayer film. In conclusion, MgHA sustained release film can realize the continuous release of bioactive drugs, which provides a new route to accelerate the repair of bone defects.

As a promising solution to the reduction of fuel consumption and CO2emissions in road transport sector, hybrid electric powertrains are confronted with complex control techniques for the evaluation of the minimal fuel consumption, particularly the excessively long computation time of the design-parameter optimization in the powertrain's early design stage. In this work, a novel and simple GRaphical-Analysis-Based method of fuel Energy Consumption Optimization (GRAB-ECO) is developed to estimate the minimal fuel consumption for parallel hybrid electric powertrains in light- and heavy-duty application. Based on the power ratio between powertrain's power demand and the most efficient engine power, GRAB-ECO maximizes the average operating efficiency of the internal combustion engine by shifting operating points to the most efficient conditions, or by eliminating the engine operation from poorly efficient operating points to pure electric vehicle operation. A turning point is found to meet the requirement of the final state of energy of the battery, which is charge-sustaining mode in this study. The GRAB-ECO was tested with both light- and heavy-duty parallel hybrid electric vehicles, and validated in terms of the minimal fuel consumption and the computation time. Results show that GRAB-ECO accurately approximates the minimal fuel consumption with less than 6% of errors for both light- and heavy-duty parallel hybrid electric powertrains. Meanwhile, GRAB-ECO reduces computation time by orders of magnitude compared with PMP-based (Pontryagin's Minimum Principle) approaches.

Purpose: Pneumatic tourniquets are used in total knee arthroplasty (TKA) for surgical field visualization and improved cementation; however, their use is controversial. This study aimed to assess the effects of tourniquet application on enhanced recovery post-TKA.Methods: A prospective randomized single-blinded trial assessed tourniquet’s effects on postoperative pain, swelling, and early outcome in TKA. One-hundred and two patients with knee osteoarthritis were randomized to full-course (FC) and second half-course (SHC) application (n= 51/group). Tumor necrosis factor-alpha (TNF-α), C-C motif chemokine ligand 2(CCL-2), pentraxin-3 (PTX-3), prostaglandin E-2 (PGE-2), superoxide dismutase-1 (SOD-1), and myoglobin (Mb) were assessed by enzyme-linked immunosorbent assay, while the visual analog scale (VAS), range of motion (ROM), and thigh circumference growth rate were recorded.Results: Average tourniquet duration significantly differed between the SHC (37.5 ± 5.1 min) and FC (66.4 ± 7.2 min) groups (p< 0.01); VAS and thigh circumference growth rate in the SHC group were much lower compared with the FC group, while ROM was higher within 48 h of tourniquet removal (p< 0.01). Blood TNF-α, PTX3, CCL2, PGE2, SOD-1, and Mb were lower in the SHC group than the FC group (p< 0.01). Additionally, intraoperative blood loss was significantly elevated in the SHC group than the FC group (p< 0.01), with lower postoperative blood loss in the drain (p= 0.001). Postoperative drainage volume was reduced in the SHC group compared with the FC group (p< 0.01); five and two patients in the FC and SHC groups required blood transfusion, respectively (p= 0.025). Hospital stay tended to be shorter in the SHC group (p= 0.023), and no tourniquet-related complications were recorded.Conclusion: Improved therapeutic outcome was observed in the SHC group, indicating patients should routinely undergo TKA with SHC tourniquet application.