IHUP: An Integrated High-Throughput Universal Phenotyping Software Platform to Accelerate Unmanned-Aerial-Vehicle-Based Field Plant Phenotypic Data Extraction and Analysis.

With the threshold for crop growth data collection having been markedly decreased by sensor miniaturization and cost reduction, unmanned aerial vehicle (UAV)-based low-altitude remote sensing has shown remarkable advantages in field phenotyping experiments. However, the requirement of interdisciplinary knowledge and the complexity of the workflow have seriously hindered researchers from extracting plot-level phenotypic data from multisource and multitemporal UAV images. To address these challenges, we developed the Integrated High-Throughput Universal Phenotyping (IHUP) software as a data producer and study accelerator that included 4 functional modules: preprocessing, data extraction, data management, and data analysis. Data extraction and analysis requiring complex and multidisciplinary knowledge were simplified through integrated and automated processing. Within a graphical user interface, users can compute image feature information, structural traits, and vegetation indices (VIs), which are indicators of morphological and biochemical traits, in an integrated and high-throughput manner. To fulfill data requirements for different crops, extraction methods such as VI calculation formulae can be customized. To demonstrate and test the composition and performance of the software, we conducted case-related rice drought phenotype monitoring experiments. In combination with a rice leaf rolling score predictive model, leaf rolling score, plant height, VIs, fresh weight, and drought weight were efficiently extracted from multiphase continuous monitoring data. Despite the significant impact of image processing during plot clipping on processing efficiency, the software can extract traits from approximately 500 plots/min in most application cases. The software offers a user-friendly graphical user interface and interfaces for customizing or integrating various feature extraction algorithms, thereby significantly reducing barriers for nonexperts. It holds the promise of significantly accelerating data production in UAV phenotyping experiments.

Bioinspired Hybrid Nanostructured PEEK Implant with Enhanced Antibacterial and Anti-inflammatory Synergy.

Implant-associated infections and excessive immune responses are two major postsurgical issues for successful implantation. However, conventional strategies including antibiotic treatment and inflammatory regulation are always compromised due to the comodification of various biochemical agents and instances of functional interference. It is imperative to provide implant surfaces with satisfactory antibacterial and anti-inflammatory properties. Here, a dual-effect nanostructured polyetheretherketone (PEEK) surface (NP@PDA/Zn) with bionic mechano-bactericidal nanopillars and immobilized immunomodulatory Zn2+ is designed. The constructed hybrid nanopillars display remarkable antibacterial performance against Gram-negative and Gram-positive strains through the synergy of physical and chemical bactericidal effects imposed by nanopillars and Zn2+. Meanwhile, the immunoregulatory property is evaluated through the investigation of macrophage polarization both in vitro and in vivo, and the results reveal that NP@PDA/Zn could downregulate the expression of M1-related cytokines and decrease the M1 macrophage recruitment to lower the inflammatory response. Notably, the surface exhibited exceptional biocompatibility with discerning biocidal activity between bacterial and mammalian cells and antioxidant performance that effectively scavenges ROS, minimizing potential cytotoxicity. Taken together, NP@PDA/Zn presents a convenient and promising strategy of combining synergistic bactericidal activity and inflammatory regulation without any mutual interference, which can support the development of multifunctional implant-associated materials.

Cortistatin prevents glucocorticoid-associated osteonecrosis of the femoral head via the GHSR1a/Akt pathway.

Long-term use of glucocorticoids (GCs) is known to be a predominant cause of osteonecrosis of the femoral head (ONFH). Moreover, GCs can mediate apoptosis of various cell types by exaggerating oxidative stress. We have previously found that Cortistatin (CST) antagonizes oxidative stress and improves cell apoptosis in several conditions. In this study, we detected that the CST expression levels were diminished in patients with ONFH compared with femoral neck fracture (FNF). In addition, a GC-induced rat ONFH model was established, which impaired bone quality in the femoral head. Then, administration of CST attenuated these ONFH phenotypes. Furthermore, osteoblast and endothelial cells were cultured and stimulated with dexamethasone (Dex) in the presence or absence of recombinant CST. As a result, Dex induced impaired anabolic metabolism of osteoblasts and suppressed tube formation in endothelial cells, while additional treatment with CST reversed this damage to the cells. Moreover, blocking GHSR1a, a well-accepted receptor of CST, or blocking the AKT signaling pathway largely abolished the protective function of CST in Dex-induced disorder of the cells. Taken together, we indicate that CST has the capability to prevent GC-induced apoptosis and metabolic disorder of osteoblasts in the pathogenesis of ONFH via the GHSR1a/AKT signaling pathway.

M2 macrophage-derived exosomal miR-486-5p influences the differentiation potential of bone marrow mesenchymal stem cells and osteoporosis.

BACKGROUND: An imbalance between osteogenesis and adipogenesis in bone marrow mesenchymal stem cells (BMMSCs) can cause osteoporosis. Macrophage-derived exosomes (MD-Exos) and microRNAs (miRNAs) enriched in exosomes participate in the differentiation of BMMSCs. METHODS: Bioinformatics methods were used to analyze differentially expressed miRNAs. We cocultured M2 macrophages and BMMSCs to examine the biological function of exosomal microRNA-486-5p (miR-486-5p) on BMMSCs differentiation. Gain-of-function experiments related to osteogenesis were designed to investigate the effects of exosomes carrying miR-486-5p on an ovariectomized (OVX) mice model and the direct impact of miR-486-5p on BMMSCs. A dual luciferase experiment was performed to demonstrate the target gene of miR-486-5p. RESULTS: Bioinformatics analysis identified high expression of miRNA-486 in M2 macrophage-derived exosomes (M2D-Exos). The in vitro results demonstrated that M2 macrophage-derived exosomal miR-486-5p enhanced osteogenic capacity but inhibited the adipogenesis of BMMSCs. The direct effect of miR-486-5p on BMMSCs showed the same effects. Animal experiments revealed that exosomal miR-486-5p rescued bone loss of OVX mice. SMAD2 was characterized as a target gene of miR-486-5p. Pathway analysis showed that M2 macrophage-derived exosomal miR-486-5p stimulated osteogenic differentiation via the TGF-ß/SMAD2 signalling pathway. CONCLUSIONS: Taken together, M2 macrophage-derived exosomal miR-486-5p influences the differentiation potential of BMMSCs through the miR-486-5p/SMAD2/TGF-ß signalling pathway and osteoporosis.

WTAP-mediated m6A modification modulates bone marrow mesenchymal stem cells differentiation potential and osteoporosis.

An imbalance in the differentiation potential of bone marrow mesenchymal stem cells (BMSCs) is an important pathogenic mechanism underlying osteoporosis (OP). N6-methyladenosine (m6A) is the most common post-transcriptional modification in eukaryotic cells. The role of the Wilms' tumor 1-associated protein (WTAP), a member of the m6A functional protein family, in regulating BMSCs differentiation remains unknown. We used patient-derived and mouse model-derived samples, qRT-PCR, western blot assays, ALP activity assay, ALP, and Alizarin Red staining to determine the changes in mRNA and protein levels of genes and proteins associated with BMSCs differentiation. Histological analysis and micro-CT were used to evaluate developmental changes in the bone. The results determined that WTAP promoted osteogenic differentiation and inhibited adipogenic differentiation of BMSCs. We used co-immunoprecipitation (co-IP), RNA immunoprecipitation (RIP), methylated RNA immunoprecipitation (MeRIP), RNA pulldown, and dual-luciferase assay to explore the direct mechanism. Mechanistically, the expression of WTAP increased during osteogenic differentiation and significantly promoted pri-miR-181a and pri-miR-181c methylation, which was recognized by YTHDC1, and increased the maturation to miR-181a and miR-181c. MiR-181a and miR-181c inhibited the mRNA expression of SFRP1, promoting the osteogenic differentiation of BMSCs. Our results demonstrated that the WTAP/YTHDC1/miR-181a and miR-181c/SFRP1 axis regulated the differentiation fate of BMSCs, suggesting that it might be a potential therapeutic target for osteoporosis.

WTAP-Mediated m6A RNA Methylation Regulates the Differentiation of Bone Marrow Mesenchymal Stem Cells via the miR-29b-3p/HDAC4 Axis.

N6-methyladenosine (m6A) methylation, a well-known modification with new epigenetic functions, has been reported to participate in the progression of osteoporosis (OP), providing novel insights into the pathogenesis of OP. However, as the key component of m6A methylation, Wilms tumor 1-associated protein (WTAP) has not been studied in OP. Here we explored the biological role and underlying mechanism of WTAP in OP and the differentiation of bone marrow mesenchymal stem cells (BMMSCs). We demonstrated that WTAP was expressed at low levels in bone specimens from patients with OP and OVX mice. Functionally, WTAP promoted osteogenic differentiation and inhibited adipogenic differentiation of BMMSCs in vitro and in vivo. In addition, microRNA-29b-3p (miR-29b-3p) was identified as a downstream target of WTAP. M6A modifications regulated by WTAP led to increased miR-29b-3p expression. WTAP interacted with the microprocessor protein DGCR8 and accelerated the maturation of pri-miR-29b-3p in an m6A-dependent manner. Target prediction and dual-luciferase reporter assays identified the direct binding sites of miR-29b-3p with histone deacetylase 4 (HDAC4). WTAP-mediated m6A modification promoted osteogenic differentiation and inhibited adipogenic differentiation of BMMSCs through the miR-29b-3p/HDAC4 axis. Furthermore, WTAP-mediated m6A methylation negatively regulates osteoclast differentiation. Collectively, our study first identified a critical role of WTAP-mediated m6A methylation in BMMSC differentiation and highlighted WTAP as a potential therapeutic target for OP treatment.

Arginine methylation of PPP1CA by CARM1 regulates glucose metabolism and affects osteogenic differentiation and osteoclastic differentiation.

BACKGROUND: The imbalance between osteoblasts and osteoclasts may lead to osteoporosis. Osteoblasts and osteoclasts have different energy requirements, with aerobic glycolysis being the prominent metabolic feature of osteoblasts, while osteoclast differentiation and fusion are driven by oxidative phosphorylation. METHODS: By polymerase chain reaction as well as Western blotting, we assayed coactivator-associated arginine methyltransferase 1 (CARM1) expression in bone tissue, the mouse precranial osteoblast cell line MC3T3-E1 and the mouse monocyte macrophage leukaemia cell line RAW264.7, and expression of related genes during osteogenic differentiation and osteoclast differentiation. Using gene overexpression (lentivirus) and loss-of-function approach (CRISPR/Cas9-mediated knockout) in vitro, we examined whether CARM1 regulates osteogenic differentiation and osteoblast differentiation by metabolic regulation. Transcriptomic assays and metabolomic assays were used to find the mechanism of action of CARM1. Furthermore, in vitro methylation assays were applied to clarify the arginine methylation site of PPP1CA by CARM1. RESULTS: We discovered that CARM1 reprogrammed glucose metabolism in osteoblasts and osteoclasts from oxidative phosphorylation to aerobic glycolysis, thereby promoting osteogenic differentiation and inhibiting osteoclastic differentiation. In vivo experiments revealed that CARM1 significantly decreased bone loss in osteoporosis model mice. Mechanistically, CARM1 methylated R23 of PPP1CA, affected the dephosphorylation of AKT-T450 and AMPK-T172, and increased the activities of phosphofructokinase-1 and pructose-2,6-biphosphatase3, causing an up-regulation of glycolytic flux. At the same time, as a transcriptional coactivator, CARM1 regulated the expression of pyruvate dehydrogenase kinase 3, which resulted in the inhibition of pyruvate dehydrogenase activity and inhibition of the tricarboxylic acid cycle, leading to a subsequent decrease in the flux of oxidative phosphorylation. CONCLUSIONS: These findings reveal for the first time the mechanism by which CARM1 affects both osteogenesis and osteoclast differentiation through metabolic regulation, which may represent a new feasible treatment strategy for osteoporosis.

The Influence of Different Injection Hole Designs of Augmented Pedicle Screws on Bone Cement Leakage and Distribution Patterns in Osteoporotic Patients.

OBJECTIVE: To compare cement distribution and leakage for 2 bone cement-augmented screws with different designs of injection holes in patients and the impact of screw locations and bone mineral density (BMD) on the results. METHODS: This study recruited 40 patients who underwent instrumentation with cement-augmented screws. Screw holes of group A were 4 holes located in the distal one third of screws, while screw holes of group B were 6 holes located in distal, middle, and proximal sites. Postoperative computed tomography images were obtained to evaluate the rate and type of cement leakage and the distribution pattern of cement. The lateral or center position of screw tip, BMD, and T-score were also analyzed for their influence on the results. RESULTS: Of 192 screws, 80 (41.7%) exhibited cement leakage on postoperative computed tomography. The incidence of cement distribution in the posterior half and type B leakage in group B was significantly higher compared with group A. In group A, the probability of cement distribution in the posterior half was significantly increased when the screw was laterally inserted. For both groups, the higher incidence of cement distribution in the posterior half was correlated with lower BMD and T-score. CONCLUSIONS: Our results showed that screws with injection holes closer to the screw tip had higher incidences of distribution in the anterior half of the body and lower incidences of type B leakage. Patients with lower BMD and T-scores should be closely monitored, and a more centered position is recommended for screw insertion.

Exosomal MiR-199a-5p Inhibits Tumorigenesis and Angiogenesis by Targeting VEGFA in Osteosarcoma.

Background: Osteosarcoma (OS) is the most common primary bone malignancy in children and adolescents. microRNAs have been found to play a vital role in tumor angiogenesis. Here, we investigated the effects of miR-199a-5p on tumor growth and angiogenesis in osteosarcoma. Furthermore, the underlying molecular mechanisms and signaling pathways were explored. Methods: The datasets were extracted from the Gene Expression Omnibus and the differentially expressed miRNAs (DEmiRNAs) were screened out by the GEO2R online platform. The potential target genes were predicted using the miRTarBase database. The predicted target genes were further analyzed by Gene Ontology and pathway enrichment analysis and a regulatory network of DEmiRNAs and their target genes was constructed. In addition, the effects of osteosarcoma cell derived exosomal miR-199a-5p on the proliferation, migration and neovascularization of HUVECs were evaluated by conducting EdU assays, Transwell experiments and tube formation assays. A dual-luciferase reporter assay was performed to detect whether VEGFA was the direct target of miR-199a-5p. Furthermore, in vivo xenograft models were established to further investigate the intrinsic role of miR-199a-5p in osteosarcoma tumorigenesis and angiogenesis. Results: A total of 149 DE-miRNAs were screened out, including 136 upregulated miRNAs and 13 downregulated miRNAs in human osteosarcoma plasma samples compared with normal plasma samples. A total of 1313 target genes of the top three upregulated and downregulated miRNAs were predicted. In the PPI network, the top 10 hub nodes with higher degrees were identified as hub genes, such as TP53 and VEGFA. By constructing the miRNA-hub gene network, we found that most of hub genes could be potentially modulated by miR-663a, miR-199a-5p and miR-223-3p. In addition, we found that the expression level of miR-199a-5p in exosomes derived from osteosarcoma cells was remarkably higher than the osteosarcoma cells, and the exosomes derived from osteosarcoma cells were transported to HUVECs. Overexpression of miR-199a-5p could significantly inhibited HUVEC proliferation, migration and neovascularization, whereas downregulation of miR-199a-5p expression exerted the opposite effect. Moreover, the in vivo results verified that overexpression of miR-199a-5p in osteosarcoma cells could suppress the growth and angiogenesis of tumors. Conclusion: Our results demonstrated that miR-199a-5p could be transported from osteosarcoma cells to HUVECs through exosomes, subsequently targeting VEGFA and inhibiting the growth and angiogenesis of osteosarcoma. Therefore, miR-199a-5p may act as a biomarker in the diagnosis and treatment of osteosarcoma.

Bionic Tiger-Bone Powder Improves Bone Microstructure and Bone Biomechanical Strength of Ovariectomized Rats.

OBJECTIVE: To study the curative effect of bionic tiger-bone powder on osteoporosis in ovariectomized rats and investigate its mechanism. METHODS: Overall, a 120 female Wistar rats were randomly divided into Sham (sham-operated group), ovariectomy (OVX, ovariectomized group), TB (bionic tiger-bone powder treatment group after ovariectomy) and TB + VD groups (bionic tiger-bone powder + vitamin D treatment group after ovariectomy). The osteoporotic rat model was established 3 months after ovariectomy, and rats were intragastrically administrated with the corresponding drugs. Serum and bone tissue samples were collected from 10 rats in each group at weeks 4, 12 and 24 after intragastric administration. The bone microstructure of L6 vertebrae was analyzed by MicroCT, the biomechanical strength of left femurs was measured by the three-point bending test, and serum bone metabolism markers (P1NP and CTX) were detected by ELISA. Changes in bone collagen were analyzed by Masson's trichrome staining and hydroxyproline detection, and members of the BMP2/SMAD/RUNX2 and OPG/RANKL/RANK signal pathways were detected by immunoblotting. RESULTS: Compared with the OVX group, the serum level of P1NP in the TB and TB + VD groups was higher (P < 0.05), while the CTX level was lower (P < 0.05). Bone collagen fiber structures in the TB and TB + VD groups were repaired, and the collagen content was significantly higher than that in the OVX group (P < 0.05). In the TB group, BMP-2, P-SMAD1/5, RUNX2 and OPG levels were increased in bone tissue (P < 0.01), RANKL levels were decreased (P < 0.01), and the bone microstructure and biomechanical strength were improved. CONCLUSION: Bionic tiger-bone powder promotes osteogenesis by activating the BMP2/SMAD/RUNX2 signaling pathway, suppresses osteoclasts by downregulating the OPG/RANK/RANKL signaling pathway, increases bone collagen content, and improves bone microstructure and bone biomechanical strength.

Ortho-silicic Acid Inhibits RANKL-Induced Osteoclastogenesis and Reverses Ovariectomy-Induced Bone Loss In Vivo.

Numerous experiments in vitro and in vivo have shown that an appropriate increase intake of silicon can facilitate the synthesis of collagen and its stabilization and promote the differentiation and mineralization of osteoblasts. In this study, we examined whether ortho-silicic acid restrains the differentiation of osteoclast through the receptor activator of nuclear factor κB ligand (RANKL)/receptor activator of nuclear factor κB (RANK)/osteoprotegerin (OPG) signaling pathway by investigating its effect in vitro and in vivo. Bone marrow macrophage (BMM) cells were isolated and cultured with or without ortho-silicic acid, and then TRAP staining and immunofluorescence were performed to detect the differentiation of osteoclast. The RANKL-induced osteoclast marker gene and protein expression including c-Fos, nuclear factor of activated T cells cl (NFATcl), tumor necrosis factor receptor-associated factor 6 (TRAF6), nuclear factor kappa B P50 (NF-κB P50), NF-κB P52, RANK, integrin ß3, cathepsin K (CTSK), DC-STAMP, and TRAP were quantitatively detected by western blot and RT-PCR. Ovariectomized (OVX) rats were injected with ortho-silicic acid (OVX+Si group) and normal saline (OVX group), and sham-operated rats were injected with normal saline (Sham group). And micro-CT, H&E, and TRAP staining, ELISA, and western blot were performed. Ortho-silicic acid could inhibit the differentiation of osteoclast, and the marker genes and proteins were decreased. The OVX-induced bone loss could be reversed by ortho-silicic acid. Our finding demonstrated that ortho-silicic acid suppresses RANKL-induced osteoclastogenesis and has potential value as a therapeutic agent for OVX-induced bone loss.

Ortho-silicic acid enhances osteogenesis of osteoblasts through the upregulation of miR-130b which directly targets PTEN.

AIMS: Osteoporosis is considered a common skeletal disease. Ortho-silicic acid has been found to enhance the osteogenic differentiation of osteoblasts. However, the molecular mechanism of osteogenesis induced by ortho-silicic acid is still undefined totally. MicroRNAs (miRs) play a key role in osteogenesis of osteoblasts. This study investigated the role of miR-130b in promoting osteogenesis induced by ortho-silicic acid. MAIN METHODS AND KEY FINDINGS: In this study, we found ortho-silicic acid enhanced osteogenesis of osteoblasts in vitro and promoted preventing and treating osteoporosis in vivo. Furthermore, the expression of miR-130b increased under application of ortho-silicic acid. In vitro, experiments demonstrated miR-130b overexpression or inhibition significantly promoted or suppressed osteogenic differentiation of osteoblasts under application of ortho-silicic acid, respectively. Consistently, downregulation of miR-130b in ovariectomy (OVX) rats dropped off the beneficial effect of ortho-silicic acid against bone loss. Mechanistically, we identified phosphatase and tensin homologue deleted on human chromosome 10 (PTEN) as the direct target of miR-130b during osteogenesis induced by ortho-silicic acid. SIGNIFICANCE: In conclusion, our findings reveal that ortho-silicic acid promotes the osteogenesis of osteoblasts mediated by the miR-130b/PTEN signaling axis, which identifies a new target to prevent and treat osteoporosis.