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With the understanding of microRNA (miRNA or miR) functions in tumor initiation, progression, and metastasis, efforts are underway to develop new miRNA-based therapies. Very recently, we demonstrated effectiveness of a novel humanized bioengineered miR-124-3p prodrug in controlling spontaneous lung metastasis in mouse models. This study was to investigate the molecular and cellular mechanisms by which miR-124-3p controls tumor metastasis. Proteomics study identified a set of proteins selectively and significantly downregulated by bioengineered miR-124-3p in A549 cells, which were assembled into multiple cellular components critical for metastatic potential. Among them, plectin (PLEC) was verified as a new direct target for miR-124-3p that links cytoskeleton components and junctions. In miR-124-3p-treated lung cancer and osteosarcoma cells, protein levels of vimentin, talin 1 (TLN1), integrin beta-1 (ITGB1), IQ motif containing GTPase activating protein 1 (IQGAP1), cadherin 2 or N-cadherin (CDH2), and junctional adhesion molecule A (F11R or JAMA or JAM1) decreased, causing remodeling of cytoskeletons and disruption of cell–cell junctions. Furthermore, miR-124-3p sharply suppressed the formation of focal adhesion plaques, leading to reduced cell adhesion capacity. Additionally, efficacy and safety of biologic miR-124-3p therapy was established in an aggressive experimental metastasis mouse model in vivo. These results connect miR-124-3p−PLEC signaling to other elements in the control of cytoskeleton, cell junctions, and adhesion essential for cancer cell invasion and extravasation towards metastasis, and support the promise of miR-124 therapy.

Background: In recent years, decision support tools (DSTs) in various fields of medicine have emerged to aid clinicians and patients in the process of shared decision-making (SDM). This scoping review aims to identify the existing DSTs for selecting treatments in lower extremity arterial disease and to evaluate their effectiveness in facilitating SDM., Methods: The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines for scoping reviews were followed. A literature search was conducted across MEDLINE, EMBASE, and Cochrane databases, along with the Decision Aid Library Inventory, for studies published between January 2000 and June 2023. Articles reporting the development and/or clinical application of a DST specific to lower extremity arterial disease were included. A narrative synthesis of the results was performed and findings were presented in tabular formats., Results: Five studies and 5 unique DSTs were included. Presenting formats included websites, booklets, brochures, and pocket cards. Overall, a high degree of heterogeneity was observed across all DSTs in their format, content, and delivery. A widespread acceptability and satisfaction were reported among patients and clinicians. However, their effect at improving SDM remains uncertain due to the lack of standardized outcome metrics., Conclusions: The development and implementation of DSTs for lower limb arterial disease treatment discussion remain in the early stages. This review lays the foundation for future studies to continue exploring optimal strategies for DST development and their role in supporting SDM., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Among bone diseases, osteoporosis-like skeleton, such as trabecular thinning, fracture and so on, is the main pathological change of cadmium-induced osteoporosis(Cd-OP), accompanied by brittle bone and increased fracture rate. However, the mechanism underlying cadmium-induced osteoporosis has remained elusive. Compound Lurong Jiangu Capsule (CLJC) is an experienced formula for the treatment of bone diseases, which has the effect of tonifying kidney and strengthening bones, promoting blood circulation and relieving pain., Objective: Network pharmacology and molecular docking technology combined with experiments were used to investigate the potential mechanism of CLJC in treating Cd-OP., Method: The active compounds and corresponding targets of each herb in CLJC were searched in the TCMSP and BATMAN-TCM databases. The DisGeNet, OMIM, and GeneCards databases searched for Cd-OP targets. The relationship between both of them was visualized by establishing an herb-compound-target network using Cytoscape 3.9.1 software. Gene ontology (GO), and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analyses were performed after determining the intersection of the targets from CLJC and Cd-OP. What's more, molecular docking was performed to validate the results. All of them were aim to obtain hud signaling pathways for further study. Finally, BAX, BCL-2, and CASPASE-3 were screened and selected for further experiments, which included bone imaging and reconstruction analysis (Micro-CT), hematoxylin-eosin Staining (HE), and western blot (WB)., Results: 106 common targets from CLJC and Cd-OP targets were identified. KEGG pathway analysis suggested that multiple signaling pathways, such as the pathways in cancer, may play roles in treatment. Verification of the molecular docking was successful. Here we showed that Cd-OP displayed Tb.Th and Tb.N significantly reduced and even broke, irregular proliferation of bone cortex, uneven and loose trabecular bone arrangement, changed in apoptosis-related proteins, such as significant upregulation of CASPASE-3, BAX protein and significant downregulation of BCL-2 protein in vivo , while CLJC rescued these phenotypes., Conclusion: This study revealed that CLJC can reduce the expression of apoptosis-related proteins, and multiple components and multiple targets inhibit Cd-OP through apoptosis signaling pathway., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhou, Huang, Cao, Yu, Li, Liang, Leng, Jin, Yu and Liu.)

The detection of ascorbic acid (AA), dopamine (DA), and uric acid (UA) is not only of great significance in the areas of biomedicine and neurochemistry but also helpful in disease diagnosis and pathology research. Due to their diverse structures, designability, and large specific surface areas, metal-organic frameworks (MOFs) have recently caught considerable attention in the electrochemical field. Herein, a family of heterometallic MOFs with amino modification, MIL-125(Ti-Al)- x NH 2 ( x = 0%, 25%, 50%, 75%, and 100%), were synthesized and employed as electrochemical sensors for the detection of AA, DA, and UA. Among them, MIL-125(Ti-Al)-75%NH 2 exhibited the most promising electrochemical behavior with 40% doping of carbon black in 0.1 M PBS (pH = 7.10), which displayed individual detection performance with wide linear detection ranges (1.0-6.5 mM for AA, 5-100 μM for DA and 5-120 μM for UA) and low limits of detection (0.215 mM for AA, 0.086 μM for DA, and 0.876 μM for UA, S/N = 3). Furthermore, the as-prepared MIL-125(Ti-Al)-75%NH 2 /GCE provided a promising platform for future application in real sample analysis, owing to its excellent anti-interference performance and good stability.

A critical challenge of existing cancer vaccines is to orchestrate the demands of antigen-enriched furnishment and optimal antigen-presentation functionality within antigen-presenting cells (APCs). Here, a complementary immunotherapeutic strategy is developed using dendritic cell (DC)-tumor hybrid cell-derived chimeric exosomes loaded with stimulator of interferon genes (STING) agonists (DT-Exo-STING) for maximized tumor-specific T-cell immunity. These chimeric carriers are furnished with broad-spectrum antigen complexes to elicit a robust T-cell-mediated inflammatory program through direct self-presentation and indirect DC-to-T immunostimulatory pathway. This chimeric exosome-assisted delivery strategy possesses the merits versus off-the-shelf cyclic dinucleotide (CDN) delivery techniques in both the brilliant tissue-homing capacity, even across the intractable blood-brain barrier (BBB), and the desired cytosolic entry for enhanced STING-activating signaling. The improved antigen-presentation performance with this nanovaccine-driven STING activation further enhances tumor-specific T-cell immunoresponse. Thus, DT-Exo-STING reverses immunosuppressive glioblastoma microenvironments to pro-inflammatory, tumoricidal states, leading to an almost obliteration of intracranial primary lesions. Significantly, an upscaling option that harnesses autologous tumor tissues for personalized DT-Exo-STING vaccines increases sensitivity to immune checkpoint blockade (ICB) therapy and exerts systemic immune memory against post-operative glioma recrudesce. These findings represent an emerging method for glioblastoma immunotherapy, warranting further exploratory development in the clinical realm., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

The challenge of wound infections post-surgery and open trauma caused by multidrug-resistant bacteria poses a constant threat to clinical treatment. As a promising antimicrobial treatment, photothermal therapy can effectively resolve the problem of drug resistance in conventional antibiotic antimicrobial therapy. Here, we report a deep-penetration functionalized cuttlefish ink nanoparticle (CINP) for photothermal and immunological therapy of wound infections. CINP is decorated with zwitterionic polymer (ZP, namely sulfobetaine methacrylate-methacrylate copolymer) to form CINP@ZP nanoparticles. Natural CINP is found to not only exhibit photothermal destruction of methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli), but also trigger macrophages-related innate immunity and enhance their antibacterial functions. The ZP coating on the surface of CINP enables nanoparticles to penetrate into deeply infected wound environment. In addition, CINP@ZP is further integrated into the thermosensitive Pluronic F127 gel (CINP@ZP-F127). After in situ spraying gel, CINP@ZP-F127 is also documented notable antibacterial effects in mice wound models infected with MRSA and E. coli. Collectively, this approach combining of photothermal therapy with immunotherapy can promote delivery efficiency of nanoparticles to the deep foci of infective wounds, and effectively eliminate wound infections., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Neuroma formation after peripheral nerve transection often leads to severe neuropathic pain. Regenerative peripheral nerve interface has been shown to reduce painful neuroma in the clinic. However, no reports have investigated the underlying mechanisms, and no comparative animal studies on regenerative peripheral nerve interface and other means of neuroma prevention have been conducted to date. In this study, we established a rat model of left sciatic nerve transfection, and subsequently interfered with the model using the regenerative peripheral nerve interface or proximal nerve stump implantation inside a fully innervated muscle. Results showed that, compared with rats subjected to nerve stump implantation inside the muscle, rats subjected to regenerative peripheral nerve interface intervention showed greater inhibition of the proliferation of collagenous fibers and irregular regenerated axons, lower expressions of the fibrosis marker α-smooth muscle actin and the inflammatory marker sigma-1 receptor in the proximal nerve stump, lower autophagy behaviors, lower expressions of c-fos and substance P, higher expression of glial cell line-derived neurotrophic factor in the ipsilateral dorsal root ganglia. These findings suggested that regenerative peripheral nerve interface inhibits peripheral nerve injury-induced neuroma formation and neuropathic pain possibly via the upregulation of the expression of glial cell line-derived neurotrophic factor in the dorsal root ganglia and reducing neuroinflammation in the nerve stump., Competing Interests: None

Bone homeostasis is regulated by hormones such as parathyroid hormone (PTH). While PTH can stimulate osteo-progenitor expansion and bone synthesis, how the PTH-signaling intensity in progenitors is controlled is unclear. Endochondral bone osteoblasts arise from perichondrium-derived osteoprogenitors and hypertrophic chondrocytes (HC). We found, via single-cell transcriptomics, that HC-descendent cells activate membrane-type 1 metalloproteinase 14 (MMP14) and the PTH pathway as they transition to osteoblasts in neonatal and adult mice. Unlike Mmp14 global knockouts, postnatal day 10 (p10) HC lineage-specific Mmp14 null mutants (Mmp14ΔHC) produce more bone. Mechanistically, MMP14 cleaves the extracellular domain of PTH1R, dampening PTH signaling, and consistent with the implied regulatory role, in Mmp14ΔHC mutants, PTH signaling is enhanced. We found that HC-derived osteoblasts contribute ~50% of osteogenesis promoted by treatment with PTH 1-34, and this response was amplified in Mmp14ΔHC. MMP14 control of PTH signaling likely applies also to both HC- and non-HC-derived osteoblasts because their transcriptomes are highly similar. Our study identifies a novel paradigm of MMP14 activity-mediated modulation of PTH signaling in the osteoblast lineage, contributing new insights into bone metabolism with therapeutic significance for bone-wasting diseases., Competing Interests: TC, PC, AY, MK, ZT, KT, ZZ No competing interests declared, KC Senior editor, eLife, (© 2023, Chu et al.)

Systematic administration of antibiotics to treat infections often leads to the rapid evolution and spread of multidrug-resistant bacteria. Here, an in situ-formed biotherapeutic gel that controls multidrug-resistant bacterial infections and accelerates wound healing is reported. This biotherapeutic gel is constructed by incorporating stable microbial communities (kombucha) capable of producing antimicrobial substances and organic acids into thermosensitive Pluronic F127 (polyethylene-polypropylene glycol) solutions. Furthermore, it is found that the stable microbial communities-based biotherapeutic gel possesses a broad antimicrobial spectrum and strong antibacterial effects in diverse pathogenic bacteria-derived xenograft infection models, as well as in patient-derived multidrug-resistant bacterial xenograft infection models. The biotherapeutic gel system considerably outperforms the commercial broad-spectrum antibacterial gel (0.1% polyaminopropyl biguanide) in pathogen removal and infected wound healing. Collectively, this biotherapeutic strategy of exploiting stable symbiotic consortiums to repel pathogens provides a paradigm for developing efficient antibacterial biomaterials and overcomes the failure of antibiotics to treat multidrug-resistant bacterial infections., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

This study aims to evaluate the anti-tumor and analgesic activities of Compound Kushen Injection(CKI) based on zebrafish model in vivo and investigate the anti-tumor mechanism. To be specific, zebrafish tumor xenotransplantation model was established by microinjection of murine LPC H12 cells into yolk sac. Then the high-dose CKI(H-CKI), medium-dose CKI(M-CKI), low-dose CKI(L-CKI) groups, and the model group were set. The anti-tumor activity of CKI was evaluated with the tumor area growth fold and integral absorbance(IA) growth fold 72 h after administration. The peripheral pain and central pain in zebrafish were respectively induced with acetic acid(AA) and phorbol myristate acetate(PMA). Zebralab ViewPoint system was employed to monitor behavioral trajectory of zebrafish, and movement times, movement time, movement distance, and movement velocity were used to evaluate the analgesic activity of CKI. Finally, real-time fluorescence quantitative polymerase chain reaction(RT-qPCR) was performed to detect the expression levels of apoptosis-related B lymphocyte tumor-2(Bcl-2) and phosphatidylinositol-3-kinase(PI3 K)/protein kinase B(Akt or PKB) pathway-related genes, for the verification of the anti-tumor mechanism. Compared with the model group, M-CKI and H-CKI significantly reduced the growth folds of tumor area and IA, relief the peripheral pain and central pain. The mechanism was that CKI can up-regulate the expression of cysteine aspartic acid specific protease-3(caspase-3, Casp3) and caspase-9(Casp9), down-regulate the expression of phosphoinositide 3-kinase(PI3 K) and Akt, and significantly reduce the expression of Bcl-2, hypoxia-inducible factor-1α(HIF-1α), and vascular endothelial growth factor(VEGF). In conclusion, CKI has significant inhibitory effect on tumor growth and pain, which is related to the PI3 K/Akt signaling pathway. The pathway mediates cell apoptosis, suppresses tumor growth, and alleviates tumor pain.

Genome-derived microRNAs (miRNAs or miRs) control post-transcriptional gene expression critical for various cellular processes. Recently, we have invented a novel platform technology to achieve high-yield production of fully humanized, bioengineered miRNA agents (hBERAs) for research and development. This study is aimed to produce and utilize a new biologic miR-34a-5p (or miR-34a) molecule, namely, hBERA/miR-34a, to delineate the role of miR-34a-5p in the regulation of mitochondrial functions in human carcinoma cells. Bioengineered hBERA/miR-34a was produced through in vivo fermentation production and purified by anion exchange fast protein liquid chromatography. hEBRA/miR-34a was processed to target miR-34a-5p in human osteosarcoma and lung cancer cells, as determined by selective stem-loop reverse transcription quantitative polymerase chain reaction analysis. The mitochondrial inner membrane protein MPV17 like 2 ( MPV17L2 ) was validated as a direct target for miR-34a-5p by dual luciferase reporter assay. Western blot analysis revealed that bioengineered miR-34a-5p effectively reduced MPV17L2 protein outcomes, leading to much lower levels of respiratory chain Complex I activities and intracellular ATP that were determined with specific assay kits. Moreover, Seahorse Mito Stress Test assay was conducted, and the results showed that biologic miR-34a-5p sharply reduced cancer cell mitochondrial respiration capacity, accompanied by a remarkable increase of oxidative stress and elevated apoptotic cell death, which are manifested by greater levels of reactive oxygen species and selective apoptosis biomarkers, respectively. These results demonstrate the presence and involvement of the miR-34a-5p-MPV17L2 pathway in the control of mitochondrial functions in human carcinoma cells and support the utility of novel bioengineered miRNA molecules for functional studies.

Tumor stemness has been reported to play important roles in cancers. However, a comprehensive analysis of tumor stemness remains to be performed to investigate the specific mechanisms and practical values of stemness in soft tissue sarcomas (STS). Here, we applied machine learning to muti-omic data of patients from TCGA-SARC and GSE21050 cohorts to reveal important roles of stemness in STS. We demonstrated limited roles of existing mRNAsi in clinical application. Therefore, based on stemness-related signatures (SRSs), we identified three stemness subtypes with distinct stemness, immune, and metabolic characteristics using consensus clustering. The low-stemness subtype had better prognosis, activated innate and adaptive immunity (e.g., infiltrating B, DC, Th1, CD8+ T, activated NK, gamma delta T cells, and M1 macrophages), more enrichment of metabolic pathways, more sites with higher methylation level, higher gene mutations, CNA burdens, and immunogenicity indicators. Furthermore, the 16 SRS-based stemness prognostic index (SPi) was developed, and we found that low-SPi patients with low stemness had better prognosis and other characteristics similar to those in the low-stemness subtype. Besides, low-stemness subtype and low-SPi patients could benefit from immunotherapy. The predictive value of SPi in immunotherapy was more accurate after the addition of MSI into SPi. MSI low SPi low patients might be more sensitive to immunotherapy. In conclusion, we highlighted mechanisms and practical values of the stemness in STS. We also recommended the combination of MSI and SPi which is a promising tool to predict prognosis and achieve precise treatments of immunotherapy in STS., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gu, Qu, Peng, Yu, Jiang, Qi and Yu.)

Herein, a N-rich metal-organic framework (MOF) with four kinds of cages, Zn 4 (ade) 2 (TCA) 2 (H 2 O) ( NENU-1000 , Hade = adenine, H 3 TCA = 4,4',4″-tricarboxytriphenylamine, NENU = Northeast Normal University), was prepared by the mixed-ligand strategy. Cationic dyes can be selectively absorbed by NENU-1000 at proper concentrations, but not neutral and anionic dyes, which perhaps can be assigned to the N-rich neutral framework of NENU-1000 . When NENU-1000 was introduced to a relatively lower concentration of cationic dye solutions (e.g., rhodamine B or basic red 2), the colors of these systems faded quickly. Furthermore, the faded solutions can be used for the detection of methanol and other small alcohol molecules with either the naked eye or common UV-vis spectra. The effect of the length of carbon chain, the position of the -OH group, and the number of the hydroxyl group of the alcohols was explored for the color development rate. In addition, the performance of NENU-1000 in iodine sorption and release was also studied.

With the understanding of microRNA (miRNA or miR) functions in tumor initiation, progression, and metastasis, efforts are underway to develop new miRNA-based therapies. Very recently, we demonstrated effectiveness of a novel humanized bioengineered miR-124-3p prodrug in controlling spontaneous lung metastasis in mouse models. This study was to investigate the molecular and cellular mechanisms by which miR-124-3p controls tumor metastasis. Proteomics study identified a set of proteins selectively and significantly downregulated by bioengineered miR-124-3p in A549 cells, which were assembled into multiple cellular components critical for metastatic potential. Among them, plectin (PLEC) was verified as a new direct target for miR-124-3p that links cytoskeleton components and junctions. In miR-124-3p-treated lung cancer and osteosarcoma cells, protein levels of vimentin, talin 1 (TLN1), integrin beta-1 (ITGB1), IQ motif containing GTPase activating protein 1 (IQGAP1), cadherin 2 or N-cadherin (CDH2), and junctional adhesion molecule A (F11R or JAMA or JAM1) decreased, causing remodeling of cytoskeletons and disruption of cell-cell junctions. Furthermore, miR-124-3p sharply suppressed the formation of focal adhesion plaques, leading to reduced cell adhesion capacity. Additionally, efficacy and safety of biologic miR-124-3p therapy was established in an aggressive experimental metastasis mouse model in vivo . These results connect miR-124-3p-PLEC signaling to other elements in the control of cytoskeleton, cell junctions, and adhesion essential for cancer cell invasion and extravasation towards metastasis, and support the promise of miR-124 therapy., (© 2021 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.)

Ethnopharmacological Relevance: Curcumae Rhizoma-Sparganii Rhizoma (CR-SR), an ancient and classical herbal couple, has been extensively used for tumor treatment in clinic of traditional Chinese medicines (TCMs)., Aim of the Study: The study aimed to uncover the anti-tumor active materials of CR-SR water decoction (CR:SR = 1:1) via an integrated approach of spectrum-effect relationship, molecular docking, and ADME evaluation., Materials and Methods: The anti-tumor activities toward A549, HepG2, Hela, BGC-823, and MCF-7 cells of the different polar elution fractions (DPEFs) of CR, SR, and CR-SR were determined by Cell Counting Kit-8 (CCK-8) assay. Likewise, the DPEFs' combinations of CR and SR were also tested. The chemical fingerprints of these fractions were profiled by HPLC. Meanwhile, HPLC-ESI-Q-TOF-MS/MS was applied for the identification of chemical components. The main effect-related compounds were screened out by spectrum-effect relationship and molecular docking method. The oral bioavailability and druggability of these active components were subsequently evaluated. Finally, five monomeric compounds were validated experimentally using HepG2 cells., Results: The 80% ethanol elution fraction of CR, SR, and CR-SR showed strong anti-tumor effects toward five cells. Also, the combinations with the 80% ethanol elution fraction of CR and SR showed stronger tumor inhibition effects among the DPEFs' combinations of CR and SR. By spectrum-effect relationship, HPLC-MS, and molecular docking analysis, 24 main effect-related compounds seemed to have potential anti-tumor effects. ADME evaluation showed rutin performed low oral bioavailability and druggability. Therefore, we suppose that 23 compounds (including 4 unknown compounds) are the primary anti-tumor active components of CR-SR water decoction. Among them, zederone, curcumol, chlorogenic acid, calycosin, and curcumenol were validated successfully with good tumor inhibition effects., Conclusions: In summary, this study demonstrated that the multi-components of CR-SR contribute to its anti-tumor effects. It established a rapid and useful strategy to explore the active material basis of traditional Chinese herbal couples with a multi-technology integrated approach in practice, including chromatography, mass spectrometry, machine algorithm models, online databases, and in vitro cell experiments., (Copyright © 2021. Published by Elsevier B.V.)

Alcoholic liver disease (ALD) is a major health issue globally due to the consumption of alcoholic beverages. Thymus quinquecostatus Celak is a food additive and an edible herb that is widely used in Asia and possesses hepatoprotective activity, but the underlying mechanisms behind this protective activity are not completely understood. The purpose of this study was to investigate the hepatoprotective effects of Thymus quinquecostatus Celak extract (TQE) against ALD as well as the underlying mechanism based on gut microbiota and the gut-liver axis. TQE supplementation markedly alleviated chronic alcohol-induced liver injury in C57 mice. TQE also ameliorated gut barrier dysfunction induced by alcohol. Consequently, the activation of the lipopolysaccharide (LPS) translocation-mediated TLR4 pathway and the subsequent inflammatory response and ROS overproduction in the liver were suppressed. Meanwhile, alcohol-induced gut microbiota dysbiosis was also corrected by TQE. To further investigate the contribution of gut dysbiosis correction to the beneficial effects of TQE on ALD, a fecal microbiota transplantation study was conducted. TQE-manipulated gut microbiota transplantation markedly counteracted the alcohol-induced gut dysbiosis in the recipient mice. In parallel with gut dysbiosis correction, liver damage was partly ameliorated in the recipient mice. Gut barrier dysfunction, endotoxemia, TLR4 pathway induction as well as downstream inflammatory response and ROS overproduction were also partly suppressed due to gut dysbiosis correction in alcohol-fed recipient mice. In summary, these results suggest that gut dysbiosis correction contributes to the hepatoprotective effects of TQE against alcohol through the gut-liver axis.

Understanding pharmacokinetic (PK)-pharmacodynamic (PD) relationships is essential in translational research. Existing PK-PD models for combination therapy lack consideration of quantitative contributions from individual drugs, whereas interaction factor is always assigned arbitrarily to one drug and overstretched for the determination of in vivo pharmacologic synergism. Herein, we report a novel generic PK-PD model for combination therapy by considering apparent contributions from individual drugs coadministered. Doxorubicin (Dox) and sorafenib (Sor) were used as model drugs whose PK data were obtained in mice and fit to two-compartment model. Xenograft tumor growth was biphasic in mice, and PD responses were described by three-compartment transit models. This PK-PD model revealed that Sor (contribution factor = 1.62) had much greater influence on overall tumor-growth inhibition than coadministered Dox (contribution factor = 0.644), which explains the mysterious clinical findings on remarkable benefits for patients with cancer when adding Sor to Dox treatment, whereas there were none when adding Dox to Sor therapy. Furthermore, the combination index method was integrated into this predictive PK-PD model for critical determination of in vivo pharmacologic synergism that cannot be correctly defined by the interaction factor in conventional models. In addition, this new PK-PD model was able to identify optimal dosage combination (e.g., doubling experimental Sor dose and reducing Dox dose by 50%) toward much greater degree of tumor-growth inhibition (>90%), which was consistent with stronger synergy (combination index = 0.298). These findings demonstrated the utilities of this new PK-PD model and reiterated the use of valid method for the assessment of in vivo synergism. SIGNIFICANCE STATEMENT: A novel pharmacokinetic (PK)-pharmacodynamic (PD) model was developed for the assessment of combination treatment by considering contributions from individual drugs, and combination index method was incorporated to critically define in vivo synergism. A greater contribution from sorafenib to tumor-growth inhibition than that of coadministered doxorubicin was identified, offering explanation for previously inexplicable clinical observations. This PK-PD model and strategy shall have broad applications to translational research on identifying optimal dosage combinations with stronger synergy toward improved therapeutic outcomes., Competing Interests: No author has an actual or perceived conflict of interest with the contents of this article., (Copyright © 2021 by The Author(s).)

Corylin, a flavonoid isolated from the fruit of Psoralea corylifolia , has an osteogenic effect on osteoblasts in vitro and bone micromass ex vivo. However, the effect and mechanism of corylin in regulating osteoclastogenesis remain unknown. By using murine bone marrow macrophages as the osteoclast precursor, corylin was found to inhibit the receptor activator of nuclear factor (NF) κB ligand (RANKL)-induced osteoclast differentiation via down-regulating osteoclastic marker genes. In parallel, F-actin formation and osteoclast migration were diminished in corylin-treated cultured osteoclasts, and subsequently the expressions of osteoclastic proteins were suppressed: the suppression of protein expression was further illustrated by transcriptomic analysis. Furthermore, corylin inhibited the nuclear translocation of p65, giving rise to a restraint in osteoclastic differentiation through the attenuation of transcription factors nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and nuclear factor of activated T cells c1 (NFATc1). There was no obvious change in apoptosis when the RANKL-induce osteoclasts were cultured in the presence of corylin. The finding supports the potential development of corylin as an osteoclast inhibitor against osteoporosis.

Rationale: Noncoding RNAs (ncRNAs) such as microRNAs (miRs or miRNAs) play important roles in the control of cellular processes through posttranscriptional gene regulation. However, ncRNA research is limited to utilizing RNA agents synthesized in vitro . Recombinant RNAs produced and folded in living cells shall better recapitulate biologic RNAs. Methods: Herein, we developed a novel platform for in vivo fermentation production of humanized recombinant ncRNA molecules, namely hBERAs, carrying payload miRNAs or siRNAs. Target hBERAs were purified by anion exchange FPLC method. Functions of hBERA/miRNAs were investigated in human carcinoma cells and antitumor activities were determined in orthotopic osteosarcoma xenograft spontaneous lung metastasis mouse models. Results: Proper human tRNAs were identified to couple with optimal hsa-pre-miR-34a as new fully-humanized ncRNA carriers to accommodate warhead miRNAs or siRNAs. A group of 30 target hBERAs were all heterogeneously overexpressed (each accounting for >40% of total bacterial RNA), which facilitated large-scale production (8-31 mg of individual hBERAs from 1L bacterial culture). Model hBERA/miR-34a-5p and miR-124-3p were selectively processed to warhead miRNAs in human carcinoma cells to modulate target gene expression, enhance apoptosis and inhibit invasiveness. In addition, bioengineered miR-34a-5p and miR-124-3p agents both reduced orthotopic osteosarcoma xenograft tumor growth and spontaneous pulmonary metastases significantly. Conclusion: This novel ncRNA bioengineering technology and resulting recombinant ncRNAs are unique additions to conventional technologies and tools for basic research and drug development., Competing Interests: Competing Interests: A.-M.Y., M.-J.T. and P.Y.H. are inventors on US Patent Application No. 62/674,939 filed on May 22, 2018 as well as US Patent No. 10,619,156 issued on April 14, 2020 and European Patent No. 3150980 granted on May 20, 2020 that are related to this work. A.-M.Y. is a founder of AimRNA, Inc. that intends to license the intellectual property., (© The author(s).)

Objectives: To develop and validate the Preoperative Risk Evaluation for Partial Nephrectomy (PREP) score to predict the probability of major postoperative complications after partial nephrectomy (PN) based on patient comorbidities., Patients and Methods: The Premier Healthcare Database was used to identify patients who had undergone elective PN. Through review of International Classification of Diseases ninth revision codes, we identified patient comorbidities and major surgical complications (Clavien-Dindo Grade III-V). Multivariable logistic regression was used to identify predictors of major complications. We used half of the set as the training cohort to develop our risk score and the other half as a validation cohort., Results: From 2003 to 2015, 25 451 PNs were performed. The overall rate of major complications was 4.9%. The final risk score consisted of 10 predictors: age, sex, congestive heart failure, coronary artery disease, chronic obstructive pulmonary disease, chronic kidney disease, diabetes, hypertension, obesity, and smoking. In the training cohort, the area under the receiver operating characteristic curve (AUC) was 0.75 (95% confidence interval [CI] 0.73-0.78), while the AUC for the validation cohort was 0.73 (95% CI 0.70-0.75). The predicted probabilities of major complication in the low- (≤10 points), intermediate- (11-20 points), high- (21-30 points), and very high-risk (>30 points) categories were 3% (95% CI 2.6-3.2), 8% (95% CI 7.2-9.2), 24% (95% CI 20.5-27.8), and 41% (95% CI 34.5-47.8), respectively., Conclusions: We developed and validated the PREP score to predict the risk of complications after PN based on patient characteristics. Calculation of the PREP score can help providers select treatment options for patients with a cT1a renal mass and enhance the informed consent process for patients planning to undergo PN., (© 2020 The Authors BJU International © 2020 BJU International Published by John Wiley & Sons Ltd.)

Objective: The purpose of this article was to compare the effectiveness and safety of single-stage (laparoscopic cholecystectomy [LC] plus laparoscopic common bile duct exploration [LCBDE]) with two-stage (LC plus endoscopic retrograde cholangiopancreatography (ERCP)/endoscopic sphincterotomy [EST]) in management for concomitant gallstones and common bile duct (CBD) stones., Materials and Methods: Systematic review and meta-analysis of randomised controlled trials (RCTs) comparing outcomes following single-stage with two-stage management for concomitant gallstones and CBD stones published from 1990 to 2017 in PubMed, Embase and the Science Citation Index. The primary outcomes were stone clearance from the CBD, post-operative morbidity and mortality. The secondary outcomes were retained stone, conversion to other procedures, length of hospital stay and total operating time. Pooled risk ratio (RR) or weighted mean differences (WMD) with 95% confidence intervals (95% CIs) were calculated using either the fixed effects model or random effects model., Results: Eleven RCTs studies were included in this analysis. These studies included a total of 1338 patients: 666 underwent LC + LCBDE and 672 underwent LC + ERCP/EST. The meta-analysis showed that no significant difference was noted between the two groups regarding CBD stone clearance (RR: 1.06; 95% CI: 0.99-1.14; P= 0.12), post-operative morbidity (RR: 1.03; 95% CI: 0.79-1.34; P= 0.81), mortality (RR: 0.30; 95% CI: 0.06-1.41; P= 0.13), retained stone (RR: 0.91; 95% CI: 0.57-1.47; P= 0.71), conversion to other procedures (RR: 0.80; 95% CI: 0.55-0.16; P= 0.23), length of hospital stay (WMD: 1.24, 95% CI: 3.57-1.09, P= 0.30), total operating time (WMD: 25.42, 95% CI: 22.38-73.22, P= 0.30)., Conclusion: Single-stage is efficient and safe in the treatment of patients with concomitant gallstones and CBD stones while avoiding the second procedure. In selected patients, single-stage management for concomitant gallstones and CBD stones might be considered as the preferred approach. However, the findings have to be carefully interpreted due to the existence of heterogeneity, in addition, patient's condition, operator's experience also should be taken into account in making treatment decisions., Competing Interests: None

Corylin is a naturally occurring flavonoid isolated from the fruit of Psoralea corylifolia L. (Fabaceae), which is a Chinese medicinal herb in treating osteoporosis. Although a variety of pharmacological activities of corylin have been reported, its osteogenic action and the underlying mechanism in bone development remain unclear. In the present study, the involvement of bone-specific genes in corylininduced differentiated osteoblasts was analyzed by RT-PCR, promoter-reporter assay, and Western blotting. In cultured osteoblasts, corylin-induced cell differentiation and mineralization, as well as increased the expressions of vital biological markers for osteogenesis, such as Runx2, Osterix, Col1, and ALP. Corylin was proposed to have dual pathways in triggering the osteoblastic differentiation. First, the osteogenic function of corylin acted through the activation of Wnt/β-catenin signaling. The nuclear translocation of β-catenin of cultured osteoblasts, as determined by flow cytometry and confocal microscopy, was triggered by applied corylin, and which was blocked by DKK-1, an inhibitor of Wnt/β-catenin signaling. Second, the application of corylin-induced estrogenic response in a dose-dependent manner, and which was blocked by ICI 182 780, an antagonist of estrogen receptor. Furthermore, the activation of Runx2 promoter by corylin was abolished by both DKK-1 and ICI 182,780, indicating that the corylin exhibited its osteogenic effect via estrogen and Wnt/β-catenin signaling pathways. In addition, corylin regulated the metabolic profiles, as well as the membrane potential of mitochondria, in cultured osteoblasts. Corylin also stimulated the osteogenesis in bone micromass derived from mesenchymal progenitor cells. This study demonstrated the osteogenic activities of corylin in osteoblasts and micromass, suggesting that corylin has the potential to be developed as a novel pro-osteogenic agent in targeting for the treatment of osteoblast-mediated osteoporosis., (© 2020 Federation of American Societies for Experimental Biology.)

Being the second most common type of primary bone malignancy in children and adolescents, Ewing Sarcoma (ES) encounters the dilemma of low survival rate with a lack of effective treatments. As an emerging approach to combat cancer, RNA therapeutics may expand the range of druggable targets. Since the genome-derived oncolytic microRNA-34a (miR-34a) is down-regulated in ES, restoration of miR-34a-5p expression or function represents a new therapeutic strategy which is, however, limited to the use of chemically-engineered miRNA mimics. Very recently we have developed a novel bioengineering technology using a stable non-coding RNA carrier (nCAR) to achieve high-yield production of biocompatible miRNA prodrugs, which is a great addition to current tools for the assessment of RNA therapeutics. Herein, for the first time, we investigated the biochemical pharmacology of bioengineered miR-34a-5p prodrug (nCAR/miR-34a-5p) in the control of ES using human ES cells and xenograft mouse models. The bioengineered nCAR/miR-34a-5p was precisely processed to mature miR-34a-5p in ES cells and subsequently suppressed cell proliferation, attributable to the enhancement of apoptosis and induction of G2 cell cycle arrest through downregulation of SIRT-1, BCL-2 and CDK6 protein levels. Furthermore, systemic administration of nCAR/miR-34a-5p dramatically suppressed the ES xenograft tumor growth in vivo while showing biocompatibility. In addition, the antitumor effect of bioengineered nCAR/miR-34a-5p was associated with a lower degree of tumoral cell proliferation and greater extent of apoptosis. These findings demonstrate the efficacy of bioengineered miR-34a-5p prodrug for the treatment of ES and support the development of miRNA therapeutics using biocompatible bioengineered miRNA prodrugs., (Copyright © 2020 Li, Yuan, Tu, Hu, Li, Yi, Li, Zhao, Cheng, Yu, Jian and Yu.)

MicroRNAs (miRNAs or miRs) are small noncoding RNAs derived from genome to control target gene expression. Recently we have developed a novel platform permitting high-yield production of bioengineered miRNA agents (BERA). This study is to produce and utilize novel fully-humanized BERA/miR-328-3p molecule (hBERA/miR-328) to delineate the role of miR-328-3p in controlling nutrient uptake essential for cell metabolism. We first demonstrated successful high-level expression of hBERA/miR-328 in bacteria and purification to high degree of homogeneity (>98%). Biologic miR-328-3p prodrug was selectively processed to miR-328-3p to suppress the growth of highly-proliferative human osteosarcoma (OS) cells. Besides glucose transporter protein type 1, gene symbol solute carrier family 2 member 1 (GLUT1/ SLC2A1 ), we identified and verified large neutral amino acid transporter 1, gene symbol solute carrier family 7 member 5 (LAT1/ SLC7A5 ) as a direct target for miR-328-3p. While reduction of LAT1 protein levels by miR-328-3p did not alter homeostasis of amino acids within OS cells, suppression of GLUT1 led to a significantly lower glucose uptake and decline in intracellular levels of glucose and glycolytic metabolite lactate. Moreover, combination treatment with hBERA/miR-328 and cisplatin or doxorubicin exerted a strong synergism in the inhibition of OS cell proliferation. These findings support the utility of novel bioengineered RNA molecules and establish an important role of miR-328-3p in the control of nutrient transport and homeostasis behind cancer metabolism., (© 2019 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.)

Wound infection has been a persistent problem that is common and costly. Thermosensitive hydrogel has been demonstrated to be a suitable dressing candidate due to its high moldability, easy administration and ability to maintain a moist topical environment at the wound bed. In the present study, a novel thermosensitive hydrogel was successfully prepared and characterized to have a porous inner structure and a sustained curcumin-releasing profile. The wound healing ability of the hydrogel was investigated in a wound infection model in rats. On analysis, it was observed that the hydrogel complex-dressed wounds exhibited a faster wound closure rate compared with gauze-covered wounds, which was paralleled with improved histological outcomes that were observed. Additionally, the results of in vitro antimicrobial, anti-oxidant, western blot analysis and reverse transcription-quantitative polymerase chain reaction assays indicated that the hydrogel complex had distinct anti-oxidative, antimicrobial and anti-nuclear factor-κB-signaling capacities. These results suggest that this novel hydrogel may be a suitable candidate for facilitating the healing of infected cutaneous wounds in rats.

The nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a transcription factor in the regulation of many oxidative enzymes and efflux transporters critical for oxidative stress and cellular defense against xenobiotics. NRF2 is dysregulated in patient osteosarcoma (OS) tissues and correlates with therapeutic outcomes. Nevertheless, research on the NRF2 regulatory pathways and its potential as a therapeutic target is limited to the use of synthetic small interfering RNA (siRNA) carrying extensive artificial modifications. Herein, we report successful high-level expression of recombinant siRNA against NRF2 in Escherichia coli using our newly established noncoding RNA bioengineering technology, which was purified to >99% homogeneity using an anion-exchange fast protein liquid chromatography method. Bioengineered NRF2-siRNA was able to significantly knock down NRF2 mRNA and protein levels in human OS 143B and MG63 cells, and subsequently suppressed the expression of NRF2-regulated oxidative enzymes [heme oxygenase-1 and NAD(P)H:quinone oxidoreductase 1] and elevated intracellular levels of reactive oxygen species. In addition, recombinant NRF2-siRNA was effective to sensitize both 143B and MG63 cells to doxorubicin, cisplatin, and sorafenib, which was associated with significant downregulation of NRF2-targeted ATP-binding cassette (ABC) efflux transporters (ABCC3, ABCC4, and ABCG2). These findings support that targeting NRF2 signaling pathways may improve the sensitivity of cancer cells to chemotherapy, and bioengineered siRNA molecules should be added to current tools for related research., (Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.)

Recently, hypoxia inducible factor-1 (HIF-1) was reported to be correlated with isocitrate dehydrogenase 1 (IDH-1) in several types of tumors. However, the expression and significance of HIF-1 and IDH-1 in osteosarcoma is still unknown. In the present study, the expression levels of IDH-1 and HIF-1α in 35 formalin-fixed paraffin-embedded sections from osteosarcoma patients were investigated by immunohistochemistry. The expression levels of IDH-1 and HIF-1α in human osteosarcoma cell lines (MG-63 and 143B) were further detected by western blotting under normal and hypoxic conditions. In addition, HIF-1α was downregulated via lentiviral vector‑mediated RNA interference (RNAi) in the MG-63 human osteosarcoma cell line. The results revealed that HIF-1α was negatively correlated with IDH-1 in the osteosarcoma tissues. Both in MG-63 and 143B cell lines, the expression of HIF-1α was increased while IDH-1 was decreased under a hypoxic condition compared to normal conditions. HIF-1α downregulation promoted IDH-1 expression in the MG-63 cell line under either normal or hypoxic conditions. In conclusion, our findings suggest that HIF-1α inhibits IDH-1 in osteosarcoma and consequently increases the incidence of osteosarcoma.

Metastasis is a major cause of mortality for cancer patients and remains as the greatest challenge in cancer therapy. Driven by multiple factors, metastasis may not be controlled by the inhibition of single target. This study was aimed at assessing the hypothesis that drugs could be rationally combined to co-target critical DNA, RNA and protein molecules to achieve "saturation attack" against metastasis. Independent actions of the model drugs DNA-intercalating doxorubicin, RNA-interfering miR-34a and protein-inhibiting sorafenib on DNA replication, RNA translation and protein kinase signaling in highly metastatic, human osteosarcoma 143B cells were demonstrated by the increase of γH2A.X foci formation, reduction of c-MET expression and inhibition of Erk1/2 phosphorylation, respectively, and optimal effects were found for triple-drug combination. Consequently, triple-drug treatment showed a strong synergism in suppressing 143B cell proliferation and the greatest effects in reducing cell invasion. Compared to single- and dual-drug treatment, triple-drug therapy suppressed pulmonary metastases and orthotopic osteosarcoma progression to significantly greater degrees in orthotopic osteosarcoma xenograft/spontaneous metastases mouse models, while none showed significant toxicity. In addition, triple-drug therapy improved the overall survival to the greatest extent in experimental metastases mouse models. These findings demonstrate co-targeting of DNA, RNA and protein molecules as a novel therapeutic strategy for the treatment of metastasis.

Osteosarcoma (OS) is the most common primary malignant bone tumor in children, and microRNA-34a (miR-34a) replacement therapy represents a new treatment strategy. This study was to define the effectiveness and safety profiles of a novel bioengineered miR-34a prodrug in orthotopic OS xenograft tumor mouse model. Highly purified pre-miR-34a prodrug significantly inhibited the proliferation of human 143B and MG-63 cells in a dose dependent manner and to much greater degrees than controls, which was attributed to induction of apoptosis and G2 cell cycle arrest. Inhibition of OS cell growth and invasion were associated with release of high levels of mature miR-34a from pre-miR-34a prodrug and consequently reduction of protein levels of many miR-34a target genes including SIRT1, BCL2, c-MET, and CDK6. Furthermore, intravenous administration of in vivo-jetPEI formulated miR-34a prodrug significantly reduced OS tumor growth in orthotopic xenograft mouse models. In addition, mouse blood chemistry profiles indicated that therapeutic doses of bioengineered miR-34a prodrug were well tolerated in these animals. The results demonstrated that bioengineered miR-34a prodrug was effective to control OS tumor growth which involved the induction of apoptosis and cell cycle arrest, supporting the development of bioengineered RNAs as a novel class of large molecule therapeutic agents.

Phloem-mobile insecticides are efficient for piercing and sucking insect control. Introduction of sugar or amino acid groups to the parent compound can improve the phloem mobility of insecticides, so a glycinergic-fipronil conjugate (GlyF), 2-(3-(3-cyano-1-(2,6-dichloro-4-(trifluoromethyl)phenyl)-4-((trifluoromethyl)sulfinyl)-1H-pyrazole-5-yl)ureido) acetic acid, was designed and synthesized. Although the "Kleier model" predicted that this conjugate is not phloem mobile, GlyF can be continually detected during a 5 h collection of Ricinus communis phloem sap. Furthermore, an R. communis seedling cotyledon disk uptake experiment demonstrates that the uptake of GlyF is sensitive to pH, carbonyl cyanide m-chlorophenylhydrazone (CCCP), temperature, and p-chloromercuribenzenesulfonic acid (pCMBS) and is likely mediated by amino acid carrier system. To explore the roles of amino acid transporters (AATs) in GlyF uptake, a total of 62 AAT genes were identified from the R. communis genome in silico. Phylogenetic analysis revealed that AATs in R. communis were organized into the ATF (amino acid transporter) and APC (amino acid, polyaminem and choline transporter) superfamilies, with five subfamilies in ATF and two in APC. Furthermore, the expression profiles of 20 abundantly expressed AATs (cycle threshold (Ct) values <27) were analyzed at 1, 3, and 6 h after GlyF treatment by RT-qPCR. The results demonstrated that expression levels of four AAT genes, RcLHT6, RcANT15, RcProT2, and RcCAT2, were induced by the GlyF treatment in R. communis seedlings. On the basis of the observation that the expression profile of the four candidate genes is similar to the time course observation for GlyF foliar disk uptake, it is suggested that those four genes are possible candidates involved in the uptake of GlyF. These results contribute to a better understanding of the mechanism of GlyF uptake as well as phloem loading from a molecular biology perspective and facilitate functional characterization of candidate AAT genes in future studies.

Isocitrate dehydrogenase 2 (IDH2) is a mitochondrial NADP-dependent isocitrate dehydrogenase. It is considered to be a novel tumor suppressor in several types of tumors. However, the role and related mechanism of IDH2 in osteosarcoma remain unknown. The expression and significance of IDH2 were investigated by immunohistochemistry in formalin-fixed paraffin sections from 44 osteosarcoma patients. IDH2 was downregulated via lentiviral vector‑mediated RNA interference (RNAi) in the Saos-2 and MG-63 human osteosarcoma cell lines. The effect of IDH2 downregulation on human osteosarcoma was studied in vitro by MTT, flow cytometry and invasion assays. Nuclear factor-κB (NF-κB) and matrix metalloproteinase-9 (MMP-9) assays were also used to study the likely molecular mechanism of IDH2 downregulation on the malignant progression of osteosarcoma cells. The results revealed that the expression of IDH2 was inversely correlated with pathological grade and metastasis in osteosarcoma. IDH2 downregulation promoted a pro-proliferative effect on the Saos-2 and MG-63 osteosarcoma cell lines. IDH2 downregulation accelerated cell cycle progression from S to G2/M phase. The pro-proliferative effect induced by IDH2 downregulation may be ascribed to increased NF-κB activity via IκBα phosphorylation. The invasive activity of osteosarcoma cells was also significantly promoted by IDH2 downregulation and may result from elevated MMP-9 activity. In conclusion, IDH2 downregulation may exacerbate malignant progression via increased NF-κB and MMP-9 activity and may implicate the potential biological importance of IDH2 targeting in osteosarcoma cells. Downregulation of IDH2 exacerbates the malignant progression of osteosarcoma cells via increased NF-κB and MMP-9 activation.

Osteosarcoma (OS) is the most common form of primary malignant bone tumor and prevalent among children and young adults. Recently we have established a novel approach to bioengineering large quantity of microRNA-34a (miR-34a) prodrug for miRNA replacement therapy. This study is to evaluate combination treatment with miR-34a prodrug and doxorubicin, which may synergistically suppress human OS cell growth via RNA interference and DNA intercalation. Synergistic effects were indeed obvious between miR-34a prodrug and doxorubicin for the suppression of OS cell proliferation, as defined by Chou-Talalay method. The strongest antiproliferative synergism was achieved when both agents were administered simultaneously to the cells at early stage, which was associated with much greater degrees of late apoptosis, necrosis, and G2 cell cycle arrest. Alteration of OS cellular processes and invasion capacity was linked to the reduction of protein levels of miR-34a targeted (proto-)oncogenes including SIRT1, c-MET, and CDK6. Moreover, orthotopic OS xenograft tumor growth was repressed to a significantly greater degree in mouse models when miR-34a prodrug and doxorubicin were co-administered intravenously. In addition, multiple doses of miR-34a prodrug and doxorubicin had no or minimal effects on mouse blood chemistry profiles. The results demonstrate that combination of doxorubicin chemotherapy and miR-34a replacement therapy produces synergistic antiproliferative effects and it is more effective than monotherapy in suppressing OS xenograft tumor growth. These findings support the development of mechanism-based combination therapy to combat OS and bioengineered miR-34a prodrug represents a new natural miRNA agent., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: The authors conducted a meta-analysis to compare the effectiveness and safety of conservative and operative treatment for distal radius fracture., Methods: PubMed, EMBASE, and the Cochrane Library were searched to identify the relevant studies published up to February of 2015. All randomized controlled trials published to compare the conservative and operative treatment were included in the study. Results were pooled using meta-analysis to compare the efficacy and safety of conservative and operative treatment for distal radius fracture., Results: The databases were derived from seven qualified studies that included a total of 523 patients in which 269 cases adopted conservative treatment while 253 cases adopted operative treatment. Overall, compared with the conservative treatment- treated the distal radius fracture, operative therapies resulted in significantly better radiographic (P<0.05), however, no significant differences of the functional outcomes and complication rate were observed between the two methods (P>0.05)., Conclusion: Surgical treatment seems to be more effective distal radius fracture compared with conservative treatment when the radiographic outcomes were analyzed, and no significant differences were deteched in the functional outcomes and complication rate.

Objective: This study aims to investigate the neuroprotective effect of Rho kinase inhibitor fasudil hydrochloride in ischemia/reperfusion injury N2a neuron., Methods: In vitro, N2a cells induced by ischemia and ischemia-reperfusion were treated with fasudil hydrochloride, cell damage was analyzed by MTT. On the other hand, the cytoskeleton of N2a cells was scanned through immunofluorescence techniques by Confocal Laser Microscopy which stained with FITC-phalloidin for F-actin visualization., Results: The activation of ROCK-II increased significantly in the damaged local during the following phase of ischemia/reperfusion injury. Ischemia induced a striking reorganization of actin cytoskeleton with a weakening of fluorescent intensity of the peripheral filament actin bands and formation of the long and thick stress fibers, but pretreatment of Fasudil hydrochloride could reversed the changes of ultra-structure on the cellular surface. MTT assay showed that Fasudil hydrochloride could prolong the survival time of the N2a cells after mimic ischemia-reperfusion for 24 h., Conclusions: The activation of ROCK-II has an exceptional hoist after ischemia/reperfusion injury, it is likely to induce the collapse of the growth cone through MLC-P. Fasudil hydrochloride could promote axonal growth on inhibitory of ROCK activity.

Objective: To assess the determinants of urolithiasis in patients with primary hyperparathyroidism (PHPT) before and after parathyroidectomy (PTX)., Methods: Institutional Research Ethics approval was obtained. A retrospective review was performed for patients presenting with PHPT to the stone, surgical oncology, and otolaryngology clinics at 2 tertiary-care centers from January 2006 to November 2011. Demographic, clinical, and surgical data were collected together with 24-hour urine collections before and after PTX., Results: Of 332 patients undergoing PTX, 255 (68.2% female patients) had PHPT. Mean age was 60.3 years (range, 18-91). Before PTX, renal calcification was detected in 51 (20%) patients, nephrolithiasis in 48 (18.8%), and nephrocalcinosis in 3 (1.2%) patients. Compared with PHPT patients without stones, PHPT patients with stones were significantly younger (56.4 vs 61.3 years, P=.02), less likely to be female (54.9% vs 71.9%, P=.03), and had significantly lower levels of vitamin D (19.7 vs 23.5 ng/mL, P=.03). Nine patients (3.5%) developed stones after PTX and were found to have significantly higher post-PTX total serum calcium levels when compared with those without stones. Although hypercalciuria was detected in 62% of pre-PTX stone formers, none of those who tested had post-PTX hypercalciuria (P<.001). On multivariate regression analysis, post-PTX stone formation was associated with male gender (adjusted odds ratio [95% confidence interval]: 6.8 [5.3-7.2], P=.01) and post-PTX hypercalcemia (adjusted odds ratio [95% confidence interval]: 1.48 [1.33-2.12], P=.02)., Conclusion: Pre-PTX urolithiasis was associated with younger age, male gender, and lower levels of vitamin D, whereas post-PTX urolithiasis was independently predicted by male gender and hypercalcemia., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Severe traumatic wounds are challenging to manage during surgery. The introduction of vacuum-assisted closure (VAC) is a breakthrough in wound management. The aim of the present study was to investigate the effect of VAC on cytokines in wounds during the management of severe traumatic wounds following initial debridement. VAC and conventional wound care (CWC) were independently applied to severe traumatic wounds on pigs. The expression levels of intercellular adhesion molecule-1 (ICAM-1), migration inhibitory factor (MIF), vascular endothelial growth factor (VEGF), basic fibroblast growth factor, collagen I and human fibroblast collagenase 1 were detected by quantitative polymerase chain reaction and western blotting. VAC significantly increased the expression of ICAM-1, MIF, VEGF and collagen I compared with that induced by CWC at the protein and mRNA levels. Therefore, the results of the present study indicate that VAC therapy is an effective method for treating severe traumatic wounds, as it increases the expression of cytokines in wounds. VAC significantly increases the expression of ICAM-1, MIF, VEGF and collagen I to manage severe traumatic wounds.

Angiogenesis is involved in the wound healing process. Increased angiogenesis and blood flow constitute a major mechanism of negative pressure wound therapy (NPWT), which has been shown to facilitate the healing of infected wounds. However, the effect on the expression of angiogensis‑related growth factor remains unknown. The goal of the current study was to investigate the angiogenic factor levels prior to and following NPWT in infected wounds. A total of 20 patients with infected wounds treated with NPWT were included in the study. Patients acted as their own control; the postoperative measurements of patients were considered as the experimental group, while preoperative measurements were considered as the controlled group. Blood flow was recorded prior to and during NPWT. A total of 10 angiogensis‑related growth factors were detected using a protein biochip array to analyze the change in protein levels prior to NPWT, and on the third day during NPWT. All wounds were successfully reconstructed by skin grafting or using local flaps following NPWT. NPWT resulted in significantly increased blood flow in the wound. There was a significant increase in vascular endothelial growth factor (VEGF), EGF, platelet‑derived growth factor and angiotesin‑2 following NPWT, while basic fibroblast growth factor decreased significantly. NPWT affects the local expression of angiogenesis‑associated growth factors, which represents another mechanism to explain how NPWT accelerates wound healing.

The ideal timing of wound closure for open tibial fractures is debatable. This study aimed to compare outcomes of primary and delayed wound closure in severe open tibial fractures initially treated with internal fixation and vacuum-assisted wound coverage (VAC). Data of 80 patients with Gustilo-Anderson type IIIA and IIIB open tibial fractures treated with primary internal fixation, VAC, either primary wound closure (PWC) or delayed wound closure (DWC), and external fixation were reviewed retrospectively, and outcomes and complications compared. Patients were divided into three groups, including a PWC group (n = 27), DWC group (n = 22), and a control group (n = 31) that had received external fixation. Among all patients, the median age was 38 years (IRQ 32-47 years), and 67.5% were male. Injuries included 33 Gustilo-Anderson type IIIA and 47 type IIIB. Among injuries, 83% (66/80) were high-energy trauma, 63.8% were contaminated and median injury severity score (ISS) was 14 points. Significant differences were found between groups in fixation methods (p < 0.001). No significant differences were observed between groups in rates of deep infection, osteomyelitis, amputation and nonunion at 6 and 12 months (all p > 0.05), although all rates were markedly lower in the PWC group. The outcomes of PWC performed in conjunction with primary internal fixation and VAC for the treatment of Gustilo-Andersen type IIIA and IIIB open tibial fractures are similar to or better than those of DWC with primary internal fixation and VAC., (Copyright © 2014 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.)

The aim of this study was to investigate the efficiency of negative pressure wound therapy (NPWT) combined with open bone graft (OBG; NPWT-OBG) for the treatment of bone and soft tissue defects with polluted wounds in an animal model. All rabbits with bone and soft tissue defects and polluted wounds were randomly divided into two groups, the experimental group (NPWT with bone graft) and the control group (OBG). The efficacy of the treatment was assessed by the wound conditions and healing time. Bacterial bioburdens and bony calluses were evaluated by bacteria counting and X-rays, respectively. Furthermore, granulation tissue samples from the wounds on days 0, 3, 7 and 14 of healing were evaluated for blood vessels and vascular endothelial growth factor (VEGF) levels. Wounds in the experimental group tended to have a shorter healing time, healthier wound conditions, lower bacterial bioburden, improvement of the bony calluses and an increased blood supply compared with those in the control group. With NPWT, wound infection was effectively controlled. For wounds with osseous and soft tissue defects, NPWT combined with bone grafting was demonstrated to be more effective than an OBG.

This meta-analysis was performed to evaluate the efficiency and the safety of absorbable implants. Five major electronic databases (PubMed, Embase, Cochrane Library, SinoMed and Wanfang Data) were systematically searched for randomized controlled trials (RCTs) from their establishment to November 2012. Studies on absorbable implants and metal implants for ankle fractures were selected. The meta-analysis was performed using RevMan 5.1. Ten studies with 762 patients were included and analyzed. Compared with metal implants, absorbable implants used for the internal fixation of ankle fractures produce similar radiographic and functional outcomes (P= 0.52). Normally, removal of the internal fixation is unnecessary (P<0.0001) and the incidence of palpable implants is lower (P=0.02) for absorbable implants. No statistically significant difference was observed between the two groups with regard to foreign body reactions (P=0.07), infection (P= 0.69), osteoarthritis (P= 0.39), pain (P= 0.06), refracture (P=0.67), skin necrosis (P=0.99), deep vein thrombosis (P=0.21) and nerve injury (P=0.94). Absorbable implants used in ankle fractures rarely require reoperation and result in similar functional outcomes and complications compared with metal implants. These characteristics make them efficient and reasonably safe for the treatment of ankle fractures.

The aim of this work is to explore the feasibility and therapeutic effect of repairing rabbit articular cartilage defects using thermo-sensitive chitosan/poly (vinyl alcohol) composite hydrogel engineered Ad-hTGF-β1-transfected bone marrow mesenchymal stem cells. Rabbit's bone marrow stromal cells (BMSCs) were obtained and cultured in vitro and transfected with a well-constructed Ad-hTGF-β1 vector, the cartilage phenotype of the transfected cells was tested by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Twenty-four New Zealand white rabbits with articular cartilage defects were randomly divided into four groups: group A was treated with CS/PVA gel and transfected BMSCs; group B received CS/PVA gel and un-transfected BMSCs; group C was treated with CS/PVA gel alone and group D was the untreated control group. Experimental animals of each group were killed at 16 weeks after operation. General observation, Masson's trichrome staining and collagen II immunohistological staining of the specimens were performed to evaluate the repair effect. The Wakitani scoring method was used to evaluate the repair effect. RT-PCR and Western blot confirmed that the hTGF-β1 gene was expressed in BMSCs and triggered the expression of specific markers of cartilage differentiation such as aggrecan mRNA and Collagen II in BMSCs after transfection with Ad-hTGF-β1. Sixteen weeks after operation, the defects in group A had smooth and flat surfaces, and the defects appeared to have completely healed, exhibiting almost the same color and texture as the surrounding cartilage. Masson's trichrome staining showed that the cell arrangement and density of regenerated cartilage tissue in group A was not significantly different from that of normal cartilage tissue. The immunohistochemical staining of Col II showed a strong expression in group A and weak expression in group B, but no expression in groups C and D. According to the Wakitani score, the difference between experimental group A and all of the other groups was statistically significant (P < 0.01). To conclude, as a thermosensitive and injectable scaffold material, CS/PVA gel engineered with BMSCs transfected with hTGF-β1 can effectively repair rabbit articular cartilage defects.

Background: Aberrant mitogen/extracellular signal-regulated kinase 5 (MEK5)-extracellular signal-regulated protein kinase 5 (ERK5)-mediated signalling has been implicated in a number of tumour types including prostate cancer (PCa). The molecular basis of ERK5-driven carcinogenesis and its clinical relevance remain to be fully characterised., Methods: Modulation of ERK5 expression or function in human PCa PC3 and PC3-ERK5 (stably transfected with ERK5) cells was performed using siRNA-mediated knockdown or the MEK inhibitor PD18435 respectively. In vitro significance of ERK5 signalling was assessed by assays for proliferation, motility, invasion and invadopodia. Expression of matrix metalloproteinases/tissue inhibitors of metalloproteases was determined by Q-RT-PCR. Extracellular signal-regulated protein kinase 5 expression in primary and metastatic PCa was examined using immunohistochemistry., Results: Reduction of ERK5 expression or signalling significantly inhibited the motility and invasive capability of PC3 cells. Extracellular signal-regulated protein kinase 5-mediated signalling significantly promoted formation of in vivo metastasis in an orthotopic PCa model (P<0.05). Invadopodia formation was also enhanced by forced ERK5 expression in PC3 cells. Furthermore, in metastatic PCa, nuclear ERK5 immunoreactivity was significantly upregulated when compared with benign prostatic hyperplasia and primary PCa (P=0.013 and P<0.0001, respectively)., Conclusion: Our in vitro, in vivo and clinical data support an important role for the MEK5-ERK5 signalling pathway in invasive PCa, which represents a potential target for therapy in primary and metastatic PCa.

Background: To detect the expression of isocitrate dehydrogenase 1 (IDH1) and transformation-related protein 53 (p53) in osteosarcoma and analyze the correlation between them and the clinico-pathological features., Methods: The expressions of IDH1 and p53 were detected in human osteosarcoma cell lines (MG-63 and U2OS) by immunocytochemistry, Real-time PCR and Western Blotting. The expressions of IDH1 and p53 in formalin-fixed paraffin-embedded tissue sections from 44 osteosarcoma patients were determined by immunohistochemistry, and the correlation between them and clinicopagthological features were analyzed. None of these patients received chemotherapy prior to surgery., Results: IDH1 is detected in osteosarcoma cell lines and biopsies. IDH1 expresses higher in U2OS cells with wild type p53 than in MG-63 cells with mutation p53. IDH1 correlates with histological Rosen grade and metastasis negatively. P53 correlates with histological Rosen grade, metastasis and overall survival in clinical osteosarcoma biopsies. Osteosarcoma patients with High IDH1 expression have a very high p53 expression., Conclusion: IDH1 may correlate with p53 and be a candidate biomarker for osteosarcoma correlate with histological Rosen grade and metastasis.

Objective: To examine the effect of TNF-related apoptosis-inducing ligand (TRAIL) and in combination with doxorubicin (ADM) to xenografted tumors in nude mice and to explore its potential mechanism., Methods: MG-63 cells (5 x 10(6)/ml) were suspended in 0.2 ml RPMI-1640 and inoculated subcutaneously into the lower limb of nude mice. Treatment groups were given TRAIL of different concentration or combination of TRAIL and ADM intraperitoneally. Normal saline was administrated in the control group. Anti-tumor effects were estimated by tumor volumes. Serum alkaline phosphatase (ALP) was detected by ALP kits. Induction of apoptosis in xenografted tumors was confirmed by TUNEL (TdT-mediated dUTP nick end labeling) assay. Expression of Bax was detected by immunohistochemical assay. Expression of TRAIL receptors was detected by RT-PCR assay., Results: Growth curve of tumors indicated that tumors carried by TRAIL-treated mice grew more slowly than that with normal saline and 2 microg TRAIL was more effective, Also tumors treated with combination of TRAIL and ADM grew more slowly than any other group. ALP activities of each group were moderately different but significance was not reached. TUNEL showed that there were more apoptotic cells in the combination group than any other group. Immunohistochemical assay showed that expression of Bax was up-regulated in the combination group. RT-PCR showed that expression of TRAIL-R2 mRNA was up-regulated in the combination group., Conclusion: TRAIL can induce an effective apoptosis of osteosarcoma cells in vivo in a dose-dependent fashion. ADM can enhance the effect of TRAIL-mediated apoptosis. And up-regulations of Bax and TRAIL-R2 may be the involved mechanism.

Purpose: The goal of this research was to determine if P58(IPK), a member of the Hsp40 family that inhibits eukaryotic initiation factor 2alpha (eIF2alpha), inhibits endoplasmic reticulum (ER) stress and leads to downregulated expression of vascular endothelial growth factor (VEGF) and decreased apoptosis in human retinal capillary endothelial cells (HRCECs)., Methods: Recombinant vectors were constructed using P58 in adeno-associated virus type 2 (rAAV2-P58 (IPK)) and P58 RNA in the plasmid pGIPZ (pGIPZ-P58(IPK)). The four experimental groups were: (1) non-transfected/non ER stressed control; (2) non-transfected/ER stressed; (3) rAAV2-P58(IPK)-transfected/ER stressed; and (4) pGIPZ- P58(IPK) RNAi transfected/ER stressed. ER stress was induced by treating cells with tunicamycin. Expression of P58(IPK) was determined in transfected cells. Expressions of the following factors were assessed: vascular endothelial growth factor (VEGF), C/EBP homologous protein (CHOP), activating transcription factor 4 (ATF4), and glucose-regulated protein 78 (GRP78). Apoptosis levels were also determined., Results: Significantly increased expression of P58(IPK) was detected in cells transfected with rAAV2-P58(IPK) (0.63+/-0.02) as compared to those transfected with pGIPZ-P58(IPK) RNAi (0.23+/-0.01). P58(IPK) expression was not different between the control transfected cells (rAAV2-GFP and pGIPZ-GFP). Following ER stress, expression levels of ATF-4, GRP78, CHOP, and VEGF in cells overexpressing P58(IPK) were not different from those in unstressed control cells. This inhibitory effect of P58(IPK) on the expression of ER stress-related factors was suppressed in cells transfected with pGIPZ-P58(IPK) RNAi. Apoptosis was significantly increased in cells transfected with pGIPZ-P58(IPK) RNAi but not with rAAV2-P58(IPK)., Conclusions: The study demonstrates that P58(IPK) inhibits ER stress and plays an important role in maintaining balance and stability of the ER in HRCECs.

Objective: To investigate the relationship between the mRNA expression levels of p53-mediating DNA damage and repair genes in the peripheral blood lymphocytes of workers and their exposures to benzene in their working environment., Methods: The mRNA expression levels of p53 and related genes were determined by SYBR Green I chimeric fluorescence quantitative real-time RT-PCR analysis in peripheral blood lymphocytes of 72 workers, who were classified into group A (46 direct exposure to benzene) and group B (26 indirect exposure to benzene) based on their positions, and 29 controls. The differences of gene expression levels were analyzed by software REST 2005. Meanwhile, the peripheral blood leukocytes, hemoglobin and platelet of workers and controls were counted. Benzene content was measured in the samples of toluene, used as raw material, and spraying agents and benzene, toluene and xylene concentrations in the air of workplaces were monitored., Results: There were no significant differences in the mRNA expression levels of p53, Ku80, Ape1 and Mdm-2 between group A or group B and control group (P > 0.05). The expression up-regulation of p21 mRNA was found, but without significant difference (P > 0.05). However, the mRNA expression levels of Rad51, Bcl-2, Bax, Xpa and Xpc in group A and Rad51 in group B were downregulated significantly (P < 0.05 or P < 0.01). Moreover, both the counts of white blood cell, hemoglobin and platelet in group A were (4.93 +/- 1.27) x 10(9)/L, (123.97 +/- 11.80) g/L and (124.02 +/- 41.22) x 10(9)/L respectively and platelet in group B (135.80 +/- 39.44) x 10(9)/L were significantly lower than in control group (P < 0.05 or P < 0.01)., Conclusion: The mRNA expression levels of some p53-mediating DNA damage and repair genes are downregulated in the workers chronically exposed to low benzene concentration. The working environment impacts on health of group A workers are greater than the ones of group B.