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BACKGROUND: Bevacizumab resistance poses barriers to targeted therapy in clear cell renal cell carcinoma (ccRCC). Whether there exist epigenetic targets that modulate bevacizumab sensitivity in ccRCC remains indefinite. The focus of this study is to explore the role of UCHL1 in ccRCC. METHODS: Both in vitro and in vivo experiments were utilized to investigate the roles of UCHL1 in ccRCC. In vivo ubiquitination assays were performed to validate the posttranslational modification of KDM4B by UCHL1. Luciferase reporter and chromatin immunoprecipitation (ChIP) assays were utilized to explore KDM4B/VEGFA epigenetic regulations. RESULTS: UCHL1 was increased in ccRCC and associated with unfavorable survival outcomes in patients. UCHL1 was required for ccRCC growth and migration. Mechanistically, the wild-type UCHL1, but not C90A mutant, mediated the deubiquitination of KDM4B and thereby stabilized its proteins. KDM4B was up-regulated in ccRCC and potentiated cell growth. UCHL1 depended on KDM4B to augment ccRCC malignancies. Targeting UCHL1 suppressed tumor growth, colony formation, and migration abilities, which could be rescued by KDM4B. Furthermore, KDM4B was directly bound to the promoter region of VEGFA, abolishing repressive H3K9me3 modifications. KDM4B coordinated with HIF2α to activate VEGFA transcriptional levels. UCHL1-KDM4B axis governs VEGFA levels to sustain the angiogenesis phenotypes. Finally, a specific small-molecule inhibitor (6RK73) targeting UCHL1 remarkably inhibited ccRCC progression and further sensitized ccRCC to bevacizumab treatment. CONCLUSION: Overall, this study defined an epigenetic mechanism of UCHL1/KDM4B in activating VEGF signaling. The UCHL1-KDM4B axis represents a novel target for treating ccRCC and improving the efficacy of anti-angiogenesis therapy.

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Artificial joint revision surgery, as an increasingly common surgery in orthopedics, often requires patient-specific prostheses to repair the bone defect. Porous tantalum is a good candidate due to its excellent abrasion and corrosion resistance and good osteointegration. Combination of 3D printing technology and numerical simulation is a promising strategy to design and prepare patient-specific porous prostheses. However, clinical design cases have rarely been reported, especially from the viewpoint of biomechanical matching with the patient's weight and motion and specific bone tissue. This work reports a clinical case on the design and mechanical analysis of 3D-printed porous tantalum prostheses for the knee revision of an 84-year-old male patient. Particularly, standard cylinders of 3D-printed porous tantalum with different pore size and wire diameters were first fabricated and their compressive mechanical properties were measured for following numerical simulation. Subsequently, patientspecific finite element models for the knee prosthesis and the tibia were constructed from the patient's computed tomography data. The maximum von Mises stress and displacement of the prostheses and tibia and the maximum compressive strain of the tibia were numerically simulated under two loading conditions by using finite element analysis software ABAQUS. Finally, by comparing the simulated data to the biomechanical requirements for the prosthesis and the tibia, a patient-specific porous tantalum knee joint prosthesis with a pore diameter of 600 µm and a wire diameter of 900 µm was determined. The Young's modulus (5719.32 ± 100.61 MPa) and yield strength (172.71 ± 1.67 MPa) of the prosthesis can produce both sufficient mechanical support and biomechanical stimulation to the tibia. This work provides a useful guidance for designing and evaluating a patient-specific porous tantalum prosthesis.

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ETHNOPHARMACOLOGICAL RELEVANCE: Fuzheng Xiaoai Decoction 1 (FZXAD1) is a clinical experience prescription for the treatment of cancer patients at an advanced stage. FZXAD1 has been used for more than 10 years in the clinic and can effectively improve the deficiency syndrome of cancer patients. However, its mechanisms need further clarification. AIM OF THE STUDY: To check the effects of FZXAD1 in colon 26 (C26) cancer cachexia mice and try to clarify the mechanisms of FZXAD1 in ameliorating cancer cachexia symptoms. MATERIALS AND METHODS: An animal model of cancer cachexia was constructed with male BALB/c mice bearing C26 tumor cells. Food intake, body weight and tumor size were measured daily during the animal experiment. Tissue samples in different groups including tumor and gastrocnemius muscle, were dissected and weighed at the end of the assay. Serum biochemical indicators such as total protein (TP), glucose (GLU) and alkaline phosphatase (ALP) were also detected. Network pharmacology-based analysis predicted the possible targets and signaling pathways involved in the effects of FZXAD1 on cancer cachexia therapy. Western blotting assays of the gastrocnemius muscle tissues from C26 tumor-bearing mice were then used to confirm the predicted possible targets of FZXAD1. RESULTS: The results of animal experiments showed that FZXAD1 could ameliorate cancer cachexia by alleviating the muscle wasting as well as kidney atrophy and increasing the body weight of cancer cachexia mice. AKT1, MTOR, MAPK3, HIF1A and MAPK1 were predicted as the core targets of FZXAD1. Western blotting confirmed the prediction that FZXAD1 increased the expression levels of phosphorylated Akt and mTOR in the muscle tissues. In addition, FZXAD1 treatment obviously ameliorated the increased levels of HIF-1α and phosphorylated Erk1/2 in C26 tumor-bearing mice. CONCLUSION: FZXAD1 effectively ameliorated cancer cachexia in an animal model of mice, which is consistent with its efficacy in the treatment of cancer patients. The mechanisms of FZXAD1 might be mainly based on its alleviating effects on muscle atrophy by activating the Akt-mTOR pathway and thus helping to maintain body weight.

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Magnetic separation is an effective method to recover iron from steel slag. However, the ultra-fine tailings generated from steel slag become a new issue for utilization. The dry separation processes generates steel slag powder, which has hydration activity and can be used as cement filler. However, wet separation processes produce steel slag mud, which has lost its hydration activity and is no longer suitable to be used as a cement filler. This study investigates the potential of magnetically separated steel slag for carbonation curing and the potential use of the carbonated products as an artificial reef. Steel slag powder and steel slag mud were moulded, carbonation-cured and seawater-cured. Various testing methods were used to characterize the macro and micro properties of the materials. The results obtained show that carbonation and hydration collaborated during the carbonation curing process of steel slag powder, while only carbonation happened during the carbonation curing process of steel slag mud. The seawater-curing process of carbonated steel slag powder compact had three stages: C-S-H gel formation, C-S-H gel decomposition and equilibrium, which were in correspondence to the compressive strength of compact increasing, decreasing and unchanged. However, the seawater-curing process of carbonated steel slag mud compact suffered three stages: C-S-H gel decomposition, calcite transfer to vaterite and equilibrium, which made the compressive strength of compact decreased, increased and unchanged. Carbonated steel slags tailings after magnetic separation underwent their lowest compressive strength when seawater-cured for 7 days. The amount of CaO in the carbonation active minerals in the steel slag determined the carbonation consolidation ability of steel slag and durability of the carbonated steel slag compacts. This paper provides a reference for preparation of artificial reefs and marine coagulation materials by the carbonation curing of steel slag.

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OBJECTIVES: This study aims to investigate the underlying mechanisms of KAT2A/MCT1 axis in renal cell carcinoma (RCC), providing potential therapeutic targets. METHODS: We obtained the expression data of KAT2A and MCT1 from The Cancer Genome Atlas Kidney Clear Cell Carcinoma (TCGA-KIRC) and International Cancer Genome Consortium (ICGC) databases. Differential analysis was conducted via the limma package. The CCK8 assay, soft agar assay, clone formation assay, and patients-derived organoid models were used to detect cell growth. The transwell and wound-healing assays were utilized to detect cell migration. The in vitro and in vivo assays were further conducted to assess the oncogenic roles of KAT2A. The transcriptome sequencing and chromatin immunoprecipitation (ChIP) sequencing were conducted to screen KAT2A downstream targets. The dose-effect curves were used to detect the 50% inhibiting concentration (IC50) of AZD3965. Data analysis was performed in the Graphpad Prism (Version 8.3.0) and R software (Version 3.6.1). RESULTS: Our study found that KAT2A was highly expressed in RCC versus normal samples. Prognostic analysis indicated that a high KAT2A was an independent biomarker and associated with poor survival outcomes. KAT2A could promote RCC proliferation and distal metastasis in vitro and in vivo. Transcriptome analysis and ChIP-seq were combined to find that KAT2A mainly regulated the glycolytic process. Validation and rescue assays revealed that MCT1 was the downstream target of KAT2A, and KAT2A depended on MCT1 to promote RCC malignant phenotypes. Lastly, MCT1 inhibitor (AZD3965) was effective to treat KAT2A-induced RCC progression. CONCLUSION: Our study indicated that KAT2A was an oncogenic chromatin modifier that promotes RCC progression by inducing MCT1 expression. We proposed that MCT1 inhibitor (AZD3965) was useful for suppressing RCC.

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Osteoblasts actively generate cell traction force (CTF) to sense chemical and mechanical microenvironments. Fluid shear stress (FSS) is a principle mechanical stimulus for bone modeling/remodeling. FSS and CTF share common interconnected elements for force transmission, among which the role of the protein-material interfacial force (Fad) remains unclear. Here, we found that, on the low Fad surface (5.47 ± 1.31 pN/FN), CTF overwhelmed Fad to partially desorb FN, and FSS exacerbated the desorption, resulting in disassembly of the actin cytoskeleton and focal adhesions (FAs) to reduce CTF and establishment of a new mechanical balance at the FN-material interface. Contrarily, on the high Fad surface (27.68 ± 5.24 pN/FN), pure CTF or the combination of CTF and FSS induced no FN desorption, and FSS promoted assembly of actin cytoskeletons and disassembly of FAs, regaining new mechanical balance at the cell-FN interface. These results indicate that Fad is a mechanical regulator for transmission of CTF and FSS, which has never been reported before.

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The transmission of cell traction force (CTF) to underlying biomaterials is essential for adhered cells to measure and respond to their mechanical microenvironment. Given that the protein layer adsorbed on materials lies between the cells and materials, we hypothesize that the interfacial strength of protein-material interfaces (i.e., the adsorption force of proteins, Fad) should have an important role in regulating the transmission of CTF. To test this hypothesis, rat mesenchymal stem cells (rMSCs) were cultured on poly(dimethyl siloxane) (PDMS) substrates with different Fad of fibronectin (FN), and the transmission of CTF was observed by immunofluorescence staining of FN and deformation of PDMS. As revealed, FN on substrates with low Fad is more liable to be desorbed by CTF, which prevents the transmission of CTF to substrates. In contrast, high Fad facilitates the transmission of CTF from rMSCs to the FN layer and PDMS substrates so that rMSCs can perceive the mechanical properties of substrates. We further demonstrated that the divergent transmission of CTF on low and high Fad substrates regulates the lineage specifications of rMSCs. Our study confirms the important role of Fad in CTF transmission and provides a new perspective to gain insights into cell-material interactions and cell fates, which may help to guide the design of better biomaterials.

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PURPOSE: Cisplatin plus gemcitabine (GEM) is a standard regimen for the first-line treatment of advanced non-small cell lung cancer. The aim of this study was to prepare biocompatible and biodegradable polymeric prodrugs and construct nanoparticles (NPs) with layer-by-layer (LbL) technique. METHODS: Platinum (Pt) (IV) complex with a carboxyl group was conjugated to the amino group of chitosan (CH), resulting in a CH-Pt conjugation with positive charge. GEM with amino group was conjugated to the carboxyl group of hyaluronic acid (HA), resulting in a HA-GEM conjugation with negative charge. Novel LbL NPs consisting of the CH-Pt core and the HA-GEM layer, named as HA-GEM/CH-Pt NPs, were constructed. The physicochemical properties of the HA-GEM/CH-Pt NPs were investigated. In vitro cytotoxicity against human non-small lung cancer cells (NCl-H460 cells) was investigated, and in vivo antitumor efficiency was evaluated on mice bearing NCl-H460 cells xenografts. RESULTS: HA-GEM/CH-Pt NPs have a size of about 187 nm, a zeta potential value of -21 mV and high drug encapsulation efficiency of 90%. The drug release of HA-GEM/CH-Pt NPs exhibited a sustained behavior. HA-GEM/CH-Pt NPs could significantly enhance in vitro cytotoxicity and in vivo antitumor effect against lung cancer animal model compared to the single-drug-loaded NPs and free drug solutions. CONCLUSION: The results demonstrated that the HA-GEM/CH-Pt NPs might be a promising system for the synergetic treatment of lung carcinoma.

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A simple, rapid and specific high performance liquid chromatography-electrospray ionization tandem mass spectrometry method was developed for simultaneous determination of dabigatran etexilate (BIBR 1048 MS), the intermediate metabolite (BIBR 1087 SE) and dabigatran (BIBR 953 ZW). In this method, a stacked protein precipitation with methanol was performed in Sirocco 96-well filtration plates to extract analytes using only 50µL plasma. The analysis was performed on an Ultimate TM XB-C18 (4.6×50mm, 5µm) column using gradient elution with a mobile phase composed of methanol containing 0.01% formic acid and pure water at a flow rate of 0.3mL/min. The gradient was set to 90% methanol containing 0.01% formic acid for the first 1.0min, after which it dropped to 10%, and then was kept at 10% for the next 5min followed by an additional 1.0min at the initial composition of 90% methanol containing 0.01% formic acid for equilibration. Detection was performed on a triple-quadrupole mass spectrometer electrospray ionization interface in positive ion mode. Linear calibration curves were obtained over the concentration ranges of 1-500ng/mL for all analytes. The validated LC-MS/MS method for its selectivity, sensitivity, linearity, precision, accuracy, recovery, matrix effect and stability had been successfully applied to a pharmacokinetic study of analytes in rat plasma following a single oral administration of 15mg/kg dabigatran etexilate.

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Fermented soybean foods have been shown to reduce incidence of diabetes and improve insulin sensitivity. 6-Hydroxydaidzein (6-HD) is a bioactive ingredient isolated from fermented soybean. In this study, we examined the effects of 6-HD on adipocyte differentiation and insulin-stimulated glucose uptake, as well as the mechanisms involved. In our experiments, 6-HD enhanced 3T3-L1 adipocyte differentiation and insulin-stimulated glucose uptake in a dosage-dependent manner. In addition, 6-HD increased peroxisome proliferator-activated receptor gamma (PPARγ) gene expression and PPARγ transcriptional activity. 6-HD increased CCAAT/enhanced binding protein alpha (C/EBPα) expression as well. Although having no effects on glucose transporter type 4 (GLUT4) gene expression, 6-HD facilitated GLUT4 protein translocation to the cell membranes. Our results indicate that 6-HD exhibited the actions of promoting adipocyte differentiation and improving insulin sensitivity by increasing the expression of C/EBPα and facilitating the translocation of GLUT4 via the activation of PPARγ, suggesting that 6-HD can be promising in diabetes management.

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A comprehensive two-dimensional (2D) separation is one that employs two separation dimensions (columns) and draws on all of the available resolving power from each of the dimensions of separate the components in a sample. In this study, a comprehensive 2D chromatography approach was developed for the separation and identification of membrane permeable compounds in a famous traditional Chinese medicine of Schisandra chinensis. The first dimensional column was the immobilized liposome chromatography (ILC) column, which mimics the biological membranes and can be used to study drug-membrane interactions in liquid chromatography. Using an automatic ten-port switching valve equipped with two sample loops, the section of the first-dimension was introduced in the second-dimension consist of a silica monolithic column. More than 40 components in Schisandra chinensis were resolved by using the developed separation system and among them 14 compounds were identified interacting with the ILC column based on their retention action, UV and mass data. With this comprehensive 2D-HPLC system, the three-dimensional chromatographic fingerprints of Schisandra chinensis were preliminarily established and processed by using principal component analysis and hierarchical clustering analysis. The obtained information can distinguish the unacceptable samples of the quality control. The result demonstrated that the 2D biochromatography system has been demonstrated to have more advantages of finding strong binding bioactive components, providing an enhanced peak capacity, good sensitivity and powerful resolution biological fingerprinting analysis of complex TCMs, which was a useful means to control the quality of and to clarify the membrane permeability of the compounds in Schisandra chinensis.

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Activity-guided fractionation of the stems of Marsdenia tenacissima led to the isolation of five new pregnane glycosides, namely marstenacissides E (1), F (2), G (3), H (4), and I (5). Their structures were determined on the basis of (1)H and (13)C NMR, COSY, TOCSY, ROESY, and FABMS experiments.

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The prevalence of metabolic syndrome has increased in modern society and the condition is proving to be a common precursor of cardiovascular disease. The aim of the present study was to investigate whether astragaloside IV, a major active constituent of Astragalus membranaceus (Fisch) Bge., is able to prevent the development of hypertension and endothelial dysfunction in fructose-fed rats. Rats were fed with 10% fructose in their drinking water for 8 weeks. From the beginning of week 5, two groups of fructose-fed rats were treated with 0.5 or 2 mg/kg, i.p., astragaloside IV. Another group of fructose-fed rats, injected with the same volume of vehicle (dimethylsulfoxide, DMSO) from week 5, served as the control group. At the end of the treatment period, blood pressure, blood glucose, glucose tolerance, blood insulin and lipids were determined. In addition, in vitro experiments were conducted at the end of the eight week treatment period to evaluate endothelium-dependent aortic vasorelaxation, as well as myocardial and aortic tissue levels of nitrate and nitrite (NOx) and cGMP. Fructose-fed rats developed clustering signs of metabolic syndrome, such as increased bodyweight, mild hypertension, hyperinsulinaemia, hypertriglyceridaemia, impaired glucose tolerance and impaired endothelium-dependent vasorelaxation. Administration of astragaloside IV reduced blood pressure and triglyceride levels in fructose-fed rats and high dose of astragaloside IV also improved glucose tolerance and endothelium-dependent vasorelaxation. The astragaloside IV-induced improvement in vasorelaxation was associated with increased levels of aortic NOx and cGMP and was abrogated by blockade of nitric oxide synthase with NG-nitro-l-arginine methyl ester (l-NAME). On the basis of its favourable effects on lipid metabolism, endothelium-dependent vasorelaxation and the nitric oxide-cGMP-related pathway, astragaloside IV may be useful in ameliorating food-induced metabolic syndrome.

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A simple, sensitive and rapid method for analysis of six lignans in rat plasma after oral administration of Schisandra chinensis extracts, utilizing liquid chromatography tandem mass spectrometry (LC-MS), was established and validated. Plasma samples were prepared by one-step protein precipitation using acetonitrile and the analytes were separated on an SB-C18 column (100 mm × 3.0 mm, 3.5 µm) with the mobile phase of acetonitrile-water at a flow-rate of 0.8 mL/min. Analytes were determined in a single-quadrupole mass spectrometer in the selected ion monitoring (SIM) mode using electrospray source with positive mode. The method was proved to be rapid, sensitive and reproducible, and it was successfully applied to the pharmacokinetic studies of six lignans in rat plasma after oral administration of Schisandra chinensis extracts. In this research, the pharmacokinetics of deoxyschisandrin was also studied following oral administration of the pure deoxyschisandrin. It was found that most of the pharmacokinetic parameters of deoxyschisandrin in the extract were changed significantly compared with those in monomer. The content assay also revealed that the concentrations of the lignan in the extract increased in vivo compared with the pure monomer. Some ingredients in the extract may increase the dissolution of deoxyschisandrin, delay its elimination and enhance its bioavailability in rat.

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AIM: To investigate the effects of sodium danshensu on vessel function in isolated rat aortic ring. METHODS: Thoracic aortae from normal rats were isolated and equilibrated in organ bath with Krebs-Henseleit buffer and ring tension was recorded. Effects of sodium danshensu on basal tonus of the vessel and its effects on vessel contraction and relaxation with or without endothelium were observed. RESULTS: In thoracic arteries under basal tonus, sodium danshensu (0.3-3 g/L) produced a dose-dependent transient contraction. In phenylephrine-precontracted thoracic arteries with or without endothelium, low concentration (0.1-0.3 g/L) of sodium danshensu produced a weak contraction, while high concentrations (1-3 g/L) produced a pronounced vasodilator after a transient vasocontraction. Pre-incubation with sodium danshensu could inhibit vessel contraction induced by phenylephrine and potassium chloride in a concentration-dependent way. Sodium danshensu inhibited phenylephrine- and CaCl(2)-induced vasoconstriction in Ca(2+)-free medium. Pre-incubation with tetraethylammonium, a non-selective K(+) channel blocker, and apamin, a small-conductance calcium-activated K(+) channel blocker partially antagonized the relaxation response induced by sodium danshensu. However, iberiotoxin (big-conductance calcium-sensitive K(+) channel blocker), barium chloride (inward rectifier K(+) channel blocker), and glibencalmide (ATP-sensitive K(+) channel blocker) had no influence on the vasodilation effect of sodium danshensu. CONCLUSION: Sodium danshensu showed a biphasic effects on vessel tension. While low dosage of sodium danshensu produced small contraction possibly through transient enhancement of Ca(2+) influx, high dosage produced significant vasodilation mainly through promoting the opening of non-selective K(+) channels and small-conductance calcium-sensitive K(+) channels in the vascular smooth muscle cells.

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AIM: To separate and identify the chemical constituents of the aril of Torreya grandis cv. Merrilli. METHODS: Three lignins were isolated by chromatography and their chemical structures were elucidated by IR, EI-MS, 1HNMR, 13CNMR, DEPT and 2D-NMR spectral methods. RESULTS: Three lignins were identified as pinonesinol, dihydrodehydrodiconiferylalcohol and (7,8-cis-8,8'-trans)-2',4'dihydroxyl-3, 5-dimethoxy-lariciresinol. CONCLUSION: These compounds were isolated from this plant for the first time, and compound III is a new compound.

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Four new pregnane glycosides, named marstenacissides A (1), B (2), C (3), and D (4), have been isolated from the stems of Marsdenia tenacissima. Their structures were established on the basis of chemical and spectral methods.

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