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Background: Tripterygium wilfordii Hook. F. (TwHF), a traditional Chinese medicine, is widely used in treatment of rheumatoid arthritis (RA), etc.. Due to multiorgan toxicity, particularly hepatotoxicity, the application of TwHF is restricted. In order to clarify the hepatotoxic substances, zebrafishes, hepatocytes and macrophages were used for screening based on hepatotoxic injury patterns. It provided a basis for further hepatotoxic mechanism of TwHF. Methods: First, 12 compounds were selected according to the chemical categories of TwHF. The fluorescence area and fluorescence intensity of zebrafishes liver were observed and calculated. The viability of two hepatocyte lines were detected by CCK8. TNF-α and IL-1β mRNA expression of BMDM was used to evaluate macrophage activation, a factor of potential indirect hepatotoxicity. Finally, hepatotoxic characteristics of 4 representative components were verified by mice in vivo. Results: Parthenolide, Triptolide, Triptonide, Triptobenzene H, Celastrol, Demethylzeylasteral, Wilforlide A, Triptotriterpenic acid A and Regelidine can significantly reduce the fluorescence area and fluorescence intensity of zebrafish liver. The viability of L-02 or AML-12 was significantly inhibited by Parthenolide, Triptolide, Triptonide, Celatrol, Demethylzeylasteral, Triptotriterpenic acid A. Parthenolide, Triptolide, Triptonide, Celatrol, Demethylzeylasteral and Triptobenzene H significantly increased TNF-α and IL-1β mRNA levels of BMDM, while Triptophenolide, Hypodiolide and Wilforine significantly reduced. Triptotriterpenic acid A, Celastrol and Triptobenzene H at dose of 10 mg/kg significantly increased the levels of mice serum ALT and AST, and aggravated liver inflammation. Conclusions: Parthenolide, Triptolide, Triptonide, Celatrol, Demethylzeylasteral, Triptotriterpenic acid A and Triptobenzene H might be main hepatotoxic components of TwFH. Among them, Triptotriterpenic acid A only presented a direct hepatotoxicity. Triptobenzene H showed indirect liver damage by activating macrophages. Parthenolide, Triptolide, Triptonide, Celatrol, Demethylzeylasteral could directly and indirectly cause liver injury.

Background: Metastasis is the most common cause of lethal outcome in various types of cancers. Although the cell proliferation related metabolism rewiring has been well characterized, less is known about the association of metabolic changes with tumor metastasis. Herein, we demonstrate that metastatic tumor obtained a mesenchymal phenotype, which is obtained by the loss of tumor suppressor NDRG2 triggered metabolic switch to glutamine metabolism. Methods: mRNA-seq and gene expression profile analysis were performed to define the differential gene expressions in primary MEC1 and metastatic MC3 cells and the downstream pathways of NDRG2. NDRG2 regulation of Fbw7-dependent c-Myc stability were determined by immunoprecipitation and protein half-life assay. Luciferase reporter and ChIP assays were used to determine the roles of Akt and c-Myc in mediating NDRG2-dependent regulation of ASCT2 in in both tumor and NDRG2-knockout MEF cells. Finally, the effect of the NDRG2/Akt/c-Myc/ASCT2 signaling on glutaminolysis and tumor metastasis were evaluated by functional experiments and clinical samples. Results: Based on the gene expression profile analysis, we identified metastatic tumor cells acquired the mesenchymal-like characteristics and displayed the increased dependency on glutamine utilization. Further, the gain of NDRG2 function blocked epithelial-mesenchymal transition (EMT) and glutaminolysis, potentially through suppression of glutamine transporter ASCT2 expression. The ASCT2 restoration reversed NDRG2 inhibitory effect on EMT program and tumor metastasis. Mechanistic study indicates that NDRG2 promoted Fbw7-dependent c-Myc degradation by inhibiting Akt activation, and subsequently decreased c-Myc-mediated ASCT2 transcription, in both tumor and NDRG2-knockout MEF cells. Supporting the biological significance, the reciprocal relationship between NDRG2 and ASCT2 were observed in multiple types of tumor tissues, and associated with tumor malignancy. Conclusions: NDRG2-dependent repression of ASCT2 presumably is the predominant route by which NDRG2 rewires glutaminolysis and blocks metastatic tumor survival. Targeting glutaminolytic pathway may provide a new strategy for the treatment of metastatic tumors.

The present study aimed to investigate the cardioprotective role of berberine in sepsis-induced cardiac dysfunction and consider the underlying mechanisms. C57BL/6J mice were randomized into four groups, namely, Control, lipopolysaccharide (LPS), LPS + berberine and LPS + N

BACKGROUND Studies in ApoE knockout mice have shown that pseudolaric acid B (PB) can act as an immunomodulatory drug and attenuate atherosclerosis progression by modulating monocyte/macrophage phenotypes. Our previous study demonstrated that high salt intake could shift the phenotype of monocytes/macrophages to an inflammatory phenotype, and that this shift was related to hypertension and hypertensive left ventricular (LV) remodeling. However, no comprehensive assessment of the effects of PB on hypertensive LV remodeling has been conducted. MATERIAL AND METHODS In this study, RAW264.7 macrophages cultured with different concentrations of NaCl were used to investigate the modulating effects of PB on macrophage phenotype. Furthermore, N-nitro-L-arginine methyl ester hypertensive mice were used to investigate the modulating effects of PB on monocyte phenotype. LV remodeling was investigated by echocardiography. LV morphologic staining (for cardiomyocyte hypertrophy and collagen deposition) was performed at the time of sacrifice. RESULTS The results showed that PB significantly improved the viability of RAW264.7 cells, suppressed their phagocytic and migration abilities, and inhibited their phenotypic shift to M1 macrophages. In addition, the blood pressure of PB-treated mice was significantly decreased relative to that of control mice. Furthermore, after PB treatment, the percentage of Ly6Chi monocytes was significantly decreased while that of Ly6Clo monocytes was apparently increased. Moreover, PB preserved LV function and alleviated myocardial fibrosis and cardiomyocyte hypertrophy as measured at the end of the experimental period. The transfer of monocytes from PB-treated mice to hypertensive mice achieved the same effects. CONCLUSIONS Together, these findings indicate that PB exerts its protective effects on hypertensive LV remodeling by modulating monocyte/macrophage phenotypes and warrants further investigation.

The construction scale of the shield tunnel underpass is expanding day by day. In order to study the safety influence and deformation control of the shield tunnel underpass on the existing tunnel, the LSSVM model is established. Based on the collected soil storage pressure, foam volume, simultaneous grouting volume and other six shield construction parameters and corresponding sample data of the tunnel bottom displacement, the horizontal displacement and settlement displacement of the existing tunnel bottom caused by the approach construction are predicted. Taking a subway project as an example, the research results show that the prediction model of shield tunneling under the existing tunnel bottom level and settlement displacement has strong generalization ability and rapid and accurate prediction effect. This method can provide reference for similar projects.

Poor frost resistance of concrete will accelerate the deterioration of concrete structure and shorten the service life of concrete. This paper predicts the frost resistance of concrete and selects the initial index of factors that affect the frost resistance of concrete. The random forest algorithm is introduced to remove the unimportant indicators in the initial indicator system of the concrete mix ratio and determine the optimal indicator combination. The relative dynamic elastic modulus is used as the output index of random forest to predict the frost resistance of concrete. The proposed random forest model provides a reasonable and effective method for concrete frost resistance prediction.

Destruction of endothelial cells (ECs) function is involved in the structural and functional pathophysiological processes of preeclampsia (PE). Vascular endothelial injury may pre-exist for several years in women that develop PE and may pose increased risks for hypertension, coronary artery disease, and type-2 diabetes mellitus. Previous findings showed that Elabela (ELA), the endogenous ligand of the apelin (APJ) receptor expressed mainly on ECs, may play a protective role in early pregnancy and prevent PE. However, the exact functional role and molecular mechanisms of ELA are unclear. Here, we aimed to classify whether and how ELA improves EC function via the ELA-APJ axis. Two human umbilical vein endothelial cell (HUVEC) lines, namely HUVECs and EA.hy926, were treated with ELA, and then their cellular activities were studied by performing CCK-8 tests, scratch-wound analysis, and tube-formation assays. Doses of ELA exceeding 0.01 μmol/L markedly improved the cell viability, migration, and tube formation ability of HUVECs and EA.hy926 cells. Western blot analysis indicated that the above effects caused by ELA were related to upregulation of the APJ receptor and activation of PI3K/Akt signalling. Further verification tests were performed using the PI3K inhibitor wortmannin, and the results illustrated that inhibiting PI3K/Akt signalling blocked the positive effects of ELA on EC function and APJ receptor expression. Taken together, our findings indicate that ELA may alter EC function via the ELA-APJ axis and PI3K/Akt signalling and that ELA shows promise for use in endothelial dysfunction therapy for preventing and treating PE.

Real-time ultrasound elastography (RTE) is used to examine liver fibrosis and benign and malignant lesions, but its use for the diagnosis of liver trauma has not been examined. The purpose of this study was to examine the use of RTE for the evaluation of differential penetrating liver trauma in a rabbit model.Eighty New Zealand rabbits were divided into 2 groups. In one group, a single incision (type "-" lesion) was made, and in the other group a hash mark incision (type "#" lesion) was made (about 0.5cm in depth; 1.0-2.0cm in length). RTE was performed at 10, 30, and 60min after injury.There were no differences in mean RTE scores between the 2 types of lesions at 10 and 30min. However, the mean values for the 2 types of lesions increased from 10min to 60min (type '-' lesion: 0.88±0.32 to 2.06±0.88; type '#' lesion: 0.89±0.34 to 2.63±1.16). At 60min, the mean elasticity score in the type '#' lesion group was significantly higher than in the type '-' lesion group (P.001). Strain ratios were not different between the groups at each time point, but in each group the values decreased from the 10min time point to the 60min time point (P-value for the trends,.001).RTE may be able to distinguish mild or severe penetrating liver trauma at 60min or more after injury.

Endophytic fungi are symbiotic with plants and possess multienzyme systems showing promising metabolite potency with region selectivity and stereoselectivity. The aim of this study was to use these special microorganisms as an in vitro model to mimic the potential mammalian metabolites of a natural iridoid gentiopicroside (GPS, compound 1). The fungi isolated from a medicinal plant, Dendrobium candidum Wall. ex Lindl., were screened for their biotransformation abilities with GPS as the substrate, and one strain with high converting potency was identified as Penicillium crustosum 2T01Y01 on the basis of the sequence of the internal transcribed spacer of the ribosomal DNA region. Upon the optimized incubation of P. crustosum 2T01Y01 with the substrate, seven deglycosylated metabolites were detected by ultraperformance liquid chromatography/quadrupole time of flight mass spectrometry (UPLC/Q-TOF MS). Preparative-scale biotransformation with whole cells of the endophytic fungus resulted in the production of five metabolites, including three novel ones, 5α-(hydroxymethyl)-6β-methyl-3,4,5,6-tetrahydropyrano[3,4- c ]pyran-1(8 H )-one (compound 2), ( Z )-4-(1-hydroxybut-3-en-2-yl)-5,6-dihydropyran-2-one (compound 3), and ( E )-4-(1-hydroxybut-3-en-2-yl)-5,6-dihydropyran-2-one (compound 4), along with two known ones, 5α-(hydroxymethyl)-6β-methyl-1 H ,3 H -5,6-dihydropyrano[3,4- c ]pyran-1(3 H )-one (compound 5) and 5α-(hydroxymethyl)-6α-methyl-5,6-dihydropyrano[3,4- c ]pyran-1(3 H )-one (compound 6), aided by nuclear magnetic resonance and high-resolution mass spectral analyses. The other two metabolites were tentatively identified by online UPLC/Q-TOF MS as 5-hydroxymethyl-5,6-dihydroisochromen-1-one (compound 7) and 5-hydroxymethyl-3,4,5,6-tetrahydroisochromen-1-one (compound 8), and compound 8 is a new metabolite. To test the metabolic mechanism, the β-glucosidase activity of the fungus P. crustosum 2T01Y01 was assayed with ρ-nitrophenyl-β- d -glucopyranoside as a probe substrate, and the pathway of GPS biotransformation by strain 2T01Y01 is proposed. In addition, the hepatoprotective activities of GPS and metabolite compounds 2, 5, and 6 against human hepatocyte line HL-7702 injury induced by hydrogen peroxide were evaluated.

Background The most common reason for malignant tumor treatment failure is recurrence and metastasis. Metastasis-associated in colon cancer-1 (MACC1) was originally identified as a metastatic and prognostic biomarker for colon cancer and later other solid tumors. Kangai 1 (KAI1), a marker of suppressor of metastasis, is also associated with metastasis and poor prognosis in many tumors. However, the prognostic value of either MACC1 or KAI1 in gastric adenocarcinoma (GAC) is unclear. In this study, we explored the relationship between MACC1 and KAI1 expression, as well as their respective correlation with clinicopathological features, to determine if either could be helpful for improvement of survival prognosis in GAC patients. Methods The expression levels of both MACC1 and KAI1 in 325 whole-tissue sections of GAC were examined by immunohistochemistry. Clinical data was also collected. Results MACC1 was significantly overexpressed in GAC tissues when compared to levels in normal gastric tissues; KAI1 was significantly down-expressed in GAC tissues when compared to levels in normal gastric tissues. Investigation of association between MACC1 and KAI1 protein levels with clinicopathological parameters of GAC indicated association between the expression of each with tumor grade, lymph node metastasis, invasive depth, and TNM stages. The overall survival time of patients with MACC1- or KAI1-positive GAC tumors was significantly shorter or longer than that of those who were negative. Importantly, multivariate analysis suggested that positive expression of either MACC1 or KAI1, as well as TNM stage, could be independent prognostic factors for overall survival in patients with GAC. Conclusions MACC1 and KAI1 may represent promising metastatic and prognostic biomarkers, as well as potential therapeutic targets, for GAC.

Hepatic fibrosis is a common pathological process of chronic liver diseases and would lead to cirrhosis, and Fuzheng Huayu (FZHY) is an effective Chinese herbal product against liver fibrosis. This study observes FZHY influence on proteome of fibrotic liver with differential proteomic approach and aims to understand FZHY multiple action mechanisms on liver fibrosis. The liver fibrosis models were induced with intraperitoneal injection of dimethylnitrosamine for 4 weeks in rats and divided into model control (model) and FZHY-treated (FZHY) groups, while normal rats were used as normal control (normal). After model establishment, rats in FZHY groups were administered 4 g/kg wt of FZHY for 4 weeks, and normal and model groups were given the same volume of saline. The liver proteins in the above 3 groups were separated by two-dimensional gel electrophoresis (2-DE), the differentially expressed spots were analyzed and compared between normal and model or model and FZHY groups, and then the proteins were identified with mass spectrum analysis and validated partially with western blot and real-time PCR. 1000~1200 spots were displayed on each 2D gel, and a total of 61 protein spots were found with significant intensity difference between normal control or FZHY and model control. 23 most obviously differential spots were excised, and in-gel digestion and 21 peptide mass fingerprints (PMF) were obtained with MALDI-TOF MS analysis, and 14 proteins were identified through protein database searching. Among 14 differentially expressed proteins, 8 proteins in normal and FZHY groups had the same tendency of differential expression compared with the ones in model group. And one of them, vimentin, was validated by western blot and real-time PCR analyses. Our study reveals 12 proteins responsible for fibrogenesis induced by DMN in rats, and among them, 8 proteins in fibrotic liver were regulated by FZHY, including aldehyde dehydrogenase, vimentin isoform (CRA_b), gamma-actin, vimentin, fructose-bisphosphate aldolase B, aldo-keto reductase, S-adenosylhomocysteine hydrolase isoform, and HSP90. It indicates that the action mechanism of FZHY antiliver fibrosis may be associated with modulation of proteins associated with metabolism and stress response, as well as myofibroblast activation. The study provides new insights and data for exploring the liver fibrogenesis pathophysiology and FZHY action mechanism against liver fibrosis.

Considerable research have described important regulatory mechanisms that provide immune homeostasis by limiting excessive immune responses and preventing loss of tolerance (1, 2). Naturally occurring regulatory T cells (Tregs) specifically express the transcription factor known as forkhead box 3 (Foxp3), a member of the forkhead/winged-helix family of transcription factors that is essential for the development, maintenance and function of Tregs. Importantly, Foxp3 expression is sufficient to confer suppressive activity to conventional non-Tregs and critical for TCR+ T cells to differentiate to Tregs in the thymus (3). Thus, functional deficiency of Tregs, an abnormality in Foxp3 expression and/or mutations in the Foxp3 gene locus results in increased susceptibility to several autoimmune diseases. Humans that have mutations in the Foxp3 gene develop IPEX (immune dysregulation, polyendocrinopathy and enteropathy, X-linked syndrome), a severe autoimmune disease, resulting in fatal lymphoproliferation at a very early age. A significantly reduced suppressive function of Tregs has been found in patients with polyglandular syndrome type II (APS-II) (4), rheumatoid arthritis (RA) (5), type-I diabetes (6), myasthenia graves (7, 8) and autoimmune hepatitis (9). Patients with multiple sclerosis (MS) not only lose functional suppression (10) but also exhibit decreases in Foxp3 mRNA and protein expressions (11). Primary biliary cirrhosis (PBC) is a progressive autoimmune liver disease characterized by immune-mediated destruction of intrahepatic small bile ducts (12). It has been suggested that the loss of Tregs plays a key role in susceptibility to PBC (13). For example, we have demonstrated a significant decrease in the frequency of Tregs in patients with PBC as well as daughters and sisters of PBC patients (14). We also reported a male child who developed liver dysfunction in association with serum anti-PDC-E2 antibodies at the age of five, who had a complete deficiency of the alpha subunit of the IL-2 receptor (IL-2Rα, CD25) in peripheral blood lymphocytes (15). CD25 is a critical component for functional Treg development and expansion. Mice deficient in IL-2 receptor also demonstrated PDC-like lesions in the liver (16). In agreement with our findings, Ae2-deficient mice that develop features resembling PBC, the proportion of CD4+CD25+FoxP3+ Tregs was reduced (17). Based on the above, we hypothesized that genetic components which are essential for the development and homeostasis of Foxp3+ Tregs is responsible for the loss of self-tolerance in PBC. Hence, we took advantage of Scurfy (Sf) mice which have a Foxp3 gene mutation that results in a deficiency of functional Tregs (18, 19) and examined levels of autoantibodies, cytokines and liver histology. We report herein that Sf mice have serologic, histologic and cytokine features characteristic of autoimmune cholangitis, similar to patients with PBC.

Gentiopicroside (GPS), the main bioactive component in Gentiana scabra Bge., has attracted our attention owing to its high bioactivity, especially the treatment of hepatobiliary disorders. The aglycone form of GPS, a typical secoiridoid glycoside, is considered to be more readily absorbed than its parent drug. This study aimed to identify and characterize the metabolites after GPS incubated with β-glucosidase in buffer solution at 37°C. Samples of biotransformed solution were collected and analyzed by ultraperformance liquid chromatography (UPLC)/quadrupole-time-of-flight mass spectrometry (Q-TOF MS). A total of four metabolites were detected: two were isolated and elucidated by preparative-HPLC and NMR techniques, and one of those four is reported for the first time. The mass spectral fragmentation pattern and accurate masses of metabolites were established on the basis of UPLC/Q-TOF MS analysis. Structure elucidation of metabolites was achieved by comparing their fragmentation pattern with that of the parent drug. A fairly possible metabolic pathway of GPS by β-glucosidase was proposed. The hepatoprotective activities of metabolites M1 and M2 were investigated and the results showed that their hepatoprotective activities were higher than that of parent drug. Our results provided a meaningful basis for discovering lead compounds from biotransformation related to G. scabra Bge. in traditional Chinese medicine.

Multiple reports have suggested that reactive oxygen species (ROS) are implicated in hepatic fibrosis and that they are capable of causing hepatocyte apoptosis in hepatic fibrosis by causing oxidative damage to the liver. Thus, the study of antioxidant compounds may shed light on the treatment of hepatic fibrosis. The aim of the current study was to investigate the protective effects of Cordyceps polysaccharide (CPS), a major antioxidative component of Cordyceps militaris, on hydrogen peroxide (H2O2)-induced cell apoptosis. The data showed that CPS markedly inhibited H2O2-induced mitochondrial dysfunction, lowered cell viability, increased the apoptotic rate, boosted ROS production, decreased mitochondrial membrane potential (MMP), reduced the intracellular adenosine triphosphate (ATP) level, increased the Bax/Bcl-2 ratio and promoted cytochrome C (Cyt C) release. These results indicated that CPS protected HL-7702 cells, which are used as the main model of hepatic fibrosis, against H2O2-induced mitochondrial dysfunction by decreasing ROS production and regulating mitochondrial apoptotic signaling through the Cyt C, Bax and Bcl-2 apoptosis-related proteins.

In this study, we investigated the anti-angiogenic effect of the Chinese herbal decoction Danggui Buxue Tang (DBT;Radix AstragaliandRadix Angelicae sinensisin 5 : 1 ratio) in a rat model of liver fibrosis, in order to elucidate its mechanisms of action against liver fibrosis. Liver fibrosis was induced with CCl4and high-fat food for 6 weeks, and the rats were treated with oral doses of DBT (6 g raw herbs/kg/d) and N-Acetyl-L-cysteine (NAC; 0.1 g/kg/d). The results showed that both DBT and NAC attenuated liver fibrosis and neo-angiogenesis. Furthermore, DBT and NAC improved SOD activity but decreased MDA content and 8-OH-dG in fibrotic livers, with DBT being more effective than NAC. DBT decreased the expression of VEGF, Ang1 and TGF-β1 and their signaling mediators, whereas NAC had no effect on VEGF and VEGFR2 expression. Both DBT and NAC reduced HIF-1αgene and protein expression in fibrotic livers, with DBT being more effective. These data clearly demonstrate that the anti-fibrotic properties of DBT are related to its ability to inhibit angiogenesis and its anti-angiogenic mechanisms are associated with improving oxidative stress, regulating the expression and signaling of angiogenic factors, and especially modulating HIF-1αin fibrotic livers.

The Transfer of Taohe Water Project (TTWP) in its reasoning stage did not put in place a lot of schemes for the selection of water price. This article applies the theory of two-part water pricing and designs 12 selective schemes, after the analyzing, determines the three specific water prices of village life, agricultural irrigation and city water consumption. Further analysis shows that the scope of water price of rural life water consumption and city water consumption are affordable; meanwhile, the water price of agricultural irrigation consumption exceeds what the people can afford, and there is a need of government subsidy.

The fragmentation behaviors of the angular- and linear-type coumarins from Peucedanum praeruptorum Dunn and P. decursivum (Miq.) Maxim were simultaneously investigated by HPLC-DAD/ESI-MSn. For more structural identification, the fragment ions were analyzed and some possible fragmentation pathways were proposed. Different positions and numbers of the substituent also led to different fragment behaviors. Two types of coumarins from P. praeruptorum and P. decursivum were structurally elucidated by these techniques. In addition, UV spectra were applied to support the MS analysis. This is the first time that the two types of coumarins from herbal extracts have been differentiated by HPLC-DAD/ESI-MSn. The method further illustrated the importance of the ESI-MSn technique in the identification of different types of coumarins and was applied for the rapid differentiation of the two herbs.