Protective Effect of Silibinin on Lipopolysaccharide-Induced Endotoxemia by Inhibiting Caspase-11-Dependent Cell Pyroptosis.

OBJECTIVE: To explore the protective effect and the underlying mechanism of silibinin (SIB), one of the active compounds from Silybum marianum (L.) Gaertn in endotoxemia. METHODS: Mouse peritoneal macrophage were isolated via intraperitoneally injection of BALB/c mice with thioglycolate medium. Cell viability was assessed using the cell counting kit-8, while cytotoxicity was determined through lactate dehydrogenase cytotoxicity assay. The protein expressions of interleukin (IL)-1 α, IL-1 ß, and IL-18 were determined by enzyme-linked immunosorbent assay. Intracellular lipopolysaccharide (LPS) levels were measured by employing both the limulus amoebocyte lysate assay and flow cytometry. Additionally, proximity ligation assay was employed for the LPS and caspase-11 interaction. Mice were divided into 4 groups: the control, LPS, high-dose-SIB (100 mg/kg), and low-dose-SIB (100 mg/kg) groups (n=8). Zebrafish were divided into 4 groups: the control, LPS, high-dose-SIB (200 εmol/L), and low-dose-SIB (100 εmol/L) groups (n=30 for survival experiment and n=10 for gene expression analysis). The expression of caspase-11, gasdermin D (GSDMD), and N-GSDMD was determined by Western blot and the expressions of caspy2, gsdmeb, and IL-1 ß were detected using quantitative real-time PCR. Histopathological observation was performed through hematoxylineosin staining, and protein levels in bronchoalveolar lavage fluid were quantified using the bicinchoninicacid protein assay. RESULTS: SIB noticeably decreased caspase-11 and GSDMD-mediated pyroptosis and suppressed the secretion of IL-1 α, IL-1 ß, and IL-18 induced by LPS (P<0.05). Moreover, SIB inhibited the translocation of LPS into the cytoplasm and the binding of caspase-11 and intracellular LPS (P<0.05). SIB also attenuated the expression of caspase-11 and N-terminal fragments of GSDMD, inhibited the relative cytokines, prolonged the survival time, and up-regulated the survival rate in the endotoxemia models (P<0.05). CONCLUSIONS: SIB can inhibit pyroptosis in the LPS-mediated endotoxemia model, at least in part, by inhibiting the caspase-11-mediated cleavage of GSDMD. Additionally, SIB inhibits the interaction of LPS and caspase-11 and inhibits the LPS-mediated up-regulation of caspase-11 expression, which relieves caspase-11-dependent cell pyroptosis and consequently attenuates LPS-mediated lethality.

High-throughput screening of ancient forest plant extracts shows cytotoxicity towards triple-negative breast cancer.

According to the World Health Organization, women's breast cancer is among the most common cancers with 7.8 million diagnosed cases during 2016-2020 and encompasses 15 % of all female cancer-related mortalities. These mortality events from triple-negative breast cancer are a significant health issue worldwide calling for a continuous search of bioactive compounds for better cancer treatments. Historically, plants are important sources for identifying such new bioactive chemicals for treatments. Here we use high-throughput screening and mass spectrometry analyses of extracts from 100 plant species collected in Chinese ancient forests to detect novel bioactive breast cancer phytochemicals. First, to study the effects on viability of the plant extracts, we used a MTT and CCK-8 cytotoxicity assay employing triple-negative breast cancer (TNBC) MDA-MB-231 and normal epithelial MCF-10A cell lines and cell cycle arrest to estimate apoptosis using flow cytometry for the most potent three speices. Based on these analyses, the final most potent extracts were from the Amur honeysuckle (Lonicera maackii) wood/root bark and Nigaki (Picrasma quassioides) wood/root bark. Then, 5 × 106 MDA-MB-231 cells were injected subcutaneously into the right hind leg of nude mice and a tumour was allowed to grow before treatment for seven days. Subsequently, the four exposed groups received gavage extracts from Amur honeysuckle and Nigaki (Amur honeysuckle wood distilled water, Amur honeysuckle root bark ethanol, Nigaki wood ethanol or Nigaki root bark distilled water/ethanol (1:1) extracts) in phosphate-buffered saline (PBS), while the control group received only PBS. The tumour weight of treated nude mice was reduced significantly by 60.5 % within 2 weeks, while on average killing 70 % of the MDA-MB-231 breast cancer cells after 48 h treatment (MTT test). In addition, screening of target genes using the Swiss Target Prediction, STITCH, STRING and NCBI-gene database showed that the four plant extracts possess desirable activity towards several known breast cancer genes. This reflects that the extracts may kill MBD-MB-231 breast cancer cells. This is the first screening of plant extracts with high efficiency in 2 decades, showing promising results for future development of novel cancer treatments.

Liang-Ge-San: a classic traditional Chinese medicine formula, attenuates acute inflammation via targeting GSK3ß.

Sepsis is a serious life-threatening health disorder with high morbidity and mortality rates that burden the world, but there is still a lack of more effective and reliable drug treatment. Liang-Ge-San (LGS) has been shown to have anti-inflammatory effects and is a promising candidate for the treatment of sepsis. However, the anti-sepsis mechanism of LGS has still not been elucidated. In this study, a set of genes related to inflammatory chemotaxis pathways was downloaded from Encyclopedia of Genes and Genomes (KEGG) and integrated with sepsis patient information from the Gene Expression Omnibus (GEO) database to perform differential gene expression analysis. Glycogen synthase kinase-3ß (GSK-3ß) was found to be the feature gene after these important genes were examined using the three algorithms Random Forest, support vector machine recursive feature elimination (SVM-REF), and least absolute shrinkage and selection operator (LASSO), and then intersected with possible treatment targets of LGS found through the search. Upon evaluation, the receiver operating characteristic (ROC) curve of GSK-3ß indicated an important role in the pathogenesis of sepsis. Immune cell infiltration analysis suggested that GSK-3ß expression was associated with a variety of immune cells, including neutrophils and monocytes. Next, lipopolysaccharide (LPS)-induced zebrafish inflammation model and macrophage inflammation model was used to validate the mechanism of LGS. We found that LGS could protect zebrafish against a lethal challenge with LPS by down-regulating GSK-3ß mRNA expression in a dose-dependent manner, as indicated by a decreased neutrophils infiltration and reduction of inflammatory damage. The upregulated mRNA expression of GSK-3ß in LPS-induced stimulated RAW 264.7 cells also showed the same tendency of depression by LGS. Critically, LGS could induce M1 macrophage polarization to M2 through promoting GSK-3ß inactivation of phosphorylation. Taken together, we initially showed that anti-septic effects of LGS is related to the inhibition on GSK-3ß, both in vitro and in vivo.

Identification of an effective fraction from Ampelopsis Radix with anti-dengue virus activities in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Dengue virus (DENV) infection is a global public health issue without effective therapeutic interventions. Chinese medicine with heat-clearing and detoxifying properties has been frequently used in the treatment of viral infection. Ampelopsis Radix (AR) is a traditional Chinese medicine for clearing heat and detoxification that has been widely used in the prevention and treatment of infectious diseases. However, no studies on the effects of AR against viral infection have been reported, thus far. AIM OF THE STUDY: To explore the anti-DENV activities of the fraction (AR-1) obtained from AR both in vitro and in vivo. MATERIALS AND METHODS: The chemical composition of AR-1 was identified by liquid chromatography-tandem MS (LC‒MS/MS). The antiviral activities of AR-1 were studied in baby hamster kidney fibroblast BHK-21 cells, ICR suckling mice and induction of interferon α/ß (IFN-α/ß) and IFN-γ R-/- (AG129) mice. RESULTS: Based on LC‒MS/MS analysis, 60 compounds (including flavonoids, phenols, anthraquinones, alkaloids and other types) were tentatively characterized from AR-1. AR-1 inhibited the cytopathic effect, the production of progeny virus and the synthesis of viral RNA and proteins by blocking DENV-2 binding to BHK-21 cells. Moreover, AR-1 significantly attenuated weight loss, decreased clinical scores and prolonged the survival of DENV-infected ICR suckling mice. Critically, the viral load in blood, brain and kidney tissues and the pathological changes in brain were remarkably alleviated after AR-1 treatment. Further study on AG129 mice showed that AR-1 obviously improved the clinical manifestations and survival rate, reduced viremia, attenuated gastric distension and relieved the pathological lesions caused by DENV. CONCLUSIONS: In summary, this is the first report that AR-1 exhibits anti-DENV effects both in vitro and in vivo, which suggests that AR-1 may be developed as a therapeutic candidate against DENV infection.

In-situ growth of nitrogen-doped carbonized polymer dots on black phosphorus for electrochemical DNA biosensor of Escherichia coli O157: H7.

Sensitive and accurate detection technology for pathogenic bacteria is of great social and economic significance in foodborne disease and food safety. In this paper, a novel portable electrochemical DNA biosensor for the detection of specific DNA sequence of Escherichia coli (E. coli) O157: H7 was constructed. To enhance the performance of the electrochemical sensor, a functionalized nitrogen-doped carbonized polymer dots in-situ grown on few-layer black phosphorus (N-CPDs@FLBP) was synthesized and used as the modifier on the surface of screen-printed electrode. Combining gold nanoparticles as immobilization matrix and methylene blue as electrochemical indicator, the analytical performance of this electrochemical DNA biosensor was evaluated using standard complementary ssDNA sequence in the linear concentration range from 1.0 × 10-19 to 1.0 × 10-6 mol/L with a low detection limit as 3.33 × 10-20 mol/L (3 σ). Furthermore, the portable electrochemical DNA biosensor was proposed based on polymerase chain reaction amplification for the detection of the E. coli O157: H7 genomic DNA from chicken meat, which verified the feasibility for practical samples detection. The research has great theoretical and practical significance for the development of electrochemical biosensor of pathogenic bacteria.

Refined-JinQi-JiangTang tablet ameliorates hypertension through activation of FGF21/FGFR1 axis in fructose-fed rats.

The aim of this study was to investigate the therapeutic effect of JQ-R on metabolic hypertension and its correlation with Fibroblast growth factor 21/Fibroblast growth factor receptors 1(FGF21/FGFR1) pathway. In this study, fructose-induced metabolic hypertension rats were used as hypertension models to detect the regulation effect of JQ-R on hypertension. The effects of JQ-R on blood glucose, blood lipids, serum insulin levels and other metabolic indicators of rats were also measured. The effects of JQ-R on FGF21/FGFR1 signaling pathway in model animals were detected by Real-time quantitative PCR and Western blotting. The results showed that JQ-R significantly reduce the blood pressure of model rats in a dose-dependent manner. Meanwhile, fasting insulin, fasting blood glucose, insulin resistance index, total cholesterol and triglyceride levels were significantly decreased, and glucose and lipid metabolism abnormalities were also significantly improved. JQ-R induces these changes along with FGFR1 phosphorylation, which was also detected in JQ-R treated FGF21 knockout mice. These results suggest that JQ-R can reduce blood pressure and improve glucose and lipid metabolism in fructose-induced hypertension rats. Activation of FGF21/FGFR1 signaling pathway to regulate downstream blood pressure and glucolipid metabolism-related pathways may be one of the important mechanisms of JQ-R in regulating blood pressure.

Endophytic Fungi from Dalbergia odorifera T. Chen Producing Naringenin Inhibit the Growth of Staphylococcus aureus by Interfering with Cell Membrane, DNA, and Protein.

This study focused on the antibacterial effects of the endophytic fungi producing naringenin from Dalbergia odorifera T. Chen against Staphylococcus aureus. The antibacterial activity was measured by the inhibition diameters, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC). The time-killing curve was also used to evaluate its antibacterial efficacy. The results of antibacterial activity determinations showed that endophytic fungi secondary metabolites can inhibit the growth of five pathogenic bacteria (S. aureus, Escherichia coli, Salmonella enteritidis, Pseudomonas aeruginosa, and Bacillus subtilis) and the most sensitive strain was S. aureus that had the MIC and MBC values of 0.13 and 0.50 mg/mL, respectively. The membrane permeability study was measured by a DNA leakage assay and electrical conductivity assay. Furthermore, the whole-cell protein lysates and DNA fragmentation assay was evaluated. The morphology of S. aureus treated with the endophytic fungi products was observed by scanning electron microscopy (SEM). The probable antibacterial mechanism of endophytic fungi secondary metabolites was the increased membrane permeability that leads to leaks of nucleic acids and proteins. SEM results further confirmed that the extracts can interfere with the integrity of S. aureus cell membrane and further inhibit the growth of bacteria, resulting in the death of bacteria. This study provides a new perspective for the antibacterial functions of endophytic fungi secondary metabolites for biomedical applications.

Alkali and Phosphorus Resistant Zeolite-like Catalysts for NOx Reduction by NH3.

At present, the deactivation of selective catalytic reduction (SCR) catalysts caused by the coexistence of alkali metal and phosphorus (P) remains an urgent problem and lacks corresponding strategies against catalyst poisoning. Herein, a novel zeolite-like Ce-Si5Al2Ox catalyst derived from an ultrasmall nanozeolite EMT precursor was synthesized without organic templates at ambient temperature. This catalyst was able to maintain above 95% NOx conversion in the 270-540 °C temperature range. Moreover, 1 wt % potassium (K) and 5 wt % P loading had no influence on the SCR performance of the Ce-Si5Al2Ox catalyst at 300-480 °C. It was demonstrated that cerium (Ce) was highly dispersed in the amorphous aluminum (Al) silicate derived from EMT zeolites and expressed high catalytic performance. Besides, a large number of acid sites were reserved to absorb ammonia allowing effective participation in the SCR reaction and capturing alkali metals, thus improving the SCR performance and K resistance. Additionally, the strong interaction between Ce and aluminosilicate decreased cerium phosphate production, preventing deactivation of the catalysts. Thus, this novel low-cost zeolite-like Ce-Si5Al2Ox catalyst with a highly active ion-exchanged metal phase and abundant surface acid sites paves a way for designing new efficient and poisoning-resistant SCR catalysts for practical applications.

Arteriovenous Graft for Hemodialysis: Effect of Cryotherapy on Postoperative Pain and Edema.

BACKGROUND: Arteriovenous grafting offers an alternative for patients whose vessels are unsuitable for arteriovenous fistula. However, as a result of subcutaneous tunnel dissection, postoperative pain and edema of the operated limb present early after surgery. As a traditional therapeutic approach, cryotherapy has the ability to suppress postoperative pain and edema. AIMS: The purpose of the study was to investigate the feasibility of cryotherapy after arteriovenous graft surgery to decrease perioperative medication usage. DESIGN: This study was a randomized controlled trial. SETTING: A large integrated health care facility in South China. PARTICIPANTS/SUBJECTS: A total of 85 hemodialysis patients who received arteriovenous graft surgery from March 2011 to February 2017 were enrolled. METHODS: The participants were divided into an intervention group and a control group according to the postoperative management. Ice packs were applied covering the operative forearm for 120 minutes after wound closure in the intervention group. General information, pain score, analgesic consumption, wound inflammation, forearm edema, and participant satisfaction were compared between the two groups. RESULTS: Cryotherapy-treated patients required less analgesia (26.19% vs. 48.84%, p < .05), reported lower pain score from 30 minutes to 48 hours postoperative (p < .05), less wound inflammation (11.90% vs. 25.58%, p < .05), and higher participant satisfaction (8.92 ± 0.57 vs. 6.52 ± 0.63, p < .05), whereas the incidence of forearm edema was equivalent (p > .05). No adverse events were reported in either group. CONCLUSIONS: Cryotherapy is a preferable intervention for patients after arteriovenous graft implantation as a result of its favorable cost, convenience, and fewer side effects.

Cryotherapy Relieves Pain and Edema After Inguinal Hernioplasty in Males With End-Stage Renal Disease: A Prospective Randomized Study.

CONTEXT: Tension-free hernioplasty under local anesthetic infiltration is a reasonable choice for end-stage renal disease patients with hernia. OBJECTIVES: The purpose of the study was to investigate the feasibility of cryotherapy after hernioplasty surgery to relieve pain and scrotal edema. METHODS: This was a prospective, randomized, and controlled trial held in a large integrated health care facility in South China. One hundred sixty-nine male patients on hemodialysis and scheduled for hernioplasty were enrolled between March 2013 and February 2017. The participants were divided into an intervention group and a control group. In the intervention group, ice packs were applied after surgery. Demographic information, vital signs, pain score, opioid consumption, wound inflammation, scrotal edema, and patient satisfaction were compared between the two groups. The primary outcome was pain score. RESULTS: Cryotherapy-treated patients required less opioid consumption (5.95 vs. 15.29 mg; P < 0.05), reported lower pain scores from 30 minutes to 48 hours after operation (P < 0.05), less wound inflammation (11.90 vs. 32.94%; P < 0.05), lower incidence of scrotal edema in the first and second days (P < 0.05), and higher patient satisfaction (8.95 vs. 6.50 cm; P < 0.05), with stable vital signs throughout the monitoring period (P > 0.05). CONCLUSION: Owing to its favorable cost, convenience, and low frequency of adverse effects, cryotherapy is useful for end-stage renal disease populations after hernioplasty to relieve pain and scrotal edema.

TPL-2-mediated activation of MAPK downstream of TLR4 signaling is coupled to arginine availability.

The innate immune response is influenced by the nutrient status of the host. Mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated kinase 1 (ERK1) and ERK2, are activated after the stimulation of macrophages with bacterial lipopolysaccharide (LPS) and are necessary for the optimal production of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha). We uncovered a role for the extracellular nutrient arginine in the activation of ERK1/2 in LPS-stimulated macrophages. Arginine facilitated the activation of MAPKs by preventing the dephosphorylation and inactivation of the MAPK kinase kinase tumor-promoting locus 2 (TPL-2). Starvation of mice decreased the concentration of arginine in the plasma and impaired the activation of ERK1/2 by LPS. Supplementation of starved mice with arginine promoted the subsequent activation of ERK1/2 and the production of TNF-alpha in response to LPS. Thus, arginine is critical for two aspects of the innate immune response in macrophages: It is the precursor used in the generation of the antimicrobial mediator nitric oxide, and it facilitates MAPK activation and consequently cytokine production.