Evaluation of drug interactions of Saposhnikoviae Radix and its major components with astragaloside IV and paeoniflorin using in vitro and in vivo experiments.

Saposhnikoviae Radix (SR) may enhance the pharmacodynamics of Huangqi Chifeng Tang (HQCFT) in the treatment of cerebral infarction according to our previous research, but the underlying mechanism is unknown. Herein, an in vivo pharmacokinetic assay in rats and in vitro MDCK-MDR1 cell assays were used to investigate the possible mechanism of SR, its main components, and its interactions with Astragali Radix (AR) and Paeoniae Radix (PR). An ultrahigh-performance liquid chromatography-tandem mass spectrometry (UPLC‒MS/MS)-based analytical method for quantifying astragaloside IV (ASIV) and paeoniflorin (PAE) in microdialysis and transport samples was developed. The pharmacokinetic parameters of SR were determined using noncompartmental analyses CCK-8 assays were used to detect the cytotoxicity of ASIV, PAE, cimifugin (CIM), prim-o-glucosylcimifugin (POG) and their combinations. Moreover, drug transport was studied using MDCK-MDR1 cells. Western blotting was performed to measure the protein expression levels of P-GP and MRP1. Claudin-5, ZO-1, and F-actin expression was determined via immunohistochemical staining of MDCK-MDR1 cells. harmacokinetic studies revealed that, compared with those of Huangqi Chifeng Tang-Saposhnikoviae Radix (HQCFT-SR), the Tmax of ASIV increased by 11.11 %, and the MRT0-t and Tmax of PAE increased by 11.19 % and 20 %, respectively, in the HQCFT group. Transport studies revealed that when ASIV was coincubated with 28 µM CIM or POG, the apparent permeability coefficient (Papp) increased by 71.52 % and 50.33 %, respectively. Coincubation of PAE with 120 µM CIM or POG increased the Papp by 87.62 % and 60.95 %, respectively. Moreover, CIM and POG significantly downregulated P-gp and MRP1 (P < 0.05), inhibited the expression of Claudin-5, ZO-1, and F-actin (P < 0.05), and affected intercellular tight junctions (TJs). In conclusion, our study successfully established a selective, sensitive and reproducible UPLC‒MS/MS analytical method to detect drug‒drug interactions between SR, AR and PR in vivo and in vitro, which is beneficial for enhancing the therapeutic efficacies of AR and PR. Moreover, this study provides a theoretical basis for further research on the use of SR as a drug carrier.

[Mechanism of acteoside in prevention and treatment of gouty arthritis based on liver metabolomics].

This study aims to reveal the effect of acteoside on gouty arthritis(GA) in rats based on liver metabolomics. The ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS) was employed to search for the potential biomarkers and metabolic pathways. SD rats were randomly assigned into blank, model, colchicine(0.3 mg·kg~(-1)), and high-, medium-, low-dose(200, 100, and 50 mg·kg~(-1), respectively) acteoside groups(n=7). The rats were administrated once a day for 7 continuous days. Monosodium urate(MSU) was used to induce GA model in rats during administration. The degree of joint swelling and pathological changes of synovial tissue in rats were observed, and the levels of interleukin(IL)-1ß, IL-18 and tumor necrosis factor(TNF)-α in the synovial tissue of rats were measured. UPLC-Q-TOF-MS was employed to collect rat liver data, and Progenesis QI and EZ info were used for data analysis. Human Metabolomics Database(HMDB) and Kyoto Encyclopedia of Genes and Genomes(KEGG) were employed to predict the potential biomarkers and metabolic pathways. The results showed that acteoside alleviated joint swelling, reduced synovial tissue damage, and lowered the levels of inflammatory cytokines in GA rats. A total of 19 common biomarkers were identified, 17 of which can be regulated by acteoside. Seven metabolic pathways were enriched, such as glycerophospholipid metabolism, linoleic acid metabolism, and taurine and hypotaurine metabolism, among which glycerophospholipid metabolism was strongly disturbed. The metabolomics analysis suggested that acteoside may down-regulate the expression of inflammatory cytokines and alleviate the symptoms of GA rats by regulating glycerophospholipid metabolism, linoleic acid metabolism, and taurine and hypotaurine metabolism. The findings provide a reference for future research and development of acteoside.

[Analysis of the effect of midazolam on pain in a rat model of lumbar disc herniation based on the p38 MAPK signaling pathway].

OBJECTIVE: To investigate the effect of midazolam on pain in lumbar disc herniation model rats based on p38 MAPK signaling pathway. METHODS: Fifty SPF-grade Sprague-Dawley healthy rats, half male and half female, were selected and randomly divided into normal group, model group, and low-dose, medium-dose, high-dose groups. Model group and low-dose, medium-dose, high-dose groups were initially modeled for lumbar disc herniation. Intraperitoneal injection of saline was performed in rats of normal and model groups; and in the low-dose, medium-dose, and high-dose groups, intraperitoneal injection of midazolam was performed with doses of 30, 60, and 90 mg/kg, respectively. Interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), 5-hydroxytryptamine (5-HT), ß-endorphin (ß-EP), substance P (SP), neuropeptide Y (NPY) were detected in the serum of rats by enzyme-linked immunoassay. The expression of p38 MAPK and matrix metalloproteinase-3(MMP-3) protein were detected by Western blot in the tissues of rats of each group. RESULTS: The levels of TNF-α, IL-1ß and ß-EP were higher and the level of 5-HT was lower in the model group than in the normal group(P<0.05);the levels of TNF-α, IL-1ß and ß-EP were lower and the level of 5-HT was higher in the low-dose, medium-dose and high-dose groups than in the model group(P<0.05). The levels of SP and NPY increased in the model group compared with the normal group (P<0.05) and the levels of SP and NPY decreased in the low-dose, medium-dose and high-dose groups compared with the model group (P<0.05). The expression of p38 MAPK and MMP-3 increased in the model group compared with the normal group (P<0.05); the expression of p38 MAPK and MMP-3 decreased in the low-dose, medium-dose and high-dose compared with the model group(P<0.05). CONCLUSION: Midazolam may ameliorate the immune inflammatory response in rats with a model of lumbar disc herniation, possibly regulated through the p38MAPK signaling pathway.

Effects of Total Saponins from Dioscorea Nipponica Makino on Monosodium Urate-Induced M1-Polarized Macrophages through Arachidonic Acid Signaling Pathway: An in vitro Study.

OBJECTIVE: To investigate and reveal the underlying mechanism of the effect of total saponins from Dioscoreae nipponica Makino (TSDN) on the arachidonic acid pathway in monosodium urate (MSU) crystal-induced M1-polarized macrophages. METHODS: M1 polarization of RAW264.7 cells were induced by 1 µ g/mL lipopolysaccharide (LPS). The methylthiazolyldiphenyl-tetrazolium bromide method was then used to screen the concentration of TSDN. MSU (500 µ g/mL) was used to induce the gouty arthritis model. Afterwards, 10 µ g/L TSDN and 8 µ mol/L celecoxib, which was used as a positive control, were added to the above LPS and MSU-induced cells for 24 h. The mRNA and protein expressions of cyclooxygenase (COX) 2, 5-lipoxygenase (5-LOX), microsomal prostaglandin E synthase derived eicosanoids (mPGES)-1, leukotriene B (LTB)4, cytochrome P450 (CYP) 4A, and prostaglandin E2 (PGE2) were tested by real-time polymerase chain reaction and Western blotting, respectively. The enzyme-linked immunosorbent assay was used to test the contents of M1 markers, including inducible nitric oxid synthase (NOS) 2, CD80, and CD86. RESULTS: TSDN inhibited the proliferation of M1 macrophages and decreased both the mRNA and protein expressions of COX2, 5-LOX, CYP4A, LTB4, and PGE2 (P<0.01) while increased the mRNA and protein expression of mPGES-1 (P<0.05 or P<0.01). TSDN could also significantly decrease the contents of NOS2, CD80, and CD86 (P<0.01). CONCLUSION: TSDN has an anti-inflammation effect on gouty arthritis in an in vitro model by regulating arachidonic acid signaling pathway.

Mechanism of Action of Shenerjiangzhi Formulation on Hyperlipidemia Induced by Consumption of a High-Fat Diet in Rats Using Network Pharmacology and Analyses of the Gut Microbiota.

Shenerjiangzhi formulation (SEJZ) is a new traditional Chinese medicine formulation (patent number: CN110680850A). SEJZ contains Eleutherococcus senticosus (Rupr. and Maxim.), Maxim (Araliaceae; E. senticosus radix and rhizome), Lonicera japonica Thunb (Caprifoliaceae; Lonicera japonica branch, stem), Crataegus pinnatifida Bunge (Rosaceae; Crataegus pinnatifida fruit), and Auricularia auricula. SEJZ has been designed to treat hyperlipidemia. Despite the therapeutic benefits of SEJZ, its underlying mechanism of action is not known. We explored the efficacy of SEJZ against hyperlipidemia by integrating network pharmacology and 16S rRNA gene sequencing and elucidated its mechanism of action. First, SEJZ targets were found through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform and from the literature. Hyperlipidemia-related therapeutic targets were obtained from GeneCards, Online Mendelian Inheritance in Man, and DrugBank databases. Then, Search Tool for the Retrieval of Interacting Genes/Proteins and Cytoscape were applied for the analyses and construction of a protein-protein interaction (PPI) network. The Kyoto Encyclopedia of Genes and Genomes database was employed to identify signaling pathways that were enriched. Second, the therapeutic effects of SEJZ against hyperlipidemia induced by consumption of a high-fat diet in rats were evaluated by measuring body weight changes and biochemical tests. SEJZ treatment was found to alleviate obesity and hyperlipidemia in rats. Finally, 16S rRNA gene sequencing showed that SEJZ could significantly increase the abundance of short-chain fatty acid-producing bacteria, restore the intestinal barrier, and maintain intestinal-flora homeostasis. Using PICRUSt2, six metabolic pathways were found to be consistent with the results of network pharmacology: "African trypanosomiasis", "amoebiasis", "arginine and proline metabolism", "calcium signaling pathway", "NOD-like receptor signaling pathway", and "tryptophan metabolism". These pathways might represent how SEJZ works against hyperlipidemia. Moreover, the "African trypanosomiasis pathway" had the highest association with core genes. These results aid understanding of how SEJZ works against dyslipidemia and provide a reference for further studies.

[Formation of neutrophil extracellular traps and their effects on autoimmune diseases].

Neutrophil extracellular traps(NETs) are networks of extracellular fibers primarily composed of DNA, histones, granular proteins, and cytoplasmic proteins and released to the outside of cells by neutrophils under the stimulation of bacteria, fungi, viruses, parasites, etc. NETs are generated in two forms, suicidal NETs and vital NETs, according to different stimuli. NETs have both anti-inflammatory and pro-inflammatory effects. On the one hand, they can play the anti-microbial role to resist inflammation by capturing, fixing, and killing invading pathogens, which is a special way for neutrophils to exert host defenses. On the other hand, in case of excessive formation or insufficient elimination, they can cause tissue damage directly, and also promote the release of inflammatory factors by recruiting other pro-inflammatory cells or proteins to further expand the inflammatory response, which is related to the pathologies of many diseases. In autoimmune diseases, NETs as important sources of autoantigens, can act as danger-associated molecular patterns( DAMPs) and activate the nucleotide-binding oligomerization domain leucine-rich repeats containing pyrin domain 3(NLRP3) inflammasome and complement system, thereby breaking self-tolerance and accelerating autoimmune inflammation. In addition, NETs can also activate other immune cells(such as B cells, antigen-presenting cells, and T cells) and regulate the acquired immune response. The present study reviewed the correlation of NETs with diseases such as systemic lupus erythematosus(SLE), rheumatoid arthritis(RA), and gouty arthritis(GA) to reveal the effect of dynamic balance between formation and clearance of NETs in autoimmune diseases and provide a theoretical basis for the investigation of underlying mechanisms and targeted therapies of traditional Chinese medicine.

Saposhnikoviae Radix Enhanced the Angiogenic and Anti-Inflammatory Effects of Huangqi Chifeng Tang in a Rat Model of Cerebral Infarction.

Huangqi Chifeng Tang (HQCFT), a traditional Chinese formula of three herbs, has been used to treat cerebral infarction (CI). Saposhnikoviae Radix (SR) was designed as a guiding drug for HQCFT to improve its angiogenic and anti-inflammatory effects. In this study, TTC staining was used to detect the area of CI. H&E staining was used to detect the histopathologic changes in the cerebral tissue. Western blotting was performed to detect the protein expression of NLRP3, caspase 1, IL-1ß, IL-6, TNF-α, MMP-9, VEGF, and VEGFR2 in cerebral tissue. Immunohistochemistry was used to detect the protein expression of MMP-9, VEGF, and VEGFR2. The contents of HIF-1α, NLRP3, caspase 1, IL-1ß, IL-6, and TNF-α in the serum were determined by ELISA. Our study showed that HQCFT and HQCFT-SR could improve the pathological condition and reduce the infarcted area of the brain tissue in a rat model. In addition, HQCFT and HQCFT-SR significantly decreased the expression levels and serum contents of NLRP3, caspase 1, IL-1ß, IL-6, and TNF-α; increased the expression levels of the VEGF and VEGFR2 proteins; and obviously reduced the serum content of HIF-1α. Importantly, the cytokines in brain tissue and serum from the HQCFT group exhibited better efficacy than those from the HQCFT-SR group. HQCFT exerted significant angiogenic and anti-inflammatory effects in rats subjected to middle cerebral artery occlusion (MCAO); these effects can be attributed to the guiding and enhancing effect of SR.

Effect of MT2A on apoptosis and proliferation in HL60 cells.

Although accumulating evidence has revealed that metallothioneins (MTs) and its family member MT2A are strongly linked to the risk of various solid tumors, researches on the occurrence and development of acute myeloid leukemia (AML) have rarely been investigated. Here, we constructed a lentiviral vector with MT2A over-expression and the interfering plasmids with MT2A expression inhibition to study the influence of MT2A on the bioactivities of HL60 cells. After cells were infected with a lentiviral vector containing the MT2A gene, both transcription and translation levels of MT2A were significantly increased in the over-expressed group in comparison with control groups. In vitro experiments, all results demonstrated that cell reproductive capacity was inhibited, but cell apoptosis rate was significantly increased. Together, the expression of apoptosis-related protein Bcl2 was remarkably reduced, while a high expression level of Bax protein was detected. Further experiments revealed that up-regulation of MT2A induced cell apoptosis and promoted G2/M phase arrest. The mechanism may be associated with down-regulated p-IκB-α and cyclinD1 expression and up-regulated IκB-α expression in the nuclear factor-kappaB (NF-κB) pathway. On the contrary, MT2A expression was down-regulated by interfering plasmids. We found that cell proliferative potential was notably increased in the interfering group compared with the negative and untreated group. What's more, MT2A may be closely related to AML cell proliferation and function via the NF-κB signal pathway.

Prediction of Prognostic Biomarkers and Construction of an Autophagy Prognostic Model for Colorectal Cancer Using Bioinformatics.

OBJECTIVE: The incidence of colorectal cancer is increasing every year, and autophagy may be related closely to the pathogenesis of colorectal cancer. Autophagy is a natural catabolic mechanism that allows the degradation of cellular components in eukaryotic cells. However, autophagy plays a dual role in tumorigenesis. It not only promotes normal cell survival and tumor growth but also induces cell death and suppresses tumors survival. In addition, the pathogenesis of various conditions, including inflammation, neurodegenerative diseases, or tumors, is associated with abnormal autophagy. The present work aimed to examine the significance of autophagy-related genes (ARGs) in prognosis prediction, to construct an autophagy prognostic model, and to identify independent prognostic factors for colorectal cancer (CRC). METHODS: This study discovered a total of 36 ARGs in CRC cases using The Cancer Genome Atlas (TCGA) and Human Autophagy-dedicated (HADd) databases along with functional enrichment analysis. Then, an autophagy prognostic model was constructed using univariate Cox regression analysis, and the key prognostic genes were screened. Finally, independent prognostic markers were determined through independent prognostic analysis and clinical correlation analysis of key genes. RESULTS: Of the 36 differentially expressed ARGs, 13 were related to prognosis, as determined by univariate Cox regression analysis. A total of 6 key genes were obtained by a multivariate Cox regression analysis. Independent prognostic values were shown by 3 genes, namely, microtubule-associated protein 1 light chain 3 (MAP1LC3C), small GTPase superfamily and Rab family (RAB7A), and WD-repeat domain phosphoinositide-interacting protein 2 (WIPI2) by independent prognostic analysis and clinical correlation. CONCLUSIONS: In this study, molecular bioinformatics technology was employed to determine and construct a prognostic model of autophagy for colon cancer patients, which revealed 3 autophagy-related features, namely, MAP1LC3C, WIPI2, and RAB7A.

Immobilization and enzymatic properties of glutamate decarboxylase from Enterococcus faecium by affinity adsorption on regenerated chitin.

Glutamate decarboxylase (GAD, EC 4.1.1.15) is an important enzyme in gamma-aminobutyric acid biosynthesis and DL-glutamic acid resolution. In this study, the Enterococcus faecium-derived GAD was successfully immobilized by regenerated chitin (RC) via specific adsorption of cellulose-binding domain (CBD). The optimal binding buffer was 20 mmol/L phosphate buffer saline (pH 8.0), and the RC binding capacity was 1.77 ± 0.11 mgcbd-gad/grc under this condition. The ratio of wet RC and crude enzyme solution used for immobilization was recommended to 3:50 (g/mL). To evaluate the effect of RC immobilization on GAD, properties of the immobilize GAD (RC-CBD-GAD) were investigated. Results indicated RC-CBD-GAD was relatively stable at pH 4.4-5.6 and temperature - 20-40 °C, and the optimal reaction pH value and temperature were pH 4.8 and 50 °C, respectively. When it was reacted with 5 mmol/L of follow chemical reagents respectively, the activity of RC-CBD-GAD was hardly affected by EDTA, KCl, and NaCl, and significantly inactivated by AgNO3, MnSO4, MgSO4, CuSO4, ZnSO4, FeCl2, FeCl3, AlCl3, CaCl2, and Pb(CH3COO)2. The apparent Km and Vmax were 28.35 mmol/L and 147.06 µmol/(gRC-CBD-GAD·min), respectively. The optimum time for a batch of catalytic reaction without exogenous pH control was 2 h. Under this reaction time, RC-CBD-GAD had a good reusability with a half-life of 23 cycles, indicating that it was very attractive for GABA industry. As a novel, efficient, and green CBD binding carrier, RC provides an alternative way to protein immobilization.

Zirconia-Pillaring in Layered HNb3 O8 and HNbMoO6.

In this work, the pillaring of two layered niobium-based oxides (HNb3 O8 and HNbMoO6 ) with zirconia was investigated in detail. Two novel zirconia-pillared layered metal oxides, zirconia-pillared layered HNb3 O8 and zirconia-pillared layered HNbMoO6 , have been successfully prepared. Both pillared products exhibited a higher thermal stability exceeding 673 K. For the pillaring of layered HNb3 O8 , two different pre-expanding agents, 1-dodecanamine and 1,12-dodecanediamine, were alternatively used, and two kinds of zirconium-pillaring solutions containing zirconium(IV) polyoxocations obtained through two different ways were employed. The 1,12-dodecanediamine-pre-expanded layered intermediate was applicable and 1-dodecanamine-pre-expanded one was not applicable to the intercalation of zirconium(IV) polyoxocations in interlayer regions of the layered niobium-based oxides. More interestingly, the zirconium-pillaring solutions prepared by using an appropriate amount of diethylene glycol as stabilizing agent was advantage for constructing the ordered zirconia-pillared product, whereas the zirconium-pillaring solutions obtained in the case of absence of diethylene glycol seemed not to conduct well an ions-exchange reaction with the alkylamine-pre-expanded layered intermediates. The order degree of zirconia-pillared layered transition metal oxides was closely related to the host sheets. The zirconia-pillared layered HNb3 O8 contained more defects, while the zirconia-pillared layered HNbMoO6 had fewer defects.

Anti-inflammation effects of the total saponin fraction from Dioscorea nipponica Makino on rats with gouty arthritis by influencing MAPK signalling pathway.

BACKGROUND: Dioscorea nipponica Makino is widely used in traditional Chinese medicine to treat gouty arthritis. METHODS: Sixty male Wistar rats were divided into six groups: the normal group, model group, colchicine group (COL) and three total saponin groups (RDN) (high dose [160 mg/kg], middle dose [80 mg/kg] and low dose [40 mg/kg]). HE staining was used to detect the histopathologic changes of the synovial tissue of joint. Immunohistochemical method was used to detect the protein expressions of P-38, p-P38, JNK, p-JNK, ERK1/2, p-ERK1/2, MEK1/2, p-MEK1/2, MKK4, p-MKK4, ICAM1, VCAM1, and PPARγ in the synovial tissue of joint. Realtime PCR and WB methods were used to detect the mRNA and protein expressions of PPARγ and AdipoR2 in the synovial tissue of joint. The contents of CXCL1 and ADP in the blood serum were measured by Elisa method. RESULTS: Our study showed that RDN could improve the situation of the synovial tissue, reduce the protein expressions of MKK4, p-MEK1/2, p-JNK, p-ERK1/2, ICAM1. They could also decrease the content of CXCL1 and increase the content of ADP in the blood serum. CONCLUSION: RDN has good effect of anti-inflammation. This is in part realized by influencing MAPK signalling pathway. It provides a new visual angle to reveal the mechanism of RDN to treat GA.

Enhancing effect of macroporous adsorption resin on gamma-aminobutyric acid production by Enterococcus faecium in whole-cell biotransformation system.

Gamma-aminobutyric acid (GABA) biosynthesis depended to a great extent on the biotransformation characterization of glutamate decarboxylase (GAD) and process conditions. In this paper, the enhancing effect of D101 macroporous adsorption resin (MAR) on the GABA production was investigated based on the whole-cell biotransformation characterization of Enterococcus faecium and adsorption characteristics of D101 MAR. The results indicated that the optimal pH for reaction activity of whole-cell GAD and pure GAD was 4.4 and 5.0, respectively, and the pH range retained at least 50% of GAD activity was from 4.8 to 5.6 and 4.0-4.8, respectively. No substrate inhibition effect was observed on both pure GAD and whole-cell GAD, and the maximum activity could be obtained when the initial L-glutamic acid (L-Glu) concentration exceeded 57.6 mmol/L and 96.0 mmol/L, respectively. Besides, GABA could significantly inhibit the activity of whole-cell GAD rather than pure GAD. When the initial GABA concentration of the reaction solution remained 100 mmol/L, 33.51 ± 9.11% of the whole-cell GAD activity was inhibited. D101 MAR exhibited excellent properties in stabilizing the pH of the conversion reaction system, supplementing free L-Glu and removing excess GABA. Comparison of the biotransformation only in acetate buffer, the GABA production, with 50 g/100 mL of D101 MAR, was significantly increased by 138.71 ± 5.73%. D101 MAR with pre-adsorbed L-Glu could significantly enhance the production of GABA by gradual replenishment of free L-Glu, removing GABA and maintaining the pH of the reaction system, which would eventually make the GABA production more economical and eco-friendly.

[Modulation effect of Acanthopanax senticosus polysaccharides through inflammatory cytokines in protecting immunological liver-injured mice].

The aim of this paper was to discuss the protective effect and mechanism of Acanthopanax senticosus polysaccharides( ASPs) on immunological liver injury caused by conanavalin A( Con A). BALB/c mice were randomly divided into seven groups: control group,model group( Con A),low-,medium-,and high-dose( 36. 25,72. 5,145 mg·kg~(-1)) ASPs groups,bifendate( 200 mg·kg~(-1),positive drug) group and pyrrolidinedithiocarbamate( PDTC,NF-κB inhibitor,200 mg·kg~(-1)) group. ASPs groups and bifendate group were given with corresponding drugs by ig administration once daily for 7 d. Control group,model group and PDTC group were given with normal saline by ig administration once daily for 7 d. After the last ig administration,PDTC was given in DTC group by iv administration( 200 mg·kg~(-1)); 0. 5 h after that,Con A( 20 mg·kg~(-1)) was injected via the tail vein to induce immunological liver injury in all the mice except normal control group. The mice were killed 8 h later and their liver tissues were collected for histopathological examination. The contents of nitric oxide( NO),superoxide dismutase( SOD),malondialdehyde( MDA),reduced glutathione( GSHPX),interleukin( IL-1ß) and tumor necrosis factor( TNF-α) in liver tissues were detected by kit assay. Western blot method was used to detect TNF-α,intercellular cell adhesion molecule-1( ICAM-1),inducible nitric oxide synthase( i NOS) and nuclear factor( NF-κB) protein expression in liver tissues. As compared with model group,ASPs not only could reduce the activity of MDA,NO,IL-1ß and TNF-α,but also increase the content of GSH-PX and SOD; at the same time,the protein expression levels of TNF-α,ICAM-1,i NOS and NF-κB were reduced in liver tissues; in addition,inflammatory cell infiltration was alleviated,hepatocyte cytoplasm was loose and swollen,and nuclear condensation and staining were improved. ASPs has a protective effect on immunological liver injury,and the mechanism may be associated with regulating secretion of inflammatory cytokines and the expression of adhesion factor through NF-κB signaling pathway.

Anti-inflammatory Effect of Total Saponin Fraction from Dioscorea nipponica Makino on Gouty Arthritis and Its Influence on NALP3 Inflammasome.

OBJECTIVE: To investigate the mechanism of Chinese herbal medicine Dioscorea nipponica for the treatment of monosodium urate crystals-induced gouty arthritis (GA) in rats. METHODS: Sixty male Wistar rats were divided into 6 groups: normal, model, indomethacin and three total saponin (900, 300 and 100 mg/kg) groups. The liver, kidney and serum levels of lysosomal enzymes, antioxidant capacities, and inflammatory factors were measured. In addition, the mRNA and protein levels of the NALP3 inflammasome components in the mononuclear cells of rats' peripheral blood were analyzed using real-time polymerase chain reaction and Western blotting methods, respectively. RESULTS: Total saponins groups could reduce the activities of ß-galactosidase, ß-N acetyl glucosamine enzyme, ß-glucuronidase, acid phosphatase, and malonaldehyde as well as the contents of TNF-α, IL-1ß and IL-8 (all P<0.05). They could also increase the activities of glutathione peroxidase and total superoxide dismutase (both P<0.05). Further studies showed that total saponins groups of high, middle and low doses could all increase the mRNA and protein levels of caspase-1, adapter apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) and NALP3 in the mononuclear cells of peripheral blood (all P<0.05). CONCLUSION: Dioscorea nipponica may treat GA by regulating lysosomal enzymes, antioxidant capacities and the NALP3 inflammasome.

High throughput metabolomics-proteomics investigation on metabolic phenotype changes in rats caused by Radix Scrophulariae using ultra-performance liquid chromatography with mass spectrometry.

Radix Scrophulariae, a traditional Chinese herb, is used to treat various diseases, including H2O2-induced apoptosis in cardiomyocytes, HaCaT cells, hyperuricaemia, and depression. This study screened metabolites, proteins and common pathways to better understand both the therapeutic effects and side effects of this herb. Methods: Untargeted metabolomics based on UPLC-TOF-MS, coupled with proteomics based on nano-UPLC-Q-Exactive-MS/MS, were used to investigate the effects of R. Scrophulariae in rats. Fifty-one identified metabolites in urine samples and 76 modulated proteins in liver tissue were potential biomarkers for R. Scrophulariae treatment. The biomarkers and common pathways involved were steroid hormone biosynthesis, drug metabolism-cytochrome p450, drug metabolism-other enzymes, pentose and glucuronate interconversions, and starch and sucrose metabolism. Some biomarkers were beneficial for treating diseases such as cancer, tuberculosis and isovaleric acidaemia, while other biomarkers caused side effects. Metabolomic and proteomic analyses of R. Scrophulariae-treated rats provided valuable information on the biological safety and efficacy of using R. Scrophulariae clinically.

Acanthopanax senticosus Protects Structure and Function of Mesencephalic Mitochondria in A Mouse Model of Parkinson's Disease.

OBJECTIVE: To investigate the neuro-protective effects of Acanthopanax senticosus Harms (EAS) on mesencephalic mitochondria and the mechanism of action, using a mouse model of Parkinson's disease (PD). METHODS: The chemical fingerprint analysis of the extract of Acanthopanax senticosus Harms (EAS) was performed using the ultra performance liquid chromatograph and time of flight mass spectrometry. Thirty mice were randomly divided into the control group, the MPTP model group, and the EAS treated group with MPTP (MPTP+EAS group, 10 in each group). The MPTP model group and the MPTP+EAS group received MPTP-HCl (30 mg/kg i.p) once a day for 5 days. The control group received an equal volume of saline (20 mL/kg i.p) once a day for 5 days. Induced by 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine hydrochloride daily (MPTP-HCl, 30 mg/kg) for 5 days, the PD mice were treated with EAS at 45.5 mg/kg daily for 20 days. The behavioral testing of mice was carried out using the pole-climbing test. The integrity and functions of neurons were examined in mesencephalic mitochondria in a PD mouse model, including nicotinamide adenine dinucleotide dehydrogenase ubiquinone flavoprotein 2 (NDUFV2), mitochondrially encoded nicotinamide adenine dinucleotide dehydrogenase 1 (MT-ND1), succinate dehydrogenase complex subunit A (SDHA), and succinate dehydrogenase cytochrome b560 subunit (SDHC). RESULTS: After treatment with EAS, the behavioral changes induced by MPTP were attenuated significantly (P<0.05). EAS protected the mesencephalic mitochondria from swelling and attenuated the decreases in their membrane potential (both P<0.05), which was supported by an ultra-structural level analysis. The changes in reactive oxygen species (ROS), malonic dialdehyde (MDA), oxidative phosphorylation (OXPHOS) system 4 subunits levels and PD-related proteins expressions (parkin, Pink1, DJ-1, α-synuclein, and Lrrk2) reverted to near normal levels (all P<0.05), based on the results of immune-histological and Western blotting observations. CONCLUSIONS: The neuro-protective effects of EAS are linked to protecting mice against MPTP-induced mitochondrial dysfunction and structural damage. Therefore, EAS is a promising candidate for the prevention or treatment of mitochondrial neurodegenerative disorders, such as PD.

[Inhibitory effect of Scutellariae Radix on hepatic fibrosis based on urinary metabonomic].

Urinary metabolomics combined with histological progression were utilized to evaluate the therapeutic effect of Scutellariae Radix decoction and baicalin on hepatic fibrosis (HF) and explore their mechanisms, intervention targets and metabolic pathways. HF rat model was established through subcutaneous injection of CCl4 for 8 weeks. Meanwhile, different doses of Scutellariae Radix decoction and baicalin were administered. Histomorphology of liver tissue was observed and scored by HE and Masson. Urinary metabonomic analysis based on UPLC-Q-TOF-MS was made for the changes of urinary potential biomarkers among different groups at different time points of HF. Finally, it was found that Scutellariae Radix decoction could improve HF by regulating L-tryptophan, 3-methyldioxyindole, 5-hydroxyindoleacetylglycine, kynurenic acid, 4-(2-amino-3-hydroxyphenyl)-2,4-dioxobutanoic acid, methylmalonic acid and L-leucine. However, baicalin could improve HF by regulating L-tryptophan, 3-methyldioxyindole, 5-hydroxyindoleacetylglycine, 4-(2-amino-3-hydroxyphenyl)-2,4-dioxobutanoic acid, kynurenic acid, and methylmalonic acid. These metabolites involved in tryptophan metabolism and valine, leucine and isoleucine degradation pathways. These results indicated that Scutellariae Radix had the multi-target and multi-pathway characteristics in the treatment of HF. Additionally, low-dose Scutellariae Radix decoction and baicalin are showed better efficacies, with no statistically significant difference between them in histomorphology.

[Liver metabonomics of acute gouty arthritis treated by Diosocorea nipponica].

To explore the prevention and protection effect of Diosocorea nipponica (DNM)) on acute gouty arthritis (AGA) rats based on liver metabonomics, and find potential biomarkers and related pathways. AGA model rats were induced by monosodium urate crystal suspension. UPLC-TOF-MS coupled with pattern recognition technique was employed to find out the potential biomarkers and related metabolic pathways. Eleven common potential biomarkers were identified. Among the potential intervention targets in normal rats given by DNM, 4 biomarkers were up-regulated, and the other 4 targets were down regulated. Among the potential intervention targets in AGA rats given by DNM, 5 metabolites were up-regulated by MSU and 5 metabolites were down regulated. The abnormal expression levels of adenosine monophosphate, 5-methyltetrahydrofolic acid, oxidized glutathione, hypoxanthine, docosahexaenoic acid, glutathione, uridine diphosphate glucose and inosine could be corrected by DNM extract. Three pathways were founded with greatest correlation, including purine metabolism, starch and sucrose metabolism and glutathione metabolism. Therefore, it could be inferred that D. nipponica has the effect for anti-acute gouty arthritis by intervening endogenous metabolites from the liver under physiological condition and acute gouty arthritis condition.

Identification of key metabolic changes during liver fibrosis progression in rats using a urine and serum metabolomics approach.

Reversibility of hepatic fibrosis is an intrinsic response to chronic injury, and with on-going damage, fibrosis can progress to its end-stage consequence, cirrhosis. Non-invasive and reliable biomarkers for early detection of liver fibrosis are needed. Based on the CCl4-induced liver fibrosis rat model, urinary and serum metabolic profiling performed by LC-QTOF-MS associated with histological progression were utilized to identify liver fibrosis-specific potential biomarkers for early prediction and to reveal significant fibrotic pathways and their dynamic changes in different stages of liver fibrosis. Finally, nine differential metabolites in urine and ten in serum were selected and identified involving the most relevant metabolic pathways. Perturbations of tryptophan, valine, leucine, isoleucine, and citrate (TCA) cycle metabolites, along with sphingolipid and glycerophospholipid metabolites, occurred from the onset of liver fibrosis. Furthermore, dysregulation of valine and bile acid biosynthesis metabolites occurred in the intermediate and advanced stages. More importantly, among these metabolites, urinary kynurenic acid, 5-hydroxyindoleacetyl glycine, 4-(2-amino-3-hydroxyphenyl)-2,4-dioxobutanoic acid and serum sphinganine, sphingomyelin, L-leucine, L-tryptophan, and LysoPC(17:0) changed at all time points and may serve as potential early biomarkers for the diagnosis of hepatic fibrosis and as therapeutic targets. Overall, this work evaluates the potential of these metabolites for the early detection of liver fibrosis.