Paeoniflorin Ameliorates Colonic Fibrosis in Rats with Postinfectious Irritable Bowel Syndrome by Inhibiting the Leptin/LepRb Pathway.

Postinfectious irritable bowel syndrome (PI-IBS) is a highly prevalent gastrointestinal disorder associated with immune dysregulation and depression- and anxiety-like behaviors. Through traditional medicine, the active ingredient of Paeoniae Radix called paeoniflorin (PF) was previously found to prevent the symptoms of PI-IBS. However, there is limited information on the effects of PF on intestinal function and depression- and anxiety-like symptoms in PI-IBS animal models. Here, we aimed to determine the effects of PF treatment on the symptoms of PI-IBS in a rat model. The PI-IBS rat model was established via early postnatal sibling deprivation (EPSD), trinitrobenzenesulfonic acid (TNBS), and chronic unpredictable mild stress (CUMS) stimulation and then treated with different dosages of PF (10, 20, and 40 mg/kg) and leptin (1 and 10 mg/kg). The fecal water content and body weight were measured to evaluate the intestinal function, while the two-bottle test for sucrose intake, open field test (OFT), and elevated plus maze test (EMT) were performed to assess behavioral changes. The serum leptin levels were also measured using an enzyme-linked immunosorbent assay. Furthermore, the expressions of leptin and its receptor, LepRb, were detected in colonic mucosal tissues through an immunohistochemical assay. The activation of the PI3K/AKT signaling pathway and the expression of brain-derived neurotrophic factor (BDNF) were also detected via western blotting. After the experimental period, the PI-IBS rats presented decreased body weight and increased fecal water content, which coincided with elevated leptin levels and heightened depression- and anxiety-like behaviors (e.g., low sucrose intake, less frequency in the center areas during OFT, and fewer activities in the open arms during EMT). However, the PF treatment ameliorated these observed symptoms. Furthermore, PF not only inhibited leptin/LepRb expression but also reduced the PI3K/AKT phosphorylation and BDNF expression in PI-IBS rats. Notably, cotreatment with leptin (10 mg/kg) reduced the effects of PF (20 mg/kg) on colonic fibrosis, leptin/LepRb expression, and PI3K/AKT activation. Therefore, our findings suggest that leptin is targeted by PF via the leptin/LepRb pathway, consequently ameliorating the symptoms of PI-IBS. Our study also contributes novel insights for elucidating the pharmacological action of PF on gastrointestinal disorders and may be used for the clinical treatment of PI-IBS in the future.

[Prediction of anti-liver fibrosis effect of Piperis Longi Fructus based on network pharmacology].

Network pharmacology and liver fibrosis(LF) model in vitro were used to analyze the underly mechanism of anti-liver fibrosis effect that induced by Piperis Longi Fructus and its major active compounds. TCMSP and TCMIP were used to search for the chemical constituents of Piperis Longi Fructus, as well as the oral bioavailability(OB), drug-likeness(DL), intercellular permeability of intestinal epithelial cells(Caco-2) and Drug-likeness grading were set as limiting conditions. The related target genes of Piperis Longi Fructus were queried by TCMSP database, while related targets of LF were screened by GeneCards databases. Interaction network was constructed using Cytoscape 3.7.1. These above data were imported into STRING database for PPI network analysis. Enrichment of gene ontology(GO) and pathway analysis(KEGG) within Bioconductor database were utilized to note functions of related targets of Piperis Longi Fructus. Finally, the core targets and pathways were preliminarily verified by in vitro experiments. The effects of piperlongumine(PL), the major active component of Piperis Longi Fructus, on proliferation of rat liver stellate cells(HSC-T6) and expression of α smooth muscle actin(α-SMA) and collagen Ⅰ were investigated. The major factors TNF-α of tumor necrosis factor(TNF) pathway and NF-κB p65, IL-6 protein expressions of LF process were examined. A total of 12 active compounds such as PL were obtained by analyzing the bioavailability and drug-like properties, which inferred to 48 targets. The functional enrichment analysis of GO obtained 1 240 GO items, mainly involving in process of biology and molecular function. A total of 99 signaling pathways were enriched in the KEGG pathway enrichment analysis, including TNF signaling pathway, cGMP-PKG signaling pathway, calcium signaling pathways. CCK-8 assay showed that PL inhibited proliferation of HSC-T6 induced by transforming growth factor-ß1(TGF-ß1). Western blot analysis found that treated with PL suppressed the protein expressions of α-SMA, collagen Ⅰ, TNF-α and p65 in HSC-T6. Enzyme linked immunosorbent assay(ELISA) showed that PL inhibited the expressions of TNF-α and IL-6 in the cluture supertant of HSC-T6 cells. In conclusion, PL could play an anti-liver fibrosis role by regulating TNF/NF-κB signaling pathway. This study provided the mechanism basis of anti-LF effects induced by Piperis Longi Fructus and its major active compounds, which might help for the further study of the mechanism and key targets of Piperis Longi Fructus.

[Identification on Solanum lyratum by X-ray diffraction Fourier fingerprint and similarity analysis].

OBJECTIVE: To provide an X-ray diffraction (XRD) method for identifying different medicinal parts of Solanum lyratum. METHODS: Analyzed X-ray diffraction Fourier patterns of different parts of Solanum lyratum, and the similarity degree of different fingerprint was calculated and analyzed according to the position (2 theta value)of peaks searched. RESULTS: Different X-ray diffraction Fourier patterns of different medicinal parts of Solanum lyratum were obtained. CONCLUSION: This method can be used for identifying different medicinal parts of Solanum lyratum. The results of similarity calculation further proves the feasibility of this method.

Comparison of the antioxidant activity amongst Gingko biloba extract and its main components.

OBJECTIVE: To compare the antioxidant activity amongst the extract of Ginkgo biloba (EGb) and its main components, flavonoids and terpenoids. METHODS: The induction of EGb, flavonoids and terpenoids on a typical antioxidant enzyme, glutamate cysteine ligase catalytic subunit (GCLC), in cell lines was detected by Western-blot. The effects of EGb, flavonoids and terpenoids on superoxide anion radical (O2*(-)), hydroxyl radical (OH*), rat erythrocyte hemolysis and lipid peroxidation of rat liver homogenate were determined by respective activity methods. RESULTS: EGb and flavonoids but not terpenoids were demonstrated significantly to induce the antioxidant enzyme (GCLC), directly scavenge O2*(-), OH* and inhibit rat erythrocyte hemolysis and lipid peroxidation of rat liver homogenate. Compared these antioxidant activities between EGb and flavonoids, the activities of flavonoids were weaker than those of EGb, which contains similar dose of flavonoids. CONCLUSION: EGb has stronger antioxidant activities than flavonoids, but terpenoids did not show antioxidant activity in this research.

Extract of Ginkgo biloba induces glutathione-S-transferase subunit-P1 in vitro.

The extract of Ginkgo biloba (EGb), containing 24% flavone glycosides and 6% terpenoids, is widely used to treat early-stage Alzheimer's disease, vascular dementia, peripheral claudication and vascular tinnitus. Its remarkable antioxidant activity has recently been demonstrated in both cell lines and animals. Glutathione-S-transferases (GSTs) are a class of important detoxification enzymes in the antioxidant system and GST-P1 is the major GST isoform highly expressed in human tissues. Over expression of GST-P1 protected prostate cells from cytotoxicity and DNA damage by the heterocyclic amine carcinogen, while inhibition of expression of GST-P1 by transfecting GST-P1 antisense cDNA or targeted deletion of GST-P1 has been found to sensitize cells to cytotoxic chemicals. It is obvious that induction of GST-P1 expression should be a promising alternative for chemoprevention. The present study aimed to investigate the induction effect of EGb on GST-P1 in HepG2 and Hep1c1c7 cell lines and found that GST-P1 was increased both at the expression and enzyme activity levels.

[Studies on fingerprints of Bupleurum chinense and vinegar-baked Bupleurum chinense from Hubei province].

OBJECTIVE: To establish the HPLC Fingerprint of Bupleurum chinense from GAP provenance of Hubei province Baokang, and discuss the changes before and after processing. METHODS: Using HPLC detection method equipped with Hypersil ODS column (5 microm, 4.6 mm x 250 mm), using acetonitrile-water as mobile phase gradient elution, flow as 1.0 ml/min, UV detector set at 203 nm, column temperature was at 25 degrees C. RESULTS: Similarity of processed Bupleurum chinense and Bupleurum chinense exceeded 0.95, and there were 13 characteristic fingerprints of processed Bupleurum chinense, 15 characteristic fingerprints of Bupleurum chinense. CONCLUSION: The HPLC Fingerprint of Bupleurum chinense from GAP provenance of Hubei province Baokang are established, and the difference between Bupleurum chinense before and after processing is studied initially.

Extract of Ginkgo biloba induces glutamate cysteine ligase catalytic subunit (GCLC).

The extract of Ginkgo biloba (EGb), containing 24% flavone glycosides and 6% terpenoids, is widely used to treat early-stage Alzheimer's disease, vascular dementia, peripheral claudication and vascular tinnitus. Its marked antioxidant activity has recently been demonstrated in both cell lines and animals. Glutathione (GSH) plays an important role in the antioxidant system by conjugating to xenobiotics to facilitate their export from cells. Glutamate cysteine ligase (GCL) is the rate-limiting enzyme for GSH synthesis and its catalytic subunit (GCLC) determines this de novo synthesis. Thus, induction of GCLC is a strategy to enhance the antioxidant capability in cells. The present study aimed to investigate the induction effect of EGb on GCLC in HepG2 and Hep1c1c7 cell lines. Real-time PCR, Western blot and enzyme activity assay were used to detect induction and it was found that GCLC was induced by EGb in these two cell lines. It is suggested that the antioxidant activity of EGb is (or is partly) through the induction of GCLC.

[Study on the inclusion technology of volatile oil in Shiquandabu pills].

OBJECTIVE: To extract volatile oil from Shiquandabu pills and study technological process of its inclusion compound with beta-Cyclodextrin (beta-CD) to improve its stability. METHODS: The study was carried out with steam distillation and orthogonal design. The process conditions were studied by determining the utilization and ratio of oil from Shiquandabu pills volatile oil and extract ratio of in-clusion compound. The most important factor was the ratio of oil to beta-CD. and the inclusion compound was identified by FT-IR spectra and XRD. RESULTS: The optimum preparation conditions for inclusion were best-established as oil: beta-CD was 1:8, the inclusion time and temperature were for 1.0 hours. at 20 degrees C. CONCLUSION: Tne method can be used for increasing volatile oil's stability and its solubility. It is suitable for production of medicinal preparation.

[The extraction and sulfation of polysaccharide from Polyporus umbellatus].

OBJECTIVE: To prepare and sulfate of polysaccharide from Polyporus umbellatus, and to compare the results of joined dispersant or not. METHODS: After purified, SEC-LLS was used to determine the molecular weight of polysaccharide. The polysaccharide was sulfated with the method of chlorosulfonic acid pyridine. The structures of polysaccharide and sulfated polysaccharides was determinated by FI RD analyse. RESULTS: The results showed that the distribute of molecular weight was narrow. It was a water-soluble glycosaminoglycan with a molecular weight of nearly 1. 6 x 10(5) Del. And sulfated polysaccharide was received by sulfation. CONCLUSION: A kind of beta-D-glucopyranoside was obtained with a molecular weight of nearly 1. 6 x 10(5) Del. After added dispersant, the production of sulfated polysaccharide increased, but the molecular weigh and the content of sulfate group was reduced.

[HPCE determination of matrine in matrine liposome].

OBJECTIVE: To establish an HPCE method for the determination of matrine in Matrine Liposome. METHODS: HPCE was used. The best condition was achieved with a fused silica capillary tube( 150 cm x 75 microm), 0.25 mol/L phosphate buffer (pH 6.8), at constant voltage of 20 kV and temperature at 25 degrees C. The UV detection wavelength was 206 nm. RESULTS: The calibration curve was linear in the range of 0.3-6.025 mg/ml for matrine (0.9995). The average recovery of matrine was 97.3%. CONCLUSION: This methods is simple, quick, accurate and suitable for the quality control for Matrine Liposome.

Extract of Ginkgo biloba induces phase 2 genes through Keap1-Nrf2-ARE signaling pathway.

The standard extract of Ginkgo biloba (EGb) has been demonstrated to possess remarkable antioxidant activity in both cell lines and animals. However, the molecular mechanism underlying this effect is not fully understood. Phase 2 enzymes play important roles in the antioxidant system by reducing electrophiles and reactive oxygen species (ROS). We demonstrated that EGb induced typical phase 2 genes: glutamate cysteine ligase catalytic subunit (GCLC) and glutathione-S-transferase subunit-P1 (GST-P1), by real-time PCR. To investigate the molecular mechanism of this induction, we used quinone oxidoreductase 1 (NQO1) -- Antioxidant response element (ARE) reporter assay and found that EGb activated the activity of the wild type but not the one with ARE mutated. It indicated that EGb induced these genes through ARE, a cis-acting motif located in the promoter region of nearly all phase 2 genes. Since nuclear factor erythroid 2-related factor 2 (Nrf2) binds ARE to enhance the expression of phase 2 genes, we detected the Nrf2 content in nucleus and found an accumulation of Nrf2 stimulated by EGb. In a further test of Kelch-like ECH-associated protein 1 (Keap1), the repression protein of Nrf2 in the cytosol under resting condition, we found that Keap1 content was inhibited by EGb and then more Nrf2 would be released to translocate into nucleus. Thus, EGb was testified for the first time to induce the phase 2 genes through the Keap1-Nrf2-ARE signaling pathway, which is (or part of) the antioxidant mechanism of EGb.

[Studies on identification of gypsum fibrosum by X-ray].

OBJECTIVE: To establish a new identification and analysis method of gypsum fibrosum, gypsum fibrosum praeparatum, pulvis talci and gypsum fibrosum blended with pulvis talci. METHODS: Powder X-ray diffraction fingerprint spectra method. RESULTS: Experiments and analysis were carried out on four samples. The standard X-ray diffraction fingerprint spectra and characteristic diffraction peaks of each sample were obtained. CONCLUSION: The experimental result indicates that X-ray diffraction fingerprint spectra can be used to identify and analyze gypsum fibrosum, gypsum fibrosum praeparatum, pulvis talci and gypsum fibrosum blended with pulvis talci.

[Study on the preparation of matrine-containing nanocell].

OBJECTIVE: To Study the preparation and optimization of matrine-containing nanocell. METHODS: Using the film-ultrasonic method and global optimization algorithm to prepare and formulate matrine-containing nanocell. UV spectrophotometry was used to determine the entrapment efficiency of the astragalan. Laser beacon was used to determine the diameter distribution of the nanocells. Using the scanning probe microscope to observe the surface topography of the nanocell. Transmission electron microscope was used to inspect the core-shell structure of the nanocell. RESULTS: The matrine-containing nanocells appeared to be round, integrated, well separated and uniform in size with high entrapment efficiency, the average particle size of the matrine-containing nanocells was 300 nm and the astragalan entrapment efficiency was above 90%. Preparation technology had fine reproducibility. CONCLUSION: Using global optimization algorithm can prepare and optimize matrine-containing nanocell.

[Identification on Spora Lygodii by X-ray diffraction fingerprint spectra method].

OBJECTIVE: To set up a new identification and analysis method of Spora Lygodii. METHOD: To get the extracts of Spora Lygodii using methanol, chloroform and petroleum ether as solvent and the extracts were identified, analyzed by X-ray diffraction Fourier fingerprint spectra. RESULTS: Experiments and analysis were carried out on three samples. The standard X-ray diffraction Fourier fingerprint spectra and characteristic diffraction peaks were obtained. There were some differences among the spectra of the extracts, but the characteristic diffraction peaks were obvious. CONCLUSION: The experimental result indicated that X-ray diffraction Fourier fingerprint spectra can be used to identify and analyze the Chinese traditional herb Lygodium japonicum (Thunb.) Sw.