OBJECTIVES: To investigate the effect of Haematococcus pluvialis (HP) on bleomycin (BLM)-induced pulmonary fibrosis in mice and on TGF-β1-induced human fetal lung fibroblasts (HFL1). METHODS: Thirty male C57BL/6 mice were randomly divided into control group, BLM-induced pulmonary fibrosis model group, low- and high-dose HP treatment groups (3 and 21 mg/kg, respectively), and 300 mg/kg pirfenidone (positive control) group. The effects of drug treatment for 21 days were assessed by examining respiratory function, lung histopathology, and expression of fibrosis markers in the lung tissues of the mouse models. In TGF-β1-induced HFL1 cell cultures, the effects of treatment with 120, 180 and 240 μg/mL HP or 1.85 μg/mL pirfenidone for 48 h on expression levels of fibrosis markers were evaluated. Transcriptome analysis was carried out using the control cells and cells treated with TGF-β1 and 240 μg/mL HP. RESULTS: HP obviously alleviated BLM-induced lung function damage and fibrotic changes in mice, evidenced by improved respiratory function, lung tissue morphology and structure, inflammatory infiltration, and collagen deposition and reduced expressions of fibrotic proteins. HP at the high dose produced similar effect to PFD. In TGF-β1-induced HFL1 cells, treatment with 240 μg/mL HP significantly reduced the mRNA and protein expression levels of α-SMA and FN. Transcriptome analysis revealed that multiple key genes and pathways mediated the protective effect of HP against pulmonary fibrosis. CONCLUSIONS HP alleviates pulmonary fibrosis in both the mouse model and cell model, possibly as the result of the synergistic effects of its multiple active components.
Animals ; Pulmonary Fibrosis/chemically induced* ; Bleomycin/adverse effects* ; Mice, Inbred C57BL ; Male ; Mice ; Fibroblasts/drug effects* ; Lung/pathology* ; Transforming Growth Factor beta1/pharmacology* ; Myofibroblasts/drug effects* ; Humans ; Pyridones

ADC189 is a novel drug of cap-dependent endonuclease inhibitor. In our study, its antiviral efficacy was evaluated in vitro and in vivo, and compared with baloxavir marboxil and oseltamivir. A first-in-human phase I study in healthy volunteers included single ascending dose (SAD) and food effect (FE) parts. In the preclinical study, ADC189 showed potent antiviral activity against various types of influenza viruses, including H1N1, H3N2, influenza B virus, and highly pathogenic avian influenza, comparable to baloxavir marboxil. Additionally, ADC189 exhibited much better antiviral efficacy than oseltamivir in H1N1 infected mice. In the phase I study, ADC189 was rapidly metabolized to ADC189-I07, and its exposure increased proportionally with the dose. The terminal elimination half-life (T_1/2) ranged from 76.69 to 98.28 hours. Of note, food had no effect on the concentration, clearance, and exposure of ADC189. It was well tolerated, with few treatment-emergent adverse events (TEAEs) reported and no serious adverse events (SAEs). ADC189 demonstrated excellent antiviral efficacy both in vitro and in vivo. It was safe, well-tolerated, and had favorable pharmacokinetic characteristics in healthy volunteers, supporting its potential for single oral dosing in clinical practice.
Humans ; Antiviral Agents/therapeutic use* ; Animals ; Male ; Adult ; Mice ; Female ; Endonucleases/antagonists & inhibitors* ; Influenza, Human/drug therapy* ; Young Adult ; Dibenzothiepins/pharmacology* ; Oseltamivir/pharmacology* ; Middle Aged ; Triazines/pharmacology* ; Thiepins/pharmacology* ; Influenza B virus/drug effects* ; Influenza A Virus, H1N1 Subtype/drug effects* ; Pyridines/pharmacology* ; Morpholines ; Pyridones

OBJECTIVE: To investigate the effect of pirfenidone for reducing urethral stricture following urethral injury in rats and explore the possible mechanism. METHODS: Thirty male SD rats were randomly assigned into negative control group, positive control group and pirfenidone group (n=10). In pirfenidone and positive control groups, the rats were subjected to incision of the posterior urethral cavernous body followed by daily intraperitoneal injection of pirfenidone (100 mg/kg) and an equivalent volume of solvent, respectively. The rats in the negative control group were given intraperitoneal injections of solvent without urethral injury. At two weeks after modeling, retrograde urethrography was performed for observing urethral stricture, and the injured urethral tissues were harvested for HE staining, Masson staining, immunohistochemical staining and Western blotting for detecting the protein expressions of α-SMA and TGF-β1. The mRNA expressions of the inflammatory factors TNF-α, IL-6, and IL-1β were detected using qRT-PCR. RESULTS: The body weight of the rats in pirfenidone group was significantly decreased compared with that in the other two groups (P < 0.05). Retrograde urethrography showed significant narrowing of the urethra in the positive control group but not in the pirfenidone group. HE staining of the injured urethral tissues showed obvious proliferation of urethral epithelial cells with narrow urethral cavity and increased inflammatory cells in positive control group. The pathological findings of the urethra were similar between pirfenidone group and the negative control group. Masson staining revealed obviously reduced collagen fibers and regular arrangement of the fibers in pirfenidone group as compared to the positive control group. Compared with those in the negative control group, the expressions of α-SMA and TGF-β1 were significantly increased in the positive control group, and pirfenidone treatment significantly inhibited their expressions (P < 0.05 or 0.01). Pirfenidone also significantly inhibited the mRNA expressions of TNF-α, IL-6, and IL-1β in the injured urethral tissue (P < 0.05 or 0.01). CONCLUSION Pirfenidone can prevent urethral fibrosis and stricture after urethral injury possibly by inhibiting the TGF-β1 pathway and inflammatory response.
Animals ; Female ; Humans ; Interleukin-6/metabolism* ; Male ; Pyridones/pharmacology* ; RNA, Messenger/metabolism* ; Rats ; Rats, Sprague-Dawley ; Solvents ; Transforming Growth Factor beta1/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Urethral Stricture/pathology*

OBJECTIVETo explore the therapeutic effect of enhancer of Zeste homolog 2 (EZH2) inhibitor GSK343 on periodontitis by regulating microphage differentiation.

METHODSMacrophage RAW264.7 cells were divided into the blank (A group), control (B group), lipopolysaccharide (LPS) stimulation (C group), and LPS+GSK343 (D group) groups. Phenotype transformations was determined through Western blot analysis and enzyme-linked immunosorbent assay by detecting the differentiation of phenotypic biological markers, including tumor necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS), interleukin-10 (IL-10), and Arginase-1 (Arg-1). Metergasis was identified by performing a phagocytosis test on Escherichia coli (E. coli).

RESULTSMacrophage RAW264.7 cells produced classical phenotypic biomarkers (M1) TNF-α and iNOS under LPS stimulation. The expression levels of IL-10 and Arg-1 increased after adding GSK343 into the culture medium. GSK343 also induced the conversion of M1 macrophages into M2 macrophages. Macrophage RAW264.7 cells exerted a phagocytic effect on E. coli, and this effect was enhanced after adding LPS into the culture medium. GSK343 regulated the macrophage RAW264.7 phagocytosis of E. coli.

CONCLUSIONSGSK343 possibly participates in the regulation of macrophage differentiation and, consequently, in the latent treatment of periodontitis.

Arginase ; Cell Differentiation ; Enhancer of Zeste Homolog 2 Protein ; Enzyme Inhibitors ; pharmacology ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; Indazoles ; pharmacology ; Interleukin-10 ; Lipopolysaccharides ; Macrophages ; Nitric Oxide Synthase Type II ; Periodontitis ; Phagocytosis ; Pyridones ; pharmacology ; Tumor Necrosis Factor-alpha

To investigate the effect of a novel EZH2 inhibitor GSK126 on cell growth, apoptosis and migration of prostate cancer cells.Prostate cancer PC-3 and DU145 cells were treated with GSK126 at different doses. Cell growth was detected by sulforhodamine assay. Cell apoptosis was assayed by Annexin V-/PI kit. Transwell chamber and wound healing assays were conducted to detect cell migration. The mRNA level was detected by quantitative PCR, and protein expression was detected by Western blot analysis.GSK126 showed significant effect on cell growth and apoptosis when the dose was higher than 50 μmol/L. Wound healing assay revealed that scratch space in PC-3 cells was significantly increased in a dose-dependent manner in GSK126-treated groups[(247.2±24.4),(347.2±19.2) and (410.5±18.1) μm in low, medium and high dose (5.0, 20.0, 50.0 μmol/L), respectively] as compared with the control group[(171.3±17.8) μm](all<0.05). Transwell assay showed that migrated PC-3 cells in control group was 322.0±17.9,while those in GSK126-treated groups were 198.3±15.4 (low),82.7±6.2 (medium) and 30.2±4.1 (high), and the differences between the control group and GSK126-treated groups were significant(all<0.05). In addition, GSK126 up-regulated E-cadherin mRNA expression and down-regulated N-cadherin and Vimentin mRNA expression, whereas had no significant effect on Snail, Fibronectin and VEGF-A mRNA expression. The protein expression of E-cadherin was elevated but VEGF-A protein did not change in GSK126-treated groups. Similar results were exhibited in DU145 cell.GSK126 can significantly inhibit cell migration and invasion in prostate cancer PC-3 and DU145 cells, which may be resulted from its effect on epithelial-mesenchymal transition. GSK126 may be used as a potential anti-prostate cancer dug in clinic.
Apoptosis ; drug effects ; Cadherins ; analysis ; drug effects ; metabolism ; Cell Line, Tumor ; drug effects ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Down-Regulation ; drug effects ; Drug Screening Assays, Antitumor ; methods ; Enhancer of Zeste Homolog 2 Protein ; analysis ; drug effects ; metabolism ; Fibronectins ; analysis ; drug effects ; metabolism ; Humans ; Indoles ; pharmacology ; Male ; Prostatic Neoplasms ; chemistry ; genetics ; physiopathology ; Pyridones ; pharmacology ; RNA, Messenger ; Up-Regulation ; drug effects ; Vascular Endothelial Growth Factor A ; analysis ; drug effects ; Vimentin ; analysis ; drug effects ; metabolism

Axl encodes the tyrosine-protein kinase receptor, participating in the proliferation and migration of many cells. This study examined the role of Axl in functions of endothelial progenitor cells (EPCs). Axl was detected by RT-PCR and Western blotting in both placentas and EPCs from normal pregnancy and preeclampsia patients. The Axl inhibitor, BMS777-607, was used to inhibit the Axl signalling pathway in EPCs. Cell proliferation, differentiation, migration and adhesion were measured by CCK-8 assay, cell differentiation assay, Transwell assay, and cell adhesion assay, respectively. Results showed the expression levels of Axl mRNA and protein were significantly higher in both placentas and EPCs from preeclampsia patients than from normal pregnancy (P<0.05). After treatment with BMS777-607, proliferation, differentiation, migration and adhesion capability of EPCs were all significantly decreased. Our study suggests Axl may play a role in the function of EPCs, thereby involving in the pathogenesis of preeclampsia.
Adult ; Aminopyridines ; pharmacology ; Blood Pressure ; Case-Control Studies ; Cell Adhesion ; drug effects ; Cell Differentiation ; drug effects ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Female ; Fetal Blood ; cytology ; enzymology ; Gene Expression Regulation ; Gestational Age ; Human Umbilical Vein Endothelial Cells ; drug effects ; enzymology ; pathology ; Humans ; Placenta ; metabolism ; physiopathology ; Pre-Eclampsia ; blood ; genetics ; physiopathology ; Pregnancy ; Primary Cell Culture ; Protein Kinase Inhibitors ; pharmacology ; Proto-Oncogene Proteins ; antagonists & inhibitors ; genetics ; metabolism ; Pyridones ; pharmacology ; RNA, Messenger ; antagonists & inhibitors ; genetics ; metabolism ; Receptor Protein-Tyrosine Kinases ; antagonists & inhibitors ; genetics ; metabolism ; Stem Cells ; drug effects ; enzymology ; pathology

Fibrosis can occur in different organs with high incidence rate and great danger. It still lacks effective drugs for prevention of fibrosis. Fluorofenidone is a newly developed drug with anti-fibrotic activity, which provides a new hope for treating the progressive fibrotic diseases. Recent studies have shown that fluorofenidone is a multifunctional small molecule with anti-inflammatory, antioxidative and anti-apoptotic eff ects. It can inhibit the activation and proliferation of myofibroblasts, promote the degradation of extracellular matrix and regulate the cellular signal transmission. Fluorofenidone can be applied to attenuate the progression of renal, hepatic and pulmonary fibrosis.
Apoptosis ; Extracellular Matrix ; Fibrosis ; Humans ; Kidney ; pathology ; Liver Cirrhosis ; Pulmonary Fibrosis ; Pyridones ; pharmacology ; Signal Transduction

OBJECTIVETo develop a LC-MS/MS method for determination of deferiprone in cell lysate and to study the potential interaction between deferiprone and hOCTs or hOAT1 transporters in vitro.

METHODSThe determination was performed on an Agilent Eclipse Plus C18 column(3.5 μm, 2.1 mm×50 mm).The gradient mobile phase was composed of solvent A:0.1% formic acid in water, and B:0.1% formic acid in acetonitrile. The mass spectrometer with an electrospray interface was operated in positive ion mode with multiple reaction monitoring (MRM) scan mode monitored the ion pair of deferiprone at m/z 140→96, or phenacetin at m/z 180→110. The effects of deferiprone on the accumulation of typical substrates of hOCTs and hOAT1 were evaluated by MDCK-hOCTs and MDCK-hOAT1 cells respectively. The accumulation of deferiprone was also investigated in MDCK-hOCTs cells and mock cells with or without typical inhibitors.

RESULTSThe standard curve was linear over the range of 5-300 nmol/L. The assay recovery of deferiprone was above 94%, and the intra-day precision (RSD) was less than 2.0%. The accumulation of MPP(+) in MDCK-hOCTs cells with 300 μmol/L deferiprone were 73.5%, 87.1% and 70.4%, respectively. The uptake of deferiprone in MDCK-hOCTs and mock cells did not show significant difference. Deferiprone of 100 μmol/L did not significantly affect the accumulation of 6-CF in MDCK-hOAT1 cell.

CONCLUSIONThe method is sensitivity and suitable for the determination of deferiprone in cell lysate. Deferiprone can significantly inhibit hOCT1 and hOCT3, but has no effects on hOCT2 and hOAT1. hOCTs may not play a major role in the transport of deferiprone.

Animals ; Chromatography, Liquid ; Dogs ; Humans ; Madin Darby Canine Kidney Cells ; Organic Anion Transporters ; drug effects ; Pyridones ; pharmacology ; Tandem Mass Spectrometry

The aim of this study was to investigate the effects of H3K27 methylation inhibitor EPZ005687 on the apoptosis, proliferation and cell cycle of U937 cells and normal CD34⁺ cells. The U937 cells and normal CD34⁺ cells were treated with different concentration of EPZ005687 at different time points. The apoptosis rate was determined by Annexin V/PI staining. The cell proliferation and cell cycle was determined using WST-1 assay and 7-AAD assay, respectively. The activity of H3K27 methylation was detected by chemiluminescent immunoassay. The results showed that the EPZ005687 induced an obvious apoptosis of U937 cells. The apoptotic rate was 3.96% ± 0.79%,5.74% ± 0.73%,13.34% ± 1.77% and 25.24% ± 2.55% in U937 cells treated with 0.5, 1, 5 and 10 µmol/L EPZ005687 for 48 hours, respectively. However, EPZ005687 had rare effect on normal bone marrow(NBM) CD34⁺ cells. The apoptotic rate was 3.64% ± 0.62%,4.28% ± 0.99%,6.18% ± 1.19% and 7.56% ± 1.34% after U937 cells were treated with 0.5, 1, 5 and 10 µmol/L EPZ005687 for 48 hours, respectively. EPZ005687 inhibited obviously the proliferation of U937 cells but had weak effect on the proliferation of NBMCD34⁺ cells. The inhibitory effect of EPZ005687 on U937 cells was time-dependent after treated with 0.5, 1, 5 and 10 µmol/L EPZ005687 from 12 to 96 hours. EPZ005687 induced G1 phase blocking (G1%, 64.18% ± 13.27% vs 49.43% ± 12.54%) and decreased the percentage of cells in S phase (9.67% ± 2.61% vs15.26% ± 5.58%) in U937 cells. However, EPZ005687 had no effect on the cell cycle of NBMCD34⁺ cells. In addition, EPZ005687 produced obviously depletion of H3K27 methylation in U937 cells (P < 0.05), but hardly had effect on the H3K27 methylation of NBMCD34⁺ cells. It is concluded that the EPZ005687 inhibites proliferation, induces apoptosis and cell cycle blocking in G1 phase in leukemia cells. This agent may have potential value in clinical application.
Antigens, CD34 ; metabolism ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Humans ; Indazoles ; pharmacology ; Methylation ; Pyridones ; pharmacology ; U937 Cells

OBJECTIVETo study the function of ABA and fluridone on the contents of penolic acids and two key synthetases (PAL and TAT).

METHODConducted 4 different concentrations in the hairy root of Salvia miltiorrhiza after culturing 18 days and treated with fluridone. One day later, harvested the hairy root and measured the activity of PAL and TAT; Treatment for 6 days, gathered and determined the contents of phenolic acids.

RESULTIn certain concentration of ABA, lower ABA could induced the production of growth and higher ABA inhibitor the growth in hairy roots of S. miltiorrhiza; ABA induced the accumulation of caffeic acid considerably, and the effect on the contents of coffee acid show positive correlation; As for the RA and LAB, the low dosage of ABA simulated the production and higher ABA inhibited the production of them; the ABA biosynthetic inhibitor fluridone can decreases ABA's the effect; The different of ABA activated the activity of PAL and TAT, but the impact were discriminating, when treatment with ABA and fluridone, the inducing were declined.

CONCLUSIONABA induced the accumulation of.

Abscisic Acid ; antagonists & inhibitors ; metabolism ; pharmacology ; Antioxidants ; analysis ; metabolism ; Biomass ; Caffeic Acids ; analysis ; metabolism ; Herbicides ; pharmacology ; Hydroxybenzoates ; analysis ; metabolism ; Medicine, Chinese Traditional ; Phenylalanine Ammonia-Lyase ; drug effects ; metabolism ; Plant Roots ; drug effects ; enzymology ; growth & development ; Pyridones ; pharmacology ; Salvia miltiorrhiza ; drug effects ; enzymology ; growth & development ; Time Factors ; Tyrosine Transaminase ; drug effects ; metabolism