AMP-activated protein kinase (AMPK) is a key enzyme in the regulation of cellular energy homeostasis. Recent studies demonstrated that AMPK also plays an important role in the modulation of inflammation, an energy-intensive molecular response. The commonly used AMPK activators include 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) and A-769662. In addition, the biological activities of metformin and adiponectin are closely related to activation of AMPK. Numerous studies have shown that these AMPK activators play an effectively protective role in animal models of acute lung injury, asthma, colitis, hepatitis, atherosclerosis and other inflammatory diseases. Therefore, AMPK activators may have promising potential for the prevention and treatment of inflammation related diseases.
AMP-Activated Protein Kinases ; physiology ; Adiponectin ; pharmacology ; Aminoimidazole Carboxamide ; pharmacology ; Animals ; Enzyme Activation ; Inflammation ; enzymology ; Metformin ; pharmacology ; Pyrones ; pharmacology ; Thiophenes ; pharmacology

OBJECTIVETo establish the rat model of acute pulmonary embolism, and study the changes of vascular active substances in pulmonary embolism rats, and investigate the interventive effect of anticoagulant drugs on vascular active substances.

METHODSOne hundred and twenty-eight rats were randomly divided into four groups: control group, model group, low-molecular-weight heparin and warfarin treated group and rivaroxaban-treated group (n = 32 in each group). The method of autologous thrombosis was used to establish the animal model of acute pulmonary embolism. The animals were treated with saline or different anticoagulant drugs. The physiological and biochemical parameters were detected at different time points after embolization. The rats were killed after embolism of 24 h, 3 d, 5 d or 1 week respectively and the pathologic samples of lung tissues were collected to analyze the pulmonary pathological changes in different groups.

RESULTSRats in embolization group after blood clots injection showed shortness of breath, oral cyanosis; quicken heart rates and other symptoms. All embolization groups had pulmonary hypertension, the levels of type B natriuretic peptide (BNP) were increased significantly. The ratio of endothelin-1 (ET-1)/NO and thromboxane (TXB2) and prostacyclin (6-k-PGFla) were abnormal. After treated with effective anticoagulant drugs, the levels of BNP, ET-1, NO, TXB2 and 6-k-PGF1a were tended to the normal levels in the control group. The pulmonary hypertensions were gradually decreased. The efficacy of rivaroxaban on pulmonary embolism was the same as that of the low molecular weight heparin or warfarin.

CONCLUSIONAnticoagulation therapy can effectively improve endothelial function after pulmonary embolism, reduce pulmonary hypertension, and revise the increased BNP levels to normal levels. The efficacy of rivaroxaban is not inferior to that of low molecular weight heparin and warfarin.

Animals ; Anticoagulants ; pharmacology ; Disease Models, Animal ; Endothelin-1 ; metabolism ; Heparin, Low-Molecular-Weight ; pharmacology ; Hypertension, Pulmonary ; drug therapy ; metabolism ; Lung ; pathology ; Morpholines ; pharmacology ; Pulmonary Embolism ; drug therapy ; metabolism ; Rats ; Rivaroxaban ; Thiophenes ; pharmacology ; Warfarin ; pharmacology

OBJECTIVE: To study the expression and significance of ubiquitin-specific protease 7 (USP7) and the key factors of the Wnt signaling pathway in the lung tissue of preterm rats after hyperoxia exposure. METHODS: A total of 180 preterm neonatal Wistar rats were randomly divided into an air control group, an air intervention group, a hyperoxia control group, and a hyperoxia intervention group, with 45 rats in each group. Lung injury was induced by hyperoxia exposure in the hyperoxia groups. The preterm rats in the intervention groups were given intraperitoneal injection of the USP7 specific inhibitor P5091 (5 mg/kg) every day. The animals were sacrificed on days 3, 5, and 9 of the experiment to collect lung tissue specimens. Hematoxylin-eosin staining was used to observe the pathological changes of lung tissue. RT-PCR and Western blot were used to measure the mRNA and protein expression levels of USP7 and the key factors of the Wnt signaling pathway β-catenin and α-smooth muscle actin (α-SMA) in lung tissue. RESULTS: The air groups had normal morphology and structure of lung tissue; on days 3 and 5, the hyperoxia control group showed obvious alveolar compression and disordered structure, with obvious inflammatory cells, erythrocyte diapedesis, and interstitial edema. On day 9, the hyperoxia control group showed alveolar structural disorder and obvious thickening of the alveolar septa. Compared with the hyperoxia control group at the corresponding time points, the hyperoxia intervention group had significantly alleviated disordered structure, inflammatory cell infiltration, and bleeding in lung tissue. At each time point, the hyperoxia groups had a significantly lower radial alveolar count (RAC) than the corresponding air groups ( CONCLUSIONS Hyperoxia exposure can activate the Wnt/β-catenin signaling pathway, and USP7 may participate in hyperoxic lung injury through the Wnt/β-catenin signaling pathway. The USP7 specific inhibitor P5091 may accelerate the degradation of β-catenin by enhancing its ubiquitination, reduce lung epithelial-mesenchymal transition, and thus exert a certain protective effect against hyperoxic lung injury.
Animals ; Animals, Newborn ; Hyperoxia/physiopathology* ; Lung/physiopathology* ; Random Allocation ; Rats ; Rats, Wistar ; Thiophenes/pharmacology* ; Ubiquitin-Specific Peptidase 7/metabolism* ; Ubiquitin-Specific Proteases ; Wnt Signaling Pathway

This study is to investigate the mechanism and action characteristics of 6-chloro-3-methyl-4-(2-methyoxycarbonylthiophene-3-sulfonyl)-3, 4-dihydroquinoxa-lin-2-(1 H)-one (XU07011) against HIV-1 replication. XU07011 anti-HIV activity was tested by using VSVG/HIV pseudotype viral system and confirmed by HIV-1 live viruses' infectious assay. Time of addition was used to test HIV-1 reverse transcription process. RNA-dependent DNA polymerase activity and RNase H activity were tested by using enzyme linked immunoabsorbent assay and fluorescence method. Wild type and nine NNRTIs-resistant reverse transcriptase enzymatic models and cell-based pharmacological models were used to evaluate XU07011 bio-characteristics. The results showed that XU07011 inhibited HIV-1 replication with IC50 of (0.057 +/- 0.01) micromol x L(-1) which was comparable to nevirapine [IC50: (0.046 +/- 0.01) micromol x L(-1)]. Mechanism study data indicated that XU07011 blocked HIV-1 reverse transcription process through acting on reverse transcriptase RNA-dependent DNA polymerase with IC 50 of (1.1 +/- 0.3) micromol x L(-1). The compound showed no effect on RNase H activity. XU07011 exhibited better activities comparing with nevirapine on K103N mutated NNRTIs-resistant HIV-1 strains. This study could provide a theoretical basis for novel anti-HIV reagents development.
Anti-HIV Agents ; chemistry ; pharmacology ; Drug Resistance, Viral ; HEK293 Cells ; HIV-1 ; physiology ; Humans ; Inhibitory Concentration 50 ; Molecular Structure ; Nevirapine ; pharmacology ; Quinoxalines ; pharmacology ; RNA-Directed DNA Polymerase ; metabolism ; Ribonuclease H ; metabolism ; Thiophenes ; pharmacology ; Virus Replication ; drug effects

Methionine synthase (MS, EC2.1.1.13), a key enzyme in the folate metabolism area catalyzing methyl transfer from N5-methyltetrahydrofolate to homocysteine to give tetrahydrofolate and methionine, takes a core position in folate cycle, one-carbon-unit transfer and sculpture amino acid pathways. Cobalamin-dependent methionine synthase was purified from rat liver. The enzyme was purified 609-fold to near homogeneity by batch chromatography on DE-52, anion-exchange chromatography on Q Sepharose Fast Flow and CHT-I hydroxyapatite column and was identified by SDS-PAGE and Western blotting. The enzyme activity was determined by spectrophotometric assay. In addition, the influencing factor and optimal reaction condition were performed. The steady state kinetic of rat liver methionine synthase was similar to that of other mammalian cobalamin-dependent methionine synthase which employed a Ping-Pong mechanism. The result indicated that cobalamin-dependent methionine synthase purified from rat liver is suitable for screening and studying methionine synthase specific inhibitors.
5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase ; isolation & purification ; metabolism ; Animals ; Electrophoresis, Polyacrylamide Gel ; Folic Acid Antagonists ; pharmacology ; Liver ; chemistry ; Male ; Methotrexate ; pharmacology ; Quinazolines ; pharmacology ; Rats ; Rats, Wistar ; Tetrahydrofolates ; metabolism ; Thiophenes ; pharmacology

Animals ; Carcinoma, Hepatocellular ; enzymology ; Liver ; enzymology ; Liver Neoplasms ; enzymology ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Phenylalanine ; analogs & derivatives ; pharmacology ; Protease Inhibitors ; pharmacology ; Rats ; Rats, Wistar ; Thiophenes ; pharmacology

In this paper, duloxetine was chosen as the lead compound. The pharmacophores with 5-HT(1A) antagonism activity were used to replace the naphthyl of duloxetine. A series of duloxetine derivatives had been designed and synthesized and whose structures were confirmed with elemental analysis, MS and H NMR. All synthesized compounds were tested by tail suspension test and forced swimming test in vivo. The test results revealed that most of the compounds have shown better activity than duloxetine at the same dosage. Some of them are worth to be studied further.
Animals ; Antidepressive Agents ; chemical synthesis ; chemistry ; pharmacology ; Duloxetine Hydrochloride ; Hindlimb Suspension ; Male ; Mice ; Mice, Inbred ICR ; Molecular Structure ; Serotonin 5-HT1 Receptor Antagonists ; pharmacology ; Structure-Activity Relationship ; Swimming ; Thiophenes ; chemical synthesis ; chemistry ; pharmacology

OBJECTIVETo explore the role of bone morphogenetic protein-7 (BMP-7) in strontium ranelate (Sr)-induced osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs).

METHODSBMSCs were isolated from 4-week-old rats and cultured in vitro. The third or fourth passages of BMSCs were examined using alkaline phosphatase kit for changes in ALP activity in response to treatment with different concentrations of Sr. Calcium nodules in the induced cells were detected using alizarin red staining, and the cellular BMP-7 expression was detected by Western blotting.

RESULTSWithin the concentration range of 0.1-3.0 mmol/L, Sr dose-dependently increased ALP activity in rat BMSCs. ALP activity reached the highest level after treatment with 3 mmol/L Sr, which also significantly promoted the formation of calcium nodules. Within the range of 0.1-3.0 mmol/L, Sr dose-dependently enhanced the expression of BMP-7, and its peak expression occurred following 3 mmol/L Sr treatment. Noggin (100 ng/ml), an inhibitor of BMP-7, obviously suppressed Sr-induced over-expression of BMP-7 and reduced ALP activity and calcium nodule formation in the BMSCs.

CONCLUSIONSr promotes osteogenic differentiation of rat BMSCs by increasing the expression of BMP-7.

Animals ; Bone Density Conservation Agents ; pharmacology ; Bone Marrow Cells ; cytology ; Bone Morphogenetic Protein 7 ; genetics ; metabolism ; Cell Differentiation ; drug effects ; Cells, Cultured ; Female ; Male ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Organometallic Compounds ; pharmacology ; Osteoblasts ; cytology ; Osteogenesis ; drug effects ; Rats ; Thiophenes ; pharmacology

OBJECTIVETo investigate the protective effect of exogenous inhibitor of matrix metalloproteinases-9 (MMP-9), batimastat, in the lung injury induced by cardiopulmonary bypass (CPB) in dogs.

METHODSThirty healthy mongrel puppies were randomly divided into 3 groups: control group, low-dose group [batimastat 10 mg/(kg.d) for 3 days before operation] and high-dose group [batimastat 30 mg/(kg.d) for 3 days before operation]. The off-pump puppies' model of acute lung injury was established, and hemodynamic and respiratory parameters were monitored. The preoperative and postoperative alveolar-arterial oxygen difference (A-aDO(2)) and respiratory index (RI) were calculated. From the beginning of surgery, blood samples were taken at the time 0, 60, 120, and 270 min. Plasma concentrations of MMP-9 were measured by ELISA, and blood MMP-9 mRNA expressions were determined by RT-PCR. The myeloperoxidase (MPO) activity of centrifugal bronchoalveolar lavage fluid were measured by Colorimetry. And MMP-9 activity was determined by Gelatin zymography. Light and electronic microscope were used to observe the morphological changes of lung tissue. A small piece of left lung tissue was taken, weighed and baked to calculate the wet weight (W/D) index.

RESULTSAfter cardiopulmonary bypass, the concentrations of MMP-9 and mRNA expressions of the control group were increased significantly, and lung injury was apparent. At 270 min, the MMP-9 plasma concentration of high-dose group (17.36 +/- 1.18) microg/L was significant reducing than control group (30.47 +/- 2.22) microg/L (P < 0.05). After operation, A-aDO(2) and RI of high-dose group were significantly improved than control group (P < 0.05). The W/D index of the high-dose group (2.8 +/- 0.48) was significantly lower than that of control group (4.7 +/- 0.6) (P < 0.05). And the pathological changes of lung tissue were significantly improved in the high-dose group. However, there was no significant difference in the MMP-9 mRNA expression in three groups.

CONCLUSIONSBatimastat plays a role in the protection of the lung injury of CBP by reducing the concentration and activity of MMP-9, the degradation of the cell membrane and pulmonary neutrophil infiltration and reduction of pulmonary edema.

Acute Lung Injury ; etiology ; prevention & control ; Animals ; Cardiopulmonary Bypass ; Disease Models, Animal ; Dogs ; Lung ; pathology ; Matrix Metalloproteinase 9 ; metabolism ; Matrix Metalloproteinase Inhibitors ; Phenylalanine ; analogs & derivatives ; pharmacology ; Postoperative Complications ; prevention & control ; Thiophenes ; pharmacology

OBJECTIVETo explore whether strontium ranelate (Sr) promotes osteoblast lineage differentiation of rat bone mesenchymal stem cells (BMSCs) through the bone morphogenetic protein-2 (BMP-2)/Smad signaling pathway.

METHODSCultured rat BMSCs were exposed to different concentrations of Sr, noggin (an inhibitor of BMP-2) or Smad1 siRNA. The activity of alkaline phosphatase (ALP) in the exposed cells was detected by colorimetry, and the formation of mineralized nodules was observed with alizarin red staining. The expressions of phosphorylated (p) Smad1/5/8 and Runt-related transcription factor 2 (Runx2) in the cells were detected by Western blotting.

RESULTSExposure to Sr at 0.1 to 10 mmol/L for 1 h markedly increased the expression of p-Smad1/5/8 in the BMSCs, and the increment was the most obvious following 1 mmol/L Sr exposure. Preconditioning with 100 ng/ml noggin for 2 h inhibited Sr-induced up-regulation of p-Smad1/5/8 expressions. Exposure of the cells to 0.1 to 5 mmol/L Sr for 6 h significantly enhanced Runx2 expression, and the peak enhancement occurred following 1 mmol/L Sr exposure. Transfection of the BMSCs with Smad1 siRNA decreased the basal level of Smad1/5/8 protein expression, and also inhibited Sr-induced up-regulation of p-Smad1/5/8 and Runx2 expressions as well as Sr-induced enhancement of ALP activity and formation of mineralized nodules.

CONCLUSIONThe BMP-2/Smad pathway is involved in Sr-induced osteoblast differentiation of rat BMSCs.

Alkaline Phosphatase ; metabolism ; Animals ; Bone Marrow Cells ; cytology ; Bone Morphogenetic Protein 2 ; metabolism ; Cell Differentiation ; drug effects ; Cells, Cultured ; Mesenchymal Stromal Cells ; cytology ; Osteogenesis ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Smad1 Protein ; metabolism ; Strontium ; pharmacology ; Thiophenes ; pharmacology