Using different models of focal cerebral ischemia, the temporal and spatial rules of metabolism and energy changes in the post-ischemia brain tissue were measured by proton magnetic resonance spectroscopy (1HMRS) to provide valuable information for judging the prognosis of acute focal cerebral ischemia and carrying out effective therapy. Nine healthy Sprague-Dawly rats (both sexes) were randomly divided into two groups: The rats in the group A (n = 4) were occluded with self-thrombus for 1 h; The rats in the group B (n = 5) were occluded with thread-emboli for 1 h. The 1H MRS at 30, 40, 50, 60 min respectively was examined and the metabolic changes of NAA, Cho and Lac in the regions of interest were semiquantitatively analyzed. The spectrum integral calculus area ratio of NAA, Cho, Lac to Pcr + Cr was set as the criterion. The values of NAA.Cho in the regions of interest were declined gradually within 1 h after ischemia, especially, the ratio of Cho/(Pcr + Cr), NAA/(Pcr + Cr) at 60 min had significant difference with that at 50 min (P < 0.05). The ratio of Lac/(Pcr + Cr) began to decrease at 40 min from initial increase of Lac in both A and B groups. MR proton spectrum analysis was a non-invasive, direct and comprehensive tool for the study of cellular metabolism and the status of the biochemical energy in acute ischemia stroke.
Brain Ischemia/*diagnosis ; Energy Metabolism ; Infarction, Middle Cerebral Artery/diagnosis ; *Magnetic Resonance Spectroscopy ; Phosphorylcholine/metabolism ; Random Allocation ; Rats, Sprague-Dawley

To explore the biofilm inhibitory efficacy of perifosine against Pseudomonas aeruginosa (P. aeruginos) and its mechanisms. Twenty-fourwell plate was used to form biofilms at the bottom and crystal violet staining was used to determine the biofilm inhibitory effects of perifosine against P. aeruginosa, the wells without perifosine was set as control group. Glass tubes combined with crystal violet staining was used to detect the gas-liqud interface related bioiflm inhibitory effects of perifosine, the wells without perifosine was set as control group. Time-growth curved was used to detect the effects of perifosine on the bacteial planktonic cells growth of P. aeruginosa, the wells without perifosine was set as control group. The interaction model between perifosine and PqsE was assessed by molecular docking assay. The inhibitory effects of perifosine on the catalytic activity of PqsE was determined by detection the production of thiols, the wells without perifosine was set as control group. Binding affinity between perifosine and PqsE was detected by plasma surface resonance. The biofims at the bottom of the microplates and air-liquid interface were effectively inhibited by perifosine at the concentration of 4-8 μg/ml. There was no influence of perifosine on the cells growth of P. aeruginosa. The resuts of molecular docking assay indicates that perifosine could interacted with PqsE with the docking score of -10.67 kcal/mol. Perifosine could inhibit the catalytic activity of PqsE in a dose-dependent manner. The binding affinity between perifosine and PqsE was comfirmed by plasma surface resonance with KD of 6.65×10^-5mol/L. Perifosine could inhibited the biofilm formation of P. aeruginosa by interacting with PqsE.
Anti-Bacterial Agents/pharmacology* ; Bacterial Proteins/metabolism* ; Biofilms ; Molecular Docking Simulation ; Phosphorylcholine/analogs & derivatives* ; Pseudomonas aeruginosa/metabolism* ; Quorum Sensing

Using different models of focal cerebral ischemia, the temporal and spatial rules of metabolism and energy changes in the post-ischemia brain tissue were measured by proton magnetic resonance spectroscopy (1HMRS) to provide valuable information for judging the prognosis of acute focal cerebral ischemia and carrying out effective therapy. Nine healthy Sprague-Dawly rats (both sexes) were randomly divided into two groups: The rats in the group A (n = 4) were occluded with self-thrombus for 1 h; The rats in the group B (n = 5) were occluded with thread-emboli for 1 h. The 1H MRS at 30, 40, 50, 60 min respectively was examined and the metabolic changes of NAA, Cho and Lac in the regions of interest were semiquantitatively analyzed. The spectrum integral calculus area ratio of NAA, Cho, Lac to Pcr + Cr was set as the criterion. The values of NAA.Cho in the regions of interest were declined gradually within 1 h after ischemia, especially, the ratio of Cho/(Pcr + Cr), NAA/(Pcr + Cr) at 60 min had significant difference with that at 50 min (P < 0.05). The ratio of Lac/(Pcr + Cr) began to decrease at 40 min from initial increase of Lac in both A and B groups. MR proton spectrum analysis was a non-invasive, direct and comprehensive tool for the study of cellular metabolism and the status of the biochemical energy in acute ischemia stroke.
Animals ; Brain Ischemia ; diagnosis ; Energy Metabolism ; Infarction, Middle Cerebral Artery ; diagnosis ; Magnetic Resonance Spectroscopy ; Phosphorylcholine ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley

Both Fas and PMA can activate phospholipase D via activation of protein kinase Cbeta in A20 cells. Phospholipase D activity was increased 4 fold in the presence of Fas and 2.5 fold in the presence of PMA. The possible involvement of tyrosine phosphorylation in Fas-induced activation of phospholipase D was investigated. In five minute after Fas cross-linking, there was a prominent increase in tyrosine phosphorylated proteins, and it was completely inhibited by D609, a specific inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC). A tyrosine kinase inhibitor, genistein, can partially inhibit Fas-induced phospholipase D activation. There were no effects of genistein on Fas-induced activation of PC-PLC and protein kinase C. These results strongly indicate that tyrosine phosphorylation may in part account for the increase in phospholipase D activity by Fas cross-linking and D609 can block not only PC-PLC activity but also tyrosine phosphorylation involved in Fas-induced phospholipase D activation.
Animal ; Antibodies, Monoclonal/immunology/*pharmacology ; Antigens, CD95/immunology/*metabolism ; Bridged Compounds/*pharmacology ; Cell Line ; Cross-Linking Reagents ; Dose-Response Relationship, Immunologic ; Enzyme Activation ; Genistein/pharmacology ; Hydrolysis ; Lymphoma/pathology ; Mice ; Phospholipase C/*antagonists & inhibitors ; Phospholipase D/*metabolism ; Phosphorylation ; Phosphorylcholine/metabolism ; Solubility ; Thiones/*pharmacology ; Tumor Cells, Cultured ; Tyrosine/*metabolism ; Water/chemistry

Both Fas and PMA can activate phospholipase D via activation of protein kinase Cbeta in A20 cells. Phospholipase D activity was increased 4 fold in the presence of Fas and 2.5 fold in the presence of PMA. The possible involvement of tyrosine phosphorylation in Fas-induced activation of phospholipase D was investigated. In five minute after Fas cross-linking, there was a prominent increase in tyrosine phosphorylated proteins, and it was completely inhibited by D609, a specific inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC). A tyrosine kinase inhibitor, genistein, can partially inhibit Fas-induced phospholipase D activation. There were no effects of genistein on Fas-induced activation of PC-PLC and protein kinase C. These results strongly indicate that tyrosine phosphorylation may in part account for the increase in phospholipase D activity by Fas cross-linking and D609 can block not only PC-PLC activity but also tyrosine phosphorylation involved in Fas-induced phospholipase D activation.
Animal ; Antibodies, Monoclonal/immunology/*pharmacology ; Antigens, CD95/immunology/*metabolism ; Bridged Compounds/*pharmacology ; Cell Line ; Cross-Linking Reagents ; Dose-Response Relationship, Immunologic ; Enzyme Activation ; Genistein/pharmacology ; Hydrolysis ; Lymphoma/pathology ; Mice ; Phospholipase C/*antagonists & inhibitors ; Phospholipase D/*metabolism ; Phosphorylation ; Phosphorylcholine/metabolism ; Solubility ; Thiones/*pharmacology ; Tumor Cells, Cultured ; Tyrosine/*metabolism ; Water/chemistry

Sphingosylphosphorylcholine (SPC) induces differentiation of human adipose tissue-derived mesenchymal stem cells (hASCs) into smooth muscle-like cells expressing alpha-smooth muscle actin (alpha-SMA) via transforming growth factor-beta1/Smad2- and RhoA/Rho kinase-dependent mechanisms. 3-Hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors (statins) have been known to have beneficial effects in the treatment of cardiovascular diseases. In the present study, we examined the effects of simvastatin on the SPC-induced alpha-SMA expression and Smad2 phosphorylation in hASCs. Simvastatin inhibited the SPC-induced alpha-SMA expression and sustained phosphorylation of Smad2 in hASCs. SPC treatment caused RhoA activation via a simvastatin-sensitive mechanism. The SPC-induced alpha-SMA expression and Smad2 phosphorylation were abrogated by pretreatment of the cells with the Rho kinase inhibitor Y27632 or overexpression of a dominant negative RhoA mutant. Furthermore, SPC induced secretion of TGF-beta1 and pretreatment with either Y27632 or simvastatin inhibited the SPC-induced TGF-beta1 secretion. These results suggest that simvastatin inhibits SPC-induced differentiation of hASCs into smooth muscle cells by attenuating the RhoA/Rho kinase-dependent activation of autocrine TGF-beta1/Smad2 signaling pathway.
Amides/pharmacology ; Blotting, Western ; Cell Differentiation/*drug effects ; Cells, Cultured ; Enzyme-Linked Immunosorbent Assay ; Humans ; Immunohistochemistry ; Mesenchymal Stem Cells/*cytology/*drug effects ; Myocytes, Smooth Muscle/*cytology/*drug effects ; Phosphorylcholine/*analogs & derivatives/pharmacology ; Pyridines/pharmacology ; Simvastatin/*pharmacology ; Sphingosine/*analogs & derivatives/pharmacology ; rhoA GTP-Binding Protein/antagonists & inhibitors/metabolism