Protease Activated Receptor-2 Contributes to Heart Failure

Heart failure is a major clinical problem worldwide. Previous studies have demonstrated an important role for G protein-coupled receptors, including protease-activated receptors (PARs), in the pathology of heart hypertrophy and failure. Activation of PAR-2 on cardiomyocytes has been shown to induce hypertrophic growth in vitro. PAR-2 also contributes to myocardial infarction and heart remodeling after ischemia/reperfusion injury. In this study, we found that PAR-2 induced hypertrophic growth of cultured rat neonatal cardiomyocytes in a MEK1/2 and p38 dependent manner. In addition, PAR-2 activation on mouse cardiomyocytes increased expression of the pro-fibrotic chemokine MCP-1. Furthermore, cardiomyocyte-specific overexpression of PAR-2 in mice induced heart hypertrophy, cardiac fibrosis, inflammation and heart failure. Finally, in a mouse model of myocardial infarction induced by permanent ligation of the left anterior descending coronary artery, PAR-2 deficiency attenuated heart remodeling and improved heart function independently of its contribution to the size of the initial infarct. Taken together, our data indicate that PAR-2 signaling contributes to the pathogenesis of hypertrophy and heart failure.     

Effects of pralidoxime applied locally into the midbrain reticular formation on the hippocampal theta rhythm induced by acetylcholinesterase inhibition

After local administration into the midbrain reticular formation of an acetylcholinesterase reactivator--Pralidoxime, a significant decrease of intensity of hippocampal theta rhythm induced by previous inhibition of acetylcholinesterase by DFP was observed already after 10 min. This result suggests that cholinergic structures are localized in midbrain reticular formation and that they play a role in the origin of hippocampal theta rhythm.     

Bifunctional indole-3-acetyl transferase catalyses synthesis and hydrolysis of indole-3-acetyl-myo-inositol in immature endosperm of Zea mays

1- O -(indole-3-acetyl)- β - d -glucose: myo -inositol indoleacetyl transferase (IA- myo -inositol synthase) is an important enzyme in IAA metabolism. This enzyme catalyses the transfer of the indole acetyl (IA) moiety from 1- O -(indole-3-acetyl)- β - d -glucose to myo -inositol to form IA- myo- inositol and glucose. IA- myo -inositol synthase was purified to an electrophoretically homogenous state from maize liquid endosperm by fractionation with ammonium sulphate, anion-exchange, adsorption on hydroxylapatite, affinity chromatography on ConA-Sepharose, preparative PAGE and isoelectric focusing. We thus obtained two enzyme preparations which differ in their R _f on 8% polyacrylamide gel. The preparation of R _f 0.36 contained a single 56.4 kDa polypeptide, whereas the preparation of R _f 0.39 consisted of two polypeptides of 56.4 and 53.5 kDa. Both purified preparations of IAInos synthase also exhibited the activity of an IAInos hydrolase, showing that the dual activity was associated with a single protein. Results of gel filtration and analytical SDS-PAGE suggest that the native enzyme exists as both a monomeric (65 kDa) and homo- or heterodimeric form (110–130 kDa). Analysis of peptide maps and amino acid sequences of two 21 amino-acid peptides showed that polypeptides of 56.4 and 53.5 kDa have the same primary structure and that the 3 kDa difference in molecular mass is probably caused by different glycosylation levels. Comparison of this partial and internal amino acid sequence with sequences of other plant acyltransferases indicated similarity to several proteins which belonged to the serine carboxypeptidase-like (SCPL) acyltransferase family.     

Bronchodilatory properties of 2-decarboxy-2-hydroxymethyl prostaglandin E1

We evaluated in a double-blind study the bronchodilatory properties of 2-decarboxy-2-hydroxymethyl prostaglandin E₁ (PGE₁-carbinol), described recently as a nonirritant bronchodilator in animals. Fifteen asthmatic patients received by inhalation single doses of 1, 10, and 30 μg PGE₁-carbinol, 55 μg PGE₂, and placebo (10% ethanol in normal saline, which was also used as diluent for the PGs). Such pulmonary function tests as forced expiratory volume in 1 second, forced vital capacity, and maximal expiratory flow were monitored during 2 hours following inhalation of each compound. 10 and 30 μg PGE₁-carbinol produced significant but short-acting bronchodilation, similar to that caused by 55 μg PGE₂. One-third of the patients reported mild cough and throat irritation during and shortly after inhalation of 30 μg PGE₁-carbinol or 55 μg PGE₂. Placebo and 1 μg PGE₁-carbinol produced minimal side effects, but neither agent caused bronchodilation. In an adjunctive, unblinded trial, the same patients received 400 μg fenoterol. Fenoterol caused greater bronchodilation 15 and 30 minutes after inhalation than did the PGs in the double-blind study.     

Isomerization of 1-O-Indol-3-Ylacetyl-β-d-Glucose : Enzymatic Hydrolysis of 1-O, 4-O, and 6-O-Indol-3-YlacetyL-β-d-Glucose and the Enzymatic Synthesis of Indole-3-Acetyl Glycerol by a Hormone Metabolizing Complex

The first compound in the series of reactions leading to the ester conjugates of indole-3-acetic acid (IAA) in kernels of Zea mays sweet corn is the acyl alkyl acetal, 1-O-indol-3-ylacetyl-β-d-glucose (1-O-IAGlu). The enzyme catalyzing the synthesis of this compound is UDP-glucose:indol-3-ylacetate glucosyl-transferase (IAGlu synthase). The IAA moiety of the high energy compound 1-O-IAGlu may be enzymatically transferred to myo-inositol or to glycerol or the 1-O-IAGlu may be enzymatically hydrolyzed. Alternatively, nonenzymatic acyl migration may occur to yield the 2-O, 4-O, and 6-O esters of IAA and glucose. The 4-O and 6-O esters may then be enzymatically hydrolyzed to yield free IAA and glucose. This work reports new enzymatic activities, the transfer of IAA from 1-O-IAGlu to glycerol, and the enzymecatalyzed hydrolysis of 4-O- and 6-O-IAGlu. Data is also presented on the rate of non-enzymatic acyl migration of IAA from the 1-O to the 4-O and 6-O positions of glucose. We also report that enzymes catalyzing the synthesis of 1-O-IAGlu and the hydrolysis of 1-O, 4-O, and 6-O-IAGlu fractionate as a hormone metabolizing complex. The association of synthetic and hydrolytic capabilities in enzymes which cofractionate may have physiological significance.     

Glucose determination using immobilized polyphosphate glucokinase.

Polyphosphate glucokinase (EC 2.7.1.63, polyphosphate:glucose phosphotransferase) was covalently coupled to collagen-coated silica gel beads. The immobilized enzyme, as a packed-bed reactor, was used to determine glucose in serum and other samples. The method was based on a spectrophotometric measurement of NADPH produced by two consecutive reactions, similar to the hexokinase method. The described approach takes advantage of the greater stability of polyphosphate compared to that of ATP, the greater specificity of polyphosphate glucokinase versus that of hexokinase, and the reusability of the immobilized enzyme. Linearity, precision, and accuracy of the method were tested and found to be very good. The results were linear between 10 and 50 nmol of glucose in a 50-microliter sample and the coefficient of variation was less than 4% in five successive determinations. The recovery of glucose was about 100% after calibration of the method. The results of the measurements correlated well with those obtained with soluble polyphosphate glucokinase (r = 0.997, y = 1.036x - 0.016). The immobilized-enzyme reactor showed good operational stability during a month of use, losing about 12% of its initial activity.     

Submitochondrial localization and function of alkaline inorganic pyrophosphatase in maize seedlings.

Alkaline inorganic pyrophosphatase and Mg-ATPase are localized within the mitoplast of maize seeding mitochondria. NaF inhibited the PPase activity, whereas oligomycin and dicyclohexylcarbodiimide inhibited the Mg-ATPase activity. The mitoplast preparation synthesized PPi from Pi under conditions excluding hydrolysis of endogenous ATP. PPi synthesis was inhibited by ADP, antimycin A, NaCN and 2,4- dinitrophenol but not by oligomycin. It is suggested that PPi synthesis in the maize seedling mitochondria proceeds at the expense of the energy of electron transport chain and is independent of the ATP synthesis.