Identification and partial characterization of phospholipases in isolated adrenocortical cells. The effects of synacthen [corticotropin-(1--24)-tetracosapeptide] and calcium ions.

Phospholipase A activity was determined in homogenates and subcellular fractions of trypsin-dispersed cat adrenocortical cells. At pH 7.4 homogenate phospholipid hydrolysis was activated by added Ca2+ and inhibited by EGTA. Phospholipid degradation in the presence and absence of Synacthen was completely blocked by EGTA. Ca2+-dependent activation of a membrane-bound phospholipase may be a critical control mechanism for regulating the molecular changes taking place during stimulation by Synacthen.     

Spin-trapping evidence that graded myocardial ischemia alters post-ischemic superoxide production

The spin trapping agent 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) was used to investigate oxy-radical production in post-ischemic rat hearts previously exposed to 20, 30, or 40 minutes of global ischemia. A hydroxyl spin adduct (DMPO-OH) was identified in coronary effluent during the initial seconds of reperfusion by Electron Spin Resonance (ESR) Spectroscopy. The intensity of the ESR signal in post-ischemic effluent increased as ischemic duration was prolonged; however, regardless of the duration of ischemia, maximal spin adduct detection occurred 3 minutes after initiation of reperfusion. Superoxide dismutase inhibited the formation of DMPO-OH, suggesting that superoxide anion was initially generated and is the principle source for the production on the hydroxyl adduct. Our investigations indicate that superoxide anion is produced during the early moments of reperfusion and that its production in the post-ischemic heart is related to the severity of ischemia.     

Modulation of lipolytic activity in isolated canine cardiac sarcolemma by isoproterenol and propranolol.

Cardiac sarcolemmal preparations isolated from dog were tested for membrane-associated phospholipase A and lipoprotein lipase activities. The sarcolemma hydrolyzed 1-acyl 2¹⁴C-linoleoyl 3-glycero-phosphorylethanolamine at pH 7.0 to form predominantly ¹⁴C-lyso PE with 5 mM EDTA and ¹⁴C-free fatty acid with 5 mM Ca²⁺ suggesting the presence of both phospholipases A₁ and A₂ and/or lysophospholipase activities in these preparations. Sarcolemmal PLA activity was stimulated 300% by 10^?5 to 10^?6 M d1-isoproterenol; this stimulation was blocked by 10^?4 M d1-propranolol. Lipoprotein lipase activity associated with the sarcolemmal fraction was enhanced 10-fold by 10^?5 M d1-isoproterenol; stimulation was blocked by d1-propranolol. Thus, the activities of membrane-bound lipolytic enzymes appear to be modulated by β-adrenergic agents in canine cardiac sarcolemma and could affect lipid dependent enzymes and/or membrane permeability.     

Substance P receptor blocker,aprepitant, inhibited cutaneous and other neurogenic inflammation side effects of the EGFR1-TKI,erlotinib

Molecular and Cellular Biochemistry - Calycosin-7-O-β-d-glucoside (CG) is the component of Astragali Radix, and the aim of the present study is to investigate whether CG protects myocardium...     

Identification of free radicals in myocardial ischemia/reperfusion by spin trapping with nitrone DMPO

The spin trapping ESR technique was applied to investigate oxygen-derived radicals in ischemic and post-ischemic rat hearts. Using 5,5'-dimethyl-l-pyrroline-N-oxide, carbon-centered radicals were identified during ischemia and oxy-radical adducts (superoxide anion radical, O.-2 and hydroxyl radicals, .OH) in post-ischemic rat heart. The formation of these spin adducts was inhibited by superoxide dismutase, suggesting that superoxide plays a role in the adducts' formation. The results demonstrate that oxygen derived free radicals are important byproducts of abnormal oxidative metabolism during myocardial ischemic and reperfusion injuries.     

Inhibition of cyclic AMP-dependent protein kinase activity by the cardiotonic drugs amrinone and milrinone

Amrinone and milrinone are new cardiotonic drugs that have potent inotropic and vasodilatory properties. The mechanism of action of these agents is controversial, but the positive inotropic component is thought to be due to the inhibition of phosphodiesterase. Because amrinone and milrinone have been shown to be involved primarily in cyclic AMP-mediated processes, we examined the effect of these agents on cyclic AMP-dependent protein kinase. The results indicate that amrinone and milrinone inhibit cyclic AMP-dependent protein kinase activity by competing with ATP but not cyclic AMP binding sites. Dissociation constants (Ki) of amrinone and milrinone for ATP binding sites on protein kinase were calculated to be 100-300 microM and 842 microM, respectively. The phosphodiesterase inhibitor isobutylmethylxanthine (1 mM) had no effect on protein kinase activity. Amrinone and milrinone inhibited the catalytic subunit of protein kinase to the same degree as the holo-enzyme by competitively inhibiting the binding of ATP. Amrinone and milrinone had no effect on phospholipid-sensitive, calcium-dependent protein kinase indicating that there may be differences in the ATP binding sites on these two protein kinases. Inhibition of cyclic AMP-dependent protein kinase by amrinone and milrinone occurs at concentrations higher than those used clinically. However, because amrinone and milrinone are lipophilic drugs, they may be useful tools for the investigation of protein kinase mediated reactions.     

Antioxidant and lysosomotropic properties of acute D-propranolol underlies its cardioprotection of postischemic hearts from moderate iron-overloaded rats

The benefits of acute D-propranolol (D-Pro, non-beta-adrenergic receptor blocker) pretreatment against enhanced ischemia/reperfusion (I/R) injury of hearts from moderate iron-overloaded rats were examined. Perfused hearts from iron-dextran-treated rats (450 mg/kg/week for 3 weeks, intraperitoneal administration) exhibited normal control function, despite iron treatment that elevated plasma iron and conjugated diene levels by 8.1-and 2.5-fold, respectively. However, these hearts were more susceptible to 25 mins of global I/R stress compared with non-loaded hearts; the coronary flow rate, aortic output, cardiac work, left ventricular systolic pressure, positive differential left ventricular pressure (dP/dt), and left ventricular developed pressure displayed 38%, 60%, 55%, 13%, 41%, and 15% lower recoveries, respectively, and a 6.5-fold increase in left ventricular end-diastolic pressure. Postischemic hearts from iron-loaded rats also exhibited 5.6-, 3.48-, 2.43-, and 3.45-fold increases in total effluent iron content, conjugated diene levels, lactate dehydrogenase (LDH) activity, and lysosomal N-acetyl-beta-glucosaminidase (NAGA) activity, respectively, compared with similarly stressed non-loaded hearts. A comparison of detection time profiles during reperfusion suggests that most of the oxidative injury (conjugated diene) in hearts from iron-loaded rats occurred at later times of reperfusion (8.5-15 mins), and this corresponded with heightened tissue iron and NAGA release. D-Pro (2 microM infused for 30 mins) pretreatment before ischemia protected all parameters compared with the untreated iron-loaded group; pressure indices improved 1.2- to 1.6-fold, flow parameters improved 1.70- to 2.96-fold, cardiac work improved 2.87-fold, and end-diastolic pressure was reduced 56%. D-Pro lowered total release of tissue iron, conjugated diene content, LDH activity, and NAGA activity 4.59-, 2.55-, 3.04-, and 4.14-fold, respectively, in the effluent of I/R hearts from the iron-loaded group. These findings suggest that the enhanced postischemic dysfunction and tissue injury of hearts from iron-loaded rats was caused by excessive iron-catalyzed free radical stress, and that the membrane antioxidant properties of D-Pro and its stabilization of sequestered lysosomal iron by D-Pro may contribute to the cardioprotective actions of D-Pro.