Action of long-chain fatty acids in vitro on Ca2+-stimulatable, Mg2+-dependent ATPase activity in human red cell membranes

Abstract

Human red cell membrane Ca2+-stimulatable, Mg2+-dependent adenosine triphosphatase (Ca2+-ATPase) activity and its response to thyroid hormone have been studied following exposure of membranes in vitro to specific long-chain fatty acids. Basal enzyme activity (no added thyroid hormone) was significantly decreased by additions of 10(-9)-10(-4) M-stearic (18:0) and oleic (18:1 cis-9) acids. Methyl oleate and elaidic (18:1 trans-9), palmitic (16:0) and lauric (12:0) acids at 10(-6) and 10(-4) M were not inhibitory, nor were arachidonic (20:4) and linolenic (18:3) acids. Myristic acid (14:0) was inhibitory only at 10(-4) M. Thus, chain length of 18 carbon atoms and anionic charge were the principal determinants of inhibitory activity. Introduction of a cis-9 double bond (oleic acid) did not alter the inhibitory activity of the 18-carbon moiety (stearic acid), but the trans-9 elaidic acid did not cause enzyme inhibition. While the predominant effect of fatty acids on erythrocyte Ca2+-ATPase in situ is inhibition of basal activity, elaidic, linoleic (18:2) and palmitoleic (16:1) acids at 10(-6) and 10(-4) M stimulated the enzyme. Methyl elaidate was not stimulatory. These structure-activity relationships differ from those described for fatty acids and purified red cell Ca2+-ATPase reconstituted in liposomes. Thyroid hormone stimulation of Ca2+-ATPase was significantly decreased by stearic and oleic acids (10(-9)-10(-4) M), but also by elaidic, linoleic, palmitoleic and myristic acids. Arachidonic, palmitic and lauric acids were ineffective, as were the methyl esters of oleic and elaidic acids. Thus, inhibition of the iodothyronine effect on Ca2+-ATPase by fatty acids has similar, but not identical, structure-activity relationships to those for basal enzyme activity. To examine mechanisms for these fatty acid effects, we studied the action of oleic and stearic acids on responsiveness of the enzyme to purified calmodulin, the Ca2+-binding activator protein for Ca2+-ATPase. Oleic and stearic acids (10(-9)-10(-4) M) progressively inhibited, but did not abolish, enzyme stimulation by calmodulin (10(-9) M). Double-reciprocal analysis of the effect of oleic acid on calmodulin stimulation indicated noncompetitive inhibition. Addition of calmodulin to membranes in the presence of equimolar oleic acid restored basal enzyme activity. Oleic acid also reduced 125I-calmodulin binding to membranes, but had no effect on the binding of [125I]T4 by ghosts. The mechanism of the decrease by long chain fatty acids of Ca2+-ATPase activity in situ in human red cell ghosts thus is calmodulin-dependent and involves reduction in membrane binding of calmodulin.

Intracellular free calcium in the neutrophils of maintenance haemodialysis patients

“…In general it is known that any factor which influences the composition of the cell membrane or the calcium transporters will result in excessive altered calcium fluxes with subsequent disturbances in the intracellular calcium status (Farber, 1981; Duncan, 1991; Matthews et al ., 1993; Trump & Berezesky, 1995). In chronic renal failure, factors that may potentially influence the intracellular calcium status include the reported abnormal parathyroid hormone levels (Alexiewicz et al ., 1990; Massry & Smogorzewski, 1994), oxidative damage to cell membranes (Luciak & Trznadel, 1991; Taccone‐Gallucci et al ., 1992; Lin et al ., 1996), decreased antioxidant activity (Nagase et al ., 1996), deranged calcium‐ATPase function (Davis et al ., 1987; Zidek et al ., 1992; Taffet et al ., 1993; Takahashi & Yamaguchi, 1994; Starling et al ., 1995; Lindner et al ., 1997), deranged sodium/potassium‐ATPase function (Islam et al ., 1989) and any calcium channel agonist or antagonist present in unphysiological proportions, including calcium channel blockers.…”

Structure-activity relationships of retinoids as inhibitors of calmodulin-dependent human erythrocyte Ca2+-ATPase activity and calmodulin binding to membranes

“…60 % and 44 % as active as all-trans retinoic acid) at a concentration of 10-8 M. We have previously reported that retinol and retinal do not inhibit Ca2+-ATPase activity [14]. Although some long-chain fatty acids, such as oleic (C,8 l) and stearic (C18:0) acids inhibit Ca2+-ATPase activity [12], two short-chain fatty acids, sorbic (C6:2) and tiglic (branched C6:1) acids, were without effect on Ca2+-ATPase (results not shown).…”

“…Among these calmodulin inhibitors are N-(6-aminohexyl)-5-chloronaphthalene- 1-sulphonamide (W-7) [6], calmidazolium [7], phenothiazines such as trifluoperazine [8], Ca2+ channel blockers [9,10], and the antiarrhythmic agent amiodarone [11]. Several fatty acids, including oleic acid [12], also inhibit human erythrocyte membrane Ca21-ATPase activity. The calmodulin antagonism by fatty acids apparently does not depend upon direct interaction of fatty acids with the Ca2+-binding protein, but rather upon their interaction with the plasma membrane at or near the calmodulin-binding site [12].…”

“…Several fatty acids, including oleic acid [12], also inhibit human erythrocyte membrane Ca21-ATPase activity. The calmodulin antagonism by fatty acids apparently does not depend upon direct interaction of fatty acids with the Ca2+-binding protein, but rather upon their interaction with the plasma membrane at or near the calmodulin-binding site [12].…”

Mechanistic Bases of Neurotoxicity Provoked by Fatty Acids Accumulating in MCAD and LCHAD Deficiencies

“…Long-chain fatty acids that are routinely present in plasma of normal individuals were shown to have cytotoxic effects at high concentrations, impair ATP generation, uncouple OXPHOS, inhibit the respiratory chain, depolarize mitochondria by their protonophoric properties, induce oxidative stress, and have permeability transition. [44][45][46][47][48][49] Medium-chain fatty acid effects on mitochondrial functions are less studied, but may similarly affect mitochondrial respiration and disturb the translocation of ADP in mitochondria. 50 Therefore, it is feasible that the fatty acids that accumulate in tissues of patients with MCAD and LCHAD deficiencies (Table 3) may similarly induce cellular toxicity.…”