Serum E-selectin and erythrocyte membrane Na+K+ ATPase levels in patients with rheumatoid arthritis

We conducted this study to assess serum soluble E-selectin (sE-selectin) levels and erythrocyte membrane Na(+)K(+) ATPase activity in patients with rheumatoid arthritis (RA) and correlate the levels with disease activity. Levels of sE-selectin were measured in the serum of 20 patients with RA and 20 control subjects by an enzyme-linked immunosorbant assay. Na(+)K(+) ATPase activity was determined by a colorimetric method in RA patients and healthy controls. There were no statistically significant differences between the two groups with respect to demographic data such as age and sex (p > 0.05). The serum levels of sE-selectin, ESR and C-reactive protein (CRP) in RA patients were significantly higher than in healthy controls (p < 0.001). Erythrocyte membrane Na(+)K(+) ATPase activity was significantly lower in the RA group than in the control group (p < 0.001). Correlation analysis revealed significant positive correlations between soluble E-selectin and ESR (r = 0.457; p < 0.05) and CRP (r = 0.682; p < 0.01) levels. There were statistically significant negative correlations between erythrocyte membrane Na(+)K(+) ATPase activity and ESR (r = -0.450; p < 0.05) and CRP (r = -0.446; p < 0.05) levels. Additionally, a significant negative correlations between sE-selectin and Na(+)K(+) ATPase activity was observed (r = -0.80; p < 0.001). These results show that decreases in erythrocyte membrane Na(+)K(+) ATPase activity and increases in sE-selectin are observed in RA, and that increased levels of sE-selectin may also reflect disease status or activity.     

Phospholipid N-methylation in diabetic erythrocytes: effects on membrane Na+, K+ ATPase activity.

Phospholipid methylation was quantified in non-diabetic and streptozotocin diabetic rat erythrocytes. While the total mass of methylated lipids remained the same in both groups, the relative abundance of individual methylated lipid species differed significantly in diabetic erythrocytes. Moreover, incubation of erythrocytes membranes with S-adenosyl methionine, a substrate for methyl transferases, not only increased membrane lipid methylation but also decreased Na+, K+ ATPase activity significantly. These results suggest that phospholipid methylation may cause the observed depression of erythrocyte Na+, K+ ATPase activity in diabetes and could contribute to the altered rheology of erythrocytes in diabetes.     

The program of friend cell erythroid differentiation: Early changes in Na+/K+ ATPase function

Treatment of Friend erythroleukemia cells with several different chemical agents causes an early decrease in the ⁸⁶Rb⁺ influx mediated by Na⁺/K⁺ adenosine triphosphatase (ATPase). These agents, which induced Friend cells to differentiate, include dimethylsulfoxide (DMSO), ouabain, hypoxanthine, and actinomycin D. The magnitude of the early decrease in ⁸⁶Rb⁺ influx correlates with the proportion of cells in cultures of inducible Friend cell clones which later go on to synthesize hemoglobin. Compounds which do not incude differentiation in these cells, such as xanthine, exogenous hematin, and erythropoietin, do not cause a change in ⁸⁶Rb⁺ influx. A change in the intracellular K⁺ ion concentration does not occur during induction by DMSO because, although there is a decrease in K⁺ content per cell soon after induction, there is a parallel decrease in cell volume. These results and previous observations from this laboratory are discussed in terms of the posible involvement of the Na⁺/K⁺ ATPase in Friend cell differentiation.     

Endothelin 1 Stimulates Na+,K+-ATPase and Na+-K+-Cl− Cotransport Through ETA Receptors and Protein Kinase C-Dependent Pathway in Cerebral Capillary Endothelium

Abstract: The effect of endothelins (ET-1 and ET-3) on ⁸⁶Rb⁺ uptake as a measure of K⁺ uptake was investigated in cultured rat brain capillary endothelium. ET-1 or ET-3 dose-dependently enhanced K⁺ uptake (EC₅₀ = 0.60 ± 0.15 and 21.5 ± 4.1 nM, respectively), which was inhibited by the selective ET_A receptor antagonist BQ 123 (cyclo-d -Trp-d -Asp-Pro-d -Val-Leu). Neither the selective ET_B agonists IRL 1620 [N-succinyl-(Glu⁹,-Ala^11,15)-ET-1] and sarafotoxin S6c, nor the ET_B receptor antagonist IRL 1038 [(Cys¹¹,Cys¹⁵)-ET-1] had any effect on K⁺ uptake. Ouabain (inhibitor of Na⁺,K⁺-ATPase) and bumetanide (inhibitor of Na⁺-K⁺-Cl^? cotransport) reduced (up to 40% and up to 70%, respectively) the ET-1-stimulated K⁺ uptake. Complete inhibition was seen with both agents. Phorbol 12-myristate 13-acetate (PMA), activator of protein kinase C (PKC), stimulated Na⁺,K⁺-ATPase and Na⁺-K⁺-Cl^? cotransport. ET-1-but not PMA-stimulated K⁺ uptake was inhibited by 5-(N-ethyl-N-isopropyl)amiloride (inhibitor of Na⁺/H⁺ exchange system), suggesting a linkage of Na⁺/H⁺ exchange with ET-1-stimulated Na⁺,K⁺-ATPase and Na⁺-K⁺-Cl^? cotransport activity that is not mediated by PKC.     

Chronic taurine supplementation ameliorates oxidative stress and Na+ K+ ATPase impairment in the retina of diabetic rats

Summary.  This study evaluates the effect of 4 months supplementation with 2% and 5% taurine (w/w) on the retina of diabetic rats. In non-diabetic rats, taurine does not modify glycemia, body weight, retinal conjugated dienes (CD), lipid hydroperoxide (LP), and Na⁺K⁺ATPase activity. In diabetic rat, at 2, 4, 8, 16 weeks following the onset of diabetes, retinal CD and LP are significantly and progressively increased, while pump activity is gradually and significantly reduced. In taurine supplemented diabetic rats, glycemia is not affected but lipid peroxidation is significantly decreased. Finally, taurine preserves ATPase activity being 5% more effective than 2% taurine. We conclude that taurine supplementation ameliorates biochemical retinal abnormalities caused by diabetes, thereby suggesting that taurine may have a role in the prevention of retinal changes in diabetes. Received November 26, 2001 Accepted January 10, 2002 Published online October 3, 2002 Authors' address: Prof. Flavia Franconi, Department of Pharmacology, University of Sassari, Via Muroni 23a, I-07100 Sassari, Italy, Fax: 39-79228715, E-mail: franconi@ssmain.uniss.it     

Energy metabolism of reticulocytes: Two different sources of energy for Na+K+-ATPase activity

Total energy production in rabbit reticulocytes amounted to 136·52 ± 6·50μmol ATP h^?1ml^?1 of reticulocytes: 88·3 per cent was provided by oxidative phosphorylation, whereas only 11·7 per cent by aerobic glycolysis. Na⁺K⁺-ATPase accounted for 23 per cent, i.e. 27·65 ± 2·55μmol ATP h^?1ml^?1 of reticulocytes, in the overall energy consumption in reticulocytes of rabbits. Under basal conditions ATP for Na⁺K⁺-ATPase activity was derived exclusively from oxidative phosphorylation. However, when the activity of Na⁺K⁺-ATPase was increased due to the stimulation of adenylate cyclase by (?)-isoprenaline, the additional energy required was provided by aerobic glycolysis. These results indicate that two different compartments, one cytosolic and the other mitochondrial, provide energy for Na⁺K⁺-ATPase activity in reticulocytes.     

Effect of Hypoxia on Na+-K+-Cl− Cotransport in Cultured Brain Capillary Endothelial Cells of the Rat

Abstract: The effect of hypoxia on Na⁺,K⁺-ATPase and Na⁺-K⁺-Cl^? cotransport activity in cultured rat brain capillary endothelial cells (RBECs) was investigated by measuring ⁸⁶Rb⁺ uptake as a tracer for K⁺. RBECs expressed both Na⁺,K⁺-ATPase and Na⁺-K⁺-Cl^? cotransport activity (4.6 and 5.5 nmol/mg of protein/min, respectively). Hypoxia (24 h) decreased cellular ATP content by 43.5% and reduced Na⁺,K⁺-ATPase activity by 38.9%, whereas it significantly increased Na⁺-K⁺-Cl^? cotransport activity by 49.1% in RBECs. To clarify further the mechanism responsible for these observations, the effect of oligomycin-induced ATP depletion on these ion transport systems was examined. Exposure of RBECs to oligomycin led to a time-dependent decrease of cellular ATP content (by ～65%) along with a complete inhibition of Na⁺,K⁺-ATPase and a coordinated increase of Na⁺-K⁺-Cl^? cotransport activity (up to 100% above control values). Oligomycin augmentation of Na⁺-K⁺-Cl^? cotransport activity was not observed in the presence of 2-deoxy-d -glucose (a competitive inhibitor of glucose transport and glycolysis) or in the absence of glucose. These results strongly suggest that under hypoxic conditions when Na⁺,K⁺-ATPase activity is reduced, RBECs have the ability to increase K⁺ uptake through Na⁺-K⁺-Cl^? cotransport.     

Glutathione S‐transferase π complexes with and stimulates Na+,K+‐ATPase

Glutathione S‐transferase (GST) was found to complex with the Na⁺,K⁺‐ATPase as shown by binding assay using quartz crystal microbalance. The complexation was obstructed by the addition of antiserum to the α‐subunit of the Na⁺,K⁺‐ATPase, suggesting the specificity of complexation between GST and the Na⁺,K⁺‐ATPase. Co‐immunoprecipitation experiments, using the anti‐α‐subunit antiserum to precipitate the GST‐Na⁺,K⁺‐ATPase complex and then using antibodies specific to an isoform of GST to identify the co‐precipitated proteins, revealed that GSTπ was complexed with the Na⁺,K⁺‐ATPase. GST stimulated the Na⁺,K⁺‐ATPase activity up to 1.4‐fold. The level of stimulation exhibited a saturable dose–response relationship with the amount of GST added, although the level of stimulation varied depending on the content of GSTπ in the lots of GST received from supplier. The stimulation was also obtained when recombinant GSTπ was used, confirming the results. When GST was treated with reduced glutathione, GST activity was greatly stimulated, whereas the level of stimulation of the Na⁺,K⁺‐ATPase activity was similar to that when untreated GST was added. When GST was treated with H₂O₂, GST activity was greatly diminished while the stimulation of the Na⁺,K⁺‐ATPase activity was preserved. The results suggest that GSTπ complexes with the Na⁺,K⁺‐ATPase and stimulates the latter independent of its GST activity. Copyright © 2012 John Wiley & Sons, Ltd.     

严重烧伤早期红细胞膜胆固醇含量及Na+-K+-ATP酶活性的变化

目的 :研究严重烧伤患者早期红细胞滤过指数 (EFI)与红细胞膜胆固醇含量、Na K ATPase活性的变化。探讨其在严重烧伤早期中的相互关系及意义。方法 :采用核孔滤膜法测定红细胞滤过指数 (EFI) ,用化学修饰电极法测定红细胞膜胆固醇含量 ,应用定磷法测定红细胞膜Na K ATPase活性。结果 :4 7例严重烧伤早期患者EFI较 6 0例正常对照组下降 (P <0 .0 1) ,红细胞膜胆固醇含量、Na K ATPase活性均高于正常对照组 (P <0 .0 1) ,且红细胞膜胆固醇含量、Na K ATPase活性与EFI呈密切负相关 (rcho =- 0 .871,rATPase =- 0 .80 1,P <0 .0 1)。结论 :严重烧伤早期EFI下降 ,变形性明显减低是导致血液粘度和微循环改变的原因之一 ,红细胞膜胆固醇含量和Na K ATPase活性的变化则是引起EFI下降、变形性减低的重要因素     

Different neuronal Na+/K+‐ATPase isoforms are involved in diverse signaling pathways

Inhibition of rat neuronal Na⁺/K⁺‐ATPase α3 isoform at low (100 nM) ouabain concentration led to activation of MAP kinase cascade via PKC and PIP₃ kinase. In contrast to ouabain‐sensitive α3 isoform of Na⁺/K⁺‐ATPase, an ouabain‐resistant α1 isoform (inhibition with 1 mM of ouabain) of Na⁺/K⁺‐ATPase regulates MAP kinase via Src kinase dependent reactions. Using of Annexin V‐FITC apoptotic test to determine the cells with early apoptotic features allows to conclude that α3 isoform stimulates and α1 suppresses apoptotic process in cerebellum neurons. These data are the first demonstration showing participation of ouabain‐resistant (α1) and ouabain‐sensitive (α3) Na⁺/K⁺‐ATPase isoforms in diverse signaling pathways in neuronal cells. Copyright © 2010 John Wiley & Sons, Ltd.     

Temperature-specific inhibition of human red cell Na+/K+ ATPase by 2,450-MHz microwave radiation

The ATPase activity in human red blood cell membranes was investigated in vitro as a function of temperature and exposure to 2,450-MHz continuous wave microwave radiation to confirm and extend a report of Na+ transport inhibition under certain conditions of temperature and exposure. Assays were conducted spectrophotometrically during microwave exposure with a custom-made spectrophotometer-waveguide apparatus. Temperature profiles of total ATPase and Ca+2 ATPase (ouabain-inhibited) activity between 17 and 31 degrees C were graphed as an Arrhenius plot. Each data set was fitted to two straight lines which intersect between 23 and 24 degrees C. The difference between the total and Ca+2 ATPase activities, which represented the Na+/K+ ATPase activity, was also plotted and treated similarly to yield an intersection near 25 degrees C. Exposure of membrane suspensions to electromagnetic radiation, at a dose rate of 6 W/kg and at five temperatures between 23 and 27 degrees C, resulted in an activity change only for the Na+/K+ ATPase at 25 degrees C. The activity decreased by approximately 35% compared to sham-irradiated samples. A possible explanation for the unusual temperature/microwave interaction is proposed.     

Cardiac sarcolemmal Na+-Ca2+ exchange and Na+-K+ ATPase activities and gene expression in alloxan-induced diabetes in rats

To determine the sequence of alterations in cardiac sarcolemmal (SL) Na⁺-Ca²⁺ exchange, Na⁺-K⁺ ATPase and Ca²⁺-transport activities during the development of diabetes, rats were made diabetic by an intravenous injection of 65 mg/kg alloxan. SL membranes were prepared from control and experimental hearts 1-12 weeks after induction of diabetes. A separate group of 4 week diabetic animals were injected with insulin (3 U/day) for an additional 4 weeks. Both Na⁺-K⁺ ATPase and Ca²⁺-stimulated ATPase activities were depressed as early as 10 days after alloxan administration; Mg²⁺ ATPase activity was not depressed throughout the experimental periods. Both Na⁺-Ca²⁺ exchange and ATP-dependent Ca²⁺-uptake activities were depressed in diabetic hearts 2 weeks after diabetes induction. These defects in SL Na⁺-K⁺ ATPase and Ca-transport activities were normalized upon treatment of diabetic animals with insulin. Northern blot analysis was employed to compare the relative mRNA abundances of _--subunit of Na⁺-K⁺ ATPase and Na⁺-Ca²⁺ exchanger in diabetic ventricular tissue vs. control samples. At 6 weeks after alloxan administration, a significant depression of the Na⁺-K⁺ ATPase _-- subunit mRNA was noted in diabetic heart. A significant increase in the Na⁺-Ca²⁺ exchanger mRNA abundance was observed at 3 weeks which returned to control by 5 weeks. The results from the alloxan-rat model of diabetes support the view that SL membrane abnormalities in Na⁺-K⁺ ATPase, Na⁺Ca²⁺ exchange and Ca²⁺-pump activities may lead to the occurrence of intracellular Ca²⁺ overload during the development of diabetic cardiomyopathy but these defects may not be the consequence of depressed expression of genes specific for those SL proteins.     

Oligomycin inhibition of Na,K,ATPase

It is shown that the incomplete, uncompetitive inhibition pattern exhibited by oligomycin toward Na,K,ATPase cannot be explained by a single-cycle enzyme model. In contrast, the experimental data are easily explained in terms of a dimeric enzyme, only one subunit of which can bind oligomycin at a time, and that subunit is then rendered inactive. In a brief analysis of the model thus obtained by way of numerical examples it is shown that it may show activation at small concentrations of moderator, which disappears at higher concentrations, a property observed for the hydrolysis ofp-nitro-phenylphosphate, which is also catalyzed by Na,K,ATPase.     

久效磷对美国红鱼鳃Na^＋／K^＋-ATP酶活性和超显微结构的影响

经不同试验浓度的久效磷(0．25、0．5、1．0和2．0mg·L^-1)处理美国红鱼4d后,分别对鱼鳃Na^+／K^+—ATP酶活性和氯细胞密度进行了测定和计数,并观察了鱼鳃组织显微结构和超微结构的变化。结果表明,低浓度久效磷(0．25mg·L^-1)处理可以诱导鱼鳃氯细胞大量增生,Na^+／K^+—ATP酶活性增强,随着试验浓度的增加,久效磷对鳃组织的损伤越来越重,Na^+／K^+—ATP酶活性逐渐降低;久效磷对鱼鳃显微结构的损伤表现为鳃小片上皮细胞水肿、脱离。鳃小片基部粘连。鳃小片上皮细胞角质化;超微结构变化主要为内质网、线粒体、微小管和核膜的水肿及部分溶解,这种损伤表现为由细胞膜到细胞核的动态过程。     

Lack of inhibition of vasopressin-stimulated NA+K+ATPase by atrial natriuretic factor in the rat renal medullary thick ascending limb of Henle's loop

The anti-diuretic hormone, arginine vasopressin (AVP) stimulates the activity of Na+K+ATPase in the rat renal medullary thick ascending limb of Henle's loop (mTAL). Atrial natriuretic factor (ANF) has been suggested to exert a tubular effect on the mammalian nephron, perhaps in part, by interacting with other hormones. In the present study, we investigated the effect of rat ANF with and without AVP upon mTAL Na+K+ATPase activity using cytochemical methods. ANF alone failed to inhibit or stimulate Na+K+ATPase activity in mTAL at any of the concentrations tested (10 nmol-0.1 pmol l-1). Unlike the rat hypothalamic digitalis-like factor, ANF (10 nmol-10 fmol l-1) did not inhibit Na+K+ATPase activity after stimulation with AVP (1 fmol l-1) for either 4 or 10 min. The results suggest that ANF does not exert an effect on mTAL, either alone or in conjunction with AVP.     

Presence of a Na+-stimulated P-type ATPase in the plasma membrane of the alkaliphilic halotolerant cyanobacterium Aphanothece halophytica

Aphanothece cells could take up Na(+) and this uptake was strongly inhibited by the protonophore, carbonyl cyanide m-chlorophenylhydrazone (CCCP). Cells preloaded with Na(+) exhibited Na(+) extrusion ability upon energizing with glucose. Na(+) was also taken up by the plasma membranes supplied with ATP and the uptake was abolished by gramicidin D, monensin or Na(+)-ionophore. Orthovanadate and CCCP strongly inhibited Na(+) uptake, whereas N, N'-dicyclohexylcarbodiimide (DCCD) slightly inhibited the uptake. Plasma membranes could hydrolyse ATP in the presence of Na(+) but not with K(+), Ca(2+) and Li(+). The K(m) values for ATP and Na(+) were 1.66+/-0.12 and 25.0+/-1.8 mM, respectively, whereas the V(max) value was 0.66+/-0.05 mumol min(-1) mg(-1). Mg(2+) was required for ATPase activity whose optimal pH was 7.5. The ATPase was insensitive to N-ethylmaleimide, nitrate, thiocyanate, azide and ouabain, but was substantially inhibited by orthovanadate and DCCD. Amiloride, a Na(+)/H(+) antiporter inhibitor, and CCCP showed little or no effect. Gramicidin D and monensin stimulated ATPase activity. All these results suggest the existence of a P-type Na(+)-stimulated ATPase in Aphanothece halophytica. Plasma membranes from cells grown under salt stress condition showed higher ATPase activity than those from cells grown under nonstress condition.     

心钠素前体分子内调控对心肌Na^＋—K^＋—ATP酶的作用

目的：研究利钾尿肽及心钠素前体分子内调控作用对心肌Ｎａ＋Ｋ＋ＡＴＰ酶的作用。方法：将大鼠心肌匀浆后,分别加入利钾尿肽、心钠素以及利钾尿肽＋心钠素,用比色法测定Ｎａ＋Ｋ＋ＡＴＰ酶活性。将大鼠心脏悬挂于Ｌａｎｇｅｎｄｏｒｆ灌流装置,分别以利钾尿肽、心钠素、利钾尿肽＋心钠素为灌流液,灌注心脏,用四道生理仪观测左心室内压、左心室收缩最大速率,左心室舒张最大速率,心率及冠脉流量。结果：心钠素虽然对Ｎａ＋Ｋ＋ＡＴＰ酶有抑制作用（抑制率２６．２％）,但是,与对照无显著性差异（Ｐ＞０．０５）。利钾尿肽显著抑制酶的活性（抑制率４６．５％,Ｐ＜０．０１）这种抑制作用可被心钠素抵消（抑制率１７．６％,Ｐ＞０．０５）。利钾尿肽可以增加左心室收缩和舒张最大速率以及左室内压,而这种强心作用可因心钠素的加入而消失或减弱。结论：利钾尿肽可以抑制心肌Ｎａ＋Ｋ＋ＡＴＰ酶的活性,产生强心作用,心钠素可以抵消以上作用。     

Enhanced (Na+K)-ATPase Activity and Expression in Mouse Brain after Chronic Ethanol Administration

It is the general hypothesis that the primary mode of action of ethanol is the alteration of membrane structure and function including the conformation of receptors and ion channels essential for neurotransmission and signal transduction. However, the issue of whether ethanol affects (Na+K)-ATPase under physiological conditions remains unsettled. In this study, adult mice were treated with a daily dose of 5 g/kg of ethanol for 28 days. The RNA was isolated from brain and the (Na+K)-ATPase mRNA level was determined using Northern blot analysis. We have found an increased expression of (Na+K)-ATPase -subunit in the chronically treated alcohol group as compared with that of controls. This result was further substantiated by increased protein phosphorylation as well as increased specific activity of this enzyme in the synaptosomal plasma membrane after chronic ethanol administration. Thus we have demonstrated that ethanol may directly affect (Na+K)-ATPase in vivo, leading to the increased synthesis of this enzyme through adaptive mechanisms.     

Na+ and K+ transport in excised soybean roots

Uptake, accumulation and xylem transport of K⁺ and Na⁺ in excised roots of soybean were investigated by use of a perfusion technique. This technique permitted independent quantification of, on the one hand, entry of ions into the roots and their transport through the cortex to the xylem vessels, and on the other hand reabsorption from the xylem vessels to the neighbouring cells and the external medium. Data are consistent with a low degree of selective uptake of K⁺ over Na⁺. However, Na⁺ depletion of the xylem stream by reabsorption limits, although weakly, its translocation to the shoots. Na⁺ reabsorbed is for a great part reexcreted into the external medium. The low efficiency of these processes is discussed in relation to the Na⁺ sensitivity of soybean.