Biomedical subjects
Publications and source records attributed to N I Pokudin.
Na+/H+ exchange rate in erythrocytes of patients with atherosclerosis of lower limb arteries is less than in erythrocytes of healthy subjects. Ultraviolet exposure of the patients blood in vitro activates Na/H exchange in erythrocytes.
The rate of proton gradient-induced Na+/H+ exchange in the erythrocytes of SHR was increased by 50-60% as compared to WKY animals. No significant correlation between Na+/H+ exchange and blood pressure was revealed in F2 hybrids of SHR and WKY rats. Na+/H+ exchange rate in the erythrocytes of F2 SHR x WKY hybrids was twice as high as in SHR and three times higher than in WKY rats.
The activity of Na+, K(+)-ATPase in SHR erythrocytes treated with saponin is increased by 30-40% as compared to the Brown Norway (BN.lx) strain whereas the activity of Ca(2+)-ATPase is decreased by 20-30%. Passive permeability of SHR erythrocytes determined by 86Rb influx is increased by 20-30%. In the presence of orthovanadate erythrocytes of SHR accumulate 45Ca by 80% more than BN.lx red cells. There was no difference in Na+/H+ exchange between erythrocytes of SHR and BN.lx animals.
The activity of transport adenosine triphosphatases (ATPases) in saponin-treated erythrocytes as well as the passive membrane permeability for 86Rb+ (K+), 45Ca2+ uptake (in the presence of orthovanadate) and the rate of Na(+)-H+ exchange in intact erythrocytes were studied in spontaneously hypertensive rats (SHR), Wistar-Kyoto (WKY) and Brown-Norway (BN.lx) rats. Higher Na+,K(+)-ATPase activity, lower Ca(2+)-ATPase activity, increased passive K+ permeability and greater 45Ca2+ uptake were observed in erythrocytes from SHR compared with BN.lx rats. Similar differences in the last two parameters were also disclosed by a comparison of SHR and WKY rats. The rate of Na(+)-H+ exchange in SHR erythrocytes was greater than in WKY rats but equal to that of BN.lx rats. A genetic analysis did not reveal a significant correlation between Na(+)-H+ exchange rate and blood pressure in F2 SHR x WKY hybrids.
The rates of 86Rb influx into human and rat erythrocytes were studied in media of various tonicity. At sucrose concentrations below 0.3 mol/l, the ouabain-insensitive, furosemide-inhibited component of influx increased in rat but not in human erythrocytes; this may be explained by a rise in the rate of Na+, K+, Cl-- and/or K+, Cl-cotransport. An increase in osmolarity resulted in a reduction of this as well as of the ouabain and furosemide-insensitive component in rat erythrocytes. At the same conditions a drastic inhibition of Na+, K(+)-pump occurred both in rat and human erythrocytes. We failed to observe a lag-phase in the activation of the cotransport in rat erythrocytes; i. e. the process of activation parallels the shrinkage of cells. In rat erythrocyte ghosts, the shrinkage-induced stimulation of the cotransport was lost, and the direction of their osmotic reaction (inhibition of transport pathways) was similar to that in human erythrocyte ghosts. It is suggested that the mechanism of volume regulation of ion transport in intact cells involves a step of physical amplification via a change in interactions between the protein carcass and the lipid bilayer.
The relation of red blood cell Na+/Li+ countertransport rate to the prevalence of arterial hypertension (AH) and blood pressure was evaluated in 720 individuals aged 23-60 years. In all age groups, the rate of red blood cell Na+/Li+ countertransport was significantly higher in males than in females. Age, alcohol use, obesity in males and age, obesity, and Na+/Li+ countertransport rate in females are factors that predispose to AH, as evidenced by multivariate logistic analysis. The latter parameter is associated with obesity and diastolic blood pressure and unassociated with age, alcohol use, and smoke, as suggested by regression analysis. The distribution of Na+/Li+ countertransport rate values in the population after standardization for ruling out the impact of obesity is erroneous and shifted to the right. This allows it to be presented as a sum of two normal distributions. However, their analysis has failed to reveal statistically significant differences in blood pressure values of AH incidence. The rate of Na+/Li+ countertransport may be regarded as a biological population risk factor for AH, which is, however, less than such factors as age, obesity, and excessive alcohol use.
The present work is concerned with the role of the velocity of Na+/Li(+)-countertransport of red blood cells (as an indicator of the function of cellular membranes) in the development of arterial hypertension under conditions of a populational study. A relationship is demonstrated between the velocity of Na+/Li(+)-countertransport and the sex, arterial pressure, body mass index, and blood serum triglycerides. The high velocity of Na+/Li(+)-countertransport is encountered in 25% of the population, being responsible for about 30% of all cases of arterial hypertension. Taking into consideration the modern multifactorial theory of the genesis of arterial hypertension, the high velocity of Na+/Li(+)-countertransport should be viewed as a definite disease pattern (one of several) and as a risk factor of arterial hypertension in women (the data of the prospective part of the study).
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The intracellular levels of sodium, potassium, and free calcium, as well as red blood cell transport of 86Rb (a radioactive potassium analogue) and 45Ca were measured in patients with bronchial asthma (BA). The intracellular content of sodium was 10-20% higher in patients with BA, though the differences in the present sample (7 controls and 18 BA patients) failed to be significant (p less than 0.05). The differences were most pronounced in infection-dependent BA and not eliminated during glucocorticoid hormone therapy. The activity of the Na+, K+ pump, as assessed by the values of an ouabain-inhibited component of 86Rb entry, was increased by 10-20% (p less than 0.05) in BA patients and of Na+, K+ cotransport (an ouabain-sensitive furosemide-inhibited component of 86Rb entry), by 35-45% (p less than 0.25). The passive red blood cell membrane permeability in BA patients (an ouabain + furosemide-insensitive component of 86Rb) remained unchanged. The quin-2-loaded red blood cell entry rate for 45Ca was decreased by 5-10%. In the absence of a Ca(2+)-ATPase inhibitor (0.5 mM orthovanadate), these differences were greatly significant (p less than 0.005). In addition, the patients with BA showed a 10% reduction in free calcium levels; however, the differences were insignificant (p less than 0.25), which might be caused by Ca-pump activation. As in the experiments with Na+ level measurements, the greatest differences in red blood cell Ca(2+)-balance control were recorded in patients with infection-dependent BA and independent of glucocorticoid hormone therapy. Possible mechanisms of involvement of the changes found in monovalent cation and Ca2+ transport in the pathogenesis of BA were also discussed.
A 3-year prospective study was undertaken to evaluate the relation of red blood cell Na+/Li+ countertransport rates to the incidence of arterial hypertension. A total of 227 subjects (males and females aged 24-54 years)--a 20% representative sample of organized population--was followed up. The annual incidence of arterial hypertension was 4.3 among males and 2.7 among females. The baseline blood pressures and obesity in males and rates of Na+/Li+ countertransport in females were demonstrated to be factors associated with the incidence of arterial hypertension. The rate of red blood cell Na+/Li+ countertransport can be regarded as an independent biological risk factor for arterial hypertension.
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The rate of delta mu H+ --induced erythrocyte Na+/H+ exchange is increased in both patients with essential hypertension (EH) and spontaneously hypertensive rats (SHR). The increase of Na+,K(+)-cotransport was revealed in erythrocytes of SHR only. This alteration as well as a decrease of mean cell volume were observed in both young and old erythrocytes of SHR. The moderate shrinkage of rat (but not human) erythrocytes results in an increase of the rate of Na+,K(+)-cotransport. The more pronounced shrinkage of rat (but not human) erythrocytes induces the Na+/H+ exchange. These reactions are accompanied by phosphoinositide response. Activator of protein kinase C (TPA) increases delta mu H+ --induced Na+/H+ exchange both in human and rat erythrocytes but it does not modify phosphoinositide metabolism. No differences were observed in the rate of Na+/H+ exchange between TPA-treated erythrocytes of SHR and WKY. We assume that the activation of protein kinase C increases Na+/H+ exchange in primary hypertension. Increased Na+/H(+)-cotransport revealed in an experimental model of primary hypertension is probably due to the decrease of erythrocyte volume.
Protein kinase C activity in the lysate of erythrocytes of patients with essential hypertension (EH) and spontaneously hypertensive rats (SHR) was found to be increased by 1.6-2.0 times as compared with normotensive controls. Membrane cytoskeleton alterations observed in the erythrocytes of patients with EH and SHR were revealed in decreased average erythrocyte volume, increase of cup-shaped cell formation, and increase of basal phosphorylation of band 4.9 protein. In addition, the rate of Na(+)-H+ exchange in erythrocytes of EH patients and SHR was increased by 1.9-fold. In vitro treatment of erythrocytes of healthy donors and Wistar-Kyoto rats (WKY) with protein kinase C activator (12-O-tetradecanoylphorbol-13-acetate) leads to similar changes of cell shape, cell volume, band 4.9 protein phosphorylation and Na(+)-H+ exchange, as well as to an increase of diS-C3-(5) fluorescence. It may be assumed that alterations of these parameters revealed in primary hypertension are caused by increased activity of protein kinase C.
The heat sorption curves for human and rat erythrocyte membranes in the temperature region from 40 degrees C to 90 degrees C have been compared resulting from thermal gel analysis of these membranes. The main heat sorption peaks are located within the temperature regions from 49 degrees C to 52 degrees C (A transition), from 56 degrees C to 59 degrees C (B transition), from 62 degrees C to 65 degrees C (C transition) and from 74 degrees C to 82 degrees C (D transition). Thermoinactivation temperatures for most of the membrane proteins in human and rat erythrocytes are rather different, which suggests essential differences in the arrangement of their membrane frameworks. Addition of the protein kinase C activator TPA induces a fast increase in light scattering of human and rat erythrocyte suspension which is connected with some changes of cell shape. This reaction is completely blocked by a minimal thermal treatment of the membrane framework proteins (preincubation at the temperature of the A transition). Such treatment inhibits also the increase of Na+, K(+)-cotransport in rat erythrocytes induced by hyperosmotic shrinkage. It is assumed that the proteins of the membrane framework take part in the volume-dependent regulation of the ion-transport systems of plasma membranes.
Osmotic swelling of human and rat erythrocytes does not induce regulatory volume decrease. Regulatory volume increase was observed in shrunken erythrocytes of rats only. This reaction was blocked by the inhibitors of Na+/H+ exchange. Cytoplasmic acidification in erythrocytes of both species increases the amiloride-inhibited component of 22Na influx by five- to eight-fold. Both the osmotic and isosmotic shrinkage of rat erythrocytes results in the 10- to 30-fold increase of amiloride-inhibited 22Na influx and a two-fold increase of furosemide-inhibited 86Rb influx. We failed to indicate any significant changes of these ion transport systems in shrunken human erythrocytes. The shrinking of quin 2-loaded human and rat erythrocytes results in the two- to threefold increase of the rate of 45Ca influx, which is completely blocked by amiloride. The dependence of volume-induced 22Na influx in rat erythrocytes and 45Ca influx in human erythrocytes on amiloride concentration does not differ. The rate of 45Ca influx in resealed ghosts was reduced by one order of magnitude when intravesicular potassium and sodium were replaced by choline. It is assumed that the erythrocyte shrinkage increases the rate of a nonselective Cao2+/(Nai+, Ki+) exchange. Erythrocyte shrinking does not induce significant phosphorylation of membrane protein but increases the 32P incorporation in diphosphoinositides. The effect of shrinkage on the 32P labeling of phosphoinositides is diminished after addition of amiloride. It is assumed that volume-induced phosphoinositide response plays an essential role in the mechanism of the activation of transmembrane ion movements.
The activity of ion-transport systems and Ca2+-induced erythrocyte haemolysis were compared between patients with essential hypertension and two strains of spontaneously hypertensive rats. Previous data on the increased rate of Na+-Li+ countertransport in erythrocytes of essential hypertensives were confirmed in this study. However, identification of Na+-Li+ countertransport in rat erythrocytes remained a complicated person because of the high rate of sodium-independent efflux of Li+. The rate of Na+-H+ exchange increased by 50-80% both in spontaneously hypertensive Wistar-Kyoto rats (SHR) and in patients with essential hypertension. No difference between Milan hypertensive strain rats (MHS) and Milan normotensive strain rats (MNS) was found. The rate of Na+,K+ cotransport increased in SHR and MHS erythrocytes compared with rats of the control strains [normotensive Wistar-Kyoto rats (WKY) and MNS; 30-50 and 90-110%, respectively]. No difference in this parameter was found between patients with essential hypertension and healthy subjects. Erythrocytes of patients with essential hypertension and of SHR were characterized by a higher sensitivity of their K+ channels to the increased concentration of intracellular Ca2+. This parameter did not change in MHS erythrocytes. Ca2+-induced haemolysis increased four- to fivefold in MHS erythrocytes compared with MNS and did not change in erythrocytes of SHR and patients with essential hypertension. The conclusion from these data is that the SHR strain is a more adequate model of human essential hypertension than the MHS.
The contribution of Ca2+ channels and Na+/Ca2+ exchange to Ca2+ uptake in rat brain synaptosomes upon long- (t greater than or equal to 30 s) and short-term (t less than 30 s) depolarization by high K+ was studied by measuring the 45Ca content and free Ca2+ concentration (from Quin-2 fluorescence). At 37 degrees C, the system responsible for the K+-stimulated uptake of 45Ca (t greater than or equal to 30 s) and the Na+/Ca+ exchanger are characterized by a similar concentration dependence of external Ca2+ (Ca0(2+] and K0+ as well as by an equal sensitivity to verapamil (Ki = approximately 20-40 microM) and La2+ (Ki = approximately 50 microM). These data and the results from predepolarization suggest that the 45Ca entry into synaptosomes at t greater than or equal to 30 s is due to the activation of Na+/Ca+ exchange caused by its electrogenic component, while the insignificant contribution of Ca2+ channels can be accounted for by their inactivation. At low temperatures (2-4 degrees C) which decelerate the inactivation, the initial phase of 45Ca uptake is fully provided for by Ca2+ channels, showing a lower (as compared to the exchanger) affinity for Ca0(2+) (K0.5 greater than 1 mM)m a greater sensitivity to La3+ (Ki = approximately 0.2-0.3 microM) and verapamil (Ki = approximately 2-3 microM); these channels are fully inactivated by predepolarization with K0+, ouabain and batrachotoxin. The Ca2+ channels can be related to T-type channels, since they are not blocked by nicardipine and niphedipine.
The erythrocytes from two spontaneously hypertensive rat strains (SHR and MHS) differ from those of control strains such as WKY and MNS in showing lower values of the mean volume and higher Na,K-cotransport rates. After 4-hour incubation in the presence of orthovanadate 45Ca levels in the erythrocytes from SHR was twice as high as in those from WKY. Under the same conditions, hemoglobin crystallization occurred, this process in SHR erythrocytes took place more intensively. No difference was found in these parameters between MNS and MHS. The fractions enriched with young and old cells remained to display variations in the mean volume of cells, Na,K-cotransport rates, 45Ca levels and hemoglobin crystallization, which were recorded between SHR and WKY. The WKY-SHR hybrids of second generation indicated that only Na,K-cotransport rates of the parameters examined positively correlated with blood pressures. From the findings it was concluded that the differences in the volume of erythrocytes and their permeability for cations were not due to the changed time course of cell aging in the blood bed, but to the specific characteristics of cytoskeleton protein arrangement, which were primarily typical of a rat reticulocytic membrane.