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Biomedical subjects

M Hanano

Publications and source records attributed to M Hanano.

At least 109 records · Page 6Linked to original sources
PubMed1988

Transport of digoxin into brain microvessels and choroid plexuses isolated from guinea pig.

To characterize the efflux system of digoxin, a cardiac glycoside, from the brain to the blood through the blood-brain barrier and blood-cerebrospinal fluid (CSF) barrier, the accumulation of digoxin by the brain microvessel or the choroid plexus isolated from guinea pig brain was investigated. The accumulation of digoxin by the brain microvessel has a saturable component (Km = 0.163 microM, Vmax = 0.142 nmol/mL of tissue/min), with a nonsaturable component [Kd = 0.203 cell-to-medium (C:M) ratio/min] that was decreased by hypothermia (Q10 = 2.9), sulfhydryl reagent, and quinidine, but not by a metabolic inhibitor [2,4-dinitrophenol (DNP)]. It was concentration- and Na+-dependent. The accumulation of digoxin by the choroid plexus was also saturable (Km = 1.9 microM, Vmax = 3.8 nmol/mL of tissue/min), and was decreased by hypothermia (Q10 = 4.4), sulfhydryl reagents, ouabain, and quinidine, but not by metabolic inhibitors (DNP, KCN); it was also concentration- and Na+-dependent. The binding of digoxin to the homogenate of choroid plexus was one-tenth of digoxin accumulation by the intact choroid plexus, suggesting that digoxin is transported into the cells and bound to the cytosol fraction. The value of (Vmax/Km + Kd) multiplied by the total tissue weight of the microvessel per guinea pig is approximately 10-fold that of Vmax/Km multiplied by the tissue weight of the choroid plexus, although (Vmax/Km + Kd) per milliliter of the microvessel is half the Vmax/Km value of the choroid plexus. These findings suggest that digoxin can be excreted from both the brain and the cerebrospinal fluid to blood by a carrier-mediated diffusion system which is inhibited by quinidine, and that a main route of digoxin efflux from the brain to the blood is not through the blood-CSF barrier, but through the blood-brain barrier.

Animals↗
PubMed1988

Effects of phenothiazine derivatives on the microclimate-pH in the rat jejunum.

The effects of several phenothiazine derivatives (PTDs) and quinidine (QD) on the jejunal microclimate-pH in rats were studied using a microelectrode. Chlorpromazine, thioridazine, chlorpromazine sulfoxide (CPZSO), trifluoperazine, prochlorperazine, and QD at concentrations of 1 mM increased this microclimate-pH by 0.15-0.3 pH units, while 1 mM diethazine and 1 mM promethazine had little effect on it. The increases in the microclimate-pH caused by PTDs and QD were concentration dependent and reversible. We studied the effects of PTDs on the fluidity of intestinal brush border membranes and on the release of proteins from the intestinal tissue to the lumen. The PTD-induced changes in microclimate-pH could not be explained by either of these nonspecific effects on the membranes. Then, the effects of PTDs on Na+,K+-ATPase activity and Mg2+-ATPase activity were studied using the jejunal homogenate. Each PTD inhibited Na+,K+-ATPase and Mg2+-ATPase activity to some extent. The inhibitory effects on Na+,K+-ATPase and Mg2+-ATPase activity were compared with the PTD-induced increases in the microclimate-pH. No good correlation was obtained between the IC50 values of PTDs for Na+,K+-ATPase activity and the concentrations required to increase the microclimate-pH by 0.1 pH unit, while IC50 values of PTDs for Mg2+-ATPase activity showed a relatively good correlation, except for that of CPZSO. These findings suggest that the effects of PTDs on the microclimate-pH were not nonspecific, although the increases in the microclimate-pH caused by PTDs cannot be fully explained by the inhibitory effects of these compounds on either Na+,K+-ATPase activity or Mg2+-ATPase activity alone.

Adenosine Triphosphatases↗
PubMed1988

Membrane-limited hepatic transport of the conjugative metabolites of 4-methylumbelliferone in rats.

The hepatic transports of 4-methylumbelliferone (4-MU) and its conjugative metabolites, the glucuronide (4-MUG) and sulfate (4-MUS), were investigated in rats with various methods. The extraction ratio (E) was estimated with the multiple indicator dilution (MID) method using isolated perfused rat liver. The values of E for 4-MUG and 4-MUS were much lower (less than 0.2) than that for the parent compound, 4-MU (0.89). In addition, we examined the simulation of the outflow curves of conjugates based on the "distributed" model in which we varied the permeability between the blood and hepatocytes. When the permeability was much smaller relative to the hepatic blood flow, the simulated curve was superimposed on the dilution curve. These results suggest that the influx permeabilities of these conjugates are so low that little extraction occurs during the passage through the liver. Measuring the unidirectional uptake of these conjugates into the liver with the in vitro centrifugal filtration method using isolated hepatocytes, we determined the influx permeabilities (PSinf(total] for the total ligands. The value of PSinf(total) determined with the in vitro method was extrapolated to that per gram of liver, assuming 1 g of liver has 1.3 X 10(8) cells. The values of PSinf(total) for 4-MU, 4-MUG, and 4-MUS were 4.8, 0.06, and 0.11 mL/min/g liver, respectively. Thus, the influx permeabilities for 4-MUG and 4-MUS were much smaller than the hepatic blood flow (1.6 mL/min/g liver), confirming the results of MID method.

Animals↗
PubMed1988

Kinetic analysis of albumin-mediated uptake of warfarin by perfused rat liver.

We previously found that the uptake of warfarin in the presence of albumin by perfused rat liver could not be explained simply by the unbound warfarin concentration. The aim of the present study is to develop a kinetic model to account for this albumin-mediated uptake of warfarin. Single circulation indicator dilution studies on warfarin uptake were carried out in the isolated perfused rat liver in the absence and presence of various concentrations of bovine serum albumin (BSA) in the perfusate. A distributed model was fitted to the dilution data and the estimates of the influx, efflux, and sequestration rate constants were obtained. The results showed that the predicted concentration of the unbound warfarin is not high enough to explain the observed uptake rate; the liver cell surface appears to reduce the binding affinity of warfarin for BSA to 1/20 of that observed in vitro. A kinetic model which considers the interaction between albumin and the liver cell surface was fitted to the uptake rates of warfarin over a wide range of BSA concentration. The model gave a dissociation constant of the cell surface for albumin of 160 microM, which is comparable with those reported by others for the hepatic extractions of free fatty acids and rose bengal. Based on this kinetic model, the contributions of the unbound and bound warfarin to its hepatic uptake were estimated, and the bound warfarin was found to contribute most in the physiological albumin concentration range.

Animals↗
PubMed1988

Hepatic clearances of antipyrine, indocyanine green, and galactose in normal subjects and in patients with chronic liver diseases.

Blood clearance of antipyrine, indocyanine green, and galactose were measured to evaluate the alterations of effective hepatic blood flow and hepatic intrinsic clearances in chronic liver diseases. Galactose blood clearance, which may be taken as effective hepatic blood flow, decreased by approximately 30% in patients with cirrhosis (12.49 +/- 0.76 ml/min/kg; mean +/- SE; n = 17) compared with normal subjects (18.17 +/- 1.03 ml/min/kg; n = 5). In patients with cirrhosis, intrinsic clearances of antipyrine (0.178 +/- 0.014 ml/min/kg; n = 17) and indocyanine green (6.19 +/- 1.38 ml/min/kg; n = 7) showed 61% and 85% reduction, respectively, compared with those of normal subjects (0.462 +/- 0.048 ml/min/kg; n = 5; 41.72 +/- 7.75 ml/min/kg; n = 5). Considering that indocyanine green and antipyrine are eliminated by different hepatic mechanism, these mechanisms may not be equally sensitive to decrements in hepatic function. In addition, fractional reductions of intrinsic clearances for these compounds are thus much greater than that of effective hepatic blood flow.

Adult↗
PubMed1988

Dynamic determination of kinetic parameters for the interaction between polypeptide hormones and cell-surface receptors in the perfused rat liver by the multiple-indicator dilution method.

Hepatic elimination of epidermal growth factor (EGF) via receptor-mediated endocytosis was studied by a multiple-indicator dilution method in the isolated perfused rat liver, in which cell polarity and spatial organization are maintained. In this method EGF was given with inulin, an extracellular reference, as a bolus into the portal vein, and dilution curves of both compounds in the hepatic vein effluent were analyzed. Analysis of the dilution curve for EGF, compared with that for somatostatin, which showed no specific binding to isolated liver plasma membranes, resulted as follows: (i) both extraction ratio and distribution volume of 125I-labeled EGF decreased as the injected amount of unlabeled EGF increased; (ii) the ratio plot [ln (inulin/EGF) versus time] of the dilution curve for EGF exhibited an upward straight line initially for a short period of time (approximately equal to 10 sec), whereas the ratio plot [ln (inulin/somatostatin) versus time] of somatostatin gradually decreased. The multiple-indicator dilution method was used for other peptides also. Insulin and glucagon, known to have hepatocyte receptors, behaved similarly to EGF in shape of their ratio plots. Thus, analysis of dilution curves can reveal whether or not the cell surface has receptors for certain peptides. In addition, the dilution curves for EGF at various doses (tracer approximately equal to 30 micrograms) were analyzed simultaneously based on a kinetic model incorporating the perfusion rate, the association rate constant of EGF to surface receptors (kappa on), the dissociation rate constant of EGF from the EGF-receptor complex (kappa off), and the sequestration rate constant of the complex. The kinetic parameters [the dissociation constant (Kd = kappa off/kappa on) and the number of surface receptors] calculated by this analysis were comparable with reported values obtained by in vitro direct binding measurements at equilibrium using liver homogenates. We conclude that the multiple-indicator dilution method is a good tool for analyzing the dynamics of peptide hormones--cell-surface receptor interaction under a condition in which spatial architecture of the liver is maintained.

Algorithms↗
PubMed1988

Comparison of specific binding of human epidermal growth factor (EGF) to sinusoidal and bile canalicular membranes isolated from rat liver.

The binding characteristics of human epidermal growth factor (EGF) were compared between highly purified canalicular (CMV) and sinusoidal (basolateral) rat liver plasma membrane (SMV) preparations. The dissociation constants (2-3 nM) for these membranes were comparable, while the binding capacity for CMV was approximately half that for SMV. The binding capacity for CMV was too high to be accounted for only by the contamination with sinusoidal membranes, since the measurements of specific activities of various enzymes (Na+,K+-ATPase, alkaline phosphatase, and leucine aminopeptidase) indicated that the extents of the cross contamination with other membrane fractions were at most 10%. Although the physiological function of specific binding of EGF to bile canalicular membrane domain remains to be determined, it may have a role in biliary excretion of EGF. The specific binding of EGF to bile canalicular membranes from rat liver was identified for the first time.

Animals↗
PubMed1988

Kinetic analysis of phenytoin disposition in rats with experimental renal and hepatic diseases.

The pharmacokinetics of phenytoin (DPH) was investigated in rats with uranyl nitrate induced renal failure, and with D-galactosamine induced hepatic failure. The serum disappearance of DPH after 10 mg/kg i.v. dose followed a two-exponential decline in normal and both types of intoxicated rats. The serum disappearance half-life (t1/2) and the volume of distribution (Vd) significantly increased in both types of intoxicated rats, while the total blood clearances (CLb) significantly decreased. The serum unbound fraction (fu) of DPH significantly increased in both types of intoxicated rats. The blood to plasma concentration ratio (RB) of DPH significantly increased in the uranyl nitrate-treated rats, while that of the D-galactosamine-treated rats did not show significant alteration. The tissue to serum concentration ratios (Kp) of most of tissues studied after i.v. bolus injection of DPH increased in both types of intoxicated rats. Except for the lung of the D-galactosamine-treated rats, the tissue to serum unbound concentration ratio (Kpu) of other tissues did not show a significant alteration. This suggested that the tissue uptake and/or binding of DPH is not affected by uranyl nitrate or D-galactosamine intoxication and that the increases in Vd and Kp are due mainly to the decrease in serum protein binding. The hepatic intrinsic clearance of unbound DPH (CLuint,H) also decreased in both types of intoxicated rats. Thus, the uranyl nitrate and D-galactosamine intoxication caused the increase in fu and the decrease in CLuint,H and these results may explain the significant decrease in CLb and increases in Vd and t1/2. The tissue concentration-time courses of DPH were predicted by a physiciologically based pharmacokinetic model and good agreement between the predicted and observed values in normal and in both types of intoxicated rats were obtained for serum, liver, kidney, brain and muscle.

Acute Kidney Injury↗
PubMed1988

Kinetic analysis of the positive inotropic action (PIA) of ouabain in isolated perfused rabbit heart. Slow onset of PIA and slow binding to Na+, K+-adenosine triphosphatase.

The positive inotropic action (PIA) of ouabain was analyzed kinetically using isolated perfused rabbit heart. The input function of the ouabain concentration in the perfusate (Ci) into the heart was controlled by changing the volume of the reservoir and the rate of ouabain infusion into the reservoir fixed in front of the heart. The time courses of PIA were measured continuously with different infusion rates. The relationship between Ci and PIA clearly depended on the infusion rate in isolated perfused rabbit heart. The binding kinetics of ouabain to Na+, K+-adenosine triphosphatase (ATPase) in the cardiac homogenate showed two kinds of binding sites. The association rate constant (kappa 1), the dissociation rate constant (kappa-1) and the binding capacity of each site was estimated by the simultaneous fitting method. The occupation curve of the high affinity site corresponded well with the PIA measured in the isolated perfused heart at steady state. These results indicate that ouabain binding to the high affinity site is related to the PIA, and the slow binding process of ouabain to Na+, K+-ATPase may be one of the principal reasons for the infusion-rate dependence of ouabain PIA.

Animals↗
PubMed1988

Kinetic study on the mechanism of tissue distribution of vinblastine.

The purpose of present study was to analyze the factors involved in the tissue distribution of vinblastine (VBL). The specific binding of VBL to mouse tissue cytosol determined by a charcoal method correlated well with the tissue concentration of tubulin, a target protein for the pharmacological activity of Vinca alkaloids. The calculated tissue-to-plasma partition coefficients (Kp) in various tissues, based on the VBL binding to 100000 x g cytosols showed a good correlation with the corresponding in vivo Kp values of rats reported in the literature, however, the calculated Kp values were greatly underestimated. The total binding (including specific and non-specific bindings) to cytosols from the liver and kidney, determined with the ultrafiltration method, were approximately 5 times higher than those determined with the charcoal method for both tissues. However, the total bindings to cytosol cannot explain the high Kp values in vivo. Considering the intracellular distribution of VBL, it was found that cytosol is not the main binding component for VBL since approximately one-half of the VBL in the liver homogenate was associated with the nuclear fraction. The Kp value in the liver, calculated by considering the intracellular distribution, became close to the in vivo Kp value. It was concluded that the tissue distribution of VBL cannot be accounted for only by the binding to tubulin, and that the bindings to other intracellular components should also be included.

Animals↗
PubMed1988

Effects of phenothiazines on the Na+-H+ exchanger of the brush-border membrane from the proximal small intestine of the rabbit.

We investigated the effects of various phenothiazines on the Na+-H+ exchanger in brush-border membrane vesicles isolated from the proximal small intestine of the rabbit. The Na+-H+ exchanger activity was assayed by measuring the dimethylamiloride-sensitive Na+ uptake into these vesicles in the presence of an outward-directed H+ gradient ([pH]o greater than [pH]i). All phenothiazines that were tested inhibited the Na+-H+ exchanger and chlorpromazine sulfoxide (CPZ-SO) was the most potent inhibitor among them. CPZ-SO inhibited the dimethylamiloride-sensitive Na+ uptake in a dose-dependent manner and IC50 for this process was 90 microM. Even though CPZ-SO drastically reduced the initial rates of Na+ uptake, the equilibrium uptake of Na+ into the vesicles was not affected, indicating that the integrity of membrane vesicles was not altered by the drug. Furthermore, the inhibition of the exchanger activity by phenothiazines was not due to nonspecific dissipation of the H+ gradient, because CPZ-SO inhibited the dimethylamiloride-sensitive Na+ uptake even in the absence of a H+ gradient. The inhibition of the exchanger by CPZ-SO was freely and completely reversible. Kinetic analyses of the Na+-H+ exchanger carried out in the presence and absence of CPZ-SO revealed that the nature of the inhibition depended on the composition of the buffers used in the assay of the exchanger because the inhibition was of a mixed type in the mannitol medium, whereas it was of a competitive type in the KCl medium.

Amiloride↗
PubMed1987

Platelet-type von Willebrand disease platelet aggregating factor: a novel functional assay of von Willebrand factor.

Blood platelets from patients with platelet-type von Willebrand disease (vWD) aggregate upon the addition of human von Willebrand factor (vWf) in the absence of ristocetin or other stimulating factors. We measured quantitatively the ability of vWf to induce directly aggregation of platelet-type vWD platelets (platelet-type vWD platelet aggregating factor [PT-PAF]). Cryoprecipitate and factor VIII concentrates were used as a source of vWf of various multimeric composition. The PT-PAF activity was dependent on the multimer size of vWf, like in the case of ristocetin cofactor (RCof) activity. However, PT-PAF activity was not equivalent to RCof activity and the relative PT-PAF/RCof ratio ranged from 1.00 to 0.18 in the materials studied. The preparations containing the higher-molecular-weight multimers had higher PT-PAF/RCof ratio. These findings suggest that PT-PAF activity is a functional expression of more highly polymerized multimers of vWf as compared with RCof activity. Measurement of PT-PAF would serve as a novel functional assay of vWf.

Factor VIII↗
PubMed1987

A novel binding site for bile acids on human serum albumin.

Binding sites of bile acids on human serum albumin were studied using various probes: dansylsarcosine (site I probe), 7-anilinocoumarin-4-acetic acid (ACAA, site II probe), 5-dimethylaminonaphthalene-1-sulfonamide (DNSA, site III probe), cis-parinaric acid (probe for fatty acid binding site) and bilirubin. Bile acids competitively inhibited the binding of dansylsarcosine to human serum albumin whereas bile acids enhanced the binding of ACAA, DNSA, cis-parinaric acid and bilirubin. Considering the concentrations of bile acids required to inhibit the binding of dansylsarcosine to human serum albumin, the secondary binding site of bile acids may correspond to site I. Dissociation constants (Kd) of the primary binding sites of lithocholic and chenodeoxycholic acid to human serum albumin were approximately 0.2 and 4 microM, respectively, which was measured by equilibrium dialysis at 37 degrees C. All the bile acids and their sulfates and glucuronides inhibited the binding of chenodeoxycholic acid to human serum albumin. Lithocholic and chenodeoxycholic acid and their sulfates and glucuronides exhibited more inhibition than cholic acid and its conjugates. In conclusion, bile acids may bind to a novel binding site on human serum albumin.

Bile Acids and Salts↗
PubMed1987

Inhibition of blood-brain barrier permeability to DL-propranolol by serum from acute renal failure rats.

The effect of uranyl nitrate-induced acute renal failure on the brain uptake of DL-propranolol was investigated in rats with a series of tissue-sampling single-carotid injection techniques. When the buffer solution was used as an injection solution, the brain uptake index (BUI), the extraction ratio (ET), and the blood-brain barrier (BBB) permeability-surface area product (PSapp) and PSu (corrected PSapp for the unbound fraction) in uremic rats were significantly lower than those in control rats. These parameters for DL-propranolol were decreased significantly in both control and uremic rats receiving injection of the uremic serum. The PSu values in both of the control and uremic rats injected with either control or uremic rat serum were significantly higher than those in rats injected with the buffer solution, suggesting the presence of a protein-mediated transport mechanism; that is, the conventional assumption that the fraction of the drug which is available for the uptake in vivo is equal to the unbound fraction as measured in vitro may not hold. In contrast, the brain extraction of D-[14C]glucose, [3H]inulin and [3H]water, which show no binding to serum protein, was not affected by the coinjection of either control or uremic rat serum. On the other hand, using either the ultrafiltrate from serum (control and uremic) or supernatant fraction from heat-treated serum (control and uremic) as the injection solution, no significant difference in the PSu value for DL-propranolol was observed between control and uremic serum. These results suggest that (1) the decrease in the PSu value for DL-propranolol in uremic rats may be attributed mainly to the presence of an endogenous inhibitory substance(s) for the brain uptake or to the decrease in the exchangeable fraction in vivo in the uremic serum; (2) the decrease in the PSu value for DL-propranolol may also be partly attributed to the change in the BBB permeability and/or surface area; (3) the inhibitor for the brain uptake may be characterized as a temperature-sensitive and nonfiltrable substance(s) at physiological pH; and (4) the ability of protein-mediated transport for DL-propranolol into brain was decreased in uremic rats.

Acute Kidney Injury↗
PubMed1987

Comparison of bile acid binding to sinusoidal and bile canalicular membranes isolated from rat liver.

Simon et al. (J. Clin. Invest., 70 (1982) 401) studied cholate binding to crude liver plasma membrane vesicles and suggested that the binding may represent mainly the binding to the receptor (carrier) on the canalicular membrane. This hypothesis was supported by finding a good correlation between the number of cholate binding sites on liver plasma membrane and the maximal rate of biliary secretion (Tm) for taurocholate. We studied bile acid binding to sinusoidal and canalicular membrane vesicles isolated from rat liver by a rapid filtration technique. Scatchard analysis of the saturation kinetics showed both [3H]cholate and [3H]chenodeoxycholate bind to two classes of binding site on each membrane. However, little difference was observed between the binding to sinusoidal and canalicular membrane vesicles for each bile acid (cholate, Kd1 = 10.4 and 19.8 microM, n1 = 31.0 23.6 pmol/mg protein, Kd2 = 1.32 and 1.73 mM, n2 = 13.1 and 23.4 nmol/mg protein; and chenodeoxycholate, Kd1 = 0.207 and 0.328 microM, n1 = 36.7 and 27.4 pmol/mg protein, Kd2 = 1.16 and 2.26 mM, and n2 = 20.6 and 24.2 nmol/mg protein; numbers show the mean values sinusoidal and canalicular membrane vesicles, respectively). Chenodeoxycholate binding to sinusoidal membrane vesicles was markedly inhibited by cholate but not by Rose bengal, an organic anion dye. These studies indicate that both membranes (sinusoidal and canalicular membrane vesicles) have two kinds of binding site for bile acids, although no clear difference in the binding properties was observed between the two membranes. Consequently, the cholate binding Simon detected may represent the binding not only to canalicular membrane vesicles but also to sinusoidal membrane vesicles.

Animals↗