Uptake of palmitate by hepatocyte suspensions: facilitation by albumin?

1992 ◽  
Vol 262 (5) ◽  
pp. G883-G894 ◽  
Author(s):  
S. M. Pond ◽  
C. K. Davis ◽  
M. A. Bogoyevitch ◽  
R. A. Gordon ◽  
R. A. Weisiger ◽  
...  
Keyword(s):  
Palmitic Acid ◽  
Association Constant ◽  
Absolute Magnitude ◽  
Diffusion Reaction ◽  
Female Rat ◽  
Theoretical Predictions ◽  
Reaction Theory ◽  
Uptake Velocity ◽  
Rat Livers ◽  
Initial Uptake

Albumin-dependent uptake of unbound [3H]palmitic acid by hepatocytes isolated from female rat livers was studied and the experimental results compared with the predictions of a noncompartmental diffusion-reaction theory for the cellular uptake of protein-bound ligands. The outright theoretical predictions involve values for the parameters of the system, some newly measured (hepatocyte radii and the rate constant for the dissociation of palmitate-albumin complex) and some taken from the literature (diffusion coefficients and the equilibrium association constant for the palmitate-albumin complex). The measured unbound clearance of [3H]palmitic acid, defined as the initial uptake velocity divided by the unbound [3H]palmitic acid concentration in the medium, was enhanced 6.6-fold as the concentration of human serum albumin was increased from approximately 5 to 480 microM. This enhancement factor was predicted by the theory, according to which the enhancement reflects codiffusion of bound ligand across the unstirred layer adjacent to the cell membrane and, therefore, an increased delivery of unbound ligand to the cell surface. In contrast, the absolute magnitude of the unbound clearance was consistent with the theory only for the lowest published value for the equilibrium association constant, 15 microM-1. For higher published values (62 and 94 microM-1), the magnitude of the unbound clearance observed experimentally was severalfold higher than that predicted by the theory. If in fact the association constant exceeds 30 microM-1, the data would imply that an albumin-dependent facilitation mechanism exists which enhances the availability of palmitate to the cell over and above the enhancement predicted by the diffusion-reaction theory.

Palmitate uptake by hepatocyte suspensions: effect of albumin

1994 ◽  
Vol 267 (3) ◽  
pp. G371-G379
Author(s):  
F. J. Burczynski ◽  
Z. S. Cai
Keyword(s):  
Reaction Model ◽  
Partition Ratio ◽  
Dissociation Rate ◽  
Diffusion Reaction ◽  
Ratio Method ◽  
Albumin Concentration ◽  
Ratio Test ◽  
Female Rat ◽  
Clearance Ratio ◽  
Rat Livers

The clearance ratio test has been used to determine whether hepatocytes isolated from female rat livers facilitate the dissociation rate of albumin-palmitate complexes. This test requires knowledge of the clearance rate and unbound palmitate fraction at two protein concentrations. The unbound fractions have been estimated using the heptane-water partition ratio method. These fractions were determined to deviate from the expected linear relationship because of radiolabeled impurities that were still present after purification of the [3H]palmitic acid. With the use of experimentally determined partition ratio values, the palmitate clearance ratios were statistically greater than those predicted by the diffusion-reaction model at albumin concentration combinations above 99 microM/49 microM. Experimental clearance rates also exceeded those predicted by the diffusion-reaction model, but only at albumin concentrations greater than 198 microM. These data support the proposal that hepatocytes facilitate the dissociation of albumin-palmitate complexes at high albumin concentrations.

scholarly journals Mass spectrometric analysis of rat liver cytosolic glutathione S-transferases: modifications are limited to N-terminal processing

1995 ◽  
Vol 308 (1) ◽  
pp. 69-75 ◽  
Author(s):  
H I Yeh ◽  
C H Hsieh ◽  
L Y Wang ◽  
S P Tsai ◽  
H Y Hsu ◽  
...  
Keyword(s):  
Molecular Mass ◽  
Female Rats ◽  
Spectrometric Analysis ◽  
Affinity Column ◽  
Female Rat ◽  
Significant Difference ◽  
Cytosolic Glutathione ◽  
Glutathione S Transferases ◽  
Molecular Masses ◽  
Rat Livers

Cytosolic glutathione S-transferases (GSTs) from rat livers were purified using an S-hexylglutathione affinity column. The GST subunits were resolved by reverse-phase HPLC and their molecular masses were determined by electrospray mass spectrometry. The major hepatic GSTs detected were subunits 1, 1′, 2, 3 and 4, with molecular mass of 25,520, 25,473, 25,188, 25,782 and 25,571 Da respectively. Subunits 6, 7 and 10 are minor components, with molecular mass of 25,551, 23,308 and 25,211 Da respectively. Alternatively, the hepatic GSTs were purified using a glutathione affinity column. Subunits 1, 1′, 2, 8 and 10 were eluted from this column with GSSG, the oxidized form of glutathione. Subunit 8 has a molecular mass of 25,553 Da. The remaining proteins on the glutathione affinity column were removed with glutathione and S-hexylglutathione. Subunits 2, 3, 4 and 6 could be detected in the eluate. We could not detect any significant difference in molecular mass between GSTs isolated from male and female rat livers. Cytosolic GSTs were isolated from livers of buthionine sulphoximine-treated female rats for MS analysis. The molecular masses obtained were identical to those determined for the controls.

scholarly journals Sex-Related Liver Injury Due to Alcohol Involves Activation of Kupffer Cells by Endotoxin

2000 ◽  
Vol 14 (suppl d) ◽  
pp. 129D-135D ◽  
Author(s):  
Ronald G Thurman
Keyword(s):  
Liver Injury ◽  
Kupffer Cells ◽  
Coupling Constants ◽  
Female Rats ◽  
Male Rat ◽  
Gadolinium Chloride ◽  
Female Rat ◽  
Ethanol Treatment ◽  
Necrosis Factor Alpha ◽  
Rat Livers

Females have a greater susceptibility to ethanol-induced liver injury than males. Females who drink ethanol regularly and have been overweight for 10 years or more are at greater risk for both hepatitis and cirrhosis than males, and females develop ethanol-induced liver injury more rapidly and with less ethanol than males. Female rats on an enteral ethanol protocol exhibit injury more quickly than males and have widespread fatty changes over a larger portion of the liver lobule. Moreover, levels of plasma endotoxin, intracellular adhesion molecule-1, free radical adducts, infiltrating neutrophils and nuclear factor kappa B are doubled in female rat livers compared with male rat livers after enteral ethanol treatment. Additionally, estrogen treatment in vivo increases the sensitivity of hepatic macrophages or Kupffer cells to endotoxin. Evidence has been presented that Kupffer cells are pivotal in the development of ethanol-induced liver injury. Destroying Kupffer cells with gadolinium chloride or decreasing bacterial endotoxin by sterilizing the gut with antibiotics inhibits early inflammation due to ethanol. Similar results have been obtained with anti-tumour necrosis factor-alpha antibody. These data pointed to the hypothesis that ethanol-induced liver injury involves elevations in circulating endotoxin concentrations leading to activation of Kupffer cells, which causes a hypoxia-reoxygenation injury. This theory has been tested using pimonidazole, a 2-nitroimidazole marker, to quantify hypoxia in downstream, pericentral regions of the hepatic lobule. After chronic enteral ethanol treatment, pimonidazole binding doubles. Enteral ethanol also increases free radicals detected with electron spin resonance. Radical adducts, with coupling constants such as alpha-hydroxyethyl radical, have been shown to arise from ethanol. Importantly, hypoxia and radical production detected in bile are also decreased by the destruction of Kupffer cells with gadolinium chloride. These data support the hypothesis that Kupffer cells contribute to the vital sex differences in liver injury caused by ethanol.

EFFECT OF SEX HORMONES ON OXALATE-SYNTHESIZING ENZYMES IN MALE AND FEMALE RAT LIVERS

1999 ◽  
pp. 668-673 ◽  
Author(s):  
HIDEKI YOSHIHARA ◽  
SATOSHI YAMAGUCHI ◽  
SUNAO YACHIKU
Keyword(s):  
Sex Hormones ◽  
Female Rat ◽  
Male And Female ◽  
Rat Livers

Models of depressed hepatic mrp2 activity reveal bromosulphophthalein-sensitive passive K+ flux

2002 ◽  
Vol 80 (12) ◽  
pp. 1167-1172 ◽  
Author(s):  
Qin Li ◽  
M Martha Briggs ◽  
Doris Folkens ◽  
Ceredwyn E Hill
Keyword(s):  
Bile Acid ◽  
Rat Liver ◽  
Organic Anion ◽  
Isolated Hepatocytes ◽  
Early Event ◽  
Female Rat ◽  
Wild Type ◽  
Rat Livers ◽  
Gender Specific ◽  
Secretory Capacity

Bile acid independent flow composes up to 40% of hepatic bile secretory capacity. Apical (canalicular) efflux of non-bile-acid organic anions provides the major osmotic driving force for bile acid independent flow. Organic anion accumulation in the hepatocyte is accompanied by increases in both K+ conductance in isolated hepatocytes and passive K+ flux in the perfused rat liver, which are indicative of K+ channel activation. We used two models of disrupted canalicular anion transport to test whether organic anion stimulated K+ efflux occurs independently of anion excretion. In both wild type (wt) and mrp2 mutant (transport minus, tr–) rat liver, bromosulfophthalein (BSP; 0.5mM) caused a reversible increase in K+ flux that (i) was outwardly directed with low external K+ and (ii) depended upon the electrochemical potential for K+. K+ efflux from wt livers of both sexes was about 1.5 times larger than that from tr– livers. Further, K+ release from female rat livers was about three times higher than that from male livers, independent of phenotype. Two transcripts of the rat hepatocyte K+ channel (Kir4.2) were expressed in hepatocytes of all rats. The results demonstrate that BSP stimulates basolateral (sinusoidal) K+ channels independently of its canalicular excretion, revealing an early event in BAIF and suggesting that Kir4.2 may mediate BSP-sensitive K+ flux.Key words: liver, GY, gender-specific, male, female, transport minus, Kir4.2.

Sarcolemmal Na(+)-Ca2+ exchange activity and exchanger immunoreactivity in developing rabbit hearts

1992 ◽  
Vol 263 (5) ◽  
pp. H1506-H1513 ◽  
Author(s):  
M. Artman
Keyword(s):  
Age Groups ◽  
Rabbit Heart ◽  
Newborn Rabbit ◽  
Developing Heart ◽  
Age Related ◽  
New Zealand White Rabbits ◽  
Uptake Velocity ◽  
Sarcolemmal Vesicles ◽  
Initial Uptake ◽  
Exchange Activity

It has been postulated that as a consequence of an underdeveloped sarcoplasmic reticulum, sarcolemmal Na(+)-Ca2+ exchange assumes relatively greater importance in modulating Ca2+ fluxes in the developing heart. To explore this concept, cardiac sarcolemmal vesicles were prepared from late fetal (28-day gestation), newborn (24-48 h), immature (14-16 days), and adult New Zealand White rabbits. Na(+)-dependent Ca2+ uptake was measured by diluting Na(+)-loaded (140 mM) vesicles into Na(+)-free buffer and measuring 45Ca2+ uptake (40 microM Ca2+) by timed quenching and rapid filtration. Vesicles from all four age groups demonstrated Ca2+ uptake curves characteristic of Na(+)-Ca2+ exchange with stimulation by valinomycin and inhibition by amiloride. Initial uptake velocity (measured at 2 s and corrected for the fraction of competent vesicles) was significantly higher in fetal (23.2 +/- 5.5 nmol/mg) and newborn (26.2 +/- 5.9 nmol/mg) than in adult sarcolemmal preparations (7.3 +/- 1.2 nmol/mg). Uptake was intermediate in the 2-wk-old group (13.7 +/- 1.7 nmol/mg). The relative amounts of exchanger protein were compared by quantitating immunoreactivity using a polyclonal antibody to the Na(+)-Ca2+ exchanger. Densitometric scanning of protein slot blots demonstrated approximately 2.5 times more exchanger protein in fetal and newborn sarcolemma than in adult preparations. The relative amount of exchanger protein detected immunologically corresponded with the age-related differences observed in exchanger activity. Thus the cardiac sarcolemmal Na(+)-Ca2+ exchanger is abundant and functionally well-developed in the late fetal/early newborn rabbit heart and appears to decline postnatally.(ABSTRACT TRUNCATED AT 250 WORDS)

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