Induction of specific human growth hormone binding sites in rat liver by human growth hormone.

125I-Labeled hGH was bound to liver plasma membranes which were obtained from female rats. The binding was displaced by hGH, hPRL, bPRL, rPRL and bGH but not by rGH. This result indicated that hGH was bound to lactogenic binding sites in rat livers. After hypophysectomy, the binding was markedly decreased. Treatment of hypophysectomized rats with hGH (80 micrograms/day) for 10 days increased the binding sites for hGH. These binding sites were different from those found in normal female rat livers because of their high affinity and specificity for hGH. These results indicate that hGH induces specific binding sites for hGH in rat livers.     

Investigation of selenium distribution in subcellular fractions of human liver by neutron activation analysis

Selenium is an important and essential trace element to living systems. In the article, two methods of instrumental neutron activation analysis and hydride generation-atomic fluorescence spectrometry were applied to determine Se in biological samples and the accuracy was evaluated by several reference materials. The subcellular distribution of selenium in human liver samples, which were obtained from normal subjects who had an accidental death, was investigated by differential centrifugation combined with INAA. Selenium was mainly enriched in mitochondria, nuclei, and cytosol. Almost half of the total Se content existed in nuclei as a result of the large amount in liver and the high Se concentration. Generally, the highest Se concentration in the mitochondrial fractions of each liver sample suggested that Se had important functions in this liver component.     

Kinetics of the early subcellular distribution of cadmium in rat hepatocytes

The kinetics of the early subcellular distribution of cadmium (Cd) was characterized in primary cultures of rat hepatocytes exposed to 10, 50 and 100 M Cd in a serum-free WME medium for 10, 30 or 60 min. Our results demonstrate a time- and concentration-dependent increase in Cd content with the highest metal concentration measured in the cytosol, whereas the lowest was observed in the mitochondria. With the exception of early localization in the plasma membrane, Cd concentrations in fractions were characterized by the following decreasing order of magnitude: cytosol > low density molecules nuclei > lysosomes mitochondria. We also found evidence for: (i) a two-step process for Cd distribution in the nuclei and mitochondria; and (ii) a time-dependent slow process of transfer from the plasma membrane to the cytosol. Saturation in Cd uptake was observed at 50 M in most cell fractions at 10 and 30 min, except for the plasma membrane. The lack of apparent saturation for Cd accumulation at 60 min was not related to an increase in metallothionein synthesis. Altogether, our data provide insights into the dynamics of transfer between intracellular compartments, and allow a better identification of the organelles that are the most subjected to Cd toxicity for early exposure conditions.Deceased March 25, 2004.     

Uptake and subcellular distribution of injected transferrin in rat liver

Radioactively labelled transferrin was injected into rats intravenously and its uptake and subcellular distribution in the liver was investigated. The amount of radioactivity in the liver remained constant from 10 min after injection. It was not influenced by asialoglycoproteins. The radioactive label was identified as asialotransferrin. After subcellular fractionation by differential and zonal sucrose density-gradient centrifugation the label was enriched in a low-density endocytic compartment showing fluorescence quenching of acridine orange and N-ethylmaleimide-sensitive ATPase activity. The data fit into a model of continuous transferrin-receptor-mediated recycling through the hepatocyte's endocytic compartment.     

The subcellular distribution of fumarase isozymes in rat liver

Between 13 and 25% of the fumarase activity of rat liver was found to be cytosolic in origin the remainder being localised in the mitochondria. Electrophoretic analysis on cellulose acetate showed that mitochondria do not contain detectable levels of cytosolic isozyme or vice versa.     

Cryopreservation of pig and human liver slices

The ability to cryopreserve human liver slices would greatly enhance the opportunities to test potentially hepatotoxic drugs and environmental contaminants as well as the metabolism of these compounds. This study focused on trying to cryopreserve pig and human liver slices. Since the acquisition of human liver tissue is unpredictable and scarce, an animal model was sought to predict problems associated with cryopreservation of human tissue. The pig liver was chosen because of its anatomical and physiological resemblance to human liver. The human liver tissues that did become available were obtained through the Arizona Organ Bank and the National Disease Research Interchange and from surgical liver resections. An in vitro culture system that employed precision-cut liver slices was used in this study. Different types and concentrations of cryoprotectants, cooling rates, and culture media were all tried in an attempt to cryopreserve pig and human liver slices. The viabilities of fresh and cryopreserved liver slices were evaluated using slice K+ retention and protein synthesis. Pig liver slices following cryopreservation retained between 80 and 85% of intracellular K+ content and protein synthesis as compared to controls using 1.4 M Me2SO, a 12 degrees C/min cooling rate, and a rapid rewarming rate of direct submersion of the slice into 37 degrees C fetal calf serum. Human liver slices following cryopreservation retained between 54 and 89% of intracellular K+ content and protein synthesis as compared to controls using the same protocol as for pigs, except that lower cooling rates were giving better results. The large variation seen in cryopreserved human liver slices was due to the length of warm and cold ischemia to which the tissue was exposed before arriving at the laboratory. This study indicated that pig and human liver slices can be cryopreserved and used for future toxicological and metabolic studies.     

Binding and subcellular distribution of cyclosporine in human fibroblasts

The uptake, binding, and subcellular sites of accumulation of [³H]-cyclosporine (CS) in two human gingival fibroblast strains, GN 23 and GN 54, have been examined. GN 23 responds to CS treatment with a decrease in collagenolysis, while GN 54 does not. Binding of the drug was determined using [³H]-CS concentrations ranging from 10^?5 to 10^?8 M in the absence or presence of excess unlabeled CS (1 mM). The binding of the drug to both strains was specific and reached a plateau within 10 min, remaining at that level for up to 1 h. Scatchard analysis of the specific binding of [³H]-CS to the responsive GN 23 strain revealed two dissociation constants: K_D = 5 × 10^?8 M (1.2 × 10⁷ sites/cell) and K_D = 1.4 × 10^?6 M (2.2 × 10⁸ sites/cell). GN 54, on the other hand, had only one class of low affinity binding site (K_D = 0.47 × 10^?6 M [1.2 × 10⁸ sites/cell]). Unlabeled CS (0.01–1 mM) inhibited the binding of [³H]-CS in a dose-dependent manner to both strains, as did the calmodulin antagonist W-7, to a lesser extent. However, W-7 inhibited CS binding much more efficiently in GN 54 than in GN 23, suggesting that calmodulin may be the predominant CS receptor in GN 54. In both strains, 70% of the drug accumulated in the crude nuclear fraction after a 1 min incubation, with very little (? 4%) being membrane associated, and the remainder was in the cytosol. In GN 23, CS levels in the crude nuclear fraction reached 80% by 20 min, and remained at this level for up to 1 h. In contrast, in GN 54, at incubation times of more than 1 min, the drug did not selectively accumulate in the crude nuclear fraction, but appeared to be in equilibrium between the nuclear and cytosolic fractions. These data show that the CS resistance of human gingival fibroblasts was not due to their inability to take up and bind CS. Rather, the different effects of CS on the collagenolysis of the responder and non-responder fibroblast strains may be related to the types of CS receptors they possess and differences in the cellular metabolism of CS occurring after binding, including the subcellular sites of drug accumulation. © 1993 Wiley-Liss, Inc.     

Kinetics of growth and hydrogen uptake byMethanobacterium formicicum

Fermentation kinetics for the growth and conversion of H₂ and CO₂ to CH₄ byM. formicicum were modeled by using Monod equations. The maximum specific growth rate and H₂ uptake rate _max and q_max, were found to be 0.053 h^–1 and 0.13 mol/hg cell, respectively. The partial pressure of H₂ was found not to have a significant effect on either growth or H₂ utilization. The yield of CH₄ from H₂ was calculated as 0.27 mol/mol, which is within 7% of the theoretical value of 0.25.     

Estrogen and the subcellular distribution of luteinizing hormone releasing hormone: Rate sedimentation studies

Rate sedimentation of the 900×G supernatants (S₁) of hypothalamic homogenates from untreated male rats or ovariectomized rats with or without 5 μg estradiol benzoate (EB) revealed two populations of LHRH particles: a minor, slowly sedimenting one (peak 1) and a major, more rapidly sedimenting one (peak 2). Some LHRH-containing material also sedimented to the bottom of the gradient. The ovariectomized rats displayed more heterogeneity of particulate LHRH than did the male rats. Furthermore, the administration of EB to ovariectomized rats altered the relative sedimentation pattern of LHRH. In ovariectomized rats, hypotonic shock of S₁ prior to rate sedimentation eliminated peak 2 and post-peak 2 LHRH and increased free LHRH at the top of the gradient. Peak 1 LHRH was still present and was elevated after EB treatment. Also, EB treatment lowered the free LHRH at the top of the gradient. These data demonstrate that the administration of EB to an ovariectomized rat alters the subcellular distribution of LHRH.     

Estrogen and the subcellular distribution of luteinizing hormone releasing hormone: Rate sedimentation studies

Rate sedimentation of the 900×G supernatants (S₁) of hypothalamic homogenates from untreated male rats or ovariectomized rats with or without 5 μg estradiol benzoate (EB) revealed two populations of LHRH particles: a minor, slowly sedimenting one (peak 1) and a major, more rapidly sedimenting one (peak 2). Some LHRH-containing material also sedimented to the bottom of the gradient. The ovariectomized rats displayed more heterogeneity of particulate LHRH than did the male rats. Furthermore, the administration of EB to ovariectomized rats altered the relative sedimentation pattern of LHRH. In ovariectomized rats, hypotonic shock of S₁ prior to rate sedimentation eliminated peak 2 and post-peak 2 LHRH and increased free LHRH at the top of the gradient. Peak 1 LHRH was still present and was elevated after EB treatment. Also, EB treatment lowered the free LHRH at the top of the gradient. These data demonstrate that the administration of EB to an ovariectomized rat alters the subcellular distribution of LHRH.