scholarly journals Effect of insulin on ultrastructure and glycogenesis in primary cultures of adult rat hepatocytes

1977 ◽  
Vol 74 (3) ◽  
pp. 878-900 ◽  
Author(s):  
D Bernaert ◽  
JC Wanson ◽  
P Drochmans ◽  
A Popowski
Keyword(s):  
Endoplasmic Reticulum ◽  
Smooth Endoplasmic Reticulum ◽  
Primary Cultures ◽  
Active Form ◽  
Rat Hepatocytes ◽  
High Concentration ◽  
Trophic Effect ◽  
Adult Rat ◽  
Glycogenosis Type ◽  
High Concentrations

Insulin in the presence of high concentrations of glucose has a beneficial trophic effect on the development of primary cultures of hepatocytes. Compared to the situation observed in hormone-free control cultures, the flattening of the reaggregated hepatocytes is enhanced, and the reconstituted cell trabeculae are enlarged and tend to form a confluent monolayer after 3 days; the survival time is prolonged from 3 to 5 or 6 days. Ultrastructural modifications are also initiated by insulin; numerous glycogen particles appear after 24 h, in between the cisternae of the proliferated smooth endoplasmic reticulum. After 48 h, large amounts of glycogen are stored, and numerous polysomes are present. A small number of cells showed an increased synthesis of lipid droplets in the lumen of the smooth endoplasmic reticulum and form liposomes at the same time. After 72 h, cytolysomes filled with glycogen develop, simulating glycogenosis type II. Simultaneously, microtubules and microfilaments, closely related to numerous polysomes, appear in cytoplasmic extensions constituting undulating membranes. The biochemical data demonstrate that, in the absence of insulin, a high concentration of glucose stimulates glycogenesis and hinders glycogenolysis. This effect of glucose on polysaccharide synthesis is progressively lost. The addition of insulin to the culture induces after 48 and 72 h, a three- to fivefold increase of the glucose incorporation into glycogen, as compared to the controls. The presence of insulin is required to maintain the hepatocyte's capacity to store glycogen. Glycogen synthetase is converted into its active form under the influence of glucose. Insulin increases the rate of activation.

scholarly journals Isolation and subfractionation on ficoll gradients of adult rat hepatocytes. Size, morphology, and biochemical characteristics of cell fractions.

1975 ◽  
Vol 66 (1) ◽  
pp. 1-22 ◽  
Author(s):  
P Drochmans ◽  
J C Wanson ◽  
R Mosselmans
Keyword(s):  
Endoplasmic Reticulum ◽  
Smooth Endoplasmic Reticulum ◽  
Gradient Centrifugation ◽  
Parenchymal Cell ◽  
Liver Homogenate ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Isolated Cells ◽  
Adult Rat ◽  
Mean Diameter

The recirculating perfusion of adult rat liver with a Ca-++-free Hanks' solution produces a release of the adhesiveness of cells and a cleaving of the desmosomes. The addition of collagenase and hyaluronidase to the perfusion medium leads to complete dissociation of the liver tissue into a mixture of isolated cells and cell cords in which the hepatocytes remain connected with specific junctional differentiations, namely the gap and tight junctions. Individual cells are released by submitting the suspension of cell trabeculae to a gentle rolling. The gap junctions are ruptured at least in one of the two adjacent cells and remain generally attached to the other cell taking with them a small portion of cytoplasm. This technique of isolation of hepatocytes yields about 60-65% of the parenchymal cells contained in a liver; endothelial cells and other cells of the connective tissue are not recovered. The ultrastructural preservation of the isolated hepatocytes is excellent and the glucose-6-phosphatase activity, confined to the endoplasmic reticulum, appears unaltered in most cells. Protein, DNA and RNA recovery in the preparations of isolated hepatocytes is satisfactory, amounting to 70% of that found in liver homogenate; glycogen, the most labile component examined, is partly lost or degraded during the manipulations. Cell diameters measured by different methods confirm the preservation of the original volume of the in situ hepatocytes and the presence of more than one type of parenchymal cell. By submitting this heterogeneous cell population to an isopycnic density gradient centrifugation, two types of hepatocytes can be distinguished: the light hepatocytes, with a mean diameter of 20.5 mum and a mean density of 1.10, are characterized by an extended smooth-walled endoplasmic reticulum entrapping dispersed alpha-glycogen particles; the heavy hepatocytes, with a mean diameter of 19.0 mum and a mean density of 1.14, present a relatively reduced compartment of smooth endoplasmic reticulum, but large accumulations of glycogen. It is suggested that the cell fraction of low density is enriched in centrolobular cells and the high density fraction in perilobular hepatocytes.

Morphological and biochemical analyses of changes in rat hepatocytes related to wine and ethanol consumption

1984 ◽  
Vol 42 ◽  
pp. 232-233
Author(s):  
S.M. Geyer ◽  
C.L. Mendenhall ◽  
J.T. Hung ◽  
E.L. Cardell ◽  
R.L. Drake ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Enzyme Activity ◽  
Malic Enzyme ◽  
Smooth Endoplasmic Reticulum ◽  
Rat Hepatocytes ◽  
Mature Male ◽  
Treatment Groups ◽  
Malic Enzyme Activity ◽  
Biochemical Analyses

Thirty-three mature male Holtzman rats were randomly placed in 3 treatment groups: Controls (C); Ethanolics (E); and Wine drinkers (W). The animals were fed synthetic diets (Lieber type) with ethanol or wine substituted isocalorically for carbohydrates in the diet of E and W groups, respectively. W received a volume of wine which provided the same gram quantity of alcohol consumed by E. The animals were sacrificed by decapitation after 6 weeks and the livers processed for quantitative triglycerides (T3), proteins, malic enzyme activity (MEA), light microscopy (LM) and electron microscopy (EM). Morphometric analysis of randomly selected LM and EM micrographs was performed to determine organellar changes in centrilobular (CV) and periportal (PV) regions of the liver. This analysis (Table 1) showed that hepatocytes from E were larger than those in C and W groups. Smooth endoplasmic reticulum decreased in E and increased in W compared to C values.

scholarly journals STUDIES ON MICROPEROXISOMES II. A CYTOCHEMICAL METHOD FOR LIGHT AND ELECTRON MICROSCOPY

1972 ◽  
Vol 20 (12) ◽  
pp. 1006-1023 ◽  
Author(s):  
ALEX B. NOVIKOFF ◽  
PHYLLIS M. NOVIKOFF ◽  
CLEVELAND DAVIS ◽  
NELSON QUINTANA
Keyword(s):  
Endoplasmic Reticulum ◽  
Guinea Pig ◽  
Lipid Droplets ◽  
Smooth Endoplasmic Reticulum ◽  
Light And Electron Microscopy ◽  
Distal Tubule ◽  
Striking Difference ◽  
High Concentration ◽  
Electron Microscopic ◽  
Pig Kidney

A modification of the Novikoff-Goldfischer alkaline 3,3'-diaminobenzidine medium for visualizing peroxisomes is described. It makes possible light microscopic as well as electron microscopic studies of a recently described class of peroxisomes, the microperoxisomes. Potassium cyanide (5 x 10–3 M) is included in the medium to inhibit mitochondrial staining, the pH is 9.7 and there is a high concentration of H2O2 (0.05%). Two cell types have been chosen to illustrate the advantages of the new procedure for demonstrating the microperoxisomes: the absorptive cells in the human jejunum and the distal tubule cells in the guinea pig kidney. Suggestive relations of microperoxisomes and lipid are described in the human jejunum. The microperoxisomes are strategically located between smooth endoplasmic reticulum that radiates toward the organelles and contains lipid droplets and "central domains" of highly specialized endoplasmic reticulum which do not show the lipid droplets. The microperoxisomes are also present at the periphery of large lipid-like drops. In the guinea pig kidney tubule there is a striking difference between the thick limb of Henle and distal tubule. The distal tubule has a population of cells with large numbers of microperoxisomes readily visible by light microscopy; these cells are not present in the thick limb of Henle. Other differences between the two are also described.

Induction and regulation of connexin26 by glucagon in primary cultures of adult rat hepatocytes

1995 ◽  
Vol 108 (8) ◽  
pp. 2771-2780 ◽  
Author(s):  
T. Kojima ◽  
T. Mitaka ◽  
Y. Shibata ◽  
Y. Mochizuki
Keyword(s):  
Culture Medium ◽  
Primary Cultures ◽  
Minor Component ◽  
Rat Hepatocytes ◽  
Primary Hepatocytes ◽  
Adult Rat ◽  
Adult Rat Hepatocytes ◽  
Epidermal Growth ◽  
A Minor ◽  
Junction Proteins

In the adult rat hepatocyte, the gap junction proteins consist of a major component, connexin32 (Cx32) and a minor component, connexin26 (Cx26). Although we recently reported our success in inducing and maintaining Cx32 in adult rat hepatocytes cultured in serum-free L-15 medium supplemented with epidermal growth factor and 2% dimethyl sulfoxide, it was very difficult to induce Cx26 in the primary hepatocytes. In the present study, we found that the addition of 10(−7) M glucagon into the culture medium could dramatically induce Cx26 mRNA and protein. Although the expression of Cx32 mRNA was also influenced by glucagon, the increase of the expression was small. Immunocytochemically, Cx26-positive spots were observed between most adjacent cells and were co-localized with the Cx32-positive spots. We also examined whether 0.5 mM dibutyl cyclic AMP could induce expression of Cx26 in the cells. The effect of dexamethasone on the expression of Cx26 mRNA compared to that of Cx32 mRNA was examined. For the induction and maintenance of Cx26 mRNA, more than 10(−7) M dexamethasone was necessary in this culture. These results suggest that expression of Cx26 in hepatocytes may be regulated by the concentrations of glucagon and glucocorticoid hormones.

Thymidine uptake in primary cultures of adult rat hepatocytes

1986 ◽  
Vol 14 (1) ◽  
pp. 101-102 ◽  
Author(s):  
ANNA ZAKHAROVA ◽  
HEATHER M. WALLACE
Keyword(s):  
Primary Cultures ◽  
Rat Hepatocytes ◽  
Thymidine Uptake ◽  
Adult Rat ◽  
Adult Rat Hepatocytes

Comparison of cytochrome P-450 levels in adult rat liver, postnatal rat liver, and primary cultures of postnatal rat hepatocytes

Life Sciences ◽  
1979 ◽  
Vol 25 (16) ◽  
pp. 1413-1418 ◽  
Author(s):  
Daniel Acosta ◽  
David C. Anuforo ◽  
Reagan McMillin ◽  
William H. Soine ◽  
Robert V. Smith
Keyword(s):  
Rat Liver ◽  
Primary Cultures ◽  
Rat Hepatocytes ◽  
Adult Rat ◽  
Cytochrome P 450
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