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PMID: 2871008 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Use of a low-speed, iso-density percoll centrifugation method to increase the viability of isolated rat hepatocyte preparations.

In vitro cellular & developmental biology : journal of the Tissue Culture Association ·Vol. 22 ·No. 4 ·1986-04-00 ·Pages 201-11

Kreamer BL, Staecker JL, Sawada N, Sattler GL, Hsia MT, Pitot HC

Abstract

A simple yet effective method (iso-density percoll centrifugation) has been developed for consistently preparing isolated rat liver parenchymal cells with over 98% initial viability. The method has been applied to cells isolated by a variety of collagenase digestion techniques. This procedure involves the low-speed centrifugation (50 X g) of the initial cell suspension through a percoll medium having a density of 1.06 g/ml and results in the separation of single and viable parenchymal cells from cell aggregates, debris, and nonparenchymal cells. The enriched parenchymal cells have been shown to be superior to untreated cells by a number of criteria including: preparation homogeneity, cell morphology, maintenance of cytochrome P-450, hormonal responsiveness (measured by the induction of tyrosine aminotransferase after treatment with glucagon or dexamethasone, or both), plasma membrane integrity (determined by both trypan blue exclusion and leakage of glutamic-oxaloacetic transaminase), and the DNA repair capability after treatment with benzo[a]pyrene or 2-acetylaminofluorene.

MeSH Terms
2-Acetylaminofluorene/pharmacology Animals Benzo(a)pyrene/pharmacology Cell Membrane/physiology Cell Separation/methods Cell Survival Centrifugation, Density Gradient Cytochrome P-450 Enzyme System/metabolism DNA/biosynthesis DNA Repair Dexamethasone/pharmacology Enzyme Induction/drug effects Glucagon/pharmacology Liver/cytology,metabolism Male Rats Rats, Inbred Strains Tyrosine Transaminase/biosynthesis
Chemicals
Benzo(a)pyrene Dexamethasone DNA Glucagon Cytochrome P-450 Enzyme System 2-Acetylaminofluorene Tyrosine Transaminase
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Kreamer B L
Staecker J L
Sawada N
Sattler G L
Hsia M T
Pitot H C
References (37)
37 references, click to expand
  1. Primary culture of parenchymal liver cells on collagen membranes. Morphological and biochemical observations.
    Exp Cell Res. 1975 Aug;94(1):70-8 PMID: 243
  2. The control of glutathione and cytochrome P-450 concentrations of primary cultures of rat hepatocytes.
    Biochem Pharmacol. 1981 Oct 1;30(19):2739-42 PMID: 7295359
  3. Effects of phenobarbital and 3-methylcholanthrene pretreatments on monooxygenase activities and proportions of isolated rat hepatocyte subpopulations.
    Mol Pharmacol. 1983 Jan;23(1):235-43 PMID: 6602940
  4. Detection of chemical carcinogens by unscheduled DNA synthesis in rat liver primary cell cultures.
    Cancer Res. 1977 Jun;37(6):1845-51 PMID: 404038
  5. The separation of cells and organelles by partitioning in two-polymer aqueous phases.
    Biochem J. 1981 Apr 15;196 (1):1-10 PMID: 6171264
  6. THE CARBON MONOXIDE-BINDING PIGMENT OF LIVER MICROSOMES. I. EVIDENCE FOR ITS HEMOPROTEIN NATURE.
    J Biol Chem. 1964 Jul;239:2370-8 PMID: 14209971
  7. Development of a toxicity test system using primary rat liver cells.
    In Vitro. 1981 Nov;17(11):1004-10 PMID: 7319528
  8. Primary monolayer cultures of adult rat liver parenchymal cells suitable for study of the regulation of enzyme synthesis.
    In Vitro. 1974 May-Jun;9(6):399-413 PMID: 4157202
  9. A simplified method for the quantitative assay of small amounts of protein in biologic material.
    Anal Biochem. 1973 Feb;51(2):654-5 PMID: 4735559
  10. Separation of different cell populations of rat liver by density gradient centrifugation in a vertical rotor with self-generated Percoll gradients.
    Acta Physiol Scand. 1983 Apr;117(4):497-505 PMID: 6349254
  11. The viability of cells grown or centrifuged in a new density gradient medium, Percoll(TM).
    Exp Cell Res. 1977 Dec;110(2):449-57 PMID: 201479
  12. High-yield preparation of isolated rat liver parenchymal cells: a biochemical and fine structural study.
    J Cell Biol. 1969 Dec;43(3):506-20 PMID: 4900611
  13. The enzymatic preparation of isolated intact parenchymal cells from rat liver.
    J Cell Biol. 1967 Dec;35(3):675-84 PMID: 4294245
  14. Drug metabolism and effects of carcinogens in cultured hepatic cells.
    Pharmacol Rev. 1979 Sep;31(3):205-28 PMID: 399684
  15. Independent induction of tyrosine aminotransferase activity by dexamethasone and glucagon in isolated rat liver parenchymal cells in suspension and in monolayer culture in serum-free media.
    Exp Cell Res. 1980 Mar;126(1):237-48 PMID: 6102037
  16. Reciprocal modulation of growth and differentiated functions of mature rat hepatocytes in primary culture by cell--cell contact and cell membranes.
    Proc Natl Acad Sci U S A. 1983 Dec;80(23):7229-33 PMID: 6580642
  17. Isolation of rat peritoneal mast cells by centrifugation on density gradients of Percoll.
    J Immunol Methods. 1980;39(1-2):135-45 PMID: 7462642
  18. The repair of DNA damage: recent developments and new insights.
    J Supramol Struct Cell Biochem. 1981;16(1):91-103 PMID: 6271981
  19. Separation of intact rat hepatocytes and rat liver nuclei into ploidy classes by velocity sedimentation at unit gravity.
    Biochim Biophys Acta. 1976 Dec 21;451(2):567-82 PMID: 999869
  20. Partial separation and biochemical characteristics of periportal and perivenous hepatocytes from rat liver.
    Eur J Biochem. 1981 Sep 1;118(3):591-7 PMID: 7028482
  21. Isolation and subfractionation on ficoll gradients of adult rat hepatocytes. Size, morphology, and biochemical characteristics of cell fractions.
    J Cell Biol. 1975 Jul;66(1):1-22 PMID: 167028
  22. Maintenance of microsomal cytochromes b5 and P-450 in primary cultures of parenchymal liver cells on collagen membranes.
    Life Sci. 1976 May 15;18(10):1139-44 PMID: 933703
  23. Preparation of rat liver cells. II. Effects of ions and chelators on tissue dispersion.
    Exp Cell Res. 1973 Jan;76(1):25-30 PMID: 4630105
  24. A sensitive fluorometric assay for the determination of DNA.
    Anal Biochem. 1982 Mar 1;120(2):243-8 PMID: 6283935
  25. Use of Percoll density gradient centrifugation for preparing isolated rat hepatocytes having long-term viability.
    Anal Biochem. 1982 May 1;122(1):119-23 PMID: 6285762
  26. A rapid and simple method to quantitate chemically induced unscheduled DNA synthesis in freshly isolated rat hepatocytes facilitated by DNA retention of membrane filters.
    Mutat Res. 1983 Nov;122(2):177-85 PMID: 6656809
  27. Albumin, fibrinogen and transferrin synthesis in isolated rat hepatocyte suspensions. A model for the study of plasma protein synthesis.
    Biochem J. 1975 Jan;146(1):141-55 PMID: 1147894
  28. Characterisation of hepatocyte sub-populations generated by centrifugal elutriation.
    Eur J Biochem. 1983 Aug 15;134(3):539-45 PMID: 6411467
  29. Primary and long-term culture of adult rat liver epithelial cells.
    Methods Cell Biol. 1976;14:357-64 PMID: 187900
  30. A note on the spectrometric assay of glutamic-oxalacetic transaminase in human blood serum.
    J Clin Invest. 1955 Jan;34(1):131-3 PMID: 13221664
  31. Isolation of rat liver cells containing concanavalin-A receptor sites.
    FEBS Lett. 1974 Oct 1;47(1):11-4 PMID: 4372083
  32. Intact endoplasmic reticulum and albumin synthesis in rat liver cell suspensions.
    Exp Cell Res. 1971 Jul;67(1):27-32 PMID: 4328067
  33. The use of new methods to measure: the effect of diet and inducers of microsomal enzyme synthesis on cytochrome P-450 in liver homogenates, and on metabolism of dimethyl nitrosamine.
    Biochem Pharmacol. 1974 Apr 1;23(7):1173-80 PMID: 4823450
  34. Two phase regulation of tyrosine aminotransferase activity by insulin in primary cultured hepatocytes of adult rats.
    Biochem Biophys Res Commun. 1981 Apr 15;99(3):775-80 PMID: 6113829
  35. A rapid method for the separation of large and small thymocytes from rats and mice.
    J Biochem Biophys Methods. 1979 Dec;1(6):341-7 PMID: 552396
  36. The functional roles of mammalian DNA polymerase.
    Arch Biochem Biophys. 1979 Dec;198(2):386-96 PMID: 117754
  37. Preparation of isolated rat liver cells.
    Methods Cell Biol. 1976;13:29-83 PMID: 177845
Article Info
Journal
In vitro cellular & developmental biology : journal of the Tissue Culture Association
Abbr.
In Vitro Cell Dev Biol
ISSN
0883-8364
DOI
10.1007/BF02623304
Published
1986-04-00
Pages
201-11
Language
English
Region
United States
NLM ID
8506951
Subset
IM
Grants
NCI NIH HHS · CA-017175 · United States
NCI NIH HHS · CA-09135 · United States
NIEHS NIH HHS · ES-01737 · United States
External Links
PubMed source DOI: 10.1007/BF02623304
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