Ploidy-dependent growth and binucleation in cultured rat hepatocytes.
The proliferative activity of rat hepatocytes, cultured in the presence of epidermal growth factor (EGF) and insulin, was examined by immunostaining of S-phase cells labeled with bromodeoxyuridine (BrdU) in culture. Proliferation rates of the different hepatocellular ploidy and nuclearity classes were measured by fluorescence image cytometry or by microscope counting of immunostained cells. Effects of EGF and insulin were largely additive, the binuclear cells being more growth factor-dependent (showing less growth in the absence of factors) than the mononuclear cells. A serial warm-washing procedure was used to remove excess BrdU from the culture medium, allowing the study of hepatocellular binucleation by a BrdU pulse-chase approach. A high rate of binucleation was detected (50%, possibly suggesting a quantal mechanism), indicating that the hormones induce a binucleating (polyploidizing) type of growth similar to that normally observed in the liver of growing rats. The highest proliferative activity (labeling index) in the hepatocyte cultures was found among the diploid cells, independent of the degree of mitogenic stimulation. The labeling index was inversely correlated with ploidy, suggesting that the ability of hepatocytes to proliferate decreases with increasing polyploidization.
Mossin
L
L
Department of Tissue Culture, Norwegian Radium Hospital, Montebello, Oslo.
Blankson
H
H
Huitfeldt
H
H
Seglen
P O
PO
D000818:Animals; D001973:Bromodeoxyuridine / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002467:Cell Nucleus / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D004815:Epidermal Growth Factor / Q000494:pharmacology; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D007328:Insulin / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D008297:Male; D008856:Microscopy, Fluorescence; D011003:Ploidies; D051381:Rats; D011921:Rats, Inbred WKY
D007328:Insulin; D004815:Epidermal Growth Factor; D001973:Bromodeoxyuridine