The influence of fixation on the relative amount of cytoplasmic ribosomes in mouse epidermal basal keratinocytes. A morphometric study of so-called "dark cells" and their putative role in epidermal carcinogenesis.
The nature and significance of so-called dark keratinocytes in the epidermis during chemical carcinogenesis is still a matter of concern and debate. Based on ultrastructural observations it has been suggested that dark cells most often are shrunken cells. Reports on skin carcinogenesis, however, claim that dark cells are a sign of ongoing tumor promotion and represent those stem cells in the epidermis from which the tumors originate. It is therefore important to find out whether these cells are simply injured and shrunken cells, or vital cells of great importance for carcinogenesis. Dark cells are assumed to be rich in ribosomes. There is evidence, however, that the observed number of dark cells is highly dependent on tissue fixation. In the present ultrastructural study, morphometric methods were used to compare the effects of two different fixation procedures on the amount of cytoplasmic ribosomes in dark cells from both untreated and carcinogen-treated hairless mouse epidermis. The results show that the ultrastructural features of both dark and clear cells vary considerably with different fixation procedures. In acetone-treated controls typical dark cells are only observed when the fixative has a lower osmotic activity than the plasma. With iso-osmolal fixation typical dark cells are not observed. After an abortive two-stage carcinogenesis treatment, in which a single application of 9,10-dimethyl-1,2-benzanthracene (DMBA) in acetone was followed by a single application of 12-O-tetradecanoyl-13-acetate (TPA) in acetone, signs of cell injury could be found after both fixation procedures. With DMBA/TPA and hypo-osmolal fixation the number of dark cells seemed to increase, whereas only signs of cell injury with occurrence of some heavily altered "clear cells" dominated the picture with iso-osmolal fixation. Morphometry showed that both the numerical and the volumetric densities of cytoplasmic ribosomes in basal keratinocytes varied most significantly with the fixation procedure used. The cytoplasmic volumes did not vary in a way that could explain these differences. One might therefore assume that the number of ribosomes depends on the fixative. Large swelling artifacts occurred when a fixative with low osmotic activity was used, leading to compression of neighboring cells. Hence, an increased ribosomal density reported previously in dark cells is probably related to such cell volume artifacts and does not reflect an actually increased quantity of ribosomes.(ABSTRACT TRUNCATED AT 400 WORDS)
Glasø
M
M
Laboratory for Carcinogenesis and Cell Kinetics, Oslo University, National Hospital, Norway.
Iversen
O H
OH
Hovig
T
T
D015127:9,10-Dimethyl-1,2-benzanthracene; D000818:Animals; D003593:Cytoplasm / Q000187:drug effects*; D000078404:Epidermal Cells; D004817:Epidermis / Q000187:drug effects* / Q000648:ultrastructure; D005260:Female; D005404:Fixatives; D007633:Keratins; D008297:Male; D051379:Mice; D008812:Mice, Hairless; D008854:Microscopy, Electron; D012270:Ribosomes / Q000187:drug effects* / Q000648:ultrastructure; D012878:Skin Neoplasms / Q000139:chemically induced* / Q000473:pathology; D013755:Tetradecanoylphorbol Acetate
D005404:Fixatives; D015127:9,10-Dimethyl-1,2-benzanthracene; D007633:Keratins; D013755:Tetradecanoylphorbol Acetate