Cell size, cell cycle and transition probability in mouse fibroblasts.
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The patterns of variation of wing cell size and number were studied under developmental conditions leading to a biphasic relationship between life span and growth rate while duration of development remained constant (development on an agar-only medium with a varying added yeast amount, constant temperature (25 degrees C) and constant larval density). Across the yeast range, a 125% increase of body weight was accompanied by a roughly 30% increase in the wing linear dimensions, wing cell size and wing cell number while estimated duration of cell division and its reciprocal mitotic division rate remained constant. Furthermore, cell size (but not cell number) varied with growth rate in a similar biphasic pattern to that observed for life span. Finally, from a simultaneous examination of the covariation patterns of life span, growth rate, cell size and cell number with decreasing yeast amount, it became apparent that there was a "critical" yeast amount, approximately 125 mg/120 eggs, below which: (a) cell number abruptly started to decrease linearly from a roughly constant value; (b) the rate of the slow decrease of cell size now tripled and that of growth rate increased even more; and (c) life span which, in the upper yeast range, increased slowly with decreasing yeast, apparently reached a maximum at the critical yeast level and decreased three times faster below that level. These data taken together suggest that: (i) the decrease of all parameters (including life span) below the critical yeast level results from a presumably suboptimal or disturbed development because of and in proportion to the lack of nutrients and (ii) the increase of life span with decreasing yeast amount above the critical yeast level has not been definitely explained but some possibilities are suggested such as changes in subcellular organelle numbers, size and/or functional properties, or other changes due to a phenomenon equivalent to food restriction in rats, probably without changes in overall metabolic rate of the flies.
Cell size, cell type and calcitonin gene-related peptide (CGRP)-like immunoreactivity were compared between cutaneous and splanchnic sensory neurons by means of a combination of fluorescent tracer and immunohistochemistry. Nineteen percent of cutaneous sensory neurons and 88% of splanchnic sensory neurons were shown to contain CGRP. The former cells were larger than the latter ones, which was also confirmed by the finding that about a half of the former cells contained 200 kDa subunit of neurofilament protein, while only 8% of the latter ones were positively stained. These findings suggest that most of the visceral CGRP-IR sensory neurons are small type B.
The variability of (1) surface area projection (size) at which cells terminate DNA replication, (2) the area at which they initiate mitosis, (3) the area at which they divide, (4) the duration of G2, and (5) the duration of G2 plus mitosis (in fact, prophase + metaphase + anaphase) has been estimated in steady-state cell populations of Allium cepa root meristems. The coefficient of variation of cell area at termination of DNA synthesis was found to be 14% while the coefficient of variation of cell area at mitosis initiation was 13%. As there is also a substantial variability of G2 (the coefficient of variation was estimated to be 38%), the combination of these data indicates that cell size regulation of G2 contributes to maintaining cell size variability (and therefore DNA concentration) within certain limits. Mitosis also varies but less than G2 (the coefficient of variation of G2 + mitosis was found to be 31%). As the coefficient of variation of cell area at division (14%) is hardly larger than the coefficient of variation of cell area at initiation of mitosis, it can be suggested that coordination between cell size and mitosis duration helps to avoid a significant increase in the variability of cell size at the end of the division cycle.
The strength of the long-range electrostatic repulsion forces on HeLa cells is measured by agglutinative titration using low molecular weight polylysine (M.W. 11,000). Repulsion forces, found to be present on the smaller HeLa cells from density-inhibited suspension cultures, are weakened by incubation of the cells in hypotonic NaCl solutions. Repulsion forces, found to be absent on the larger cells from fast growing cultures, can be induced on these cells by incubation in hypertonic NaCl solutions. Both effects of anisotonicity are reversible, and disappear on restoration of the medium to normal tonicity. Induction of repulsion forces on fast growing cells is prevented by previous treatment of the cells with neuraminidase. Neuraminidase also abolishes repulsion on density-inhibited cells. It is proposed that alterations of the cell size, produced by anisotonicity or occurring during growth in isotonic suspension medium, affect mutual cell adhesiveness by modifying the strength of the repulsion forces generated by cell surface sialic acids.
Pituitary cells increase their numbers more than 3-fold during the 1st 10 days of life while maintaining the same cell size ratios. In the 25-day-old animal, the rate of cell division slows and there is a slight increase in the number of large cells. An increase in adult weight is attributed to hyperplasia and a shift to a population of larger cells.
Toad epidermis is a suitable model for studies on tissue homeostasis because cell pool size, influx into and efflux from the cell pool can be easily determined. The cell pool size was obtained by cell counting on photomicrographs, the influx (cell birth rate) was assessed by the metaphase-arrest technique, and the efflux (cell loss by moulting) assessed by counting the number of cells in the corneal layer and recording of intermoult periods. The importance of the methods for assessing these parameters per square unit of skin surface is emphasized. These parameters were studied in eight groups of ten adult male toads sacrificed at various hours of the day. There were minor variations in the cell birth rate, fluctuating around a mean of 26 cells/mm2/hr (obtained at the metaphase collection period from 11.00-14.00 hours). By summation of the cell productions during the eight metaphase collection periods of 3 hr, and extrapolation to an intermoult period (time between two moults), a calculated cell production of about 6340 cells/mm2 in 10.3 days was obtained, whereas the cell loss at each moult was only 2370 cells/mm2. Thus the cell production rate exceeds the rate of cell loss through moults by a factor of 2.7 Arguments are presented that the 'surplus' of cells produced cannot be permanently accommodated within the living epidermis. Consequently a cell deletion rate beyond that by moulting of about 4000 cells/mm2 in 10.3 days or 16 cells/mm2/hr can be calculated. These results are discussed in relation to current concepts of tissue homeostatic mechanism(s). The results are consistent with the hypothesis that controlled cell deletion may be a tissue homeostatic mechanism complementary to controlled cell divisions.
It has previously been shown that in toad epidermis the cell birth rate (Kb) exceeds the rate of cell loss through moulting (Kd) and that the 'surplus' of cells seems to be removed in a controlled manner. Assuming that the epidermis is non-expanding, a Kb/Kd ratio greater than 1 indicates that cell deletion additional to desquamation takes place. In normal toads this ratio is 2-3. Following implantation of hydrocortisone pellets into intact toads (release rate, 18 micrograms/g toad/d), the Kb/Kd ratio, over a period of 14 d of hormone treatment, had increased to about 7, due mainly to an increased Kb and to a lesser extent to a decreased Kd. No change in the epidermal cell pool size had taken place. It was previously shown that, following removal of the pars distalis of the pituitary gland, the Kb/Kd ratio decreased with time, due to a decreasing Kb and an increasing Kd, eventually leading to a decreased epidermal cell pool size. In this paper it is shown that, in pars distalisectomized toads with hydrocortisone pellets implanted, the Kb/Kd ratio is restored to control levels by a restoration of the Kb as well as the Kd. The results differ from those of previous studies in which ACTH or adrenocorticosteroids were administered discontinuously (by injection). Thus, by experimental manipulation, different Kb/Kd ratios can be obtained: low (less than 1, pars distalis ablation), medium (2-3, normal toads) and high (7, hydrocortisone implantation). The potentiality of this unique situation in analysing the important question of how the 'surplus' cells are deleted is discussed.
Following removal of the pars distalis of the pituitary gland in toads, epidermal efflux from the stratum corneum recruitment cell pool (i.e. production of corneal layers) is greatly increased. In this investigation the cell birth rate is studied by means of the metaphase arrest technique, as a function of time after pars distalis ablation. The method allows assessment of the total cell production over 14 days after the operation, to be compared with the total efflux and changes in the epidermal cell pool size. Whereas in intact toads the rate of cell production exceeds that of cell loss by moulting by a factor of 2.7, the 'surplus' of cells neither being used for formation of corneal layers nor permanently accommodated within the living epidermis, a 'balance sheet' of efflux and influx indicates that following pars distalis ablation all cells produced are also used for the (excessive) formation of corneal cell layers. The observations lend further support to the hypothesis that controlled cell deletion is a tissue homeostatic mechanism complementary to controlled cell divisions.
HeLa cells harvested from density-inhibited or fast growing suspension cultures, were incubated in NaCl solutions of different tonicity. Cell size enlargement produced by hypotonicity is accompanied by an increased sedimentation rate of the density-inhibited cells, whereas no appreciable change is observed in the sedimentation rate of fast growing cells. Hypotonicity also has no effect on the sedimentation rate of density-inhibited cells which previously had been treated with neuraminidase or trypsin. It is shown that the effect of hypotonicity on density-inhibited cells cannot be ascribed to release of cell surface sialic acids during hypotonic incubation. Several arguments are presented which indicate that the changes in sedimentation rate, as measured in the rotating suspension system, are not the direct consequence of the alterations in cell size, but rather must be attributed to differences in intercellular adhesiveness resulting from the size alterations. Analogous changes in intercellular adhesiveness and cell size are shown to occur during growth in isotonic suspension culture. The results can be explained by assuming that changes in cell size affect the intercellular adhesiveness by modifying the extent to which cell surface sialic acids counteract adhesion.
The development of tissues and organs in the post-embryogenic period has commonly been thought to be by two main consecutive processes: the multiplication of cells followed by a growth in their size. The capacity for recovery following growth restriction has been considered to depend on whether restriction occurred during the phase of cell multiplication, in which case catch-up was permanently inhibited; or later, in which case recovery could occur. This model is now no longer valid for a number of reasons, the main one being that the original assumptions about the sequence of developmental processes have been shown to be false. There certainly seems to be an early period of vulnerability during which restriction is not followed by catch-up, but this cannot now be related to the multiplication of cells.
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