Mitotic Wave Research Articles

Circadian synchronization of hepatocyte proliferation in young rats: the role played by adrenal hormones.

The circadian rhythm of hepatic cell proliferation in rats appears on the 20th day of life, when the hypothalamo-adrenal axis is mature enough for circadian activity to occur. From the 20th day to the 30th day of life, the mitotic rhythm is progressively induced by a reduction in nocturnal values, while diurnal rhythms remain unchanged. Mitotic peaks emerge at 10.00 hours. A labelling index wave occurs 8 hr before the corresponding mitotic wave, with a peak at 02.00 hours and a minimum in the evening, coincidental with the acrophase of plasma corticosterone level (activity phase). Labelled mitoses curves and metaphase accumulation after colchicine injection show that the duration of the S, G2 and M phases remain approximately constant and that the circadian variation is due to a variation in the rate of cells that enter these successive phases. During the synchronization period (from day 20 to 30), the growth fraction decreases progressively. Adrenalectomy at this time is followed by a higher cell proliferation and all rhythms disappear after 2 days. Corticosterone injected before the triggering of the rhythmic activity in 17-day-old rats immediately reduces the labelling index, while the mitotic index is decreased 10 hr later; this delay is equal to the S + G2 duration. The results are discussed. They favour the hypothesis that the circadian variation of corticosterone is responsible for the induction of a circadian variation in developmental cell proliferation by inhibition of the G1-S transition when it is higher in the evening.

Ploidy class-dependent metabolic changes in rat hepatocytes after partial hepatectomy

Glucose-6-phosphate dehydrogenase (G6PDH), succinate dehydrogenase (SDH) activity and the single-stranded RNA (ssRNA) content of isolated hepatocytes of different ploidy classes from adult male rats have been studied after partial hepatectomy using quantitative cytochemical means. The SDH activity and ssRNA content in all classes of hepatocytes are decreased during the first hours after operation followed by an increase above control values. The increase of both SDH activity and ssRNA content is significant only in the mononuclear diploid (MD) cells but not in the hepatocytes of higher ploidy classes and is related with the mitotic wave at 32 h after hepatectomy. After the mitotic wave, the values quickly return to normal levels. The G6PDH activity does not show any significant change in hepatocytes other than MD cells. In MD cells the G6PDH activity is elevated on a highly significant level up to a maximum value of 3.5 times the control value at 48 h after operation. The G6PDH activity in MD cells is returned to normal values within 14 days after operation. It is concluded that: 1. 1. The MD cells show a distinct metabolic behaviour due to their function as stem cells of liver parenchyma and retain at least some of their fetal characteristics. 2. 2. G6PDH activity is not a transformation-linked discriminant for neoplastic metabolism.

Blastoderm development in honey bee embryogenesis as seen in the scanning electron microscope

Summary Based on observations in the scanning electron microscope, we describe the successive changes at the cell surface of fertilized honey bee eggs which, over a period of 20 h, lead to blastoderm formation. Each change starts in the differentiation center located in the anterior egg half and from there spreads as a wave towards both poles. However, one of the waves is heterogeneous: the protrusions or caps of oolemma which start enveloping the preblastoderm nuclei arise in different ways in the anterior third and in the more posterior regions of the egg cell. After the formation of these protrusions, three mitotic waves pass over the peripheral nuclei. The last of these blastemal mitoses results in a two-layered arrangement of peripheral nuclei, each surrounded by an outpocketing of the egg cell basally confluent with the central yolky part. This two-layered state after some time transforms into a single layer of columnar outpocketings which increase in height by extending their borders centripetally ...

Fate-mapping in wild-typeDrosophila melanogaster

The pattern of cell proliferation and cell movements inDrosophila embryogenesis has been analysed with the aim of constructing a blastoderm fate map. Post-blastoderm cell proliferation starts at gastrulation and ends around the stage of germ band shortening. Three mitotic waves affect the embryonic cells according to a constant spatio-temporal pattern. For any of these waves mitotic activity starts at well-defined loci, which have been called mitotic centres. During the first and second mitotic waves all cells undergo mitosis, except for those of the amnioserosa, which do not proliferate at all. The third wave spares most of the ectodermal cells. Neuroblasts, progenitors of epidermal sensilla and germ line cells show their own, different pattern of proliferation.

Migration and division of cleavage nuclei in the gall midge,Wachtliella persicariae

The control of nuclear division and migration was studied in time-lapse films of the multinucleate egg cell of a gall midge by experimental alterations of the mitotic pattern. During each cleavage cycle, a wave of randomly oriented saltations of yolk particles (WROS) is seen to travel through the ooplasm. This wave proved to be an indispensable prerequisite for the accompanying anaphase wave and for the activation of the nuclear migration cytasters: WROS cycles can occur autonomously without cleavage nuclei being present, but there is no anaphase without a WROS passing the dividing nucleus. WROSs and mitotic waves can be inverted, and the WROS cycles and the cleavage cycles can be desynchronized by temperature grandients or by locally impaired gas exchange. If a nucleus is not ready for anaphase when met by a WROS, it will only divide in the course of the next WROS. WROSs thus indicate autonomous anaphase-triggering waves governing the cleavage divisions. Rhythmic ooplasmic movements continue even if the WROSs as well as the nuclear divisions are inhibited by colchinine. The characteristics of the WROSs support the hypothesis that each of them is the visible effect of a wave of calcium release (similar to that established in vertebrate eggs) which acts locally on the microtubular system and may continue even if the WROSs are suppressed. The correlations between a possible calcium release, WROS activity, microtubule disassembly and nuclear cycle are discussed.

Effect on Inhibition of Micromere Segregation on the Mitotic Pattern in the Sea Urchin Embryo: (micromeres/mitotic pattern/sea urchin).

Detergent treatment of sea urchin eggs at the mid 4-cell stage results in prevention of micromere segregation at the fourth cleavage. In these embryos not only the formation of the primary mesenchyme is suppressed, but synchrony of cell division, which is the rule during the first four cleavage cycles, continues for several cycles after the 16-cell stage while the typical mitotic phase wave that sets in after micromere segregation is abolished. These results support the hypothesis that micromeres act as coordinators of the mitotic activity of the embryo.

Visualisation des centres générateurs selon la théorie phyllotaxique de Plantefol, dans le point végétatif du grand frêne (Fraxinus excelsior)

To visualize the generative center of each foliar helix and its oriented progression, cell proliferation was studied in the ash (Fraxinus excelsior L.) shoot apex. The distribution of mitosis was analysed first with the hypothesis of the multiple foliar helices phyllotactic theory and then without any preliminary hypothesis. The cell cycles were asynchronic and mitotic duration was constant. Therefore, the parameters of both the Poisson distribution of mitosis in each apical zone and of mitotic density, evaluated by the nearest-neighbour method or by the rotation of a unit area, were indicative of the cell proliferation level. The axial zone differed from the initiating ring by virtue of its lower mitotic activity. The initiating ring was heterogeneous and involved two parts which were opposite each other and symmetrical with regard to the axis of the apex. In each half, the cell proliferation increased gradually to a generative center with a maximal mitotic activity, then decreased twice as fast. This heterogeneity showed the necessary rotation of a privileged mitotic wave, which progressed step by step in a single direction. The cartography of the cell proliferation intensity was mapped for one sample meristcm.

Developmental analysis of the grandchildless ( gs(1)N26) mutation in Drosophila melanogaster: Abnormal cleavage patterns and defects in pole cell formation

This article describes developmental analysis of gs(1)N26 mutation. gs(1)N26 is a temperature-sensitive maternal-effect mutation affecting the formation of the germ line (Y. Niki and M. Okada, Whlhelm Roux's Arch. Dev. Biol. 190, 1–10, 1981). At 25°C, the cleavage nuclei do not divide synchronously and show various degrees of retarded migration to the posterior region. Blastoderm nuclei show antero-posterior mitotic waves; posterior yolk nuclei also are reduced in number at this stage. Pole cells from only when the cleavage nuclei migrate directly to the posterior pole. In fact, the posterior region of young eggs presents the usual ultrastructural features, and it is also able to participate in the formation of pole cells, as was proven by cytoplasmic transfer experiments. Therefore the defects in blastogenesis, in particular in the formation of pole cells of gs(1)N26 embryos, appear to result from the delayed migration of cleavage nuclei to the posterior pole.

Studies of nuclear and cytoplasmic behaviour during the five mitotic cycles that precede gastrulation in Drosophila embryogenesis.

Using differential interference contrast optics, combined with cinematography, we have studied the morphological changes that the living, syncytial embryo undergoes from stage 10 through 14 of Drosophila embryogenesis, that is just prior to and during formation of the cellular blastoderm. We have supplemented these studies with data collected from fixed, stained, whole embryos. The following information has been obtained. The average duration of nuclear cycles 10, 11, 12 and 13 is about 9, 10, 12 and 21 min, respectively (25 degrees C). In these four cycles, the duration of that portion of the mitotic period that lacks a discrete nuclear envelope is 3, 3, 3 and 5 min, respectively. The length of nuclear cycle 14 varies in a position-specific manner throughout the embryo, the shortest cycles being of 65 min duration. During nuclear cycles 10 through 13, it is commonly observed in living embryos that the syncytial blastoderm nuclei enter (and leave) mitosis in one of two waves that originate nearly simultaneously from the opposite anterior and posterior poles of the embryo, and terminate in its midregion. From our preparations of quick-frozen embryos, we estimate that these mitotic waves take on average about half a minute to travel over the embryonic surface from pole to equator. The yolk nuclei, which remain in the core of the embryo when the rest of the nuclei migrate to the periphery, divide in synchrony with the migrating nuclei at nuclear cycles 8 and 9, and just after the now peripherally located nuclei at nuclear cycle 10. After cycle 10, these yolk nuclei cease dividing and become polyploid. The syncytial embryo has at least three distinct levels of cytoskeletal organization: structured domains of cytoplasm are organized around each blastoderm nucleus; radially directed tracks orient colchicine-sensitive saltatory transport throughout the peripheral cytoplasm; and a long-range organization of the core of the embryo makes possible coherent movements of the large inner yolk mass in concert with each nuclear cycle. This highly organized cytoplasm may be involved in providing positional information for the important process of nuclear determination that is known to occur during these stages.

The cell cycle of epidermal cells during reprogramming in the pupa of Galleria

The epidermal cell cycle of the pupal mesonotum of Galleria was investigated by the determination of mitotic indices, [ 3H]thymidine incorporation and flow-cytophotometric analysis during the first 48 h after pupation. Immediately after the pupal ecdysis nearly all epidermal cells are arrested in G2. Thereafter only a few mitoses occur, leading to a slow increase in the number of G1 nuclei. With the onset of a mitotic wave at a pupal age of 21 h this increase becomes more rapid. On day 2, the cell population reaches a plateau in the number of G1 (resp. G2) cells, reflecting a steady state between mitotic activity and DNA synthesis. A comparison of these cell cycle changes with known data of the time course of reprogramming and ecdysteroid titre leads to the conclusion that there is no causal relationship between DNA synthesis and cellular determination in the sense of a quantal cell cycle, and that DNA synthesis can precede the definite rise in ecdysteroid titre.

Cell cycle inHelianthus tuberosus tuber tissue in relation to dormancy

Observations are presented on the patterns of DNA synthesis and mitotic activity in medullary parenchyma cells excised from tubers ofHelianthus tuberosus in four different periods of dormancy. Dormancy break (activation) was induced byin vitro culture on media added with 2,4-dichlorophenoxyacetic acid. The cell cycle responsein vitro to different combinations of growth substances has also been investigated. The results show that remarkable changes in the timing of the first and second cell cycles and their phases occur with the progression of dormancy. With increasing time after tuber harvest, the following behaviours are observed: (i) a lengthening of the first cell cycle, chiefly due to a lengthening of the G2 phase (G2 is absent at the beginning of dormancy) and an increase in the time interval between the start of thein vitro culture and the onset of the first mitotic wave; (ii) an increased duration of the S phase; (iii) a remarkable reduction in the cell synchrony. These behaviours, as indicated also by their comparison with thein vitro response of the cell cycle to different hormonal treatments, seem to depend on the physiological status of the tubers at the time of explant. It is concluded that the analysis of the cell cycle is an useful tool for understanding some aspects of such a complex physiological situation as dormancy.

Mitotic waves and embryonic pattern formation: No correlation inCallosobruchus (Coleoptera).

In early insect embryogenesis, mitosis (which is not accompanied by cell division) often starts at one or both egg poles and spreads like a wave over the egg. Relationships between these waves and those processes which coordinate spatial cell differentiation have been proposed. One possibility is that the egg region which has a slower mitotic rate may become temporally advanced in differentiation because of its longer interphase periods, so that the egg becomes polarized (Agrell 1962). Alternatively, the mitotic waves might reflect the position of different determined states (Kauffman 1973). We investigated the mitotic waves inCallosobruchus eggs, treated to produce 20% partially reversed segment sequences (double abdomens). In normal eggs, mitotic waves move predominantly from anterior to posterior whereas in treated eggs, the reversed posterior to anterior orientation was predominant. Despite this, we concluded that mitotic waves do not reflect processes involved in the specification of segment position because the reversal of mitotic waves was more than twice as frequent as the reversal of segment sequence and because they occurred in various control experiments in which there was no reversal of segment sequence.

A quantitative analysis of mitotic gradients during early development in Calliphora erythrocephala Meig. (Diptera).

Mitotic waves during superficial cleavage and early gastrulation were analyzed quantitatively in Calliphora. Three consecutive patterns are present: (1) a monotonic anterioposterior mitotic gradient during early superficial cleavage; (2) a double mitotic gradient from the anterior and posterior poles during superficial cleavage, especially toward the end of the period; and (3) more complicated patterns with intermediate mitotic centers during the last superficial cleavage division and during early gastrulation. Mitotic gradients are absent in many eggs during early superficial cleavage, but they then become ubiquitous. The gradients are longitudinal; no transverse component was detected before gastrulation. Anterior and posterior gradient patterns are not mirror images of each other; mitotic activity always starts earlier anteriorly. The gradients are accompanied by a pronounced increase in interphase length. The mitotic gradients are compared with the morphogenetic gradients predicted in a current model for pattern specification in insect eggs.

The epidermal cell cycle during the metamorphosis of Tenebrio molitor L. (Insecta Coleoptera)

The main features of epidermal cell cycle during the metamorphosis of Tenebrio molitor have been investigated by cell density study, [ 3H]thymidine incorporation, and cytophotometry. G 1, S, and M phases are restricted to short periods during each larval stage. The pupal mitotic wave is relatively longer than the larval ones. Outside of each mitotic period, 95 to 100% of the epidermal cells are blocked in G 2 (4C DNA content). The cell density study shows that during the penultimate larval instar, numerous cells bypass the mitotic period without dividing while during the mitotic period of the last larval instar and of the pupal stage, each cell divides once and twice for some cells. After the pupal mitotic period, numerous adult cells are definitely blocked in G 2 but 20 to 30% of cells present in old adults are in G 1. Adult epidermal oenocytes differentiate during the last larval mitotic period, becoming 8 to 32C. From these results, it can be concluded that the G 2 block and the release of G 2-blocked cells into M are two critical events of each molting cycle. A possible hormonal control of these events is discussed.

Etude autoradiographique de la proliferation et de la migration cellulaires au cours de la croissance larvaire de l'appareil visuel chez Aeshna cyanea Müll. (odonata : Aeshnidea)

Sites of cell proliferation in the ocular apparatus of Aeshna cyanea have been studied by an autoradiographic method ( 3H-thymidine). This technique was used to locate at the compound eye and optic lobe levels the zones of cellular multiplication and the migration of newly formed cells. In addition, the effect of the differential growth of the compound eye on the development of the optical lobe was studied. In the compound eye, two distinct levels of cell proliferation were revealed: the first one was the ocular budding zone itself, i.e. the ocular edge adjacent to the cephalic epidermis. A second mitotic focus, where the cells gather into ommatidial columns, was demonstrated. Thus, two closely related successive mitotic waves occur in the eye of this heterometabolous insect, whereas in holometabolous insects these two phases are much more distinct. Migration of ganglionic cells from the neuroblast masses, where they are formed, to the optical ganglions, where they differentiate, was also followed by this method. In addition, a differential growth of optic lobes was revealed: at the end of the nymphal life, the development was much greater in the posterior area of the optic lobes than in the anterior, although the numbers of cells produced in the neuroblast masses were, at the beginning, identical in both areas. This observation has been interpreted in the light of the experimental results.

INTERFERENCE OF SEX‐RELATED FACTORS IN THE RESPONSE OF LIVER CELLS TO EXPERIMENTAL MITOTIC STIMULI

Stimulation of liver cell multiplication was obtained under two different experimental conditions. (1) A single injection of casein solution resulted in (a) an identical synchronized mitotic wave response in 10-day old male and female rats and (b) a significantly lower response in adult male rats compared to females, a difference which was reduced by castration of males at birth but essentially maintained if animals were operated when 10 days old. (2) Partial hepatectomy shortly after puberty resulted in active hepatocyte multiplication occurring 3 hr earlier in females were ovariectomized at birth and significantly reduced when they were spayed at a later age. Hepatocytes of castrated females entered actively into S phase 2 hr later than the sham-operated controls. Unilateral ovariectomy on the other hand indicated that during compensatory and/or hypercompensatory activity of the single ovary there was a maximum difference between the male and female rate of [3H]thymidine uptake in liver nuclei 20 hr after hepatectomy. A further kinetic study (t = 25, 30,40, 65, 90 hr) indicated no significant sex-related difference in the number of S phases per 10,000 cells. The DNA content of regenerating versus control livers was comparable in both sexes at t = 22 and 90 hr but higher in females at t = 40 and 65 hr. A possible early postnatal interference of certain hormonal mechanisms in the receptivity to mitotic stimuli is postulated and discussed.

Phase response versus positive and negative division delay in animal cells

The time of median cell division in V79 Chinese hamster cells following high serum pulses was determined for two synchronous cell generations following mitotic selection. Differences in cell cycle time for each pair of pulse and control cultures were computed and plotted as a function of time of serum pulse. This phase response curve for hamster cells with an 8.5 h cell cycle shows a characteristic biphasic pattern. Beginning 0.5 h after mitotic selection, pulses with serum produce delays in the midpoint of the subsequent mitotic waves. Delay is maximum at 1.5 h. Delays give way abruptly to advances at 2.5 h and the amount of advance then decreases as pulses are given between 3 and 5 h into the cycle. At 5 h decreasing advances become delays, with increasing delays due to serum pulses occurring between 5 and 6 h. Delays again give way abruptly to advances at 6 h and again the amount of advance decreases through the late portion of the cycle. Pulses very late in the cycle appear to generate phase delays. This biphasic response to serum is interpreted as an expression of an underlying time-keeping oscillator whose period is nominally of 4 h duration.

Fibrinolysis induced by rat glioma cells: A mitosis-triggered process?

Experimentally induced brain glioma cell lines were studied in cell cultures for their fibrinolytic activity; simultaneously the mitotic index was determined. It was found that the most intense fibrinolytic activity of glioma cells coincides with mitotic waves. After 68 hr incubation of SD/G1 cells on a [ 125I]fibrin layer, [ 125I]fibrin digestion products were examined by gel chromatography. Hydroxy-urea and actinomycin D were used as inhibitors in cell cultures, investigating their effect on the expression of fibrinolysis. These studies reveal that [ 125I]fibrin digestion products are mainly low molecular weight (< 12,000 daltons) compounds and that fibrinolysis is partially inhibited by the drugs added.

Ion exchange analysis of porphyrins in urine

Intubation of ethylene dibromide (7.5 to 10 mg. per 100 gm. of body weight) into the stomach of nonfasted Wistar rats induces DNA synthesis and cell division in the liver. The peak of DNA synthesis, as measured by 3H-methyl thymidine incorporation, was attained at or shortly after 24 hours. The mitotic waves measured with the aid of colchicine occurred at 24 to 30 hours and 48 to 54 hours after ethylene dibromide treatment. Approximately 16 per cent of liver cells entered mitosis. The increase in DNA synthesis was confirmed by autoradiography. The stimulation of liver cell mitosis occurs in nonfasted animals without apparent cell necrosis. Thus, ethylene dibromide appears to be an effective mitogen for liver under some experimental conditions. The possible relationship between the mitogenic effect at a low dosage range and necrosis at a higher range is discussed.

G2 sub-population in mouse liver induced into mitosis by lead acetate.

A single intracardiac dose of lead acetate (40 microgram lead/g body weight) induced a 25-fold increase in mitosis of mouse hepatocytes 5 hr after injection, as determined by autoradiography. The prompt appearance of a mitotic wave and the relatively large number of mitoses suggest that the mitotic cells were derived from a hepatocyte sub-population arrested in the G2 phase. The injection of lead also stimulated a small increase in labeled hepatocytes within 6 hr. Analysis of grain counts gave no evidence for unscheduled DNA synthesis. The incremental labeled cells may have originated from a small fraction of the G1 population that was ready to enter the S phase without the usual pre-synthetic delay.