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1.
The cell kinetics of the murine JB-1 ascites tumour have been investigated on days 4, 7 and 10 after transplantation of 2·5 × 106 cells. The experimental data, growth curve, percentage of labelled mitoses curves, continuous labelling curves and cytophotometric determination of single-cell DNA content have been analysed by means of a mathematical model for the cell kinetics. The important result was the existence of 8% non-cycling cells with G2 DNA content in the 10-day tumour, while only 0·2 and 0% were observed in the 7- and 4-day tumours, respectively. The doubling times determined from the growth curve were 22·8, 70 and 240 hr, respectively, in the 4-, 7- and 10-day tumours. Growth fractions of 76, 67 and 44% were calculated for the same tumour ages. The mean cell cycle time increased from 14 to 44 hr from day 4 to 7 due to a proportional increase in the mean transit time of all phases in the cell cycle. In the 10-day tumour, the mean cell cycle changed to 41 hr and T G1 decreased to 0·5 hr. The cell production rate was 4·3%/hr in the 4-day tumour, 1·2%/hr in the 7-day tumour and 1·0%/hr in the 10-day tumour. The cell loss rates in the same tumours were 1·3, 0·2 and 0·7%/hr, respectively. The analysis made it probable that the mode of cell loss was an age-specific elimination of non-cycling cells with postmitotic DNA content.  相似文献   

2.
A detailed study of the cellular proliferation kinetics in interfollicular plucked and unplucked mouse skin has been made in Swiss albino mice, using tritiated thymidine autoradiography. Diurnal variations in mitotic and labelling indices were demonstrated in both systems.
The mean cell cycle times for unplucked and plucked skin were estimated by four different methods and found to be 100 ± 10 and 47 ± 3 hr respectively. Most of the difference was due to the shortening of G1 phase after plucking. Repeated labelling at intervals shorter than the DNA synthesis times resulted in all the basal layer cells becoming labelled, so that the growth fraction was unity, in unplucked and plucked skin.
A well-defined second wave of labelled mitoses was seen at about 100 hr after labelling the unplucked (i.e. normal) mouse skin.
A double labelling technique using 14C-TdR and 3H-TdR with a single layer of emulsion gave reasonable values for the duration of the DNA synthesis phase.  相似文献   

3.
Abstract. In order to characterize the growth pattern of the human promyelocytic leukaemia cell line HL60, its kinetic parameters were studied. The doubling time was calculated from serial cell counts, the duration of the various cell cycle phases from the analysis of the labelled mitoses curve, and quiescent population from continuous labelling experiments. Proliferation in culture was exponential up to a saturation density of about 3.0 × 106 cells/ml, with a doubling time of 34.0 hr. The cell cycle duration was 24.3 ± 4.1 hr (SD), and that of the cell cycle phases was: G1, 3.8 ± 2.2 hr; S, 15.1 ± 3 hr; and G2, 5.4 ± 1.2 hr. The growth fraction was 0.85, and cell loss was restricted to the quiescent cells. The HL60 cell line, with fully characterized kinetics, provides a useful tool for the in vitro study of substances which may affect human leukaemic myelopoietic proliferation.  相似文献   

4.
A double isotope DNA labelling method has been used to determine the duration of DNA synthesis (S) in bone marrow lymphoid cells classified by their nuclear diameters in smears. Incorporation of 3H-thymidine was confined almost entirely to marrow lymphoid cells of 8·0-15·0 μm nuclear diameter (large lymphoid cells). After exposure to 3H-thymidine in vivo and 14C-thymidine 40-104 min later in vitro , the proportion of cells labelled with 3H alone to those labelled with 14C(±3H) in radioautographic smears, plotted against time indicated the efflux from S per hour. Collectively, 28·3 ± 1·1% of all large lymphoid cells were in S and the efflux from S was 15·1% per hour. With decreasing cell size (nuclear diameter) the efflux fell progressively from 28·3% per hour (11·0 μm) to 9·2% per hour (8·0-8·9 μm) and the proportion of cells in S declined from 54·9 ± 2·3% to 14·8 ± 1·6%. Influx into S, measured in vitro by reversing the sequence of isotopes, closely resembled the corresponding efflux values in vivo relative to cell size. Most DNA synthesizing marrow large lymphoid cells belonged to a subgroup with deeply basophilic cytoplasm. The results demonstrate that basophilic large lymphoid cells in the marrow are actively proliferating and have a mean S phase duration of 6·6 hr. The largest marrow lymphoid cells (11·0 μm) proliferate most rapidly (S phase, 3·5 hr; maximum cell cycle time, 6·4 hr) while S duration is prolonged progressively to 10·9 hr for the smaller cells (8·0-8·9 μm).  相似文献   

5.
A short-term in vitro method was employed to study the Mitomycin-C sensitivity of normal mouse bone marrow CFU without triggering the G0-phase cells into the proliferative cycle. Comparison was made of the toxicities of the drug against cells in different phases of the cell cycle including G0. Mitomycin-c killed CFU both in and out of the S-phase. No significant difference could be found between its toxicities against normal and proliferating CFU; along the exponential part of the survival curve 1·6 μg/ml concentration of the drug reduced survival to 10%. Although in the normal bone marrow only a few CFU are in the S-phase and are killed by the agent, presence of the sensitive G0 cells produce a significant amount of non-S-phase mortality. Among the proliferating CFU population the non-S-phase lethality is less due to the absence of G0 cells. About 75% of the S-phase cells are killed after incubation with 1 μg/ml drug; outside the S-phase, the lethality is about 40–50%. The studies indicate that the G0 cells which are situated near the G1-S boundary are almost as sensitive to the drug as other non-S-phase cells like G1 cells. The clinical significance of the findings is discussed.  相似文献   

6.
Cytophotometric determination of single-cell DNA after repeated 3H-thymidine labelling of the JB-1 ascites tumour in the plateau phase of growth showed a massive accumulation of unlabelled cells with both G1 and G2 content. Autoradiography combined with cytophotometry or colcemid block demonstrated that some of these unlabelled cells were rapidly triggered into the cell cycle when plateau tumours were transferred to new hosts. This indicated that tumour cells may be held up in non-cycling stages corresponding to both the G1 and the G2 phase of the cell cycle.  相似文献   

7.
Actinomycin D (0.5 μg/ml) did not prevent M stage cells from entering G1 stage, but blocked their progress from G1 to S stage. The position of the block was approximately 1.4 hr before S stage or just after the beginning of G1 stage. Actinomycin D in this concentration also significantly depressed uridine-3H uptake into G1 stage cells, but did not suppress leucine-3H uptake by M and G1 cells. This suggests that some proteins may be synthesized in M and G1 stage cells by messenger RNA left over from the previous cell cycle. However, entry of G1 cells into S stage would require synthesis of new messenger RNA near the beginning of G1 stage. Puromycin (10 μg/ml) did not prevent M cells from entering G1 stage, but blocked their progress from G1 to S stage. The site of blockage was about 0.7 hr before S stage or in the first two-third of G1 stage. This might be the site where the cells synthesize new G1 proteins necessary for entry to S stage.
Comparison of sensitivities of G1 and G2 stages to the two antibiotics reveals that the puromycin sensitivity of G1 cells was similar to that of G2 cells, but the actinomycin D sensitivity of G1 was greater than that of G2 cells.  相似文献   

8.
Abstract: Rat glioma mouse neuroblastoma hybrid neurotumor cells (NG108-15), synchronized by amino acid deprivation, showed a cell-cycle-dependent peak of activity of a ganglioside N-acetylgalactosaminyl transferase 14-24 h following release from the cell cycle block (S/G2 phase). Maximal expression of two typical lysosomal hydrolases, N-acetyl-β-hexosaminidase and β-galactosidase, occurred between 18 and 21 h following release (S phase), declining to G1 phase levels during the peak of N-acetylgalactosamine (GalNAc) transferase activity. In addition, glycosyltransferase activity in G2 phase cells showed an increase in apparent Vmax (suggesting the presence of more enzyme/mg of cell protein) and apparent binding affinity for uridine diphosphate N-acetylgalactosamine (UDP-GalNAc) (32 versus 14 M) when compared to transferase activity in the G1 phase. However, the opioid peptide enkephalin [D-Ala2, o-Leu5], which inhibits ganglioside GalNAc transferase activity in unsynchronized NG108-15 cultures, was much more inhibitory in whole cells 8 h after release from the cell cycle block (G1 phase) than in cells 20 h after release (G, phase), with 50% inhibition occurring at 2 ± 10-9M and 2 ± 10-7M, respectively. These results suggest that the GalNAc transferase activity is regulated in more than one way during the cell cycle, since both Vmax and Km changes are observed, and that the cyclic AMP-dependent mechanism by which opiates reduce transferase activity is receptor mediated and cell cycle dependent.  相似文献   

9.
Abstract. The durations of the cell cycle and its component phases have been determined for the basal layer of the epidermis of the skin from the upper surface of the hind foot of the rat using single pulse [3H]-thymidine labelling and the percent labelled mitosis (PLM) technique. Rats of three age groups were used, namely 7, 14 and 52 weeks. The duration of DNA synthesis (Ts) and the G2 plus M phase (Tg2± m) were comparable in 7-week and 52-week-old rats ( P > 0–1). The major difference between 7-week and 52-week-old rats was in the duration of the G1 phase (Tg1). In 7-week-old rats Tg1 was 15.0 ± 0.8 h and in 52-week-old rats Tg1 was 31.2 ± 3.5 h. A consequence of this variation was that the overall duration of the cell cycle was longer in 52-week-old rats (53.9 ± 5.3 h) than in 7-week-old rats (30.1 ± 1.3 h).
Difficulties were found in fitting a simple curve to the PLM data for 14-week-old rats. This suggests that the proliferative cell population of the epidermis of rats of this age group may be heterogeneous. A satisfactory fit to the data was obtained using a computer model which assumed that the proliferative population of the epidermis of 14-week-old rats was a mixture of cells with cell cycle parameters the same as those of the 7-week and the 52-week-old rats. These two sub-populations of relatively slowly and rapidly proliferating cells were present in the ratio of 2:1.  相似文献   

10.
THE KINETICS OF GRANULOSA CELLS IN DEVELOPING FOLLICLES IN THE MOUSE OVARY   总被引:1,自引:0,他引:1  
This investigation describes the kinetics of the granulosa cells in medium-sized follicles type 3b, 4 and 5a in ovaries of 28-day-old Bagg mice. the method of labelling with 3H-thymidine followed by high resolution autoradiography is used in the experimental work, which consist of determining percentage labelled mitosis (PLM-) and continuous labelling (CL-) curves. In order to analyse the data by computer two alternative hypotheses A and B are set up. Both include the assumptions of no cell loss, exponential growth and a resting compartment Q. In hypothesis A cells from Q re-enter the mitotic cycle via the normal DNA-synthesis compartment Sp. Hypothesis B includes beside compartment Sp a special DNA-synthesis compartment Sq where only cells from Q are synthesizing DNA, and these cells re-enter the mitotic cycle via the G2 compartment. the mean transit time in Sq is considered to be longer than the mean transit time in Sq. On the basis of the hypothesis mathematical expressions for the PLM- and CL-curves are obtained, and by means of a computer the theoretical curves are fitted to the experimental values: thereby all relevant cell kinetical parameters are estimated. Hypothesis B seems to give the best fit between the theoretical and experimental curves. the estimated parameters are: mean cycle times, μc= (56.1 hr, 56.1 hr and 22.3 hr for type 3b, 4 and 5a respectively), doubling times, T D= (96.4 hr, 118.6 hr and 59.1 hr) and the proportion of cells in Q, p Q = (0.60, 0.71 and 0.69).  相似文献   

11.
The duration of the mitotic cycle and of its components was analysed for each of the six successive generations of differentiating spermatogonia (A1, A2, A3, A4, intermediate and B), using radioautographed whole mounts of seminiferous tubules from testes of adult Sprague-Dawley rats. Cell cycles were determined from two successive waves of per cent labeled metaphases obtained during the period of 81 hr after a single dose of 3H-thymidine. Except for the A1 spermatogonia, all spermatogonial types (A2 to B) had similar cell cycle durations of 41-42.5 hr and comparable pre-DNA synthesis phases (G1) of 11-13 hr. Although the combined duration of DNA synthesis (S) and the post-synthesis phase (G2) remained identical for all the cell types including A1, there was a progressive lengthening of the S period at the expense of G2 during the process of spermatogonial maturation. This change was most marked during the transition from A1 to A3 spermatogonia when the S period increased from 14 hr to 21 hr, and the G2 phase shortened from 13 hr to 7.5 hr. This feature seems to be unique to germ cells and may be associated with an increasing amount of heterochromatin in the nucleus. Excluding the development of type A1 cells, the entire process of spermatogonial maturation lasted for 208 hr. Combined data on cell cycle times indicated that every 313 hr or 13 days, a new sequence of spermatogonial differentiation was initiated by the A1 cells. This was equivalent to the duration of one 'cycle' of the seminiferous epithelium as measured by other techniques.  相似文献   

12.
Abstract. Multivariate analysis of the expression of cyclin proteins and DNA content has opened new possibilities for the study of the cell cycle. By virtue of their cell cycle phase specificity, the expression of cyclins may serve, in addition to DNA content, as another marker of a cell's position in the cycle, and provide information about the proliferative potential of cell populations. Several applications of the methodology based on bivariate analysis of DNA content v . expression of B, E and D type cyclins are reviewed: 1 expression of cyclins by individual cells during their progression through the cycle can be studied, using exponentially growing cells without the necessity of cell synchronization or other perturbations of the cycle; 2 cells having the same DNA content but residing in different phases of the cycle (e.g. G2 diploid v. G1 tetraploid) can be distinguished; 3 cell transition from G0 to G1 and progression through G1 (e.g. mitogen stimulated lymphocytes) can be assayed; 4 the population of proliferating cells can be distinguished from noncycling cells based on dual cell labelling with a G1 and G2 cyclin antibody; 5 cyclin restriction points can serve as additional cell cycle landmarks to map the point of action of antitumour drugs; 6 unscheduled expression of cyclins (e.g. the presence of cyclin B1 during G1 and S) can be detected in several tumour transformed cell lines, possibly indicating disregulation of the machmery of cell cycle progression. The last finding 6 is of special importance, because such disregulation may be of prognostic consequence in human tumours.  相似文献   

13.
Experiments in mice on the fraction of haemopoietic stem cells in S-phase after irradiation indicated that a large fraction of the cells resting in G0 will enter S-phase after a very short interval of time.
After excluding alternative explanations it must be concluded that cells in G0 have completed all preparations for going into S-phase or, in other words, that the localization of these G0 cells in relation to other phases of the cell cycle must be between G1 and S-phase.  相似文献   

14.
Abstract. In Snell dwarf mice, the influence of short-term treatment with human growth hormone (hGH) or thyroxine on the proliferative and sulphation activity of the proximal tibial growth plate was studied. By autoradiographic methods, the [3H]methylthymidine incorporation after a single injection was measured, after 2 hr incorporation time. the labelling index was calculated and the number of labelled mitoses was counted. In addition, the distribution of the labelled nuclei over the proliferating and degenerating zones was determined by continuous labelling for 25 and 73 hr.
In untreated dwarf mice after [3H]-methylthymidine administration, the number of labelled nuclei in the growth plate is low. Labelling occurs, as expected, mainly in the cells of the proliferative zones. the number of labelled nuclei in control dwarf mice was similar after 25 and 73 hr continuous labelling. This suggests that many cells are in a resting Go or prolonged G1 phase. Both hGH and T4 treatment induce a significant increase of the number of labelled nuclei per growth plate and of the number of mitoses. Since hormonal treatment induces a small number of mitoses after 2 hr incorporation of the label, the minimal G2 phase of the cell cycle is less than 2 hr. In addition, treatment with hGH and T4 stimulates chondrocytes in the zone of proliferative and hypertrophic cells to actively incorporate [35S]-sulphate.  相似文献   

15.
Abstract. To analyse the putative role of methylation of cytosine residues in the nuclear DNA as a regulatory step during cellular ageing, we incubated ageing human amniotic fluid derived fibroblast-like cells and non-ageing NIH-3T3 cells with 5-azacytidine. BrdUrd/Hoechst and acridine orange (AO) flow cytometry was used to compare the effects of the base analogue on cell proliferation and cell differentiation. In NIH-3T3 cultures, 96 h exposures to 4 μM 5-azacytidine caused diminished cell proliferation due to cell arrest in the G1 compartments of the second and third cell cycles of serum stimulated cells. The exit from the G0/G1 compartment was not affected. The 5-azacytidine induced cell kinetic disturbances were unstable in NIH-3T3 cultures, such that pre-treated cells reverted to normal cell cycle transit within 2–3 days after termination of treatment. In contrast, 5-azacytidine pre-treated amniotic fluid derived fibroblast-like cell cultures showed persistently elevated G2 phase arrests and delayed G0/G1 phase exit kinetics, which explain the premature cessation of proliferation observed in these primary cultures. In both cell systems, 5-azacytidine exposed cultures showed elevated numbers of G1 phase cells with increased RNA content as revealed by AO flow cytometry. Again, this effect was reversible in NIH-3T3 cells but not in amniotic fluid derived fibroblast-like cells. These contrasting responses to 5-azacytidine are likely to reflect intrinsic differences in methylation patterns or de novo methylase activity between ageing cell strains and non-ageing cell lines.  相似文献   

16.
The Sr/Ca ratios in otoliths of silver Japanese eels Anguilla japonica , in Pearl River, China, indicated that both sexes did not stay in brackish water and grew in fresh water from the glass eel stage until spawning migration. This did not support the hypothesis that females tended to distribute upstream and males might be restricted to estuaries. The back-calculated total length of males at glass eel stage was not significantly different from that of females, indicating that the hypothesis that small glass eels became males and larger ones became females may not be true. The mean (±S.D.) age and total length of males at migration were 6·4±1·6 years and 48·3±4·5 cm, which were significantly smaller than for females, 8·3±1·6 years and 61·4±4·1 cm. The age of migration was related inversely to growth rate for both sexes. Growth parameters of the von Bertalanffy growth equation were K =0·21 cm year°1, L =55·7 cm and t o=-0·55 year for males and K =0·14 cm year−1, L =77·5 cm and t o=-0·60 year for females. The difference in asymptotic length ( L ) between males and females may be because females postpone migration to achieve larger size for maximizing reproductive success.  相似文献   

17.
Abstract. We describe a double labelling method for estimating the duration of DNA synthesis (Ts) and the flux of cells into and from the S phase of the cell cycle, based on labelling with tritiated thymidine ([3H]TdR) followed by bromodeoxyuridine (BrdU) and combining immunohistological detection of BrdU with conventional autoradiography. In practice, the change in size of a window of double labelled cells occurs as the time interval between the two labels increases. In mouse tongue epithelium there is a marked circadian variation in the number of cells in DNA synthesis. From 0900 to 1500 h this labelling index (LI) falls, but from 2100 to 0300 h it increases. Our results show that the circadian decrease in LI is associated with a short Ts (5·8 ± 0·3 h), a high S phase efflux and an initially low influx of cells from G: into S. Conversely, the rising circadian LI is associated with a longer Ts (9.4 ± 0.1 h), an initially low efflux and a moderate to high influx. Two time-points exist on the circadian LI curve when influx and efflux rates change abruptly. At 0100 h the efflux rate rises from low (5 cells %/h) to high (15–16 cells %/h) and simultaneously the influx rate changes from high to low. Similarly at 1300–1400 h, efflux rate falls from high (19–20 cells %/h) to low (4–8 cells %/h) values and influx rates change from low to high. This double labelling method has revealed that the duration of DNA synthesis varies across the circadian cycle, as do influx and efflux values which generally fall within a discrete range of high or low values. The timing of the changes in flux suggests the presence of two 'control' points on the circadian LI cycle that were previously unrecognized.  相似文献   

18.
Abstract The c-myc oncogene codes for a DNA binding protein that functions in a cell cycle-related manner. A useful model for studying the relationship of c-myc expression with cell cycle kinetics is the HL60 cell line. HL60 cells constitutively express high levels of c-myc mRNA; however, the level can be down-regulated as the cells are induced to differentiate. We have developed a flow cytometric assay for correlating c-myc oncoprotein levels with DNA content. C-myc oncoprotein levels were additionally correlated with c-myc mRNA levels as determined by slot blot hybridization. Dimethylsulphoxide (DMSO) and cytosine arabinoside were used to induce granulocytic and monocytic maturation respectively. Treatment of HL60 cells with DMSO leads to an increase in the per cent of cells in G1/G0 and a decrease in mean c-myc mRNA and oncoprotein levels. The cells with G1 DNA content show the greatest decrease in c-myc protein. ARA-c treatment of HL60 cells leads to a slowing and an accumulation of cells in S phase with a moderate decrease in mean mRNA and only a slight decrease in mean c-myc protein levels. These data support the hypothesis that c-myc is involved in the switch from G1 to G0.  相似文献   

19.
Values of T s provided by the double labelling method have been compared with those given by the percentage labelled mitoses curve for blast cells in the peripheral blood of a patient with plasma cell leukaemia and of rats bearing a transferable acute leukaemia. the double labelling method was carried out giving the first label (3H-thymidine) in vivo and the second label (14C-thymidine) in vitro with several values for the interval between the two labels. T s was calculated by fitting regression lines to the results obtained. Data for percentage labelled mitoses were analysed by computer. For the plasma cell leukaemia values of T s= 17.1 ± 7.0 hr and T s= 19.8 ± 3.4 hr, and for the rat leukaemia values of 8.7 ± 1.7 hr and 9.0 ± 1.7 hr (7.1 hr corrected for exponential growth) were obtained from the percentage labelled mitoses and double labelling methods respectively. It is concluded that the double labelling method is valid for the study of cell proliferation in leukaemic blast cells.  相似文献   

20.
Abstract. The cell population kinetics of the epidermis were studied in 4-month-old pigs. Mitotic figures were confined to the basal cell (L1) and the first suprabasal cell layer (L2). The mitotic index (MI) was 0.17 ± 0.04% for L1 and 0.08 ± 0.03% for L2. Labelled nuclei were distributed throughout the viable epidermis, the majority (79.1 ± 1.1%) were in L1 with 19.5 ± 1.2% in L2. The labelling indices (LI) in layers L1 and L2 were 7.1 ± 0.4% and 3.4 ± 0.1%, respectively. After labelling with two injections of tritiated thymidine [3H]TdR separated by 90 min, the LI increased to 8.2 ± 0.3% in L1 and to 4.0 ± 0.2% in L2. This increased labelling confirmed that cell proliferation occurs in both layers, L1 and L2, of the epidermis.
The cell production rate ( K ) in L1 and L2 had an upper limit of 10.7 ± 1.0 and 6.2 + 1.8 cells per 1000 cells per hour respectively. The cell flow rate per hour (cell flux), into and out of the DNA synthesis phase (S), and the duration of DNA synthesis were determined from double-labelling studies with [3H]TdR and [14C]TdR. The cell flux into and out of S was identical and was calculated as 0.6 ± 0.1%/hr (L1) and 0.5 ± 0.1%/hr (L2). Values for t S varied from 8 to 10 hr. The cell turnover times ( t T) were in the range 89–129 hr and 180–261 hr for L1 and L2, respectively.
Log normal curves were fitted to the fraction labelled mitoses data for L1 and L2. Values for t S for cells in L1 and L2 were 9.8 hr and 11.9 hr, respectively. t G2+ 1/2 t M was 7.2 hr in L1 and 9.1 hr in L2.  相似文献   

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