bluuebunny/crossref_metadata_2025 huggingface dataset:
Double labelling to obtain S phase subpopulations: application to determine cell kinetics of diploid cells in an aneuploid tumour
Authors: White R. A. , Pollack A. , Terry N. H. A. , Meistrich M. L. , Cao S.
Published: May 2008

Abstract. We studied the cell kinetics of the murine mammary carcinoma MCa‐K using iododeoxyuridine (IdUrd) and chlorodeoxyuridine (CldUrd) given at different times as independently detectable labels of S phase cells. The presence of IdUrd and CldUrd, and the amount of DNA were measured by three‐colour flow cytometry making it possible to define three subpopulations within S phase and to measure the progression through the cell cycle during the time following labelling. In DNA histograms of these subpopulations, the diploid and aneuploid cells (which had a DNA index of 1.7) are essentially completely separated. From appropriate combinations of cells labelled with IdUrd only, CldUrd only, or both, it was possible to construct separate DNA distributions for the labelled diploid and aneuploid cells at the times of administration of each label. The kinetics of the diploid and aneuploid cells could be calculated for individual tumours from these two time points without having to make corrections for the presence of the second population. The diploid and aneuploid populations had indistinguishable S and G2+ M phase durations, TS and TG2+M, of about 9 and 2h; however, the potential doubling time values for the aneuploid and diploid populations were 30.2 and 101.2h respectively.
FB1, a monoclonal antibody reacting with a keratin 14 epitope, stains only a small subset of psoriatic basal keratinocytes
Authors: Parent D. , Golstein J. , Mairesse N. , Bernerd F. , Faverlyt D. , Robben H. , Ramaekers F. , Heenen M.
Published: May 2008

Abstract

A murine monoclonal antibody, FB1, reacted with the basal keratinocytes of human stratified epithelia. One‐dimensional and two‐dimensional immunoblotting assays, performed on keratins extracted from HaCat cells and normal human keratinocytes, showed that FBI recognizes K14. When LL002, another K14 monoclonal antibody is added, the FB1 stained area in the 2D‐immunoblot seems to cover a fraction of the LL002 spot. Immunohistochemical data obtained from studies on normal human tissues supported the K14 specificity of FB1, but when compared with two other monoclonal antibodies, LL002 and RCK107 reacting with K14, some differences appeared. These differences were mainly seen in sweat glands, hair follicles, psoriatic epidermis and salivary glands. In psoriatic epidermis, FB1 showed a heterogeneous pattern of staining of the basal cell compartment. Intense reactivity was only observed at the bottom of the rete ridges. Staining diminished and finally disappeared in the basal cells above the dermal papillae. This observation supports the view that an increased germinative cell population in psoriasis involves a partially differentiated amplifying compartment in which the number of cell divisions is increased.
Gompertzian re‐evaluation of the growth patterns of transplantable mammary tumours in sialoadenectomized mice
Authors: Bassukas I. D.
Published: May 2008

Abstract

The method of the recursion formula of the Gompertz function (Bassukas & Maurer‐Schultze 1988) has been applied to analyse tumour growth data taken from the literature; namely the growth perturbation of transplantable mammary tumours in sialoadenectomized mice with or without subsequent epidermal growth factor substitution (results on two mouse strains, C3H or SHN, have been reported; Inui, Tsubura & Morii 1989). The recursion formula of the Gompertz function fits growth curves to all seven sets of data well (P > 0.05 for lack of fit test). The growth pattern of the tumours in the unperturbed hosts is Gompertzian and does not change if tumours are transplanted in sialoadenectomized mice, although the starting specific growth rate decreases in C3H mice. However, if sialoadenectomy is carried out after tumour inoculation, a complex alteration of the tumour growth evolves: tumour growth does not simply decelerate but it also shifts from the conventional Gompertzian to an exponential or even ‘hyperexpo‐nential’ growth pattern, i.e. with an accelerating specific growth rate. Some theoretical mechanisms of this alteration, as well as the differences between the present Gompertzian analysis and a previously published Verhulstian analysis of part of the same data (Leith, Harrigan & Michelson 1991), are discussed. It is concluded that the quantitative analysis of tumour growth patterns by the method of the difference equation of the Gompertz function presently applied may substantially contribute to the improvement of the interpretation of perturbations of tumour growth‐irrespective of their genesis. In contrast to the application of some a priori fixed growth function, e.g. the Verhulstian one, the present method can quantitatively interprete different growth patterns and their classification on the basis of linear regression analysis.
The effects of vinblastine in assessment of the influence of age on proliferative activity of murine palate and footpad epithelium
Authors: Berg J. H. , Hill M. W.
Published: May 2008

Abstract

Data concerning changes in the rate of cell proliferation of stratified epithelia with increasing age are conflicting. In the present study young (3‐month‐old) and old (22‐month‐old) C57B1/6NNia male mice were injected intraperitoneally with 2, 3, 4 or 8 mg vinblastine sulfate/kg body weight and killed after 1.5, 3, 4.5 or 6 h. The number of arrested metaphase figures per 1000 basal cells was counted in histological sections. Data were analysed using a multivariate analysis of variance. There was a significant difference between the accumulation of mitotic figures in footpad epidermis and palate epithelium and both tissues contained an increased number of mitotic figures with increasing periods of accumulation at all dose levels. In the footpad epidermis neither the age of the animal nor the dose of vinblastine had a significant effect on the number of mitotic figures. In contrast, for palate epithelium the accumulation of mitotic figures was significantly less in the old mice compared with the young mice and at a dose of vinblastine of 2mg/kg compared with the higher doses. There was a statistically signifycant interaction between the dose of vinblastine and its period of action. It was concluded that the different tissues manifest a differential sensitivity to vinblastine and that only palate epithelium showed a significant reduction in proliferative activity with age.
Expression of G1 and G2 cyclins measured in individual cells by multiparameter flow cytometry: a new tool in the analysis of the cell cycle
Authors: Gong J. , Li X. , Traganos F. , Darzynkiewicz Z.
Published: May 2008

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.
Negative control of cell proliferation in eukaryotes
Authors: Epifanova O. I. , Brooks R. F.
Published: May 2008

Abstract. Control over cell growth in eukaryotes is predominantly achieved by regular transition of cells from proliferation to rest and vice versa as a result of a co‐ordinated inter‐relationship between intracellular growth inhibitors and extracellular growth stimulators (mitogens). The ability to cease and resume growth on demand implies the existence of a refined intracellular regulatory network including both positive and negative control elements. We review here evidence that resting cells are able to produce molecules with antiproliferative activity, some of which behave as short‐lived repressor proteins. A number of genes coding for growth inhibitory molecules have been identified. However, it is not yet certain whether the same molecules ensure the maintenance of a resting state. It has become apparent that immediate growth arrest or growth resumption require not only a rapid production of inhibitors and stimulators but also their biochemical transformation (e.g. phosphorylation or dephosphorylation) and/ or translocation within the cell, whereby one and the same molecule can be a growth inhibitor or fulfil some other function in the cell cycle, according to circumstances or context. At present, three levels of negative cell growth control can be tentatively outlined: 1 rapid appearance of growth inhibitory molecules to bring about temporary arrest at critical checkpoints of the proliferative cycle; 2 transition of a cell to proliferative rest with continuous production of growth inhibitor(s); 3 long‐lasting maintenance of the resting state provided for by complex intracellular changes not connected with production of growth inhibitor(s).
Image analysis of in situ cell cycle related changes of PCNA and Ki‐67 proliferating antigen expression
Authors: Santisteban M. S. , Brugal G.
Published: May 2008

Abstract. This study reports on the proliferating cell nuclear antigen (PCNA) and Ki‐67 cell cycle related expression and distribution pattern analysed in the same cells. MCF‐7 cells were synchronized by mitotic detachment and triple stained for DNA, PCNA and Ki‐67. The major cell type was identified on each time sample as a function of the PCNA/Ki‐67 pattern, and both antigens as well as DNA were quantified. During G1 phase, the expression of PCNA greatly increased whereas Ki‐67 content decreased. During S phase, nuclear Ki‐67 content continuously increased especially in the second half of this phase, mainly due to the accumulation of the antigen in the nucleoli. During G2 phase, the antigen significantly passed into the nucleoplasm, its content continued to increase and reached its maximum in mitotic cells. Nuclear PCNA content mostly increased in the first part of S phase and sharply declined in mitotic cells as the antigen shifted to the cytoplasm. Cells showing PCNA positive Ki‐67 negative labelling were observed in all time samples from the beginning of the experiment. Their nuclear size, DNA content (of G1 cells), PCNA content (equivalent to the content of some late G, cells) and time occurrence (their percentage increased after the last late G1 cells had disappeared) tend to indicate that these cells have left the cycle by the end of G1 phase to enter a quiescent state. Cells coming out of mitosis split into two groups according to their Ki‐67/PCNA content. The biggest fraction was PCNA negative and Ki‐67 positive while the smallest showed positive staining for both antibodies. Cells of this second cohort slowly lost their 1–67 while their PCNA content increased as they moved through G1. Concurrently, most of the cells of the first cohort (here called Q2 and Q3 cell types) lost their Ki‐67 without increasing their PCNA content; then they joined cells of the second cohort by increasing their PCNA content at the end of G, phase. Some cells of this first cohort can also increase their PCNA and thus reach cells of the first cohort before the end of G1 phase. The existence of these two main cell cohorts suggests that cells after mitosis differ in some way that make them progress dlfferently through G1. Some cells seem to go through early G1 (G1a and late G1 (Glb) while others may come out of mitosis committed to go through the following cycle by directly entering late G1 compartment.
Expression of p53 mRNA is proliferation‐dependent in the human fibroblast cell line WI‐38
Authors: Kucsera M. , Bori Z. , Marcsek Z.
Published: May 2008

Abstract. Although alterations in the p53 tumour suppressor gene are one of the most frequent genetic lesions occurring in human cancers, the exact function and mechanism of action of normally regulated p53 in the control of cell cycle is unclear. To clanfy further the possible role of this gene in the control of cell proliferation, the cellular level of p53‐specific mRNA and its changes during density‐dependent growth, and in different proliferation states induced by serum starvation and subsequent serum‐stimulation, were followed in WI‐38 cells, a normal human diploid fibroblast cell line. Marked differences in the expression of p53 mRNA could be observed in the different proliferation states tested. The pattern of p53 expression proved to be inversely proportional to the growth‐rate of the cultures. mRNA was considerably more abundant when cells reached confluency or were arrested by serum deprivation while serum‐stimulation caused the opposite effect. These results support the hypothesis that the p53 gene plays a role in G1 control of normal cell proliferation.
Comparison of monoclonal antibodies PC 10 and MIB 1 on microwave‐processed paraffin sections
Authors: Kubbutat M. H. G. , Cattoretti G. , Gerdes J. , Key G.
Published: May 2008

Abstract. Antibodies against the proliferation‐associated nuclear antigen (PCNA) and against the Ki‐67 protein are widely used as operational proliferation markers in human tumour diagnostics. The original Ki‐67 antibody had the inherent drawback in that it could only be used when fresh‐frozen material was available. The antibody PClO was supposed to offer the advantage that it could be applied on formalin‐fixed and paraffin‐embedded tissues. However, in cases in which the formalin fixation exceeded 4 h, PC10 staining proved to be inconsistent and often failed.

The aim of this study was to compare a recently prepared Ki‐67 equivalent monoclonal antibody (MIB 1) and PC10 in routinely fixed histopathological material using antigen retrieval by microwave processing.

Antibody MIB 1 stained the nuclei of cells known to belong to the proliferative compartments in microwave‐processing paraffin sections of formalin‐fixed tissues. Quiescent cells were consistently negative for MIB 1 staining. In contrast, PC10 was positive in almost all nuclei of different tissues in microwave‐treated paraffin sections. Thus, antigen retrieval by microwave processing is beneficial for the detection of the Ki‐67 protein in paraffin sections, whereas it is not needed for the detection of the PCNA.
S‐phase cell fraction in the prognosis of Dukes' stage D colorectal carcinoma
Authors: Faranda A. , Costa A. , Silvestrini R. , Quagliuolo V. , Gennari L.
Published: May 2008

Abstract. The S‐phase cell fraction was prospectively defined as the [3H]‐thymidine labeling index ([3H]dT LI) and flow cytometric S‐phase (FCM‐S) on 52 Dukes' D colorectal cancers. FCM‐S estimates obtained by using different modeling systems were superimposible but they were weakly related to [3H]dT LI detected on the same tumour. Moreover, FCM‐S values were higher in aneuploid than in diploid tumours, whereas [3H]dT LI values were independent of DNA‐ploidy status. [3H]dT LI and FCM‐S were also related differently to some clinical and pathological features such as tumour site and histology. [3H]dT LI and FCM‐S were indicative of prognosis in terms of 2–year freedom from progression and overall survival. However, product‐limit survival analysis showed an unexpectedly better freedom from progression and overall survival for patients with high S‐phase tumours than for those with low S‐phase tumours as defined by both cell kinetic variables.
Cell cycle of neural crest cells in the early migratory stage in vivo
Authors: Paglini M. G. , Rovasio R. A.
Published: May 2008

Abstract. The fact that directional migration of neural crest cells (NCC) in vivo occurs in narrow pathways at high cell density, together with our preliminary results showing their high proliferative behaviour, supports the view that a ‘population pressure’ could be an important factor in the mechanism of early dispersion of NCC.

The purpose of this work was to establish the cranial proliferative pattern of chick embryo NCC during their early migratory stage in vivo. Growth rates and parameters of cell cycle were obtained from cell populations at several cephalic levels by means of autoradiography after labelling with [3H]dT. The labelled cell index of NCC (Forebrain, 0.288; Midbrain, 0.206; and Hindbrain, 0.134) was significantly greater than in other cells populations (e.g. for the neural tube cells: 0.085, 0.030, and 0.031, respectively). The cell generation time was the shortest in NCC (16 h), compared to ectoderm (33 h), mesoderm (58 h) and endoderm (72 h). The duration of the cell cycle phases for NCC were: M, 0.29 h; G1, 11.23 h; S, 3.40 h; and G2, 1.02 h.

These quantitative results show that NCC have the greatest proliferative rate in young chick embryos. In relation to cranial regions, the data are consistent with the idea that, in the early migratory phase of this cell population in vivo, migration is in part driven by population pressure.
Relationship between epidermal growth factor receptor levels, autophosphorylation and mitogenic‐responsiveness in normal mouse mammary epithelial cells in vitro
Authors: McIntyre B. S. , Birkenfeld H. P. , Sylvester P. W.
Published: May 2008

Mammary epithelial cells were isolated from mid‐pregnant BALB/c mice, grown within collagen gels and maintained on DME/F12 (1:1) media containing 10% bovine calf serum and 10 μ/ml insulin. Initial time‐course and dose‐response studies showed that epidermal growth factor (EGF)‐induced autophosphorylation of the EGF‐receptor (EGF‐R) in these cells was maximal 5 min after exposure to 75 ng/ml EGF. Mammary epithelial cells displaying little or no growth during their first 2 days in primary culture cells were found to contain low levels of EGF‐R. However, EGF‐induced autophosphorylation of the EGF‐R in these cells was extremely intense. Subsequent studies demonstrated that during the proliferative and plateau phases of growth, EGF‐R levels progressively increased, while conversely EGF‐induced autophosphorylation of the EGF‐R decreased over time in primary culture. These results demonstrate that EGF‐R levels and autophosphorylation do not show a direct correlation with mammary epithelial cell mitogen‐responsiveness. Intense EGF‐R autophosphorylation appears to be required for initiating growth, but sustained mammary epithelial cell proliferation occurs when EGF‐R autophosphorylation is low. This inverse relationship between EGF‐R levels and autophosphorylation may reflect changes in receptor affinity and function during the various phases of mammary epithelial cell growth in primary culture.
Labelling DNA strand breaks with BrdUTP. Detection of apoptosis and cell proliferation
Authors: Li X. , Darzynkiewicz Z.
Published: May 2008

In situ presence of numerous DNA strand breaks is a typical feature of apoptotic cells. Selective DNA strand break induction by photolysis (SBIP) at sites that contain incorporated halogenated DNA precursors has recently been proposed as a method of analysing DNA replication. Detection of DNA strand breaks, thus, enables one to identify apoptotic and/or DNA replicating cells. The current methods for DNA strand break labelling rely on the use of exogenous terminal deoxynucleotidyl transferase which either directly attaches the fluorochrome conjugated triphosphodeoxynucleotides to 3′OH ends in the breaks, or indirectly labels 3′OH ends with digoxygenin or biotin conjugated triphosphodeoxynucleotides. A limitation of these methodologies, especially restricting their routine application in the clinic, is high cost of reagents. In the present study we have tested whether relatively simple compound BrdUTP, which is approximately three orders of magnitude less expensive than dUTP conjugated to digoxygenin, can be used as marker of DNA strand breaks. Apoptosis of HL‐60 cells was induced by DNA topoisomerase I inhibitor camptothecin. The incorporated BrdUTP was detected by fluoresceinated anti‐BrdUrd MoAb. Cellular fluorescence was measured by flow cytometry as well as by Laser Scanning Cytometer (LSC). The data show that intensity of DNA strand break labelling with BrdUTP was nearly four‐ and two‐fold higher than that obtained with the indirect labelling using biotin‐ or digoxygenin‐conjugated dUTP, respectively, and over eight‐fold higher than in the case of direct labelling with the fluorochrome (fluorescein or BODIPY)‐conjugated deoxynucleotides. The increased labelling of DNA strand breaks with BrdUTP may reflect more efficient incorporation of this precursor by terminal transferase, compared to the nucleotides with bulky fluorochrome conjugates. DNA strand break labelling with BrdUTP, thus, offers a possibility of more sensitive (and at lower cost) detection of apoptotic or DNA replicating cells, compared to the alternative methods of DNA strand break labelling.
Cell proliferation in the subependymal layer of the adult mouse in vivo and in vitro
Authors: Hauke C. , Ackermann I. , Korr H.
Published: May 2008

Pulse labelling experiments with [3H] thymidine (dT) and double labelling experiments with [3H]dT and bromodeoxyuridine (BrdUrd) were carried out on cells of the subependymal layer in the brain of adult normal mice in vivo, in vivo/in vitro and in vitro. The results should (i) lead to information about cell cycle parameters of these cells in the brain of adult mice, since these cells have been studied mostly in the rat brain up to now and (ii) answer the question whether results concerning cell proliferation obtained in vivo correspond with those from brain slices incubated in vitro with or without prelabelling in vivo. In vivo an LI of 20.2 ± 2.7% (x̄± SEM) and Ts= 7.2 ± 0.7h were found. Furthermore, grain count halving experiments led to a surprisingly short cycle time (Tc) of 11.2–14.2 h. The longer Tc values (18–20 h) reported in the literature for subependymal cells in the rat brain seem to be due to evaluations of different areas around the lateral ventricle without considering the migrating behaviour of these cells which is quite different regionally. The in vitro studies (with or without prelabelling in vivo) showed a significantly reduced LI due to the fact that about 20% of the S phase cells, possibly lying in the middle of S, stopped further DNA synthesis after transfer to culture. This was shown by comparing the cell fluxes at the G1/S and S/G2 borders of in vivo vs. in vitro studies.
In vitro bromodeoxyuridine‐labelling of single cell suspensions: effects of time and temperature of sample storage
Authors: Carbajo‐Pérez E. , Carbajo S. , Ciudad J. , López A. , Dolbeare F. , Orfao A.
Published: May 2008

The present study was aimed to explore how the in vitro BrdUrd‐labelling of rat thymocytes might be affected by both the time elapsed between obtaining the sample and the beginning of the labelling (0, 15, 30 or 60 min) and the effect of the temperature of storage (4°C versus room temperature). Single cell suspensions obtained after in vivo labelling with BrdUrd were used as controls. The S phase fraction was calculated by flow cytometry both according to BrdUrd‐immunolabelling and DNA content. Immediate incubation with BrdUrd after the sample was obtained resulted in a slight decrease of the proportion of S phase cells analysed either according to DNA content or to BrdUrd‐immunolabelling. Regardless of storage‐temperature, the S phase fraction decreased in samples kept for 15 min or more before BrdUrd incubation. No BrdUrd‐positive cells were detected in samples stored for 60 min at room temperature. This effect was related to temperature since positive cells were found when the samples were kept at 4°C during the same time period. Our results suggest that during in vitro incubation a relative loss of S phase cells exists and that a delay beyond 15 min between obtaining the sample and the in vitro labelling seriusly compromises the results of this technique.
Comparison of proliferating cell nuclear antigen (PCNA) staining and BrdUrd‐labelling index under different proliferative conditions in vitro by flow cytometry
Authors: Lohr F. , Wenz F. , Haas S. , Flentje M.
Published: May 2008

PC10 is a monoclonal antibody against proliferating cell nuclear antigen (PCNA). The staining pattern in immunochemistry depends on fixation and detergent extraction treatment. The aim of this study was to validate the flow cytometric PCNA assay against Bromodeoxyuridine‐labelling index (BrdUrd‐LI) under different proliferative conditions in vitro. Expression of PCNA in methanol fixed cells with, and without, prior detergent extraction with EDTA/Triton was compared to BrdUrd‐labelling index in NIH‐3T3 fibroblasts and human Caski tumour cells in exponential phase and under confluent conditions. Serum stimulation and serum starvation conditions were studied. The results for BrdUrd‐LI and PCNA‐index after extraction showed good correlation for 3T3 fibroblasts and for Caski cells, with some differences for serum withdrawn Caski cells. There was no correlation between the number of cells that were positive for PCNA without extraction and BrdUrd‐LI. Spheroid cells with G1‐DNA‐content showed an almost synchronous recruitment and progression through the cell cycle after trypsination and replating. Tightly bound PCNA paralleled this synchronicity whereas total PCNA did not change significantly. The results demonstrate that immunochemical detection of non‐extractable PCNA‐index gives similar results as compared with BrdUrd‐labelling index under different proliferative conditions in vitro for different monolayer cell lines, whereas without extraction PCNA does not correlate with BrdUrd‐LI in these fast growing cell lines due to its long half‐life. PCNA expression parallels the progression through the cell cycle in V79 spheroids, a primitive model of tumour growth.
Modulation of rabbit articular chondrocyte (RAC) proliferation by TGF‐β isoforms
Authors: Boumediene K. , Vivien D. , Macro M. , Bogdanowicz P. , Lebrun E. , Pujol J‐P.
Published: May 2008

We have previously shown that TGF‐β1 exerts a bifunctional effect on RAC proliferation. Added to quiescent cultures, it inhibits the entry of G0/G1 cells into S phase whereas in S phase synchronized populations, it stimulates the DNA replication rate with a delayed G2+ M phase and a subsequent transient increase of cell number. As TGF‐β2 and β3 isoforms are also expressed in bone and cartilage tissues, it was of interest to study their effect on RAC proliferation, in comparison to that of TGF‐β1.

Using cell counting and tritiated thymidine incorporation, we found that all the TGF‐βs used here induced an increase of RAC proliferation rate occurring between 24 and 48 h of exposure. TGF‐β2 appeared as the most efficient form as judged from the maximum of thymidine labelling. However, TGF‐β3 induced an increase of cell number slightly higher than both TGF‐β1 and TGF‐β2 (+30% versus 20% for TGF‐β1 and β2). TGF‐β2 and β3 were able to stimulate the DNA replication rate as previously demonstrated for TGF‐β1. However, the effect occurred later for TGF‐β2 and β3 (12 h) than for TGF‐β1 (6 h). This was confirmed by flow cytometric analysis of DNA content. In addition, immunodetection by flow cytometry demonstrated that all TGF‐β isoforms enhanced endogenous expression of TGF‐β‐related peptides. The effect was shown to be associated with the cell cycle S phase and was greater for TGF‐β3 than for TGF‐β1 and β2. These findings suggest that TGF‐βs could act on RAC functions via autocrine and paracrine ways.

Taken together, these data indicate that TGF‐βs may modulate proliferation of articular chondrocytes and therefore could play a role in the activation of these cells in the early stages of osteoarthritis.
Different growth patterns of a cancer cell population as a function of its starting growth characteristics: analysis by mathematical modelling
Authors: Solyanik G. I. , Berezetskaya N. M. , Bulkiewicz R. I. , Kulik G. I.
Published: May 2008

The growth kinetics of a cancer cell population as a function of the total number of cells and the proportion of proliferating and resting cells at the beginning of the growth has been analysed by a mathematical model. The model takes into account the processes of cell division, death and transition from proliferation to rest and backwards. It is shown that a single cell population growing under the same environmental conditions has an extremely broad spectrum of growth patterns. The whole multiplicity of possible growth patterns has been determined by the inherent cellular growth characteristics of the population, while the growth pattern actually realized of the variety of growth curves depends on the total number of cells and the proportion of proliferating and resting cells at the initial moment of growth. The model is shown to provide a good prediction of experimentally measured kinetics of regrowth of tumour cells subcultured after various times of the growth in unfed cultures, and the kinetics of tumour cell growth after severe hypoxia. The role of cell transitions between proliferating and resting stages in the problem of growth control is discussed.
Effect of ebrotidine on the density of antral G‐cells in the gastric mucosa in the rat
Authors: Romero A. , Gömez F. , Villamayor F. , Sacristán A. , Ortíz J‐A.
Published: May 2008

Two groups of male rats were treated orally for 60 days with ebrotidine and cimetidine, used as reference standard, respectively. The dose used of both drugs was 500mg/kg. A third group, used as control, received 10 ml/kg of an aqueous agar suspension. After receiving the last dose, the animals were killed by inhalation of CO2 in a sealed chamber and the stomachs were removed for histological preparation.

The purpose of this study was to count antral G‐cells throughout the gastric mucosa by light microscope. A PAP system‐associated antigastrin immunohistochemical method was used for cell identification. Cell density, with respect to their location in the gastric mucosa and the treatments given, was calculated using image analysis techniques.

The results showed that ebrotidine did not cause any significant differences in the density of the population of these cells compared with the control group, while cimetidine induced a significant proliferation of antral G‐cells.
Differential effects of nitrogen mustardin vivoon the cancer and host cells in MCa‐11 tumours
Authors: Bose K. K. , Zahurak M. L. , Allison D. C.
Published: May 2008

We analysed the effects of nitrogen mustard (HN2) on the growth, cell cycle distributions, and ratios of tumour cells to host cells for MCa‐11 tumours grownin vivo.Treatment of tumour‐bearing BALB/c mice with 3 mg/kg of HN2produced a significant slowing of MCa‐11 tumour growth. Seventy‐two hours after treatmentin vivowith either 3 or 4 mg/kg of HN2, the host cells in the treated tumours showed a significantly decreased G0/G1peak and an increased G2/M peak (P< 0.01), whereas the cancer cells in the treated tumours showed significant increases in the G0/G1peak coupled with relatively decreased proportions of S and G2/M tumour cells (P< 0.001). The ratio of the total number of cancer cells to the total number of host cells in the tumours was significantly increased 72 h after HN2administration (P<0.01). Thirty‐two days after treatment with HN2, the cell cycle distributions of the host and tumour cells in the treatment and control tumours had returned to being identical, but the ratio of the total number of cancer cells to the total number of host cells remained increased in the treated tumours (P<0.01). These results show that the administrationin vivoof HN2can lead to entirely different cell cycle effects for the host and cancer cells in the same tumour, and that the partial growth arrest of MCa‐11 tumours from HN2treatment may be due in part to the preferential destruction of host cells rather than solely to a direct cytotoxic effect on the cancer cells.
Hormone gradients and cartilage cell kinetics
Authors: Vogl C. , Atchley W. R.
Published: May 2008

We present a model of growth control in mammalian cartilage growth plates by hormones. The model is based on the distribution of insulin‐like growth factors I and II (IGF‐I and IGF‐II) and their receptors, and assumes that a hormone‐receptor complex of IGF controls cells proliferation. A system of differential equations is derived and solved with simplifications in extreme cases, for the one‐dimensional time independent case. Even if opposite extremes, such as proliferation control by factors extrinsic to the cell versus intrinsic to the cell, are assumed, similar distributions of hormones and proliferating cells are produced. Hence, choice between alternative models of growth control must be based on empirical observations. On the positive side, similarities between our model for cartilage growth and other models for differentiation and proliferation are evident and might be exploited for unifying these systems on an abstract level.
Argyrophilic nucleolar organizer regions and bromodeoxyuridine and 3[H]‐thymidine labelling indices in colorectal cancer
Authors: Losi L. , Gregorio C. Di , Fante R. , Migaldi M. , Roncucci L. , Pedroni M. , De Leon M. Ponz , Trentini G. Paolo
Published: May 2008

The count of argyrophilic nucleolar organizer regions (AgNORs) has been proposed as a useful method for evaluating cell replication in human tumours. The current study was undertaken to compare AgNOR values in colorectal cancers with two better established methods for investigating cell proliferation such as bromodeoxyuridine (BrdUrd) and 3[H]‐thymidine (3[H]dT) labelling indices (LIs).

Because some concern still exists regarding accuracy and reproducibility of AgNOR quantifying methods, we carried out a control study by independently repeating the same measurements (number, area and area per silver‐stained NOR particle) in two centres with different operators and computer‐assisted image analysers on 40 colorectal carcinomas. AgNOR values recorded in the two centres were strictly correlated (r= 0.75; P < 0.001 for number; r= 0.62, P < 0.01 for area; r= 0.63, P < 0.001 for area per silver‐stained NOR particle) and the range of values were almost identical. Then, AgNOR values were compared with BrdUrd and 3[H]dT LIs, respectively obtained by in vivo incorporation and in vitro incubation in the same series of colorectal carcinomas. No correlation was found between AgNOR values and BrdUrd or 3[H]dT LIs. BrdUrd and 3[H]dT LIs were instead reciprocally significantly correlated. No evident correlation was seen between LIs or AgNOR values and clinico‐pathological parameters of the tumour. In conclusion, in colorectal neoplasms, AgNOR values did not appear to relate with more direct parameters of cell proliferation. It follows that AgNOR reliability as a biomarker of cell proliferation remains questionable.
Analysis of DNA content in multidrug‐resistant cells by image and flow cytometry
Authors: Palissot V. , Liautaud‐Roger F. , Carpentier Y. , Dufer J.
Published: May 2008

Abstract. Nuclear DNA content was assessed in multidrug‐resistant (MDR) cells by image and flow cytometry. Two human MDR cell lines (K562‐Dox and CEM‐VLB) obtained by in vitro drug selection and overexpressing mdr1 gene were compared to their respective sensitive counterparts (K562 and CCRF‐CEM) and to the MDR hamster LR73‐R cell line obtained by transfection of mouse mdr1 cDNA. Both cell lines obtained by selection displayed a decreased DNA content, as measured by image cytometry after Feulgen staining, or by flow cytometry after staining with propidium iodide, ethidium bromide, or Hoechst 33342. This decrease was not accompanied by changes in cell cycle phase distribution of cells. Moreover, image cytometry of cells stained after various hydrolysis times in 5 M HCl indicated that MDR cells displayed the same hydrolysis kinetics and sensitivity as drug‐sensitive cells with a well‐preserved stoichiometry of the Feulgen reaction. LR73‐R cells transfected with mdr1 cDNA exhibited only a very limited change in propidium iodide staining as compared with sensitive LR73 cells, suggesting that mdr1 gene overexpression alone could not account for the alterations in DNA content observed in the selected MDR cells.
Established cell lines transformed by E2F‐1 overexpression contain wild‐type p53
Authors: Sladek T. L.
Published: May 2008

Abstract. Overexpression of genes encoding E2F transcription factors can transform some cultured cell lines and cause apoptosis of others. Apoptosis due to E2F overexpression requires the presence of wild‐type p53. Cell lines in which stable E2F overexpression is possible might be expected, therefore, to contain mutant p53. In this report, it was asked whether endogenous p53 was mutant or wild‐type in four established fibroblast cell lines that this laboratory previously showed stably overexpressed and were transformed by exogenous E2F‐1. Unexpectedly, it was found that the p53 in these cells was wild‐type by the criteria of immunoprecipitation with conformation‐specific, p53 monoclonal antibodies and by transactivation of a p53‐dependent reporter gene construct in transient transfection assays. These data indicate that stable overexpression of E2F‐1 is possible in the presence of wild‐type p53 and may result in cell transformation.
Changes in the amount and distribution of prosomal subunits during the differentiation of U937 myeloid cells: high expression of p23K
Authors: Henry L. , Baz A. , Château M.‐T. , Scherrer K. , Bureau J. P.
Published: May 2008

Abstract. Prosomes (Proteasomes/Multicatalytic proteinase (MCP)‐complexes) are protein particles built of 28 subunits in variable composition, having proteinase activity. We have studied the changes in prosomal subunits p29K, p31K and the highly expressed p23K during the differentiation of U937 cells. Control cells had little prosomal subunit p31K in the cytoplasm, while p29K antigen was detected in both the nucleus and cytoplasm; more p23K antigen was found in the cytoplasm than in the nucleus. Flow cytometry demonstrated a biphasic intracellular decrease in prosomes during differentiation induced by phorbol‐myristic‐acetate (PMA) and retinoic acid plus 1,25‐dihydroxycholecalciferol (RA + VD). p23K and p29K decreased both in the cytoplasm and the nucleus of differentiated cells, though the p23K antigen was concentrated near vesicles and the plasma membrane in PMA‐induced cells. The p31K antigens disappeared from RA + VD‐induced cells, while in PMA‐induced cells, cytoplasmic labelling was unchanged and nuclear labelling was increased. Small amounts of prosomal proteins p23K and p29K were found on the outer membrane of un‐induced cells. While there was no labelling on the outer membrane of RA + VD‐induced cells, p23K protein increased on the plasma membrane of PMA‐induced cells. The prosome‐like particle protein p21K was not present to any significant extent in the intracellular compartment of control or induced cells; however, p21K was detected on the outer surface of control cells and was increased only in PMA‐induced cells. The culture medium of control and induced cells contained no p21K, p23K, p29K or p31K. RA + VD seemed to induce a general decrease of prosomal subunits within the cells and at the outer surface, whereas PMA caused a migration toward the plasma membrane and an increase at the outer surface. These changes in the distribution and type of prosomes in RA + VD‐ and PMA‐induced cells indicate that prosomes may play a part in differentiation, especially p23K which is the most highly expressed protein among those studied and presents the more important changes.
Exponential growth, random transitions and progress through the G1 phase: computer simulation of experimental data
Authors: Sennerstam R. , Strömberg J.‐O.
Published: May 2008

Abstract. At a time of increasing knowledge of gene and molecular regulation of cell cycle progression, a re‐evaluation is presented concerning a phenomenon discussed before the present expanding era of cell cycle research. ‘Random transition’and exponential slopes of α‐ and β‐curves were conceived in the 1970s and early 1980s to explain cell cycle progression. An exponential behaviour of the β‐curve was claimed as being necessary and sufficient for a ‘random transition’in the cell cycle. In our present work, similar slopes of those curves were shown to materialize when the increase in mass of single cells was set as exponential in a structured cell cycle model where DNA replication and increase in cell mass were postulated to be two loosely coupled subcycles of the cell cycle, without introducing any ‘random transition’. Findings published in the 1980s demonstrating the effect of serum depletion of 3T3 Balb‐c cells were simulated and the shallower slope of the α‐ and β‐curves found experimentally could be attributed to the reduced rate of exponential growth in cell mass, rather than to a reduced ‘transition probability’.
pp32 overexpression induces nuclear pleomorphism in rat prostatic carcinoma cells
Authors: Gusev Y. , Romantsev F. E. , Chen T. T.‐H. , Kayler A. E. , Kuhajda F. P. , Dooley W. C. , Pasternack G. R.
Published: May 2008

Abstract. Nuclear pleomorphism is an important diagnostic factor in tumour pathology. Traditionally, nuclear pleomorphism is evaluated qualitatively or semiquantitatively, often as a component of tumour grade; the molecular basis of nuclear pleomorphism, however, remains unclear. In this study, we investigated the quantitative effects on nuclear morphology of overexpressing pp32, a recently described nuclear phosphoprotein highly expressed in self‐renewing and neoplastic cell populations. Assessment of Feulgen‐stained transfected and control lines of AT3.1, a rat prostatic carcinoma cell line, using a computerized Cellular Image Analysis System (BD CAS‐200) showed that stable overexpression of human pp32 in AT3.1 cells is accompanied by marked increases in the coefficient of variation of nuclear shape, nuclear size and chromatin textures but not in DNA content. In contrast, stable transfection with control vector, with ras, or with bcl‐2 failed to affect nuclear morphology. Cell cycle analysis further showed that pp32‐related increases in variation of nuclear structure manifested principally in G1. These studies suggest that pp32 plays a role either directly or indirectly in the control of nuclear shape of G1 cells.
Pluripotential liver stem cells: facultative stem cells located in the biliary tree
Authors: Alison M. R. , Golding M. H. C. , Sarraf C. E.
Published: May 2008

Abstract. The ability of the liver to regenerate after parenchymal damage is usually accomplished by the ephemeral entry of normally proliferatively quiescent (G0) hepatocytes into the cell cycle. However, when hepatocyte regeneration is defective, arborizing ductules which are continuous with the biliary tree, proliferate and migrate into the surrounding parenchyma. In man these biliary cells have variously been referred to as ductular structures, neoductules and neocholangioles, and have been observed in many forms of chronic liver disease, including cancer. In experimental animals similar ductal cells are usually called oval cells, and their association with defective regeneration has led to the belief that these cells represent a progenitor cell population. Oval cells are thought to take over the burden of regenerative growth after substantial hepatocyte loss, suggesting that they are the progeny of facultative stem cells. The liver is not, however, generally considered as a stem cellfed hierarchy, although this is disputed by others. Despite this, the subject of oval cells has aroused intense interest as these cells may represent a target population for hepatic carcinogens, and they may be useful vehicles for ex vivo gene therapy.

This review proposes that the liver does harbour stem cells which are located throughout the biliary epithelium, and that oval cells represent the progeny of these stem cells and function as an amplification compartment for the generation of ‘new’hepatocytes. This is a conditional process which only occurs when the regenerative capacity of hepatocytes is overwhelmed and thus, unlike the intestinal epithelium, the liver is not behaving as a classical continually renewing stem cell‐fed lineage. We focus on the biliary network, not merely as a conduit for bile, but also as a cell compartment with the potential to proliferate under appropriate conditions and give rise to fully differentiated hepatocytes and other cell types.
Monoclonal antibodies Ki‐S3 and Ki‐S5 yield new data on the ‘Ki‐67’proteins
Authors: Heidebrecht H. J. , Buck F. , Haas K. , Wacker H. H. , Parwaresch R.
Published: May 2008

Abstract. The monoclonal antibody (mab) Ki‐67 has been used for about 10 years, mainly in tissue sections, to monitor proliferating cells, but so far only very little is known about the proteins it recognizes. The new mabs Ki‐S3 and Ki‐S5 detect proliferating cells in frozen and paraffin‐embedded tissues. They recognize proteins with the same molecular mass as Ki‐67 in Western blot and for the first time also in immunoprecipitation experiments. With these mabs we were able to enrich and purify the Ki‐67 proteins. Protein sequencing of four peptides of the digested proteins corresponded to the cDNA‐deduced amino acid sequence already published for the Ki‐67 proteins.

Since we were able to immunoprecipitate the Ki‐67 proteins, we performed various immunoprecipitation experiments to obtain more information about the nature of these proteins. After radiolabelling L428 cells with [35S]‐methionine we were able to immunoprecipitate the Ki‐67 proteins after only 5 min of labelling time. In turnover experiments the Ki‐67 proteins could not be detected 3 h after the end of labelling. These data indicate a halt‐life of the Ki‐67 proteins of about 90 min.

Labelling experiments with [32P]‐orthophosphate revealed that the Ki‐67 proteins are phosphorylated. After dephosphorylation was blocked with okadaic acid or cell growth was arrested by means of Colcemid, the phosphorylation of the Ki‐67 proteins was greatly increased, indicating that the Ki‐67 proteins are phosphorylated via serine and threonine, and that the phosphorylation of the Ki‐67 proteins increases in cycling cells. Labelling experiments with [3H]‐mannose and [3H]‐glucose revealed that the protein is weakly N‐glycosylated.