bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of melphalan and hyperthermia on cell cycle progression and cyclin B1 expression in human melanoma cells
Authors: Orlandi L. , Zaffaroni N. , Bearzatto A. , Costa A. , Supino R. , Vaglini M. , Silvestrini R.
Published: May 2008

We have investigated the effect of mild hyperthermia (42°C) on the cytotoxic activity of a 1 h melphalan exposure in human melanoma cell lines. Hyperthermia did not affect cell growth of any culture, but it increased, to a different extent, melphalan cytotoxicity in all cell lines, with a reduction in the IC50 of 1.7 to 2.6‐fold. Flow cytometric analysis showed that in normal temperature conditions melphalan caused S phase cell accumulation, which was evident only at 24 h in JR8, M14 and 2/21 cell lines and was still persistent at 72 h in 2/60 cells. Moreover, in all cell lines, the delay in S phase was paralleled, or followed, by an accumulation of cells in G2+ M, which was transient in JR8 and M14 cells and persisted until 72 h in 2/21 and 2/60 melanoma clones. Hyperthermia caused a stabilization and prolongation of melphalan induced G2+ M accumulation in JR8 and M14 cells. Conversely, in 2/21 and 2/60 clones, cell cycle perturbations induced by the drug were similar under normothermic or hyperthermic conditions. Specifically, in JR8, for which the maximum enhancement by hyperthermia on melphalan cytotoxicity was observed, cell accumulation in G2+ M was still present 120 h after treatment. The accumulation was accompanied by an inhibition in the G2 ‐ M transition, as demonstrated by the significant reduction in the mitotic index of cells exposed to combined treatment compared to controls. Moreover, a bivariate distribution of cells stained for DNA and cyclin B1 showed that, following melphalan and hyperthermia treatment, the fraction of cyclin B1‐expressing cells paralleled the fraction of G2+ M phase cells, thus indicating that the inability of cells to enter mitosis was not ascribable to a reduction of cyclin B1 expression. On the whole, our results indicate that hyperthermia can stabilize the G2 accumulation induced by melphalan in human melanoma cells. Such a stabilization could contribute to the enhancement of melphalan cytotoxicity by heat, even though a strict correlation was not observed between the magnitude and persistence of the cell cycle perturbations and the extent of melphalan activity.
Transient inhibition of protein synthesis induces expression of proto‐oncogenes and stimulates resting cells to enter the cell cycle
Authors: Rosenwald I. B. , Setkov N. A. , Kazakov V. N. , Chen J.‐J. , Ryazanov A. G. , London I. M. , Epifanova O. I.
Published: May 2008

There is evidence that resting cells are able to produce molecules with antiproliferative activity, some of which behave as short‐lived repressor proteins. We suggest that transient inhibition of protein synthesis in resting cells would lead to a decrease in the levels of these negative growth regulators and might, therefore, promote mitogenic responses.

We report that treatment of resting (serum‐deprived) NIH3T3 cells with cyclocheximide (CH) or puromycin induces expression of c‐fos, c‐jun and c‐myc proto‐oncogenes in a manner similar to that of platelet‐derived growth factor (PDGF). Actinomycin D (Act D) abrogates the induction of proto‐oncogene expression. Transient inhibition of protein synthesis by CH or puromycin also induces the resting NIH 3T3 and C3H 1OT1/2 cells to enter the cell cycle. Inhibition of new RNA or protein synthesis abolishes the proliferative response. These findings show that control mechanisms at both transcriptional and translational levels are operative in the resting cells treated with protein synthesis inhibitors.

Cell fusion experiments with resting and serum‐stimulated NIH 3T3 cells revealed that brief pre‐incubation of resting cells with either PDGF, CH or puromycin abrogates their ability to suppress the onset of DNA synthesis in the nuclei of stimulated cells in heterodikaryons. However, the abrogative effect of PDGF disappeared in the presence of Act D, whereas the effects of protein synthesis inhibitors did not, indicating their independence of the induction of transcription. The data suggest that the observed effects of protein synthesis inhibitors are connected with elimination of some short‐lived negative growth regulators, since a brief translational arrest is sufficient for the resumption of DNA synthesis in the nuclei of stimulated cells blocked by resting cells in heterodikaryons.
The characterization of the monoclonal antibody Th‐10a, specific for a nuclear protein appearing in the S phase of the cell cycle in normal thymocytes and its unregulated expression in lymphoma cell lines
Authors: Muto M. , Utsuyama M. , Horiguchi T. , Kubo E. , Sado T. , Hirokawa K.
Published: May 2008

A monoclonal antibody (Th‐10a) specific for the nuclear protein appearing in the S phase of the cell cycle in normal mouse thymocytes was derived by immunizing Wistar rats with a murine thymic lymphoma (TIGN), and its isotype was rat IgG2a and had κ light chain. Immunohistochemical staining of frozen sections of B10.Thy1.1 newborn thymus and embryonic intestine revealed that this monoclonal antibody reacted strongly with the nuclear proteins of subcortical thymocytes and the basal layer of the mucosa, where many cells were dividing, but not with that of the thymic medullary area. To evaluate the expression of the nuclear proteins during the cell cycle in detail, the results of an immunofluorescence analysis of the thymocytes from hydroxyurea‐treated B10 mice using Th‐10a monoclonal antibody were compared with those of DNA synthesis of these cells with the use of the FITC‐conjugated anti‐BrdUrd monoclonal antibody. The results indicated that the nuclear protein detected by Th‐10a monoclonal antibody was highly expressed in the S phase of normal thymocytes, while the cells in G1, G2 and M phases exhibited a low level of the expression. Moreover, the variations in expression of the nuclear proteins in the thymocytes at different times after hydroxyurea treatment were observed to correspond with the frequency of DNA synthesizing cells. In contrast, the high level and unregulated expression of the nuclear protein detected by Th‐10a monoclonal antibody was observed throughout the cell cycle of the mouse lymphoma cell lines examined. Since Th‐10a monoclonal antibody does not react with the nuclear proteins derived from human, hamster or rat proliferating cells, this antibody may recognize a murine specific epitope of the nuclear protein.

To further characteize the nuclear proteins, we extracted them from normal thymocytes or thymic lymphomas, and analysed them by immunoblotting or***
Aspirin, age, and proximity to lymphoid nodules influence cell proliferation parameters in rat colonic crypts
Authors: Barnes C. J. , Lee M. , Hardman W. E. , Cameron I. L.
Published: May 2008

Recent epidemiological studies have demonstrated a correlation between regular aspirin (acetylsalicylic acid) use and decrease risk for the development of fatal colorectal cancer. An increase in the size of the cell proliferation compartment in colorectal crypts has been correlated with an increased risk for the development of colon cancer in animals and in humans. To determine if acetylsalicylic acid acts to decrease the size of the cell proliferation compartment, young (3 month) and old (22 month) rats were treated intragastrically with: 1 the vehicle for acetylsalicylic acid delivery (0.25% wt/vol carboxymethylcellulose in 0.15 N HCI), 2 a single dose of acetylsalicylic acid (100 mg/kg), or 3 acetylsalicylic acid (30 mg/kg) given daily for 30 days. One day after the last treatment, colons were resected, fixed, sectioned and mounted on slides for immunohistochemical staining with a monoclonal antibody to proliferating cell nuclear antigen to assess cell proliferation parameters in the colonic crypts. The results were subjected to three way analysis of variance to assess the effects of: 1 rat age, 2 acute or chronic acetylsalicylic acid treatment, and 3 location of crypts over and away from aggregates of lymphoid nodules on the crypt proliferative parameters. Results demonstrated that: 1 acetylsalicylic acid treatment caused an overall decrease in the proliferative zone height, as measured in number of cells in the crypt column, 2 that crypts located over aggregates of lymphoid nodules had significantly higher proliferative activity than crypts located away from aggregates of lymphoid nodules, and 3 after chronic acetylsalicylic acid treatment there was a greater suppression of proliferative zone height in the crypts of old rats than in the crypts of young rats. In conclusion, acute and chronic intragastric delivery of acetylsalicylic acid caused an overall downward shift in the cell proliferation compartment of colonic crypts of young and of old rats. Whether or not acetylsalicylic acid administration will cause the same proliferative zone height response in carcinogen‐treated rats is not yet established.
Trophic effects of essential fatty acids on pig skin
Authors: Morris G. M. , Hopewell J. W. , Ross G. A. , Whitehouse E. , Wilding D. , Scott C. A.
Published: May 2008

The daily oral administration of 3 ml of two oils (So‐5407 and So‐1129) containing essential fatty acids (EFAs) for 16 weeks resulted in a transient increase in cell proliferative activity in the skin of female Large White pigs. The So‐5407 oil contained 7% gamma‐linolenic acid (GLA) whereas So‐1129 was an oil of similar composition, but with no GLA.

Hyperplasia of the epidermis was observed after the administration of both oils, and this was characterized by an increase in the size of the rete pegs. The maximum effect occurred at 4 weeks after the start of oil administration, at which time the number of viable cell layers had increased by a factor of approximately 1.5, and mean epidermal thickness (excluding the stratum corneum) was approximately 40% greater than that of the epidermis prior to oil administration. There was a marked increase in the labelling index (LI) of the basal cell layer of the epidermis in pigs receiving So‐5407. Maximum LIs were quantified at 4 weeks after the start of administration and were 18.8 ± 1.3% and 13.1 ± 1.7% for pigs receiving So‐5407 and So‐1129, respectively. After this time the LI declined prgressively and had returned to values within normal limits (P>0.1) by 8 weeks after the start of administration of both oils. A similar pattern of change in the LI was seen in the follicular epithelium, although the peak values at 4 weeks after the start of oil administration of 12.2 ± 1.8% and 10.8 ± 0.9 for the groups receiving So‐5407 and So‐1129, respectively, were lower than in the epidermis. Labelled cells were also counted in the papillary dermis and maximum values were again seen at 4 weeks after the start of oil administration. Of the two oils, So‐1129 had the greatest effect, with the number of labelled cells in the papillary dermis being a factor of three to four‐fold higher than in skin prior to oil administration, between 2 and 12 weeks after the start of administration.
Regulation of thymosin β4 mRNA levels during cell proliferation
Authors: Zalvide J. B. , Alvarez C. V. , Vidal A. , Dieguez C. , Vega F. V. , Domínguez F.
Published: May 2008

The levels of thymosin β4 mRNA were studied throughout the cell cycle of NIH 3T3 cells. In serum deprived, quiescent cells, the levels of thymosin β4 were undetectable; after serum restoration, the cells were induced to proliferate and we found a pronounced increase in thymosin β4 mRNA levels at the G1/S transition. Thymosin β4 mRNA was induced even in the presence of cycloheximide. On the other hand, cycling cells that were synchronized at different stages of the cycle by means of mitotic shake‐off after nocodazole arrest or a double thymidine block did not show any variation in the levels of thymosin β4 mRNA when they progressed synchronously through the cycle. In conclusion, the present data indicate that the thymosin β4 gene is regulated by cell proliferation but it is not a cell cycle‐regulated gene. Finally, we studied thymosin β4 mRNA stability by inhibiting thymosin β4 gene transcription with actinomycin D. Our results suggest that thymosin β4 mRNA has a pronounced stability, a fact that might be relevant to account for the presence of thymosin β4 in enucleated cells like platelets.
A quantitative method for the analysis of mammalian cell proliferation in culture in terms of dividing and non‐dividing cells
Authors: Sherley J. L. , Stadler P. B. , Stadler J. Scott
Published: May 2008

The application of the exponential growth equation is the standard method employed in the quantitative analyses of mammalian cell proliferation in culture. This method is based on the implicit assumption that, within a cell population under study, all division events give rise to daughter cells that always divide. When a cell population does not adhere to this assumption, use of the exponential growth equation leads to errors in the determination of both population doubling time and cell generation time. We have derived a more general growth equation that defines cell growth in terms of the dividing fraction of daughter cells. This equation can account for population growth kinetics that derive from the generation of both dividing and non‐dividing cells. As such, it provides a sensitive method for detecting non‐exponential division dynamics. In addition, this equation can be used to determine when it is appropriate to use the standard exponential growth equation for the estimation of doubling time and generation time.
Streaming of labelled cells in the conjunctival epithelium
Authors: Zajicek G. , Perry A. , Pe'er J.
Published: May 2008

This study examines epithelial cell streaming and turnover in normal rat bulbar conjunctiva. Twenty seven male adult random‐bred Hebrew rats weighing between 250–300 g, were injected i.p. with [3H]‐thymidine. Three rats were killed at various times, thereafter from 1 h to 28 days. The enucleated eyes were fixed in formalin, cut into 5 μ thick sections, dipped into liquid emulsion, exposed for three weeks and stained with haematoxylin and eosin. Conjunctival epithelium was scanned from the limbus and outward, using an ocular micrometer grid with 10 x 10 divisions. In each consecutive field the grid was positioned along the basement membrane which was defined as the x‐axis. The y‐axis extended from the basement membrane outward. The x, y coordinate of each nucleus with three grains or more and its grain content were recorded along the entire epithelium.

Conjunctival epithelium is divided into two cell kinetic compartments: a progenitor (P), along the basal and supra basal layer, in which cells proliferate, and a non proliferating Q‐compartment, in the layers above. One hour after labelling most of the labelled cells were in the basal and supra basal layers. From then onward labelled cells streamed along both axes. Their x‐velocity was 10·5±2·4 μ/day and the y‐velocity 9·3 ± 5·4 μ/day. Cells are eliminated at the epithelial surface which is the outer Q‐compartment boundary. Basal cell turnover was estimated from grain count dilution curves. The time it takes for the grains in a cell to reach half of their initial value was 8·3 days. It is closely related to the cell's generation time.

The present study demonstrates that conjunctival epithelium in the rat streams along two axes, x, and y: 1 The x‐axis extends along the basal layer, from the limbus and outward. 2 The y‐axis extends from the basal layer into the layers above it. Cells first stream along the x‐direction and then turn y‐ward. Since cells are ultimately exfoliated from the conjunctival surface, and since the conjunctiva maintains steady state, we propose that stem cells located in the limbus generate transitional cells that stream along the two axes. Macroscopically the limbus is circular, and the stem cells are situated around the cornea. Each stem cell and its streaming progeny can be viewed as a conjunctival epithelial unit.

We propose that conjunctival and corneal epithelium, are the descendants of an uncommitted stem cell that generates two differentiation pathways, a corneal and a conjunctival.
Levels of selected growth factors in viable and necrotic regions of xenografted HCT‐8 human colon tumours
Authors: Leith J. T. , Michelson S.
Published: May 2008

Xenografted tumours were produced in nude mice by injection of HCT‐8 human colon tumour cells. At average volumes of about 750 mm3, animals were injected with fast green vital dye, and 20 min later, tumours were excised and dissected into viable (stained) and necrotic portions (unstained). Viable and necrotic regions were then examined for cell yields, colony forming efficiencies, and levels of basic fibroblast growth factor (FGF‐2), transforming growth factors‐β1 and ‐α (TGF‐β1, TGF‐α), platelet derived growth factor (PDGF), and vascular endothelial growth factor (VEGF) using enzyme‐linked immunoassay (ELISA) procedures. Levels in the viable and necrotic regions were compared to levels in unseparated tumours. The average extent of necrosis in HCT‐8 tumours of this size was 64%. The data for cell yields, colony forming efficiencies FGF‐2, VEGF, TGF‐β1 and TGF‐α indicated that values determined in the unseparated tumours could be understood on the basis of the weighted average between viable and necrotic tissue, with the higher values occurring in the viable tissue. Low levels of FGF‐2 and VEGF were found in the necrotic portions of the tumour while no measurable levels of TGF‐β1 and TGF‐α could be determined. PDGF levels were, however, equivalent in both the viable and necrotic regions indicating that necrotic tissue could be an important reservoir for this growth factor.
Evidence for quiescent S‐ and G2‐phase cells in human colorectal carcinomas: a flow cytometric study with the Ki‐67 antibody
Authors: Zölzer F. , Speer A. , Pelzer T. , Streffer C.
Published: May 2008

The expression of certain antigens specific for proliferating cells can be determined simultaneously with cell cycle distribution by means of two‐dimensional flow cytometry. In this way, a tumour's growth potential is characterized more precisely than with any one parameter alone. Here we describe such simultaneous measurements of DNA content and labelling with the Ki‐67 antibody that distinguishes between cycling and non‐cycling cells. Having overcome a number of technical problems we were able to analyse material from 29 biopsies of human colorectal tumours.

In a number of cases, Ki‐67 negative cells were found with a DNA‐content of G0/1 only, whereas all cells with an S‐ or G2‐phase DNA‐content were Ki‐67 positive. There were other cases in which cells with an S‐ and G2‐phase DNA‐content had obviously become quiescent (Ki‐67 negative), sometimes even outnumbering the proliferating (Ki‐67 positive) cells in the respective compartments of the cycle. Generally, however, when Ki‐67 negative and positive subpopulations were analysed separately it was found that the former had a significantly lower (S + G2)‐phase fraction than the latter.

There was evidence for a correlation between Ki‐67 index and (S + G2)‐phase fraction at least in the subgroup of aneuploid tumours. Neither of the two parameters was correlated with stage according to Duke's classification or tumour size. However, a positive correlation was found between the fraction of unlabelled S‐ and G2‐phase cells and tumour size as reflected in the T category.
Threshold expression of cyclin E but not D type cyclins characterizes normal and tumour cells entering S phase
Authors: Gong J. , Traganos F. , Darzynkiewicz Z.
Published: May 2008

Complexes of cyclin‐dependent kinases (cdk) and their partner cyclins drive the cell through the cell cycle, each such complex phosphorylating a distinct set of proteins at a particular check‐point or phase of the cycle. Immunocytochemical detection of cyclins combined with measurement of cellular DNA content by flow cytometry makes it possible to relate expression of each of these proteins with the actual cell cycle position, without the necessity of cell synchronization. In the present study, we have investigated expression of E and D type cyclins in G1 cells and in cells entering S phase, in eight different human hematopoietic and solid tumour cell lines (two leukaemias, a lymphoma, three breast carcinomas, a colon carcinoma and a bladder transitional cell carcinoma) during their exponential phase of growth, as well as in normal mitogen stimulated lymphocytes. In all the cell types studied, the average level of D type cyclin expression was invariable throughout the cell cycle. A great intercellular variability, in particular of the G1 cell subpopulations, and the presence of a large fraction of G1, S and G2+ M cells that were cyclin D negative (20–40% in tumour cell lines and about 80% among lymphocytes), were other characteristic features of D type cyclin expression. In contrast to D type cyclins, the expression of cyclin E was discontinuous during the cycle, peaking at the time of cell entrance to S. Also, a well defined threshold in expression of cyclin E characterized cells that were entering S phase, and virtually no cyclin E negative cells were seen during the early portion of S phase. The data indicate that while cell entrance to S phase is unrelated to expression of D type cyclins (at the time of entrance), accumulation of cyclin E up to critical level is a prerequisite for initiation of DNA replication. The great intercellular variability in expression of D type cyclins and their invariant average level across the cell cycle suggest that these cyclins, in addition to their acknowledged function in promoting cell progression through mid‐ to late‐G1 may have other role(s), related or unrelated to the cell cycle progression. The presence of a large number of D type cyclin negative cells in all phases of the cycle suggests that during exponential growth the cells may not express this protein and yet may traverse the cycle, including G1 phase.
Regrowth resistance in cancer: why has it been largely ignored?
Authors: Preisler H. D. , Venugopal P.
Published: May 2008

In drug‐sensitive malignancies, treatment failure is a complex phenomenon resulting from both drug resistance and the rapid regrowth of malignant cells in between cycles of chemotherapy. This phenomenon has been referred to as regrowth resistance, and has been identified as a common impediment to the successful treatment of many neoplastic diseases. This phenomenon has received little attention in the past. The administration of maximally tolerated individual dosages of chemotherapeutic agents, combined with the use of biological response modifiers to slow the regrowth between courses of therapy, would be an effective approach to reduce the regrowth resistance. This also could reduce the emergence of drug resistant clones.
An analysis of the growth of the retinal cell population in embryonic chicks yielding proliferative ratios, numbers of proliferative and non‐proliferative cells and cell‐cycle times for successive generations of cell cycles
Authors: Morris V. B. , Cowan R.
Published: May 2008

Growth curves of the retinal cell population of embryonic chicks were fitted by a branching‐process model of cell population growth, thereby estimating the proliferative ratios and mean cell‐cycle times of the generations of cell cycles that underlie retinal growth. The proliferative ratio determines the proportion of cells that divides in the next generation, so the numbers of proliferative and non‐proliferative cells in each generation of cell cycles were obtained. The mean cell‐cycle times determine the times over which the generations are extant. Assuming growth starts from one cell in generation 0, the proliferative cells reach 3.6 × 106 and the non‐proliferative cells reach 1.1 × 106 by generation 23. The next four generations increase the proliferative cell numbers to 13.9 × 106 and produce 20.1 × 106 non‐proliferative cells. In the next five generations in the end phase of growth, non‐proliferative cells are produced in large numbers at an average of 13.9 × 106 cells per generation as the retinal lineages are completed. The retinal cell population reaches a maximum estimated here at 98.2 × 106 cells. The mean cell‐cycle time estimates range between 6.8 and 10.1 h in generations before the end phase of growth and between 10.6 and 17.2 h in generations in the end phase. The retinal cell population growth is limited by the depletion of the proliferative cell population that the production of non‐proliferative cells entails. The proliferative ratios and the cell‐cycle‐time distribution parameters are the likely determinants of retinal growth rates. The results are discussed in relation to other results of spatial and temporal patterns of the cessation of cell cycling in the embryonic chick retina.
In vivo labelling with halogenated pyrimidines of squamous cell carcinomas and adjacent non‐involved mucosa of head and neck region
Authors: Kotelnikov V. M. , Coon J. S. , Taylor S. , Hutchinson J. , Panje W. , Caldarelli D. D. , LaFollette S. , Preisler H. D.
Published: May 2008

The frequency and distribution of labelled cells were studied immunohistochemically in 37 squamous cell carcinomas (SCC) of head and neck after in vivo infusion of IdUrd and BrdUrd. Tumours were classified according to their labelling patterns. Low and moderate grade SCC consisted of tumour islands separated by interstitial tissue. In some tumours labelled cells only appeared near the basal layer while in others proliferative cells were evenly distributed within the neoplastic island. In anaplastic carcinomas labelled cells were distributed either randomly or around blood vessels (cord structures). While the basal layer in adjacent normal epithelium contained very few labelled cells (LI = 1.6 ± 0.2%), the LI of basal cells in tumour islands were much higher than the average LI of the tumour (47.2 ± 2.8% and 23.8 ± 1.6%, respectively). In patients who had received cytotoxic therapy up to two months before the biopsy, the LI in the basal layer of normal epithelium was 19.0 ± 3.5%. In sequential biopsies obtained 1–2 weeks after the infusion of IdUrd and BrdUrd some labelled tumour cells were found in necrotic foci and in pearl structures. Additionally, in six tumours, we found areas of cells labelled with IdUrd alone, even though the IdUrd infusion had been followed by a BrdUrd infusion 1 h later. This is in agreement with the phenomenon of intermittent tumour blood flow described earlier in experimental tumours.
Comparison of mathematical formulas used for estimation of DNA synthesis time of bromodeoxyuridine‐labelled cell populations with different proliferative characteristics
Authors: Johansson M. C. , Baldetorp B. , Bendahl P.‐O. , Fadeel I. A. , Oredsson S. M.
Published: May 2008

Abstract. Growth kinetic data of human tumours, obtained by flow cytometric analysis of cells labelled with bromodeoxyuridine (BrdUrd) might provide prognostic information and allow prediction of response to radio‐ and chemotherapy. However, the theoretical models applied for calculation of growth kinetic data are not fully evaluated. The purpose of this study was to investigate the dependence of the estimation of DNA synthesis time (Ts) on sampling time after BrdUrd labelling, using four different mathematical formulas (Begg et al., White & Meistrich, White et al. and Johansson et al.) which have been developed for the evaluation of flow cytometry‐derived data of BrdUrd‐labelled cells. In addition, we have investigated the influence of the growth kinetic properties of the cell populations using two cultured cell lines (one slow and one fast growing), and two heterotransplanted human tumours. The dependence of the estimation of Ts on sampling time was more or less pronounced, depending on the cell population examined and on the formula used. In the fast growing cell line, the estimates of Ts did not vary significantly with sampling time when using the formulas by White et al., whereas in the slow growing cell line, the estimates of Ts did not show any significant dependence on sampling time when using the formula by Johansson et al. In the tumours, the estimation of Ts depended on sampling time with all formulas used, although to different degrees. In one of the tumours, this was mainly caused by the influence of mouse cells, as we demonstrate. Our results indicate that the proliferative characteristics of a cell population should be taken into consideration when choosing a mathematical formula in order to attain Ts values that are independent of sampling time.
Cell proliferation and nuclear abnormalities are increased and apoptosis is decreased in the epidermis of the p53 null mouse after topical application of benzo[a]pyrene
Authors: Miller M. L. , Andringa A. , Cody T. , Dixon K. , Albert R. E.
Published: May 2008

Abstract. Cell proliferation and cell death in mouse epidermis are altered by topical application of benzo[a]pyrene (BaP), a procarcinogen, which yields metabolites that can form DNA adducts. The mitotic rate, nuclear abnormalities, labelling index, grain density, necrosis and apoptosis were compared in the epidermis of TSG‐p53 null (p53‐/‐) and C57BL wild‐type (wt) mice after weekly treatments with BaP to determine whether the absence of the p53 gene altered cytokinetic responses to DNA damaging agents in vivo. Acetone alone or 64 μg BaP in 50 μl acetone was applied to the clipped dorsum of mice once, or in four consecutive weekly treatments. Indices of cell proliferation and cell death were the same in both wt and p53‐/‐ mice treated only with acetone. One application of BaP depressed mitosis and slowed the rate of DNA synthesis in both genotypes. After four applications of BaP the number of keratinocytes in S phase increased substantially, while there was no further slowing in the rate of S phase in the wt and p53‐/‐ mice. Cell proliferation rates and numbers of cells with nuclear abnormalities were higher and there were fewer apoptotic cells and apoptotic bodies in the p53‐/‐ mice than in the wt mice. Numbers of ‘sunburn’cells were similar in both types.
Elevated levels of ERK2 in human breast carcinoma MCF‐7 cells transfected with protein kinase Cα
Authors: Gupta A. K. , Galoforo S. S. , Berns C. M. , Martinez A. A. , Corry P. M. , Guan K.‐L. , Lee Y. J.
Published: May 2008

Abstract. We investigated the effect of elevated levels of protein kinase Cα (PKCα) on cell proliferation in human breast carcinoma cells (MCF‐7). MCF‐7 cells transfected with either the pSV2M(2)6 vector without the insert (MCF‐7/Vector) or containing a full length cDNA encoding PKCα (MCF‐7/PKCα) were compared. MCF‐7/PKCα cells were found to have an increased proliferative rate with a doubling time of 15 h as compared to 42 h for MCF‐7/Vector cells. Flow cytometry illustrated a greater percentage of MCF‐7/PKCα cells in the S phase of the cell cycle. Western and Northern blot analyses demonstrated an increase in extracellular regulated protein kinase 2 (ERK2) gene expression in MCF‐7/PKCα cells but no alteration of this gene expression in MCF‐7/Vector cells. These results suggested that the elevated level of ERK2 which is also known as mitogen activated protein kinase is probably involved in the increase in MCF‐7/PKCα cell proliferation.
Aspirin suppresses 1,2‐dimethylhydrazine‐induced alteration of proliferative parameters in rat colonic crypts
Authors: Barnes C. J. , Lee M. , Hardman W. E. , Cameron I. L.
Published: May 2008

Abstract. Human epidemiological reports and rodent experimental research data indicate a possible chemopreventive effect of regular aspirin use for decreasing risk of colon and rectum cancer incidence and mortality. We have previously demonstrated that aspirin can significantly suppress proliferative parameters in normal rat colonic epithelium when examined 24 h following an acute or chronic course of aspirin administration. To investigate whether aspirin would effectively suppress known carcinogen‐induced changes in colonic epithelium, rats were given single s.c. injections of either aspirin (50 mg/kg bw) or saline on days 1–3 and either 1,2‐dimethylhydrazine (DMH; 12 mg base/kg bw) or DMH vehicle on day 4 of each week for eight consecutive weeks. Rats were sacrificed 4 days after the last aspirin dose and 3 days after the last DMH or DMH vehicle dose. Using the proliferative biomarkers of proliferating cell nuclear antigen positive cells per midaxial crypt section (SCC), crypt proliferative zone height (PZ), crypt differentiated zone height (DZ), and total crypt height (CH), it was found that aspirin does suppress DMH‐induced increases in SCC, PZ and CH. The findings demonstrate that aspirin has a long term (i.e. several days) protective effect against early carcinogen‐induced proliferative changes in rat colonic crypts which may help account for aspirin's chemopreventive action against colon cancer.
Increased autocrine production of insulin‐like growth factor II (IGF‐II) alters serum sensitivity of MCF‐7 human breast cancer cell proliferation
Authors: Corcoran D. , Perachiotti A. , Darbre P. D.
Published: May 2008

Abstract. Regulation of the growth of breast cancer cells is the result of a complex interaction between steroid hormones and growth factors, and in particular of oestrogen and insulin‐like growth factors (IGF). Alteration of any one mitogenic component can affect the cell response to other pathways. Previous work has shown that increased autocrine production of IGF‐II from a transfected inducible expression vector can result in reduced oestrogen sensitivity of growth of MCF‐7 human breast cancer cells. This report describes alterations to non‐oestrogen regulated pathways of cell growth following enhanced IGF‐II expression in these transfected MI7 cells. Serum sensitivity of cell growth in the absence of oestrogen was found to differ between MI7 and untransfected MCF‐7 cells, in that growth of MI7 but not MCF‐7 cells was strongly inhibited by high serum levels. Increased serum had no effect on levels of IGF‐II mRNA, IGFIR, IGFBP4 mRNA, or IGFBP secreted in MI7 cells. However, growth inhibition by serum in MI7 cells could be overcome by increasing levels of IGF‐II in the serum or by removal of IGFBP onto polycarbonate membranes. Thus, the growth inhibition by serum in MI7 cells is concluded to result from the increased levels of IGFBP added with higher serum. This would support an inhibitory role for IGFBP on growth of breast cancer cells when cell growth is being driven by IGF pathways in the absence of oestrogen, and would suggest that cellular sensitivity to such factors can depend on levels of endogenous IGF production.
Tissue kinetics in mouse tongue mucosa during daily fractionated radiotherapy
Authors: Dörr W. , Emmendörfer H. , Weber‐Frisch M.
Published: May 2008

Abstract. The purpose of the present investigation was to quantify cell flux between the distinct layers of the epithelial lining of the ventral surface of mouse tongue during daily fractionated radiotherapy.

A fraction of DNA‐synthesizing cells in control epithelium, or at various days through a course of daily fractionated radiotherapy with 3 or 4 Gy per day, was labelled with [3H]dT or BrdUrd, respectively. The labelling indices (LI) in the different epithelial layers were defined histologically after autoradiography, or immunohistochemistry, at intervals between 1 and 10 days after label administration.

In tongue epithelium of untreated mice, the minimum residence time of cells in the germinal layer is 2–3 days. Migration through the functional layers requires an additional 2–3 days before labelled cells are observed in the most superficial layer of nucleated cells. A plateau in LI is observed for several days post‐labelling in control epithelium, indicating an equilibrium between loss and proliferation of labelled cells. During fractionated radiotherapy, the minimum time from division to occurrence of labelled cells in the stratum lucidum is less than 2 days, and hence significantly shorter than in control epithelium. In contrast to untreated epithelium, no plateau in the germinal layer LI is seen, indicating that frequently both labelled daughters from dividing labelled cells are being lost from this compartment.

In conclusion, the present data support a recently described model of radiation‐induced accelerated repopulation in squamous epithelia, which postulates that the majority of damaged cells undergoes abortive’divisions resulting in two differentiating daughters.
Interleukin‐8 and de novo mammalian angiogenesis
Authors: Norrby K.
Published: May 2008

Abstract. In the rat mesenteric‐window angiogenesis assay (MWAA), the test tissue is natively vascularized, lacks significant physiological angiogenesis and its homeostasis is unperturbed by surgical intervention. Using the rat MWAA, it is shown here that interleukin‐8 (IL‐8), administered at approximately physiological doses, is able to induce de novo angiogenesis. Human recombinant IL‐8 was administered intraperitoneally at two daily doses of 25 pM, 250 pM and 2.5 nM for 5 consecutive days (days 0–4). Using microscopic, computer‐aided techniques including image analysis, the de novo angiogenic response was quantified in groups of animals on days 7, 14 and 21 in terms of the relative vascularized area (VA), a measure of the microvascular spatial extension, and the microvascular length (MVL), a measure of microvascular density or length. The total microvascular length (TMVL) was computed from VA × MVL. A statistically significant angiogenic effect was found in terms of MVL on day 7 and in terms of VA and TMVL on day 14 following the treatment with 2.5 nM, whereas MVL was significantly increased in statistical terms on day 14 following the treatment with IL‐8 at the low dose of 25 pM. Notably, IL‐8 at the intermediate dose of 250 pM did not induce a statistically significant angiogenic effect in terms of VA, MVL or TMVL on any observation occasion, thereby suggesting that the dose‐related angiogenic effect of IL‐8 may be nonlinear. This appears to be the first paper showing that IL‐8 is able to induce de novo angiogenesis in normally vascularized mammalian tissue.
Growth kinetics of the granulosa cell population in ovarian follicles: an approach by mathematical modelling
Authors: Cleément F. , Gruet M. A. , Monget P. , Terqui M. , Jolivet E. , Monniaux D.
Published: May 2008

This paper describes, from a mathematical viewpoint, the cellular changes in the granulosa of ovarian follicles during their terminal development. A dynamic model takes into account the processes of (1) cell division, (2) exit from the cell cycle towards differentiation, and (3) apoptotic cell death. Proliferative cells leave the cycle in an irreversible way. The risk of entering apoptosis applies to non‐cycling cells. Changes in the cell numbers and in the growth fraction are derived from differential equations. The transitions between the different cell states are ruled by time‐dependent rates. Numerical applications of the model concern ovulating and degenerating ovarian follicles in the ewe. The main feature of the ovulating case is the progressive exhaustion of the proliferating compartment for the benefit of the non‐cycling cells. From an initial mainly proliferative state the granulosa progressively switches to a highly differentiated state, so that the growth fraction continuously decreases. In the atretic cases, the pattern of changes in the total viable cell number is influenced by the follicular age at the onset of the apoptotic process and by the intensity of the cell death rate. As apoptosis affects the non‐cycling cells, the growth fraction is no longer strictly decreasing. The sensitivity of the model to the parameters is studied in a more general framework than the granulosa cell population.
A polymerase chain reaction assay for simultaneous detection and quantitation of proto‐oncogene and GAPD mRNAs in different cell growth rates
Authors: Goswami P. C. , Albee L. D. , Spitz D. R. , Ridnour L. A.
Published: May 2008

A reverse transcriptase followed by a polymerase chain‐reaction (RT‐PCR) assay was developed for the simultaneous detection and quantitation of proto‐oncogene (c‐fos and c‐myc) mRNAs using an internal standard mRNA glycer‐aldehyde‐6‐phosphate dehydrogenase (GAPD). Total cellular RNA was reverse transcribed and PCR amplified with oligonucleotide primers specific to GAPD and either c‐fos or c‐myc genes. In contrast to Northern blot analysis, the RT‐PCR assay is rapid and sensitive enough to quantitate specific proto‐oncogene levels from as little as 12–25 ng of total cellular RNA. The reliability of the assay was tested by measuring c‐fos and c‐myc expression in C3H 10T1/2 mouse embryo fibroblast cells under two different growth states: (a) quiescent cell entry into the proliferative cycle, and (b) plateau phase. Furthermore, the assay was used in measuring variations in c‐fos or c‐myc expression in HA‐1 hamster cells following exposure to the cellular stressing agent, nitric oxide. In serum‐stimulated cells, the RT‐PCR measurements of transient increase in c‐fos (16‐fold at 30 min) and c‐myc (10‐fold at 1 h) mRNA levels were comparable to previously reported results in the literature using a Northern blotting assay. In addition, a two‐ to fivefold increase in c‐fos mRNA levels was observed in plateau phase cells when compared to log phase growth. Furthermore, a transient increase in c‐fos mRNA levels (threefold at 2 h) was also observed following cells’ exposure to the stressing agent nitric oxide. These results suggest that the multiplex RR‐PCR assay represents a significant improvement over current methods to quantitate specific cellular mRNAs under different growth conditions or following environmental insults.
A novel growth stimulating activity from BRL‐3A cell conditioned medium
Authors: Xu S.‐Q. , Sell C. , DuBois G. C. , Baserga R.
Published: May 2008

The R– cell line is a 3T3‐like cell line originating from mouse embryos with a homozygous disruption of the type 1 insulin‐like growth factor receptor (IGF‐IR) genes. Although R– cells cannot grow at all in serum‐free medium (SFM) supplemented by several known growth factors, either singly or in combination, they are able to grow in 10% serum, albeit at a reduced rate. These findings suggested that serum contains an unknown, or unidentified, growth factor that can promote cell growth even in cells devoid of IGF‐IRs. In an effort to identify such growth factor, we searched, using R– cells, for a growth and DNA synthesis stimulating activity in SFM conditioned by different cell lines. We found that the BRL‐3A cell line secreted an activity capable of stimulating DNA synthesis and cell proliferation in R– cells. This activity (which is concentration‐dependent) can be collected and concentrated by ultrafiltration, it is heat‐labile, proteinase K‐sensitive and has a size larger than 10 kDa. Because of the resistance of R– cells to stimulation by known growth factors, we believe that this activity is due to a novel polypeptide secreted by BRL‐3A cells. Further characterization of the active component(s) is in progress.
Perceptions of health and health service utilization among homeless and housed psychiatric consumer/survivors
Authors: FORCHUK C. , BROWN S. A. , SCHOFIELD R. , JENSEN E.
Published: May 2008

Homelessness has a direct impact on health. Homeless individuals report several barriers to accessing health care. Although research exists regarding the utilization of health services for homeless and housed psychiatric consumer/survivors, few studies have compared the perceived health and service utilization of these two groups. The objective of this study was to determine whether or not differences exist between the utilization of health services and the perceptions of health of homeless and housed psychiatric consumer/survivors in London, Ontario, Canada. It was hypothesized that differences would exist between homeless and housed psychiatric consumer/survivors on all health‐related variables examined. A secondary analysis of quantitative data was conducted in a Community–University Research Alliance on Mental Health and Housing project funded by the Social Sciences and Humanities Research Council of Canada. Key findings include significant differences in the characteristics of each population, the use of health services and their perceptions of health. Implications for practice and policy are discussed.
Wechselwirkung zwischen Gelber‐Cercospora‐Substanz und Schwermetall‐Ionen
Authors: Schlösser E. , Stegemann H.
Published: May 2008

Aus Cercospora heticola konnte eine biologisch aktive, gelb gefärbte Fraktion (GF) isoliert werden (SCHLOSSER 1962). Diese crgab mit methanolisdier FeCl3Lösung eine dunkelbraun gefärbte Komplexverbindung. Aus diesem Grunde wurde die Affinität zu physiologisch bedeutsamen Schwermetallionen geprüft, wobei das Mg‐Ion wegen seiner Bedeutung für den Stoffwechsel mit einbezogen wurde. Es interessierte ferner die relative Stabilität der gebildeten Komplexe sowie ihre biologische Aktivität.
Production of Cytokinin‐like Substances by Cylindrocarpon destructans (Zins.) Scholt. Isolates Pathogenic and Nonpathogenic to Fir (Abies alba Mill.) seedlings
Authors: Strzelczyk E. , MariaKampert
Published: May 2008

The production of cytokinin‐like substances by different strains of Cylindrocarpon destructans either pathogenic or non pathogenic to fir (Abies alba Mill.) was studied. The cytokinins were extracted from the fungal culture media and separated by means of column chromatography. The biological activities in different fractions were assayed by callus growth of soybean (Glycine max Merill var. Acme). The amount of these substances was calculated on the basis of fresh weight of callus obtained from cultures containing authentic cytokinin.

Cytokinin‐like substances were detected in 5 of 6 pathogenic isolates tested and in 2 of 5 non‐pathogenic isolates.

On the basis of their occurrence in particular column fractions, most of the substances seem to be identical or similar to zeatin and zeatin riboside. In addition the fungi studied produced other, not identified cytokinin‐like substances. The synthesis of Ri P can not be excluded. However the method applied did not allow to separate Ri P from zeatin and zeatin riboside.
Detection of Four Seed‐borne Plant Viruses by the Enzyme‐Linked Immunosorbent Assay (ELISA)
Authors: Faris‐Mukhayyish S. , Makkouk K. M.
Published: May 2008

This study was conducted to evaluate the sensitivity of the ELISA technique in detecting four economically important viruses, namely barley stripe mosaic (BSMV), cucumber green mottle mosaic (CGMMV), bean common mosaic (BCMV), and squash mosaic (BSMV) viruses in single seeds as well as in batches of barley, cucumber, bean and squash seeds, respectively. Results indicated the suitability of the technique in detecting the above viruses in single germinated seeds or embryos. Accordingly, seed transmission rates of BSMV, CGMMV, BCMV and SqMV were found to be 67 %, 17%, 17% and 12%, respectively. In artificially contrived mixtures of infected: healthy seeds or embryos, BSMV, CGMMV, BCMV and SqMV were successfully detected at ratios of 1 : 500, 1 : 25, 1 : 10 and 1 : 10, respectively. Sensitivity of detection was increased in the ease of BSMV by using germinated rather than ground dry BSMV‐infected barly seeds; and in the case of SqMV, by using whole germinating emybryos rather than coleoptiles only. Trials on re‐using the enzyme‐γ‐globulin conjugate indicated that CGMMV conjugate used once can be re‐used with little loss in reactivity.