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1.
A quantitative immunochemical study was carried out of four enzymes, cathodal esterase, acid phosphatase, glucosaminidase and β-glucuronidase. In homogenates of the human diploid cell line WI-38, the relative amounts of the enzymes increased with the passage number of the culture, although great variation was found in later passages just before death of the culture. 相似文献
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《The Journal of cell biology》1979,83(2):487-498
WI-38 and SV40WI-38 cells have been synchronized using centrifugal elutriation. This technique allows for the rapid harvesting of early G1 phase cells from exponentially growing populations of both the normal and transformed cell. Using these cells, as well as WI-38 cells synchronized by serum deprivation, we have examined the effects of extracellular Ca and Mg levels on the progression of cells through G1 phase. A differential sensitivity to both Ca and Mg deprivation is observed between normal and transformed cells. The WI-38 cell requires higher levels of both ions for traversal of G1 phase and for continued proliferation as compared to the transformed cell. The temporal nature of the Ca and Mg requirements for the WI-38 cell has been examined during G1 phase. Ca is strictly required during early and late G1 phase, but not necessarily throughout mid-G1. An early as well as a late G1 Ca requirement is also found in serum-stimulated WI-38 cells. In contrast, the Mg requirement of WI-38 cells does not appear to be temporally well-defined. Mg appears to be a permissive factor, required throughout G1 phase rather than at certain prescribed intervals. On the basis of these data, it seems unlikely that these two cations exert their effects on cell growth entirely through a common competitive mechanism. Ca would appear to be involved in early serum or growth factor-mediated G1 events and later pre-S-phase events, as suggested in previous studies on other cell lines. 相似文献
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A brief exposure of quiescent (Go) WI-38 human fibroblasts to the tumor promoter TPA results in an increase in the mRNA levels of c-fos protooncogene. The same effect is produced by exposing to TPA human diploid fibroblasts WI38 synchronized in S phase by treatment with 2.5 mM hydroxyurea. Induction of c-fos mRNA in response to TPA occurs also during the progression of synchronized WI38 throughout the second and third cell cycle, but it is not associated with measurable changes in the cell cycle progression of these cells. These findings suggest that TPA induction of c-fos mRNA levels in proliferating cells is a stimulus specific rather than a function specific event. 相似文献
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Propagation of the human diploid cell WI-38 in galactose medium 总被引:2,自引:0,他引:2
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Assessment of the respiratory and glycolytic capacity of non-growing WI-38 cells shows that, in the absence and presence of added glucose, the mean rates of oxygen consumption were 247 (QO2 = 5.61) and 208 (QO2 = 4.73) mμmoles/mg dry wt/hr., respectively. Mean glucose consumption was 225 mμmoles/mg dry wt/ hr. With uniformly labeled 14C glucose as substrate, 36 mμg atoms of carbon dioxide were produced, corresponding to 15–20% of the total cellular respiration. Mean values for lactate production in the presence and absence of glucose were 345 (QLO2 = 7.85) and 196 (QLO2 = 4.45) mμmoles/mg dry wt/hr., respectively. Human diploid cells in culture age, in the sense that their ability to proliferate decreases with time during serial subcultivation. Studies of their respiratory and glycolytic capacity as a function of the aging process showed that total respiration, glucose respiration and glycolytic capacity were relatively constant for cells in the middle and late passages and indicate that aging in this sense is not directly related to the respiratory and glycolytic capacity of the cell. 相似文献
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Clone size variation in the human diploid cell strain, WI-38 总被引:3,自引:0,他引:3
By mapping the location of isolated single cells; and then counting the number of cells at each location as a function of time. it was possible to accumulate data on the growth history for each of a large group of clones. The clone size distribution, its mean and standard deviation were computed for each day in culture. Variations in schedule of medium change and time of exposure to trypsin, did not measurably affect variation in clone size. Neither could clone size variation be accounted for on the basis of (1) occurrence of nondividing cells nor (2) presence of heritable growth rate variants in the population. It is probable that clone size variation under our conditions is primarily a consequence of a highly variable interdivision time among the constituent cells. 相似文献
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Z. K. Kadanka J. D. Sparkes H. G. Macmorine 《In vitro cellular & developmental biology. Plant》1973,8(5):353-361
Summary Cultures of the human diploid cell strain WI-38 were subcultivated under conditions which would meet the requirements proposed
for the use of this strain as a substrate for the preparation of viral vaccines and would be in keeping with efficient production
procedures. For chromosomal analysis, the cultures were combined in three groups at low, intermediate, and high passage levels,
the latter being beyond those recommended for vaccine production. At all passage numbers, the incidence of aneuploid cells
was low and constant up to those passages where the finite life span was approached and the population doubling time became
markedly prolonged. At all passage levels, the incidence of gaps was higher than that of breaks but there was no significant
increase of either of these abnormalities with continuous subcultivation. Among structural abnormalities dicentrics, despiralizations
and deletions predominated. A significant increase in polyploidy occurred in the highest passage numbers, although the ratio
of polyploidy to endoreduplication was constant throughout the series. Neither heteroploid transformation nor nonrandom chromosomal
aberrations were observed. Nor was there correlation between observed aberrations and their location on the chromosomes. The
incidence of hypodiploidy was lower than reported by other investigators. At the cellular level, no morphological changes
could be associated with the distribution of chromosomal aberrations.
This study was assisted by funds provided by Canadian Public Health Research Grant 605-7-710 of the National Health Grants
Programme. 相似文献
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Shelly A Christman Byung-Whi Kong Megan M Landry Hyunggee Kim Douglas N Foster 《BMC cell biology》2006,7(1):1-12
Background
The present study was carried out to determine whether the p53 pathway played a role in the spontaneous immortalization of the SC-2 chicken embryo fibroblast (CEF) cell line that has been in continuous culture for over three years.Results
The SC-2 cell line emerged from an extended crisis period with a considerably slower growth rate than primary CEF cells. The phenotype of the SC-2 cells changed dramatically at about passage 80, appearing smaller than at earlier passages (e.g., passage 43) and possessing a small, compact morphology. This morphological change coincided with an increase in growth rate. Passage 43 SC-2 cells expressed undetectable levels of p53 mRNA, but by passage 95, the levels were elevated compared to primary passage 6 CEF cells and similar to levels in senescent CEF cells. However, the high level of p53 mRNA detected in passage 95 SC-2 cells did not correlate to functional protein activity. The expression levels of the p53-regulated p21WAF1 gene were significantly decreased in all SC-2 passages that were analyzed. Examination of the Rb pathway revealed that E2F-1 and p15INK4b expression fluctuated with increasing passages, with levels higher in passage 95 SC-2 cells compared to primary passage 6 CEF cells.Conclusion
The present study suggests that altered expression of genes involved in the p53 and Rb pathways, specifically, p53 and p21WAF1, may have contributed to the immortalization of the SC-2 CEF cell line. 相似文献18.
Introduction of wild-type p53 in a human ovarian cancer cell line not expressing endogenous p53. 总被引:5,自引:1,他引:4
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Utilizing a temperature sensitive p53 mutant (pLTRp53cGval135) which expresses mutant p53 at 37 degrees C and a wild-type like p53 at 32 degrees C, we transfected a human ovarian cancer cell line (SKOV3) which does not express endogenous p53. Among the different clones obtained, we selected three clones. Two were obtained from simultaneous transfection of p53 and neomycin resistance expression plasmids (SK23a and SK9), the other was obtained from transfection experiments utilizing the neomycin resistance gene only (SKN). Introduction of mutant p53 did not alter the morphology or growth characteristics of this ovarian cancer cell line. Upon shifting to the permissive temperature, a dramatic change in morphology and growth rate was observed in SK23a and SK9 cells that is associated with the presence of a wild-type like p53. SKN and SKOV3 cells maintained at 32 degrees C did not change morphology and only slightly reduced proliferation. Both SK23a and SK9 cells did not show evidence of apoptosis when measured up to 72 hours of maintenance at 32 degrees C. In contrast to what observed in other cell lines, SK23a and SK9 cells maintained at 32 degrees C were not blocked in G1, but they were accumulated in G2-M. This accumulation was transient and could be due either to a blockade or to a delay in the G2 progression. No down-regulation of c-myc was observed in p53 expressing clones when shifted to the permissive temperature. In these conditions gadd45 mRNA expression was highly stimulated in SK9 and SK23a cells but not in SKN cells. In both clones Gas1 mRNA was not detected either at 37 degrees C or 32 degrees C. This system represents a new and useful model for studying the effect of the absence of p53 (SKOV3 or SKN), presence of mutated p53 (SK23a and SK9 kept at 37 degrees C) or wild type p53 (SK23a and SK9 kept at 32 degrees C) on the mechanism of response of cancer cells to DNA damaging agents. 相似文献
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《Cell cycle (Georgetown, Tex.)》2013,12(19):3074-3082
In mammalian cells, ADP ribosylation factor like 2 (Arl2) has been shown to form a complex with tubulin binding cofactor D (TBC-D) and the tumor suppressor protein phosphatase 2A (PP2A). We have previously shown that alterations in Arl2 protein content were associated with corresponding modifications of the tumor suppressor PP2Ac protein content in breast cancer cells. Here, we show that modified Arl2 expression level influences sensitivity to various anticancer compounds such as taxol, navelbine, gemcitabine and doxorubicin in MCF7 derived cell lines. Modifications of Arl2 expression levels were also associated with an altered phosphorylation status and/or cellular sublocalization of certain PP2A targets such as p53, a key mediator of chemotherapy-induced apoptosis. A decreased level of Arl2 expression was associated with an increase of phospho-ser15-p53, a form which was found to be preferentially bound to microtubules. Assays using okadaic and cantharidic acid, two different PP2A inhibitors, showed an increase in microtubule-bound phospho-p53 and reduced sensitivity to chemotherapy. Our results suggest that Arl2 could, via PP2A, influence p53 phosphorylation status. Certain forms of phosphorylated p53 demonstrating increased binding to microtubules appear to be less prone to nuclear translocation after exposure to chemotherapeutic agents, thereby possibly contributing to reduced chemosensitivity. 相似文献
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Katherine H. Chen Mercedes A. Paz Paul M. Gallop 《In vitro cellular & developmental biology. Plant》1977,13(1):49-54
Summary The action of hydralazine on collagen prolyl hydroxylation was studied in a cell culture system using WI-38 fibroblasts. The
prolyl hydroxylation level was determined by a method involving the digestion of collagen by bacterial collagenase and the
examination of specific peptides. The presence of low concentrations of hydralazine (0.2 mM) in both “young” and “old” fibroblast
cultures strongly inhibited collagen prolyl hydroxylation. The degree of inhibition was greater in serum-deficient cultures.
No significant improvement in the degree of hydroxylation was observed by increasing either ascorbate or iron levels in the
hydralazine-containing cultures in which hydroxylation was inhibited. Some of the reported side effects of hydralazine seen
in patients might be related to its inhibitory effects on mixed function oxidative (MFO) hydroxylation systems. While the
ascorbate dependence of the prolyl hydroxylase system of WI-38 decreased with the “age” of the culture, hydralazine inhibition
of hydroxylation was dramatic with cultures of all “ages”.
This work was supported by NIH grants nos. AM15671, AM1675 and HD07376, and fellowship no. HD01998. 相似文献