Flowcytometric Measurement of the Cell Cycle of Experimental Tumors: Some Devices for Accurate Measurement of Proliferative Activity

Tsugita, Masashi; Tsuru, T.; Takasaki, Tomohiko; Shinomiya, Nariyoshi; Kobayashi, Shoko; Hanyu, Fujio

doi:10.1159/000226954

Cited by 8 publications

(3 citation statements)

References 2 publications

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“…Cell cycle analyses were performed employing FACS, combining propidium iodide (PI, Sigma‐Aldrich, Munich, Germany) staining and 5‐bromo‐2′‐deoxyuridine (BrdU, Sigma‐Aldrich, Munich, Germany) incorporation as described by Tsugita [19]. 1 × 10 6 LAN‐1 cells in exponential growth phase were incubated with 8 μM of nemorosone for 24 hrs (end‐point).…”

Section: Methodsmentioning

confidence: 99%

Cytotoxic activity of nemorosone in neuroblastoma cells

Díaz-Carballo

Malak

Bardenheuer

et al. 2008

J Cellular Molecular Medi

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Neuroblastoma is the second most common solid tumour during childhood, characterized by rapid disease progression. Most children with metastasized neuroblastoma die despite intensive chemotherapy due to an intrinsic or acquired chemotherapy resistance. Thus, new therapeutic strategies are urgently needed. Here, we demonstrate that the novel compound nemorosone isolated from alcoholic extracts of Clusia rosea resins by reverse phase high pressure liquid chromatography (RP-HPLC) exerts cytotoxic activity in neuroblas-toma cell lines both parental and their clones selected for resistance against adriamycin, cisplatin, etoposide or 5-fluorouracil. Cell cycle studies revealed that nemorosone induces an accumulation in G0/G1- with a reduction in S-phase population combined with a robust up-regulation of p21Cip1. Furthermore, a dose-dependent apoptotic DNA laddering accompanied by an activation of caspase-3 activity was detected. Nemorosone induced a significant dephosphorylation of ERK1/2 in LAN-1 parental cells probably by the inhibition of its upstream kinase MEK1/2. No significant modulation of signal transducers JNK, p38 MAPK and Akt/PKB was detected. The enzymatic activity of immunoprecipitated Akt/PKB was strongly inhibited in vitro, suggesting that nemorosone exerts its anti-proliferative activity at least in part by targeting Akt/PKB in the cell lines studied. In addition, a synergistic effect with Raf-1 inhibitor BAY 43-9006 was found. Finally, nemorosone induced a considerable down-regulation of N-myc protein levels in parental LAN-1 and an etoposide resistant sub-line at the same drug-concentrations.

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Section: Methodsmentioning

confidence: 99%

Cytotoxic activity of nemorosone in neuroblastoma cells

Díaz-Carballo

Malak

Bardenheuer

et al. 2008

J Cellular Molecular Medi

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“…Selective gating excluded doublet cells. Data were analyzed with Consort 30 and Cell FIT software [9]. Mean cell size in each phase was measured by encircling the GoGi or G;M fraction on the forward scatter.…”

Section: Cell Cycle Analysismentioning

confidence: 99%

Effects of Cisplatin on Cell Cycle Kinetics, Morphological Change, and Cleavage Pattern of DNA in Two Human Ovarian Carcinoma Cell Lines

Sekiguchi¹,

Suzuki²,

Tamada³

et al. 1996

Oncology

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We evaluated the effects of cisplatin on the cell kinetics, cytomorphological changes, and cleavage patterns of DNA in two lines of human ovarian carcinoma cells. KF-1 cells displayed a cell cycle arrest in the G₂M phase, while HMG cells displayed a transient cell accumulation in the S phase, without obvious G₂M arrest. Morphological changes characterized by condensation and fragmentation of chromatin, and DNA cleavage by oligonucleosome-sized DNA fragments were observed in the HMG cells but not in the KF-1 cells. The pattern of cell death in HMG cells was considered to be apoptosis, but that of KF-1 cells necrosis. These findings showed the different mechanisms of antitumor effect of cisplatin on human ovarian carcinoma cell lines, including cell kinetics and pattern of cell death.

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“…Measurements of fluorescence signal were made with a FACStar plus flow cytometer (Becton Dickinson) using a 488 nm argonion laser. Data were acquired by using selective gating to exclude doublet cells (19) and were analyzed with DNA software. The fraction ratio of cells in each cell cycle compartment was calculated by a sum of broadened rectangles (SOBR) analysis model.…”

Section: Flow-cytometric Analysis Of Cell Cycle and Dna Fragmentationmentioning

confidence: 99%

Caffeine induces S-phase apoptosis in cis-diamminedichloroplatinum-treated cells, whereas cis-diamminedichloroplatinum induces a block in G2/M

et al. 1997

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Caffeine overrides checkpoints in the G2 phase of the cell cycle by inhibiting DNA repair at this phase and increases the cytotoxicity of antitumor drugs, such as cis‐diamminedichloroplatinum (CDDP). The enhanced cell death induced by caffeine is characterized by apoptosis. In this paper, we demonstrate that this apoptotic event occurs in S phase of the cell cycle, whereas CDDP induces a block in G2/M. DNA histogram analysis revealed that caffeine reduced G2 arrest in CDDP‐treated EL‐4 cells. In a synchronous population, the ratio of cyclin B:p34cdc2 was up‐regulated just before the cells went into the apoptotic pathway. A rapid increase in DNA fragmentation was detected at 12–24 h, when marked regression of G2/M phase was observed. Moreover, the degree of DNA fragmentation in CDDP + caffeine‐treated cells was not reduced when the cell cycle was arrested at metaphase by exposure to the spindle‐inhibitor nocodazole. It is possible that execution of the apoptotic program after treatment with caffeine did not require the EL‐4 cells to reenter G1 phase. The apoptotic cell fraction in the group of CDDP + caffeine was recognized as an S population by bivariate analysis of apoptosis and DNA content. These results suggest that enhancement of the apoptotic activity of CDDP‐treated cells by caffeine is not a G1‐phase event but an S‐phase‐specific event, whereas cells were arrested in G2/M phase, and that it is regulated by G2 checkpoint‐related proteins. Cytometry 27:365–373, 1997. © 1997 Wiley‐Liss, Inc.

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Flowcytometric Measurement of the Cell Cycle of Experimental Tumors: Some Devices for Accurate Measurement of Proliferative Activity

Cited by 8 publications

References 2 publications

Cytotoxic activity of nemorosone in neuroblastoma cells

Cytotoxic activity of nemorosone in neuroblastoma cells

Effects of Cisplatin on Cell Cycle Kinetics, Morphological Change, and Cleavage Pattern of DNA in Two Human Ovarian Carcinoma Cell Lines

Caffeine induces S-phase apoptosis in cis-diamminedichloroplatinum-treated cells, whereas cis-diamminedichloroplatinum induces a block in G2/M

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