Analysis of Cell Cycle by Flow Cytometry

Pożarowski, Piotr; Darżynkiewicz, Zbigniew

doi:10.1385/1-59259-811-0:301

Cited by 397 publications

(355 citation statements)

References 14 publications

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“…Cell cycle status was analyzed with a FACScan flow cytometer (Becton Dickinson, San Diego, USA) by measuring fluorescence from cells stained with propidium iodide [20] . Data are presented as the relative fluorescence intensity of cell sub-populations in the 2-dimensional FACS profile or as the percentage of cells in a given sub-population.…”

Section: Cell Cycle Analysismentioning

confidence: 99%

Neferine inhibits angiotensin II-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1

Tong

Zhang

et al. 2010

Acta Pharmacol Sin

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Aim:To explore the effect of neferine on angiotensin II (Ang II)-induced vascular smooth muscle cell (VSMC) proliferation. Methods: Human umbilical vein smooth muscle cells (HUVSMCs) were used. Cell proliferation was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry analysis. Heme oxygenase (HO)-1 protein expression was tested by Western blot analysis. Extracellular signal-regulated protein kinase 1/2 (ERK1/2) activation was determined by using immunoblotting. Results: Pre-incubation of HUVSMCs with neferine (0.1, 0.5, 1.0, and 5.0 µmol/L) significantly inhibited Ang II-induced cell proliferation in a concentration-dependent manner and neferine 5.0 µmol/L increased HO-1 expression by 259% compared with control. The antiproliferative effect of neferine was significantly attenuated by coapplication of zinc protoporphyrin IX (ZnPP IX, an HO-1 inhibitor) with neferine. Ang II-enhanced ERK1/2 phosphorylation was markedly reversed by neferine. By inhibiting HO-1 activity with ZnPP IX, the inhibitive effect of neferine on ERK1/2 phosphorylation was significantly attenuated. Cobalt-protoporphyrin (CoPP), an HO-1 inducer, significantly decreased Ang II-induced ERK1/2 phosphorylation and inhibited Ang II-induced cell proliferation. The ERK1/2 pathway inhibitor PD98059 significantly blocked Ang II-enhanced ERK1/2 phosphorylation and inhibited cell proliferation. Conclusion: These findings suggest that neferine can inhibit Ang II-induced HUVSMC proliferation by upregulating HO-1, leading to the at least partial downregulation of ERK1/2 phosphorylation.

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Section: Cell Cycle Analysismentioning

confidence: 99%

Neferine inhibits angiotensin II-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1

Tong

Zhang

et al. 2010

Acta Pharmacol Sin

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“…Suspensions of zebrafish embryo cells, HeLa cells, or HCT-116 cells were processed for DNA content FACS analysis as described (Pozarowski and Darzynkiewicz 2004). The cells were analyzed using a FACScan machine (Becton-Dickinson), and the data were analyzed using ModFit LT software (Verity Software).…”

Section: Cell Cycle Analysismentioning

confidence: 99%

DTL/CDT2 is essential for both CDT1 regulation and the early G2/M checkpoint

Sansam¹,

Shepard²,

Lai³

et al. 2006

Genes Dev.

151

150

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Checkpoint genes maintain genomic stability by arresting cells after DNA damage. Many of these genes also control cell cycle events in unperturbed cells. By conducting a screen for checkpoint genes in zebrafish, we found that dtl/cdt2 is an essential component of the early, radiation-induced G2/M checkpoint. We subsequently found that dtl/cdt2 is required for normal cell cycle control, primarily to prevent rereplication. Both the checkpoint and replication roles are conserved in human DTL. Our data indicate that the rereplication reflects a requirement for DTL in regulating CDT1, a protein required for prereplication complex formation. CDT1 is degraded in S phase to prevent rereplication, and following DNA damage to prevent origin firing. We show that DTL associates with the CUL4-DDB1 E3 ubiquitin ligase and is required for CDT1 down-regulation in unperturbed cells and following DNA damage. The cell cycle defects of Dtl-deficient zebrafish are suppressed by reducing Cdt1 levels. In contrast, the early G2/M checkpoint defect appears to be Cdt1-independent. Thus, DTL promotes genomic stability through two distinct mechanisms. First, it is an essential component of the CUL4-DDB1 complex that controls CDT1 levels, thereby preventing rereplication. Second, it is required for the early G2/M checkpoint.

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“…48 A citometria é uma técnica estabelecida para estudar eventos intracelulares como estresse oxidativo, 49 autofagia, 50 alterações no ciclo celular, 51 apoptose, 52 entre outros. Mesmo alterações como fragmentação do DNA celular, antes analisada por eletroforese ou microscopia, também pode ser investigada por citometria de fluxo.…”

Section: Citometria De Fluxounclassified

Como estudar interações entre nanopartículas e sistemas biológicos

et al. 2016

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Recebido em 02/05/2016; aceito em 15/06/2016; publicado na web em 16/08/2016 STUDYING THE INTERACTIONS BETWEEN NANOPARTICLES AND BIOLOGICAL SYSTEMS. Although in recent years there has been an increasing amount of literature on nanotechnology and their clinical applications, it is still scarce a deep understanding of the interactions at the molecular levels between nanoparticles and cells. Studies demonstrating the underlying mechanisms of nanoparticles endocytosis, intracellular trafficking, and cellular processing are imperative to understand better how cells interact with those materials and their possible undesired effects, e.g. nanotoxicity. The rising awareness concerning nanoparticles applications and its interactions with the cellular environment is part of the new research field called Nanotoxicology. The cumulative knowledge in nanotoxicology will allow us to foresee toxic effects, establish regulations and limits for nanoparticles applications. In this work, we discuss the theoretical concepts about studying endocytosis and intracellular trafficking of nanoparticles. The nanoparticles-cell interactions are a multi-step process, which can be divided into nanoparticles' internalization, intracellular processing and triggering effects of nanomaterials on eukaryotic cells. Finally, we discuss the main techniques used to study this process: flow cytometry, use of endocytosis inhibitors and confocal microscopy.Keywords: endocytosis; intracellular trafficking; nanoparticles, nanotoxicology; nano-cell interactions. INTRODUÇÃONanotecnologia é um dos ramos mais promissores de tecnologia, sobre o qual se deposita grandes expectativas. 1 Somente em 2015, foram publicados cerca de 9400 artigos relacionados ao termo "Nanotechnology" no banco de dados Science Direct; além disso, diversos produtos têm sido testados e comercializados, por exemplo, protetores solares, cosméticos, produtos biomédicos, biosensores e sistemas para transporte e entrega de fármacos. 2 Entretanto, há uma crescente preocupação com a segurança e possíveis efeitos nocivos de nanomateriais que vêm sendo desenvolvidos, 3 pois dados e regulamentações internacionais referentes à segurança e uso desses materiais ainda são escassos, ainda estão em debate ou em fase de implementação. 4 Uma nova área da ciência destinada ao estudo de efeitos adversos de nanomateriais foi recentemente criada, a Nanotoxicologia. 5 O conhecimento acumulado nessa área tem como objetivo prever efeitos tóxicos e auxiliar nas regulamentações e diretrizes para aplicação da nanotecnologia 6 e uso de produtos a ela relacionados. A Figura 1 mostra o crescimento do número de artigos científicos que contêm o termo "nanotoxicology" e estão no banco de dados do National Center for Biotechnology Information (NCBI), subdivisão do National Institute of Health (NIH/USA).Apesar do número de artigos em nanotoxicologia ser crescente, esta é uma área ainda muito recente (com menos de uma década), e ainda não é possível determinar exatamente quais impactos os sistemas naturais e artificiais co...

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Analysis of Cell Cycle by Flow Cytometry

Cited by 397 publications

References 14 publications

Neferine inhibits angiotensin II-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1

Neferine inhibits angiotensin II-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1

DTL/CDT2 is essential for both CDT1 regulation and the early G2/M checkpoint

Como estudar interações entre nanopartículas e sistemas biológicos

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