Annexin V binding assay as a tool to measure apoptosis in differentiated neuronal cells

Schütte, B.; Nuydens, Rony; Geerts, Hugo; Ramaekers, Frans C.S.

doi:10.1016/s0165-0270(98)00147-2

Cited by 214 publications

(136 citation statements)

References 25 publications

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“…The quantitative analysis of apoptosis induced by different PTX/DOX-loaded liposomes was performed by Annexin V-FITC/PI double staining (Schutte et al, 1998). B16 cells were treated with PTX/DOX-loaded preparations (DOX/ PTX ¼ 20 mg/ml) for 6 h at 37 C. At the end of the treatment, cells were harvested, washed with cold PBS, suspended in 500 ml binding buffer and stained by 5 ml Annexin V-FITC and 5 ml PI.…”

Section: In Vitro Apoptosis Staining Assaymentioning

confidence: 99%

Targeted delivery of transferrin and TAT co-modified liposomes encapsulating both paclitaxel and doxorubicin for melanoma

et al. 2015

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The purpose of this study was to develop an efficient dual-ligand based liposomal drug delivery system with targeting specificity as well as properties that would kill melanoma cells. Liposomes modified with transferrin (Tf) and cell-penetrating peptide TAT was prepared, which encapsulated two kinds of chemotherapy drugs, paclitaxel and doxorubicin (Tf/TAT-PTX/DOX-LP). The Tf ligands specifically bind to the overexpressed Tf receptors on the surface of melanoma cells, while the TAT ligands functioned as a classical cell penetrating peptide, helping dual-ligand liposomes be internalized by melanoma cells. The effect of dual-targeting system and ''double-drug'' combination therapy were evaluated both in vitro and in vivo. In vitro, cellular uptake, intracellular distribution and tumor spheroids penetration studies demonstrated that the system could not only be selectively and efficiently penetrate melanoma cells. Besides, apoptosis staining assay and cytotoxicity showed effective anti-tumor capability and obvious synergistic effect of combination therapy of PTX and DOX. In vivo imaging and fluorescent images of tumor section further demonstrated that Tf/TAT-PTX/DOX-LP had the highest tumor distribution. The results of these experiments demonstrated that double-drug liposomal drug delivery systems (DDS) had both enhanced targeting efficiency and increased therapeutic efficacy.

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Section: In Vitro Apoptosis Staining Assaymentioning

confidence: 99%

Targeted delivery of transferrin and TAT co-modified liposomes encapsulating both paclitaxel and doxorubicin for melanoma

et al. 2015

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“…Apoptosis was detected by flow cytometry in cells stained with fluorescein isothiocyanate (FITC)-conjugated annexin V and PI [51]. Measurements were performed immediately after PI addition on a BD LSR II flow cytometer, and data were analyzed with FlowJo software.…”

Section: Maintenance and Differentiation Of Es Cells And Ethanol Treamentioning

confidence: 99%

Ethanol Alters the Balance of Sox2, Oct4, and Nanog Expression in Distinct Subpopulations During Differentiation of Embryonic Stem Cells

Ogony

Malahias

Vadigepalli

et al. 2013

Stem Cells and Development

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The transcription factors Sox2, Oct4, and Nanog regulate within a narrow dose-range embryonic stem (ES) cell pluripotency and cell lineage commitment. Excess of Oct4 relative to Sox2 guides cells to mesoendoderm (ME), while abundance of Sox2 promotes neuroectoderm (NE) formation. Literature does not address whether ethanol interferes with these regulatory interactions during neural development. We hypothesized that ethanol exposure of ES cells in early differentiation causes an imbalance of Oct4 and Sox2 that diverts cells away from NE to ME lineage, consistent with the teratogenesis effects caused by prenatal alcohol exposure. Mouse ES cells were exposed to ethanol (0, 25, 50, and 100 mM) during retinoic acid (10 nM)-directed differentiation to NE for 0-6 days, and the expression of Sox2, Oct4, and Nanog was measured in single live cells by multiparametric flow cytometry, and the cellular phenotype was characterized by immunocytochemistry. Our data showed an ethanol dose-and time-dependent asymmetric modulation of Oct4 and Sox2 expression, as early as after 2 days of exposure. Single-cell analysis of the correlated expression of Sox2, Oct4, and Nanog revealed that ethanol promoted distinct subpopulations with a high Oct4/Sox2 ratio. Ethanol-exposed cells differentiated to fewer b-III tubulin-immunoreactive cells with an immature neuronal phenotype by 4 days. We interpret these data as suggesting that ethanol diverted cells in early differentiation from the NE fate toward the ME lineage. Our results provide a novel insight into the mode of ethanol action and opportunities for discovery of prenatal biomarkers at early stages.

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“…2B). The translocation of the membrane phospholipid phosphatidylserine from the inner to the outer leaflet of the plasma membrane was considered to be an earlier marker of apoptosis than the changes in nuclear morphology (20) and was quantified by flow cytometry using an annexin V/fluorescein isothiocyanate (FITC) conjugate and propidium iodide (PI). Apoptosis was also shown to be increased by this method after a 48-h induction of DN-HNF1A (Fig.…”

Section: Expression Of Dn-hnf1a Causes Bioenergetic Dysfunction Inmentioning

confidence: 99%

AMP-activated Protein Kinase Mediates Apoptosis in Response to Bioenergetic Stress through Activation of the Pro-apoptotic Bcl-2 Homology Domain-3-only Protein BMF

Kilbride

Farrelly

Bonner

et al. 2010

Journal of Biological Chemistry

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Heterozygous loss-of-function mutations in the hepatocyte nuclear factor 1A (HNF1A) gene result in the pathogenesis of maturity-onset diabetes-of-the-young type 3, (HNF1A-MODY). This disorder is characterized by a primary defect in metabolism-secretion coupling and decreased beta cell mass, attributed to excessive beta cell apoptosis. Here, we investigated the link between energy stress and apoptosis activation following HNF1A inactivation. This study employed single cell fluorescent microscopy, flow cytometry, gene expression analysis, and gene silencing to study the effects of overexpression of dominant-negative (DN)-HNF1A expression on cellular bioenergetics and apoptosis in INS-1 cells. Induction of DN-HNF1A expression led to reduced ATP levels and diminished the bioenergetic response to glucose. This was coupled with activation of the bioenergetic stress sensor AMP-activated protein kinase (AMPK), which preceded the onset of apoptosis. Pharmacological activation of AMPK using aminoimidazole carboxamide ribonucleotide (AICAR) was sufficient to induce apoptosis in naive cells. Conversely, inhibition of AMPK with compound C or AMPK␣ gene silencing protected against DN-HNF1A-induced apoptosis. Interestingly, AMPK mediated the induction of the pro-apoptotic Bcl-2 homology domain-3-only protein Bmf (Bcl-2-modifying factor). Bmf expression was also elevated in islets of DN-HNF1A transgenic mice. Furthermore, knockdown of Bmf expression in INS-1 cells using siRNA was sufficient to protect against DN-HNF1A-induced apoptosis. Our study suggests that overexpression of DN-HNF1A induces bioenergetic stress and activation of AMPK. This in turn mediates the transcriptional activation of the pro-apoptotic Bcl-2-homology protein BMF, coupling prolonged energy stress to apoptosis activation.

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Annexin V binding assay as a tool to measure apoptosis in differentiated neuronal cells

Cited by 214 publications

References 25 publications

Targeted delivery of transferrin and TAT co-modified liposomes encapsulating both paclitaxel and doxorubicin for melanoma

Targeted delivery of transferrin and TAT co-modified liposomes encapsulating both paclitaxel and doxorubicin for melanoma

Ethanol Alters the Balance of Sox2, Oct4, and Nanog Expression in Distinct Subpopulations During Differentiation of Embryonic Stem Cells

AMP-activated Protein Kinase Mediates Apoptosis in Response to Bioenergetic Stress through Activation of the Pro-apoptotic Bcl-2 Homology Domain-3-only Protein BMF

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