Direct visualisation and quantification of cellular cytotoxicity using two colour fluorescence

Kroesen, Bart-Jan; Mesander, G; Haar, Janina; Leij, Lou de

doi:10.1016/0022-1759(92)90009-i

Cited by 64 publications

(25 citation statements)

References 18 publications

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“…Propidium iodide (PI) was used in this study to identify dead cells and extracellular DNA. PI (MW = 668.4) is an intercalating DNA-binding dye that is nonspecific for base pairs (Eilam et al 1994); it is used to stain DNA in compromised (senescent and necrotic) cells but is unable to penetrate viable cell membranes (Kroesen et al 1992). These stains could be introduced to specimens without rinsing, thus reducing possible disruption to the specimen.…”

Section: Methodsmentioning

confidence: 99%

Development of a scanning confocal laser microscopic technique to examine the structure and composition of marine snow

Holloway

Cowen

1997

Limnology & Oceanography

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A technique was developed to image marine snow particles by scanning confocal laser microscopy (SCLM). This method allows structural and compositional characterization of fully hydrated marine snow particles with minimal disruption of the particle structure. High specificity fluorescent stains permitted sequential imaging of selected polysaccharides (concanavalin A: specific for mannose and glucose polymer:;), proteins [5-(4,6-dichlorotriazin-2-yl) aminofluorescein], and DNA (propidium iodide). Chl a autofluorescence can rtlso be imaged. SCLM produced optical slices of marine snow particles that were suited for a number of image-processing applications. These applications include high-resolution fluorescently derived optical slices and composite images produced from combined optical slices. Initial observations suggest that marine snow particles found in the oligotrophic Pacific Ocean vary in composition and structure. Variations in polysaccharide and protein composition of marine snow particles may be related to particle morphology. When combined with other conventional analysis, such as epifluorescent and electron microscopy, LSCM can provide important contributions to characterizing composition and structure of marine snow.

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Section: Methodsmentioning

confidence: 99%

Development of a scanning confocal laser microscopic technique to examine the structure and composition of marine snow

Holloway

Cowen

1997

Limnology & Oceanography

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“…Apoptosis of adherent cells was directly observed, as previously described, 15 using the fluorescent DNA-binding dyes acridine orange (100 g/ml), to determine the percentage of cells that have undergone apoptosis and ethidium bromide (100 g/ml), to differentiate between viable and nonviable cells. Observed through wide-band FITC filters, cells in the early stages of apoptosis are characterized by condensed or fragmented bright green chromatin and late apoptotic cells by condensed or fragmented bright orange chromatin.…”

Section: Apoptosis Assaysmentioning

confidence: 99%

Expression of CD154 on renal cell carcinomas and effect on cell proliferation, motility and platelet‐activating factor synthesis

Bussolati

Russo

Deambrosis

et al. 2002

Intl Journal of Cancer

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CD40 activation by CD154 may trigger diverse cellular responses, ranging from proliferation and differentiation to growth suppression and cell death, in normal and malignant cells. However, the pathophysiologic role of CD154 expressed by tumor cells remains unclear. We have investigated the expression of the CD40-CD154 system in 24 primary cultures derived from renal cell carcinomas, its correlation with tumor stage and its potential functional significance. We found coexpression of CD40 and CD154 in most of the renal carcinoma cell lines. CD154, but not CD40 expression, significantly correlated with tumor stage. Moreover, renal carcinoma cell lines also released the soluble form of CD154 into the supernatant. CD40 engagement by CD154 did not affect apoptosis or survival. On the contrary, CD154 stimulated cell proliferation, motility and production of PAF, a phospholipid mediator of inflammation with angiogenic properties. Key words: CD154; platelet-activating factor; renal carcinoma; CD40; angiogenesis CD154, the ligand of CD40, is a type II integral membrane protein, related to TNF and FasL. 1 Its receptor, CD40, is a 50 kDa transmembrane glycoprotein of the TNF superfamily, which is expressed in a multitude of different cell types, including B cells, monocytes, dendritic cells, endothelial cells, fibroblasts and renal epithelial cells. 2,3 CD40 was originally identified in a bladder carcinoma cell line, 4 and it is also expressed in a number of carcinomas, such as ovarian, nasopharyngeal, bladder, lung, colon and breast carcinomas, whereas normal nonproliferating tissues are CD40-negative. 5 CD40 engagement by CD154 conveys signals regulating diverse cellular responses, ranging from proliferation and differentiation to growth suppression and apoptosis. 6,7 The CD40 cytoplasmic C terminus lacks intrinsic kinase activity and adapter proteins of the TRAF family appear to mediate the activation of CD40-signaling cascades. 8 Despite the large interest, the functional role of CD40 in cancer development remains undetermined. CD40 appears to possess an opposite effect on tumor cell survival and apoptosis. CD40-mediated signaling has been reported to induce apoptosis when expressed in certain transformed cells of mesenchymal origin 9 and in carcinomas of the breast and lung. 10,11 Conversely, Jakobson et al. 12 demonstrated that CD40 stimulation inhibits Fas-mediated apoptosis in human bladder carcinoma cells and we showed a similar protective effect of CD40 in Kaposi's sarcoma cells. 13 Moreover, immunohistochemic studies revealed that detection of CD40 in primary cutaneous malignant melanoma might have negative prognostic value, suggesting its role in tumor progression. 14 CD154 is coexpressed with CD40 in melanoma cells and some carcinomas. 10,11,14 However, the pathophysiologic role of CD40 -CD154 interaction within the tumor cell remains unclear.In the present study, we investigated expression of the CD40 -CD154 system in 24 primary cultures derived from renal clear cell carcinomas, its correlation with tumor st...

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“…To determine the mechanism of anti-ABCB5 monoclonal-antibody-mediated inhibition of tumour formation and growth, the immune effector responses ADCC and complementdependent cytotoxicity (CDC) were assessed, as described 24 . Anti-ABCB5 monoclonal antibody but not isotype control monoclonal antibody significantly induced ADCC-mediated melanoma target cell death (2.1 ± 0.4% versus 0.2 ± 0.2%, respectively, mean ± s.e.m., P < 0.05) in a melanoma subpopulation comparable in size to the ABCB5-expressing subset 7 ( Fig.…”

mentioning

confidence: 99%

Identification of cells initiating human melanomas

Schatton

Murphy²,

Frank

et al. 2008

Nature

1,300

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Tumour-initiating cells capable of self-renewal and differentiation, which are responsible for tumour growth, have been identified in human haematological malignancies 1,2 and solid cancers [3][4][5][6] . If such minority populations are associated with tumour progression in human patients, specific targeting of tumour-initiating cells could be a strategy to eradicate cancers currently resistant to systemic therapy. Here we identify a subpopulation enriched for human malignantmelanoma-initiating cells (MMIC) defined by expression of the chemoresistance mediator ABCB5 (refs 7, 8) and show that specific targeting of this tumorigenic minority population inhibits tumour growth. ABCB5 + tumour cells detected in human melanoma patients show a primitive molecular phenotype and correlate with clinical melanoma progression. In serial humanto-mouse xenotransplantation experiments, ABCB5 + melanoma cells possess greater tumorigenic capacity than ABCB5 − bulk populations and re-establish clinical tumour heterogeneity. In vivo genetic lineage tracking demonstrates a specific capacity of ABCB5 + sub-populations for selfrenewal and differentiation, because ABCB5 + cancer cells generate both ABCB5 + and ABCB5 − progeny, whereas ABCB5 − tumour populations give rise, at lower rates, exclusively to ABCB5 − cells. In an initial proof-of-principle analysis, designed to test the hypothesis that MMIC are also

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Direct visualisation and quantification of cellular cytotoxicity using two colour fluorescence

Cited by 64 publications

References 18 publications

Development of a scanning confocal laser microscopic technique to examine the structure and composition of marine snow

Development of a scanning confocal laser microscopic technique to examine the structure and composition of marine snow

Expression of CD154 on renal cell carcinomas and effect on cell proliferation, motility and platelet‐activating factor synthesis

Identification of cells initiating human melanomas

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