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Use of bioluminescent imaging to investigate the role of nuclear factor-κΒ in experimental non-small cell lung cancer metastasis

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Abstract

Nuclear factor (NF)-κB is frequently over-expressed in non-small cell lung cancer (NSCLC), but the exact role of this observation remains unclear. In this regard, activation of the transcription factor may govern distinct steps of NSCLC progression, such as carcinogenesis, angiogenesis, and metastasis. In these studies we attempted to dissect the effects of two proteins of the NF-κB pathway (p65/RelA and IκΒα) on experimental metastasis of murine NSCLC, using a novel approach of bioluminescent detection of NF-κB activation in tumor cells. Stable integration of a NF-κΒ reporter confirmed high basal activation of the transcription factor in mouse NSCLC cells in vitro and during experimental metastasis to the lungs, like human NSCLC. In the mouse model of NSCLC metastasis, NF-κΒ-dependent luciferase expression served as a reliable indicator of tumor cell delivery to the lungs, establishment of metastatic tumors, and lung tumor burden. In vitro transient p65/RelA and IκΒα gene transfer to mouse NSCLC cells resulted, respectively, in significant NF-κB activation and inhibition, without affecting cell growth. However, p65/RelA overexpression in NSCLC cells drastically reduced in vivo metastasis to the lungs, while overexpression of IκΒα had no effect. In conclusion, using bioluminescent detection of NF-κB activation in mouse lug adenocarcinoma cells, we found a negative impact of p65/RelA on NSCLC metastasis.

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Abbreviations

NF-κB:

Nuclear factor-κB

IκB:

Inhibitor of NF-κB

NSCLC:

Non-small cell lung cancer

LLC:

Lewis lung cancer

pNGL :

NF-κB.GFP.LUC reporter plasmid

MOI:

Multiplicity of infection

pfu:

Plaque forming units

Ad:

Adenoviral vector

IκBα-DN:

Dominant inhibitor of NF-κB

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

LDH:

Lactate dehydrogenase

Luc:

Luciferase

GFP:

Green fluorescent protein

βgal:

β-Galactosidase

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Acknowledgements

This work was funded by NIH HL66196 (TSB), NIH HL61419 (TSB), the Department of Veterans Affairs (TSB), the Ingram Charitable Fund (FEY) and Vanderbilt-Ingram Cancer Center (FEY). GTS was supported by a scholarship by the Greek State Scholarship Foundation (IKY).

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Authors and Affiliations

  1. Division of Allergy, Pulmonary and Critical Care Medicine, Vanderbilt University School of Medicine, Nashville, TN, USA

    Georgios T. Stathopoulos, Taylor P. Sherrill, Wei Han, Ruxana T. Sadikot, Vasiliy V. Polosukhin & Timothy S. Blackwell

  2. Department of Critical Care and Pulmonary Services and Applied Biomedical Research and Training Center “Marianthi Simou”, General Hospital “Evangelismos”, School of Medicine, University of Athens, 3 Ploutarhou Str., Athens, 10675, Greece

    Georgios T. Stathopoulos

  3. Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN, USA

    Taylor P. Sherrill & Timothy S. Blackwell

  4. Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, TN, USA

    Barbara Fingleton, Fiona E. Yull & Timothy S. Blackwell

  5. Department of Veterans Affairs, VA Tennessee Valley Healthcare System, Nashville, TN, USA

    Timothy S. Blackwell

Authors
  1. Georgios T. Stathopoulos
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  2. Taylor P. Sherrill
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  3. Wei Han
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  4. Ruxana T. Sadikot
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  5. Vasiliy V. Polosukhin
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  6. Barbara Fingleton
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  7. Fiona E. Yull
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  8. Timothy S. Blackwell
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Correspondence to Georgios T. Stathopoulos.

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Stathopoulos, G.T., Sherrill, T.P., Han, W. et al. Use of bioluminescent imaging to investigate the role of nuclear factor-κΒ in experimental non-small cell lung cancer metastasis. Clin Exp Metastasis 25, 43–51 (2008). https://doi.org/10.1007/s10585-007-9100-z

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  • DOI: https://doi.org/10.1007/s10585-007-9100-z

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