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ΔNp63 expression in four carcinoma cell lines and the effect on radioresistance—a siRNA knockdown model

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Abstract

Objectives

This study investigated the expression of ΔNp63α in carcinoma cell lines of the upper aerodigestive tract and their potential influence on radioresistance using a small interfering RNA (siRNA) knockdown approach.

Materials and methods

Four carcinoma cell lines were investigated for the expression of the ΔNp63 isoform by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) (0, 24, 48 h) with and without single dose irradiation of 6 Gy. Furthermore, all cell lines were transfected with siRNA against the ΔNp63α isoform over 24 h. Knockdown effectiveness was controlled by qRT-PCR and Western blot. Apoptotic events were evaluated by terminal transferase dUTP nick end labeling (TUNEL) assay and cross-checked by a test for cell viability (WST-1, Roche) over 48 h.

Results

All cell lines presented varying expression of the ΔNp63α isoform with and without irradiation. A sufficient knockdown rate was established by siRNA transfection. Knockdown of the ΔNp63 isoform showed an effect on radiation sensitivity proven by an increase of apoptotic events detectable by immunofluorescence (TUNEL assay) and likewise a significant reduction of formazan production (WST-1 test) in three cell lines.

Conclusions

We found overexpression of ΔNp63α with and without irradiation in three cell lines, and the knockdown of ΔNp63α led to increased apoptotic events and fewer viable cells. Thus, the overexpression of ΔNp63α might protect carcinoma cells against irradiation effects.

Clinical relevance

The present work supports the hypothesis that protein 63 might serve as a negative predictor for irradiation response and survival in a clinical setting and may be a target for future therapeutic strategies.

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Abbreviations

p63:

Protein 63

LI:

Labeling index

PCR:

Polymerase chain reaction

rtPCR:

Real-time polymerase chain reaction

HNSCC:

Head and neck squamous cell carcinoma

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Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Authors and Affiliations

  1. Department of Oral and Maxillofacial Surgery, University of Mainz, Medical Center, Augustusplatz 2, 55131, Mainz, Germany

    Maximilian Moergel & Jutta Goldschmitt

  2. Department of Radiation Oncology, University of Mainz, Medical Center, Langenbeckstr. 1, 55101, Mainz, Germany

    Marcus Stockinger

  3. Department of Oral and Maxillofacial Surgery, University of Bochum, Medical Center, In der Schornau 23-25, 44892, Bochum, Germany

    Martin Kunkel

  4. Department of Oral and Maxillofacial Surgery, Johannes Gutenberg-University, Medical Center, Augustusplatz 2, 55131, Mainz, Rheinland-Pfalz, Germany

    Maximilian Moergel

Authors
  1. Maximilian Moergel
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  2. Jutta Goldschmitt
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  4. Martin Kunkel
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Correspondence to Maximilian Moergel.

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Moergel, M., Goldschmitt, J., Stockinger, M. et al. ΔNp63 expression in four carcinoma cell lines and the effect on radioresistance—a siRNA knockdown model. Clin Oral Invest 18, 1259–1268 (2014). https://doi.org/10.1007/s00784-013-1078-0

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  • DOI: https://doi.org/10.1007/s00784-013-1078-0

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