Measuring the response of human head and neck squamous cell carcinoma to irradiation in a microfluidic model allowing customized therapy

Cheah,Ramsah; Srivastava,Rishi; Stafford,Nicholas D.; Beavis,Andrew W.; Green,Victoria; Greenman,John

doi:10.3892/ijo.2017.4118

Measuring the response of human head and neck squamous cell carcinoma to irradiation in a microfluidic model allowing customized therapy

Authors:
- Ramsah Cheah
- Rishi Srivastava
- Nicholas D. Stafford
- Andrew W. Beavis
- Victoria Green
- John Greenman
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Affiliations: Hull York Medical School, University of Hull, Hull, HU6 7RX, UK, Radiation Physics, Hull and East Yorkshire Hospitals NHS Trust, Faculty of Science and Engineering, University of Hull, Hull, HU6 7RX, UK, School of Life Sciences, University of Hull, Hull, HU6 7RX, UK
Published online on: September 5, 2017 https://doi.org/10.3892/ijo.2017.4118
Pages: 1227-1238

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Abstract

Radiotherapy is the standard treatment for head and neck squamous cell carcinoma (HNSCC), however, radioresistance remains a major clinical problem despite significant improvements in treatment protocols. Therapeutic outcome could potentially be improved if a patient's tumour response to irradiation could be predicted ex vivo before clinical application. The present study employed a bespoke microfluidic device to maintain HNSCC tissue whilst subjecting it to external beam irradiation and measured the responses using a panel of cell death and proliferation markers. HNSCC biopsies from five newly-presenting patients [2 lymph node (LN); 3 primary tumour (PT)] were divided into parallel microfluidic devices and replicates of each tumour were subjected to single-dose irradiation (0, 5, 10, 15 and 20 Gy). Lactate dehydrogenase (LDH) release was measured and tissue sections were stained for cytokeratin (CK), cleaved-CK18 (cCK18), phosphorylated-H2AX (γH2AX) and Ki‑67 by immunohistochemistry. In addition, fragmented DNA was detected using terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL). Compared with non‑irradiated controls, higher irradiation doses resulted in elevated CK18-labelling index in two lymph nodes [15 Gy; 34.8% on LN1 and 31.7% on LN2 (p=0.006)] and a single laryngeal primary tumour (20 Gy; 31.5%; p=0.014). Significantly higher levels of DNA fragmentation were also detected in both lymph node samples and one primary tumour but at varying doses of irradiation, i.e., LN1 (20 Gy; 27.6%; p=0.047), LN2 (15 Gy; 15.3%; p=0.038) and PT3 (10 Gy; 35.2%; p=0.01). The γH2AX expression was raised but not significantly in the majority of samples. The percentage of Ki‑67 positive nuclei reduced dose-dependently following irradiation. In contrast no significant difference in LDH release was observed between irradiated groups and controls. There is clear inter- and intra-patient variability in response to irradiation when measuring a variety of parameters, which offers the potential for the approach to provide clinically valuable information.

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