Assessing the effects of radiotherapy on head and neck squamous cell carcinoma using microfluidic techniques

Carr, Simon D.

September 2013

Thesis or dissertation

© 2013 Simon David Carr. All rights reserved. No part of this publication may be reproduced without the written permission of the copyright holder.

The aim of this study was to investigate how HNSCC tissue biopsies maintained in a pseudo in vivo environment within a bespoke microfluidic device, respond to radiation treatment.

Materials and Methods
35 patients with HNSCC were recruited; in addition liver tissue from 5 Wistar rats was used. A glass microfluidic device was used to maintain the tissue biopsy samples in a viable state. Rat liver was used to optimise the methodology. HNSCC was obtained from patients with T1-T3 laryngeal or oropharyngeal SCC; N1-N2 metastatic cervical lymph nodes were also obtained. Irradiation consisted of single doses of between 2 Gy and 40 Gy and a fractionated course of 5x2 Gy. Cell death was assessed in the tissue effluent using the soluble markers LDH and cytochrome c, and in the tissue by immunohistochemical detection of cleaved cytokeratin18 (M30 antibody). Radiation-induced DNA strand breaks were detected using the TUNEL assay.

A significant surge in LDH release was demonstrated in the rat liver after a single dose of 20 Gy; in HNSCC it was seen after 40 Gy, compared to the control. There was no significant difference in cytochrome c release after 5 Gy or 10 Gy. M30 demonstrated a dose-dependent increase in apoptotic index for a given increase in single dose radiation. There was a significant increase in apoptotic index between the non-irradiated HNSCC tissue and irradiated tissue and between the tissue irradiated with 1x2 Gy and 5x2 Gy. As with the apoptotic index, there was a significant increase in radiation-induced DNA breaks between the non-irradiated and the irradiated tissue and between the tissue irradiated with 1x2 Gy and 5x2 Gy.

This microfluidic technique can be used to study the effects of radiation on HNSCC tissue. The device was capable of maintaining the HNSCC in a viable state, without it undergoing significant apoptosis or DNA damage and can be used to demonstrate the relationship between radiotherapy dose and radiation-induced cell death using tissue-based cell death markers.
This study is a significant step towards achieving the ultimate goal of developing this device as a tool, capable of predicting a patient’s response to radiotherapy prior to the commencement of treatment.

Hull York Medical School, The University of Hull and University of York
Greenman, John (Professor of tumour immunology); Stafford, Nicholas
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