Radiation response of head and neck squamous cell carcinoma using early markers of apoptosis and proliferation, utilising a microfluidic approach

Srivastava, Rishi

August 2016

Thesis or dissertation

© 2016 Rishi Srivastava. All rights reserved. No part of this publication may be reproduced without the written permission of the copyright holder.

The purpose of this study was to assess the response to irradiation of HNSCC tissue samples maintained in a microfluidic device, utilising early markers of apoptosis and proliferation.

Materials and Methods
Rat liver had initially been used to optimise the conditions so HNSCC tissue biopsies could be maintained in a pseudo in vivo environment. Parameters assessed were different flow rates, oxygenated media and concentrations of sera in the medium, buffer components and infusion times with bromodeoxyuridine. HNSCC tissue samples from the oral cavity, oropharynx, larynx, metastatic lymph node and maxillary sinus were obtained. Tissue from each subsite was irradiated with doses of 5Gy, 10Gy, 15Gy and 20Gy. Cell death was measured using LDH and the morphology of the tissue was assessed using H&E staining. Apoptosis was calculated using immunohistochemical techniques to detect cytokeratin and the M30 antibody; and proliferation was assessed using the bromodeoxyuridine assay.

HNSCC tissue samples could be maintained in the microfluidic device after an initial rise in LDH levels and assessment of the tissue architecture using H&E staining. Statistically, there was no significant difference observed in LDH post irradiation. Apoptotic indices calculated for all subsites revealed variation within the same tissue with the same dose, statistically no significant difference between irradiation with different doses of the head and neck subsites. Proliferation was noted with the bromodeoxyuridine assay in three out of the five subsites but no significant pattern was observed.

The microfluidic device is capable of maintaining HNSCC tissue in a viable state and is able to withstand rigorous testing. This study did not demonstrate a dose dependent relationship and suggests HNSCC intratumour heterogeneity would account for the variation in responses with irradiation. The microfluidic technique is a very useful method in assessing a patient’s response to radiotherapy and allows a personalised treatment approach for the future.

Hull York Medical School, The University of Hull and University of York
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