PII S0360-3016(02)02867-5 CLINICAL INVESTIGATION Normal Tissue LONG-TERM ALTERATIONS OF ORAL MUCOSA IN RADIOTHERAPY PATIENTS FRANZ-JOSEF PROTT, M.D., PH.D.,* J¨ ORG HANDSCHEL, M.D., D.D.S., † OLIVER MICKE, M.D.,* CORD SUNDERK ¨ OTTER, M.D., PH.D., ‡ ULRICH MEYER, M.D., D.D.S., PH.D., † AND JOSEF PIFFKO, M.D., D.D.S., PH.D. † Departments of *Radiotherapy, † Cranio- and Maxillofacial Surgery, and ‡ Dermatology, University of Mu ¨nster, Mu ¨nster, Germany Purpose: The aim of this investigation was to describe the alterations in oral mucosa after radiotherapy. Methods and Materials: Biopsies were taken from patients before irradiation, at 60 Gy, and 6 –12 months after radiotherapy. Histomorphological evaluation of the vessels was performed, and endothelial expression of ICAM-1, VCAM-1, and E-selectin was also evaluated, as well as distribution of LFA-1–, Mac-1–, VLA-4 –, RM3/1-, 27E10-, and 25F9-bearing cells in the subepithelial tissue. Results: The expression of ICAM-1 was downregulated after radiotherapy, whereas the percentage of LFA-1– and VLA-4 – bearing cells increased. VCAM-1 remained at low levels. The subepithelial infiltration was still dominated by RM3/1-positive macrophages. The number of vessels decreased, while the lumina of the remaining vessels in the deeper connective layer increased. Conclusions: The late effects of radiotherapy are characterized by a decreased number of blood vessels and by significantly different expression patterns of the adhesion molecules studied, and of integrins and macrophage subpopulations compared to the conditions before irradiation and at 60 Gy. © 2002 Elsevier Science Inc. Mucositis, Radiotherapy, Adhesion molecules, Immunohistochemistry. INTRODUCTION Radiotherapy is indicated postoperatively in the treatment of head-and-neck cancer in the case of tumors larger than T2 and lymph node involvement. Usually, symptoms ap- pearing or extending beyond 6 months after completion of irradiation are classified as late effects (1, 2). The late alterations in irradiated oral mucosa are described as fibrosis and even changes in the small blood vessels. Therefore, an investigation is required to describe those late vascular alterations in comparison with the status before and during irradiation. Additionally, there are no studies of the late changes concerning the composition of the different subep- ithelial (SE) cell populations and the function of the endo- thelial cells expressing various types of adhesion molecules. However, adhesion molecules play a leading role in the transendothelial migration of leukocytes and thus influence the cellular composition of the tissue (3, 4) and the inflam- matory response. Interactions between integrins, expressed by leukocytes, and adhesion molecules mediate a coordi- nated sequence of events, resulting in leukocyte emigration from the bloodstream into the tissue (Fig. 1) (5). During irradiation, an increase of intercellular adhesion molecule 1 (ICAM-1) and E-selectin was found in parallel with the occurrence of leukocytes bearing specific receptors for ICAM-1 (LFA-1, Mac-1) (6). Regarding the invading cells, it was discovered that macrophages play a major role in the acute effects of radiotherapy (7). Interestingly, we found a significant correlation between the clinical stages of acute symptoms after irradiation and the increase of a macrophage subpopulation identified with the RM3/1 antibody (7). Cells reactive with the monoclonal antibodies 27E10, RM3/1, and 25F9 are also known to prevail at different stages of the inflammatory response (8 –10). The purpose of the present prospective study was to describe the late effects of radiotherapy on oral mucosa. METHODS AND MATERIALS Patients Patients (age range: 45–77 years) with squamous cell carcinoma of the head and neck were postoperatively irra- diated with 60 Gy. Forty-five specimens were taken from these patients from nontumorous buccal mucosa, 21 before irradiation (Group A), 13 samples within 1 h after 60 Gy Reprint requests to: Dr. Franz-Josef Prott, Institute of Radio- therapy Wiesbaden, Josefshospital Wiesbaden, Solmsstrasse 15, D-65189 Wiesbaden, Germany. Tel.: ++49-611-1771645; Fax: ++49-611-1771642; E-mail: prott@stahlentherapie-wiesbaden.de Presented at the RSNA Scientific Meeting, Chicago, IL, November 2001. Acknowledgments—We are grateful to Prof. Ulrich Joos and Prof. Normann Willich for their support in this investigation. Received Dec 5, 2001, and in revised form Mar 27, 2002. Accepted for publication Apr 9, 2002. Int. J. Radiation Oncology Biol. Phys., Vol. 54, No. 1, pp. 203–210, 2002 Copyright © 2002 Elsevier Science Inc. Printed in the USA. All rights reserved 0360-3016/02/$–see front matter 203