NF-kB dependent cytokine levels in saliva of patients with oral preneoplastic lesions and oral squamous cell carcinoma Nelson L. Rhodus DMD, MPH a, * , Vu Ho MD b , Craig S. Miller DMD, MS c , Sandra Myers DMD, MS a , Frank Ondrey MD, PhD b a Department of Oral Medicine, University of Minnesota, Minneapolis, MN, USA b Department of Otolaryngology, University of Minnesota, Minneapolis, MN, USA c Department of Oral Medicine, University of Kentucky, Lexington, KY, USA Accepted 22 October 2004 Abstract Previous investigations in our laboratory and others (Chen et al., 1998) have shown that the levels of certain inflammatory, proangiogenic cytokines in saliva and tissue specimens of patients with oral premalignant lesions (OPML) are elevated. We have also shown that these cytokines are elevated in tissue culture of oral squamous cell carcinoma (OSCC) cell lines. The purpose of this pilot study was to determine the level of several inflammatory, NF-kB-dependent cytokines in whole unstimulated saliva (WUS), in subjects with OPML as compared to those with diagnosed OSCC. Subjects (n = 13) with OMPL, OSCC (n = 13), and age–sex matched controls without oral lesions (C) (n = 13) were enrolled. The mean age was 58.7 years. WUS was collected by standard techniques for 5 min (Navazesh, 1993). WUS samples were centrifuged and the cytokine analysis was performed on the supernatants by ELISA as previously described by Ondrey et al. (1991). The cytokines analyzed were: TNF-alpha, interleukin-1, interleukin-6, and interleukin-8 (TNF-a, IL-1, IL-6, and IL-8). The results as analyzed by Pairwise t-tests revealed significant differences in the salivary levels of: (1) TNF-a: (mean Æ S.E.M.: TNF-a–OSSC = 28.9 Æ 14.6 * pcg/ml versus OPML = 10.5 Æ 7.4 * pcg/ml versus controls = 3.0 Æ 1.0 pcg/ml; * p < 0.01); (2) IL-1: (IL-1–OSSC = 454.4 Æ 215.8 * pcg/ml versus OPML = 255.1 Æ 124.8 * pcg/ml versus controls = 173.2 Æ 66.9 pcg/ml; * p < 0.01); (3) IL-6: (mean Æ S.E.M.: IL-6–OSSC = 88.2 Æ 43.2 * pcg/ml versus OPML = 70.8 Æ 24.3 * pcg/ml versus controls = 1.4 Æ 1.0 pcg/ml; * p < 0.001) and (4) IL-8 in saliva: (mean Æ S.E.M.: IL-8–OSSC = 3154.1 Æ 1023.2 * pcg/ml versus OPML = 1918.2 Æ 899.1 * pcg/ml versus controls = 1580.7 Æ 789.0 pcg/ml; * p < 0.001). There was a sig- nificant increase in the levels of all cytokines in the saliva of the OPML as compared to controls, and a significant difference in the cytokines of OSSC saliva compared to the OPML and controls. These results suggest that these proangiogenic, proinflammatory cytokines are elevated in the saliva of patients with OSSC and OPML as compared to controls, which may have diagnostic and/or prognostic significance. # 2004 International Society for Preventive Oncology. Published by Elsevier Ltd. All rights reserved. Keywords: Cancer; Oral; Saliva; Cytokines; Squamous cell carcinoma; Diagnosis 1. Introduction Cancer of the mouth and pharynx accounts for nearly 40,000 cases of cancer per year in the United States and is the sixth most common cancer worldwide. Over 90% of these oral–pharyngeal cancers are squamous cell carcinomas (SCC). The 5-year survival rate (approximately 50%) from oropharyngeal carcinomas has not significantly improved in the past 30 years. [1,2]. Epithelial malignancies of the oral cavity often begin as preneoplastic lesions in the form of inflammatory lesions such as leukoplakia. Leukoplakia is associated with tobacco and alcohol use and chronic inflammation with the risk of malignant transformation to SCC of approximately 5–17% [3]. Previous studies of in vitro human cell lines as well as OSCC tumors have demonstrated that certain proinflamma- tory, proangiogenic NF-kB dependent cytokines levels are increased. These include: TNF-a, IL-1, IL-6, IL-8, GM-CSF, and VEGF, which can be found in significantly, elevated quantities in the local milieu of SCC. There is evidence that www.elsevier.com/locate/cdp Cancer Detection and Prevention 29 (2005) 42–45 * Corresponding author. E-mail address: rhodu001@maroon.tc.umn.edu (N.L. Rhodus). 0361-090X/$30.00 # 2004 International Society for Preventive Oncology. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.cdp.2004.10.003