Identification of GM3 as a Marker of Therapy- Resistant Periradicular Lesions Mario L. Zuolo, DDS, Marcos S. Toledo, MS, Helio E. Nogueira, MD, Anita H. Straus, PhD, and Helio K. Takahashi, PhD The purpose of this study was to analyze the profile of glycosphingolipids (GSLs) in periradicular le- sions refractory to endodontic treatment. Sixteen periapical lesions were removed surgically from patients (experimental group) and compared with 10 samples of periodontal ligament removed from extracted intact third molars (control group). After the GSLs extraction and purification procedures were performed the neutral and acidic GSL frac- tions were analyzed by high-performance thin- layer chromatography and quantified by densitom- etry. Data reported herein show that: (i) tissues in the experimental group presented about twice as much GSLs as the control group; (ii) lesion tissues express lactoneotetraosylceramide, and lactofu- copentaosyl (IV) ceramide, whereas these neutral GSLs are absent in normal tissues; and (iii) normal tissues express GT1b, whereas lesions cells do not express this ganglioside. In contrast lesion tissues express GM3, which is conspicuously absent in normal tissues. Polymicrobial infection of the dental pulp is the main cause of pulp tissue inflammation and can cause bone destruction in the perira- dicular tissues. Most of these lesions can be successfully treated when adequate cleaning, shaping, and obturation of the root canal system are followed by definitive restoration procedures to prevent reinfection occurrence (1). However endodontic treatment or retreatment can fail, and sev- eral local and systemic factors have been related to endodontic failures. The persistence of bacteria in the root canal is the most studied local factor that can negatively affect the success rate after endodontic treatment (1, 2). On the other hand the pathological mechanisms involved in persistent periradicular lesions, the so-called therapy-resistant or refractory lesions, remain unclear until now. Recent literature has been focusing on issues that can play significant roles in endodon- tic treatment failures, such as the presence of microorganisms in the periapical tissues; the host-defense processes, the immunocom- petent cells present at the site of the lesion, and the humoral immune response against irritants (3– 6). Technical developments in immunology and molecular biology have been used to study the pathogenesis of many diseases at a molecular level. The importance of glycosphingolipids (GSLs) in several biological processes—such as cell– cell interaction, cell– bacteria interaction, tumor-associate markers, and inhibition of lymphoprolipheration— has been established by a number of stud- ies (7–10). Despite the amount of available information regarding the biological roles of GSLs no information is available associating these macromolecules with pathological mechanisms in perira- dicular lesions. Thus the objective of this study was to conduct a systematic analysis on the profile of neutral and acidic GSLs in therapy-resistant, refractory lesions. MATERIALS AND METHODS Source of Tissues EXPERIMENTAL GROUP Sixteen periradicular lesions were taken from 16 teeth at the time of therapeutic surgical treatment. All affected teeth were asymptomatic and without periodontal involvement. Also only teeth in which at least one retreatment had been performed and which had no clinical or radiographic signs of coronal microleak- age were included in this study. In all cases approximately one- third of each specimen was placed in formalin solution and sub- mitted to histological evaluation. CONTROL GROUP Ten samples of periodontal ligaments were obtained by root planning of soft tissue attached to the roots of 10 intact third molars just after surgical extraction of these teeth. After the tissues were collected they were washed with cold phosphate-buffered saline with pH 7.4, weighed, and stored at -70°C. All samples were processed within 7 days after collection. JOURNAL OF ENDODONTICS Printed in U.S.A. Copyright © 2001 by The American Association of Endodontists VOL. 27, NO. 2, FEBRUARY 2001 107