JOURNALOF ENDODONTICS Printed in U.S.A. Copyright © 1999 by The American Association of Endodontists VOL. 25, NO. 4, APRIL 1999 Comparative Effects of Two Endodontic Irrigants, Chiorhexidine Digluconate and Sodium Hypochlorite, on Macrophage Adhesion to Plastic Surfaces Juan Jose Segura, DDS, MD, PhD, Alicia Jim~nez-Rubio, Juan Miguel Guerrero, and Juan Ramon Calvo, MD, PhD This study was designed to compare the "in vitro" effect of chlorhexidine digluconate, proposed as a new irrigant solution, with sodium hypochlorite, the currently irrigant solution used to disinfect the root canal system before obturation of the canal, on substrate adherence capacity of macrophages. Inflammatory macrophages were obtained from Wistar rats and resuspended in RPMI-1640 me- dium. The adherence capacity of macrophages to a plastic surface was determined. Assays were conducted in Eppendorf tubes for 15 min of incu- bation at 37°C in an humidified atmosphere of 5% CO 2 in air. The adherence index was calculated. Chlorhexidine digluconate inhibited substrate ad- herence capacity of macrophages in all conditions tested. Half-maximal inhibition (IC50) was observed at 1:5.6 dilution. Chlorhexidine digluconate was less potent than 5.25% sodium hypochlorite (IC5o = 1:24.1 dilution) in inhibiting substrate adherence capacity of macrophages. Taking into account that substrate adherence is the first step in the phago- cytic process of macrophages and in antigen pre- sentation, chlorhexidine digluconate could inhibit macrophage function and modulate inflammatory reactions at the level of inflamed periapical tissues. Bacteria and their products play an essential role in the pathogen- esis of pulpo-periapical diseases (1). A long-standing endodontic infection allows bacteria to propagate to the entire root canal system, including ramifications, isthmuses, apical deltas, and den- tinal tubules. Therefore, a major objective in cndodontic therapy is the disinfection of the root canal system before obturation of the canal. A solution of 5.25% sodium hypochlorite is the current inigant of choice (2). Recently, 2.0% chlorhexidine gluconate irrigant has been showed to possess in vitro antimicrobial activity equivalent to that of 5.25% sodium hypochlorite (3) and to instill substantive antimicrobial activity when used as an endodontic irrigant (4). Moreover, in a clinical and laboratory study, 0.12% chlorhexidine gluconate has been shown to reduce the endodontic microbiota (5). Macrophages are the most dominating immunocompetent cells during all stages of experimentally induced periapical lesions (6). They are known to have several mediator and regulatory functions, and are involved in the entire spectrum of defense reactions (7). Macrophages interact with other cells and components of the extracellular environment by means of adhesion receptors. Adhe- sion is the first step in the phagocytic process of macrophages (8). Scavenger receptors have been reported to mediate macrophage adhesion to serum-coated plastic surfaces (9), and adhesion assays exploiting this property have been developed (10). Using these assays, sodium hypochlorite, as well as glutaraldehyde (11), cal- cium hydroxide (12), and EDTA (13), have been shown to de- crease "in vitro" substrate adherence capacity of macrophages. Subsequently, it has been argued that leakage of sodium hypochlo- rite through the apical foramen during root canal treatment could alter macrophage functions modulating reparative mechanisms and decreasing inflammatory reactions in perirradicular tissues (l 1). Several studies indicate that chlorhexidine has toxic effects on human cells and granulation tissue. Concentrations of chlorhexi- (line well below those used in clinical dentistry have been reported to cause cell injury, cell death, and inhibition of protein synthesis in human fibroblasts cultures, and HeLa cell cultures (14). Thus, a chlorhexidine digluconate (CHX) leak to perirradicular tissues could alter repair mechanisms. In this study, the effects of 5.25% sodium hypochlorite and 0.12% CHX on macrophage adhesion to plastic surfaces are com- pared. 243 MATERIALS AND METHODS Collection of Inflammatory Macrophages Peritoneal macrophages were elicited from Wistar rats by the method described previously (15) (Fig. 1). Briefly, each rat was