Reactions of Connective Tissue to Mineral Trioxide Aggregate and Amalgam Mehmet Yaltirik, PhD, Hakan Ozbas, PhD, Bilge Bilgic, MD, and Halim Issever, MD The purpose of this study was to evaluate the sub- cutaneous connective tissue reactions to ProRoot, mineral trioxide aggregate (MTA; Dentsply), and Oralloy, high-copper amalgam (Coltene). These materials were placed in polyethylene tubes and implanted into dorsal connective tissue of Wistar albino rats, and tissue biopsies were collected and histologically examined 7, 15, 30, 60, and 90 days after the implantation procedure. The presence of inflammation, predominant cell type, calcification, and thickness of fibrous connective tissue were recorded. Scores were defined as follows: 0, none or few inflammatory cells, no reaction; 1, <25 cells, mild reaction; 2, 25 to 125 cells, moderate reaction; 3, >125 cells, severe reaction. Fibrous capsule was categorized as “thin” when thickness was < 150 m and “thick” at > 150 m. Necrosis and forma- tion of calcification were recorded. Both materials were well tolerated by the tissues in a 90-day eval- uation period. One notable finding is the presence of dystrophic calcification in connective tissue ad- jacent to MTA; this finding is consistent with the hypothesis of hard tissue induction by this material. For many years, amalgam was considered the retrograde filling and perforation repair material of choice (1– 4). However, there are many disadvantages with amalgam, including the potential for mercury and other ion release, corrosion and electrolysis, delayed expansion, marginal leakage, and tattoo formation (3, 4). There- fore, various alternatives to amalgam have been suggested (2, 3). A mineral trioxide aggregate (MTA) has been developed at Loma Linda University to seal communications between the tooth and the external surfaces (5). This material was studied in a series of in vivo and in vitro investigations. The biocompatibility of MTA has been reported previously using two cell culture techniques (6). MTA displays excellent low levels of microleakage when placed into extracted teeth (7, 8). Furthermore, histologic findings in dogs have confirmed laboratory observations that this material has great potential to facilitate tissue healing (9). The aim of this study was to histopathologically examine the biocompatibility of MTA and high-copper amalgam, by implanting them into the subcutaneous connective tissue of rats for 7, 15, 30, 60, and 90 days. MATERIALS AND METHODS We used 30 male, 5– 6 month old Wistar Albino rats weighing 270  30 g. Animal care was performed in accordance with Public Health Service Policy on Human Care and Use of Laboratory Animals of American Veterinary Medical Association Panel on Euthanasia, 2000. The following materials were investigated: • Group 1: ProRoot, mineral trioxide aggregate (Dentsply; Tulsa Dental, Tulsa, Oklahoma, USA). • Group 2: Oralloy, high-copper amalgam (Coltene AG, Altstat- ten, Switzerland). • Group 3: Control group The test materials were placed in clean, sterile polyethylene tubes (BARD C.R., Bard Ireland Ltd, Galway, Ireland) with 1.1-mm inner diameter and 10-mm length. MTA and amalgam were prepared according to the recommendations of the manufac- turers and were applied with a plastic and an amalgam carrier, respectively. Thirty empty polyethylene tubes were used as controls. The dorsal skins of animals were shaved under ether anesthesia and disinfected with 5% iodine solution. The backs of animals were incised over a length of 2 cm using a no. 15 blade in a head-to-tail alignment orientation. The skin was reflected, and the implantation materials were inserted into spaces created by blunt dissection. To prevent interactions of materials, the tubes were placed at least 2 cm from each other. The skin was closed with 3/0 silk suture. The evaluations were made 7, 15, 30, 60, and 90 days after surgical implantation (10, 11). In each examination period, six animals were sacrificed by administrating high doses of anesthetics. The dorsal skin was shaved, and the tubes were excised together with connective tis- sues around them. The specimens were kept in a 10% formalin solution (37% formaldehyde; Merck Darmstadt, Germany) for 2 wk. Sections of 6 m thickness were taken from specimens placed in paraffin blocks and stained with hematoxylin & eosin and Von Kossa (22). Evaluations were made in a light microscope (62774; Carl Zeiss, Oberkachen, Germany) at 40, 125, 200, 310, and 400 magnifications (12, 13, 18). Quantitative evaluations of JOURNAL OF ENDODONTICS Printed in U.S.A. Copyright © 2004 by The American Association of Endodontists VOL. 30, NO. 2, FEBRUARY 2004 95