Hisashi Anan

Humoral Immune Responses to S-Layer-Like Proteins of Bacteroides forsythus

ABSTRACT Bacteroides forsythus is one of the important periodontopathic bacteria, and this microorganism is known to have an S-layer outside the outer membrane. The S-layer-like antigens were recently isolated from B. forsythus, and they were found to be 270- and 230-kDa proteins in the envelope fraction. In this study, these proteins were confirmed to be specific to B. forsythus by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and they were clearly recognized by sera from patients with adult and early-onset periodontitis in Western immmunoblot analysis. We compared the immunoglobulin G (IgG) responses against the purified S-layer-like antigen by enzyme-linked immunosorbent assay. IgG responses against this antigen were low in healthy control subjects, but they were significantly higher in subjects with adult and early-onset periodontitis. Together with the fact that the IgG responses against the crude extract of B. forsythus did not rise significantly in patients with periodontitis, S-layer-like proteins are considered to be specific antigens of B. forsythus and may play an important role in the progression of periodontitis.

An enzyme histochemical study of the behavior of rat bone cells during experimental apical periodontitis

Bone alkaline and tartrate-resistant acid phosphatase activities were studied in paraffin sections obtained from experimentally induced periapical lesions of rats. These enzyme activities were demonstrated despite the use of high temperatures during embedding. In the formation phase, numerous osteoblasts and periodontal ligament cells showed strong alkaline phosphatase activity. On the other hand, a number of osteoclasts and preosteoclasts showed tartrate-resistant acid phosphatase activity in the activated resorption sites of periapical lesions. It is suggested that enzyme histochemical applications to paraffin sections are a useful means for clarifying the behavior of bone cells and that bone remodeling has occurred in the periapical lesions.

An immunohistochemical study of the behavior of cells expressing interleukin-1α and interleukin-1β within experimentally induced periapical lesions in rats*

Cells expressing interleukin-1 (IL-1) were examined in rat periapical lesions using immunohistochemical methods. During the acute phase, both IL-1α- and IL-1β-expressing cells were observed adjacent to many osteoclasts that contained large cytoplasms, and consequently the periapical lesions tended to expand. However, the number of IL-1β-expressing cells was about twice that of IL-1α-expressing cells during the same period. On the other hand, there were significantly less IL-1β-expressing cells during the chronic phase and consequently a significant expansion of the periapical lesions was not seen. Furthermore, IL-1α- and IL-1β-expressing cells were located far from the bone surfaces, whereas a fibroblastic cell layer lay between the IL-1-expressing cells and the bone surfaces. Both IL-1α and IL-1β were mainly expressed by macrophages. In addition, after the percolated application of formocresol to experimentally induced periapical lesions, numerous macrophages expressing IL-1β became conspicuous within the vicinity of the extensive bone resorption sites. These results suggest that macrophages expressing IL-1β may play an important role in the activation and recrudescence of osteoclastic bone resorption, and this may be due to their localization close to the bone resorption sites noted within experimental periapical lesions.

Detection of interleukin-1β mRNA in rat periapical lesions

Cells expressing interleukin-1β (IL-1β) mRNA were demonstrated by in situ hybridization in rat periapical lesions. A great number of osteoclasts with significant tartrate-resistant acid phosphatase activity were observed on the bone surfaces, and numerous IL-1β mRNA-expressing cells were widely distributed in the periodontal ligaments. IL-1β mRNA-expressing cells were mainly observed around the blood vessels in the vicinity of the bone resorption sites and occasionally found near the osteoblasts. Immunohistochemistry and enzyme histochemistry assays showed that IL-1β mRNA-expressing cells were not bone cells, but that they had the characteristic features of macrophages. These results suggested that macrophages may contribute to the production of IL-1β and play an important role in activation of osteoclastic bone resorption.

Effects of a combination of an antibacterial agent (ofloxacin) and a collagenase inhibitor (FN-439) on the healing of rat periapical lesions.

To investigate the effects of a combination of an antibacterial agent (ofloxacin) and a collagenase inhibitor (FN-439) in the root canal treatment of apical periodontitis, we studied the healing process of experimentally induced periapical lesions in rats by using immunohistochemical methods. With a topical application of a combination of ofloxacin and FN-439 following experimentally induced periapical lesions, both neutrophils and macrophages became significantly decreased in number, while active cementogenesis and extensive bone formation were seen in the periapical region. However, the use of ofloxacin alone also demonstrated a beneficial effect on periapical inflammation and healing. Therefore, it is suggested that ofloxacin is powerful against bacterial infection whether FN-439 is added. The only observed effect of a combination of ofloxacin and FN-439 is that it may more effectively inhibit osteoclastic bone resorption and activate the remodeling of the apical periodontal tissue if this combined medicament is used in a stage of active bone destruction characterized by high production of tissue collagenase.