The Role of Cytokines Produced via the NLRP3 Inflammasome in Mouse Macrophages Stimulated with Dental Calculus in Osteoclastogenesis.

Dental calculus (DC) is a common deposit in periodontitis patients. We have previously shown that DC contains both microbial components and calcium phosphate crystals that induce an osteoclastogenic cytokine IL-1ß via the NLRP3 inflammasome in macrophages. In this study, we examined the effects of cytokines produced by mouse macrophages stimulated with DC on osteoclastogenesis. The culture supernatants from wild-type (WT) mouse macrophages stimulated with DC accelerated osteoclastogenesis in RANKL-primed mouse bone marrow macrophages (BMMs), but inhibited osteoclastogenesis in RANKL-primed RAW-D cells. WT, but not NLRP3-deficient, mouse macrophages stimulated with DC produced IL-1ß and IL-18 in a dose-dependent manner, indicating the NLRP3 inflammasome-dependent production of IL-1ß and IL-18. Both WT and NLRP3-deficient mouse macrophages stimulated with DC produced IL-10, indicating the NLRP3 inflammasome-independent production of IL-10. Recombinant IL-1ß accelerated osteoclastogenesis in both RANKL-primed BMMs and RAW-D cells, whereas recombinant IL-18 and IL-10 inhibited osteoclastogenesis. These results indicate that DC induces osteoclastogenic IL-1ß in an NLRP3 inflammasome-dependent manner and anti-osteogenic IL-18 and IL-10 dependently and independently of the NLRP3 inflammasome, respectively. DC may promote alveolar bone resorption via IL-1ß induction in periodontitis patients, but suppress resorption via IL-18 and IL-10 induction in some circumstances.

Effects of periodontal therapy on phagocytic activity of peripheral blood neutrophils - evidence for an extrinsic cellular defect.

PURPOSE: To compare phagocytic activity of peripheral blood neutrophils from subjects with and without periodontal disease and evaluate the effects of periodontal therapy in individuals with similar levels of resolution of inflammation at the end of treatment. MATERIALS AND METHODS: To compare the phagocytic activity of neutrophils, peripheral blood was collected from 27 control subjects with a healthy periodontium and 28 periodontitis subjects before and after treatment. The phagocytosis of killed Saccharomyces cerevisiae, pre-sensitised or non-sensitised with fresh serum from the donor, was quantified and a phagocytic index was calculated as the mean number of yeast cells phagocytised by the percentage of neutrophils involved in phagocytosis. RESULTS: Prior to periodontal treatment, subjects with periodontitis exhibited significantly lower neutrophil phagocytic activity than control subjects with a healthy periodontium. Periodontal treatment significantly improved in clinical periodontal status and resulted in significantly increased phagocytosis of both pre-sensitised (from 113.0 pre- to 157.0 post-treatment, P = 0.02) and non-sensitised S. cerevisiae (from 1.5 pre- to 3.5 post-treatment, P = 0.001), to levels observed in control subjects. CONCLUSIONS: The phagocytic activity of peripheral blood neutrophils from subjects with periodontal disease was lower than that of healthy controls. Subjects who underwent non-surgical periodontal treatment and strict supportive therapy for 6 months showed improved phagocytic activity in peripheral blood neutrophils. The phagocytic index values from subjects with periodontal disease after treatment achieved those found in the control group.

The relationship between the presence of tooth-borne subgingival deposits and inflammation found with a dental endoscope.

BACKGROUND: Inflammatory periodontal diseases are found in many dentate individuals, but therapists and researchers who assess disease activity have had to rely on external clinical signs and symptoms to ascertain the health of the subgingival periodontal tissues. However, by using an endoscope in the subgingival environment, the therapist can see the relationship of subgingival tooth-borne accretions to signs of inflammation in the pocket wall. This study explored those relationships via the endoscope. METHODS: Twenty-six patients with moderate to severe periodontitis were chosen. The study visit involved a standardized, masked examiner who gathered data on the external gingival index, probing depth, gingival recession, and clinical attachment level. A second standardized examiner, masked to the findings of the first, used a dental endoscope. A set of indices (endoscopic biofilm index, endoscopic calculus index, and endoscopic gingival index) specifically developed for subgingival parameters was used. A fixation stent ensured that the periodontal probe and the endoscopic explorer traveled along the same path. RESULTS: A statistically significant relationship was found between deposits of subgingival calculus covered with biofilm and inflammation of the pocket wall, as measured by color change. In >60% of the cases, this inflammation was associated only with biofilm over deposits of calculus, not biofilm alone. Only subgingival calculus was statistically significant in relation to the positive traditional gingival index. CONCLUSIONS: Deposits of subgingival calculus covered with biofilm were directly related to >60% of pocket wall inflammation as measured by increased redness of the pocket epithelium. This was in comparison to biofilm alone.

Identification of calprotectin, a calcium binding leukocyte protein, in human dental calculus matrix.

Calprotectin is a calcium binding protein produced by leukocytes, macrophages and epithelial cells, and its levels in several tissues increase during infections and in many inflamed areas, suggesting that it may be an indicator of inflammatory activity. Osteopontin is a prominent phosphorylated glycoprotein in bone matrix, having calcium binding capacity. Recently, it has been reported that calprotectin and osteopontin are present in urinary stones (pathological mineralized masses in the body), and that these proteins may be involved in their formation. Dental calculus formed by mineralization of dental plaque is an inflammatory factor which may contribute to periodontal disease. It contains many organic components involved in mineralization. We recently found osteopontin molecules in human dental calculus and suggested that the components of its matrix may be similar to those of urinary stones. In this study, we investigated the presence of calprotectin in human dental calculus by immunohistochemical and immunoblotting analyses using a specific antibody for calprotectin. After fixation and demineralization of dental calculi adhered to tooth roots, sections embedded in paraffin were immunoreacted with the antibody for calprotectin and positive immunostaining for calprotectin was observed. Dental calculus proteins were then extracted with EDTA and separated by electrophoresis on 15% polyacrylamide gels. By immunoblotting analysis, 3 or 4 bands were observed at 11, 14.5, 22-25, 28 or 36.5 kDa and these patterns corresponded to those of calprotectin subunits. When non-immune rabbit serum was used instead of calprotectin-specific antibody as a negative control, no immunoreactivity was observed. These findings indicate that calprotectin is associated not only with antibacterial action but also with calcium binding capacity during dental calculus formation.

[The role of tartar in the light of the modern clinical-epidemiological, morphological and immunological studies]. / Roliata na zubniia kamuk v svetlinata na suvremennite kliniko-epidemiologichni, morfologichni i imunologichni izsledvaniia.

After the public recognition of the dental plaque as the primary etiological factor for the diseases of gingiva and periodontium, the tartar is the object of comparatively small number of studies. The author discusses the publications during the last 25 years, grouping the results from the epidemiological, clinical, morphological and immunological studies. Throwing light on new aspects of the pathogenetic role of the tartar in maintaining and chronification of the gingival inflammation as well as in osseous resorption, puts new accent on the necessity of its removal in prophylaxis and therapy. The importance of tartar as a reservoir of antigen substances associated with the total reactivity of macro-organisms is stressed upon.

Immunoglobulins in human dental calculus demonstrated with the peroxidase-antiperoxidase (PAP) method.

IgG and IgA were demonstrated in both sub- and supragingival dental calculus. Their distribution followed mainly incremental lines in the calculus. Most likely they originated from saliva and gingival fluid. Immunoglobulins which are adsorbed to crystals in biologic tissues may slow down the growth rate of the crystals. It was thus suggested that the incremental lines may be caused by the accumulation of IgG and IgA on the surface of dental calculus with a subsequent arrest in growth.