Extracellular 37-kDa Antigenic Protein from Actinobacillus actinomycetemcomitans Induces TNF-α, IL-1β, and IL-6 in Murine Macrophages

Abstract

The extracellular antigens of Actinobacillus actinomycetemcomitans Y4 (serotype b) contain a 37-kDa protein which is a major target for IgGs from patients suffering from severe alveolar bone loss. Since the 37-kDa protein has not been studied sufficiently, our investigation focused on its characteristics, e.g., its localization, specificity, and whether it directly stimulates macrophages to produce cytokines. The 37-kDa protein was purified from the culture supernatant of the Y4 strain by means of chromatofocusing and gel filtration. The 37-kDa protein is a unique glycoprotein which forms immune complexes with monoclonal antibodies against rhamnose-fucose polysaccharide. Patients with A. actinomycetemcomitans-associated periodontitis had higher antibody titers to the purified 37-kDa protein than healthy subjects (p < 0.001). Anti-37-kDa protein antibodies recognized a 37-kDa band in the cytosolic, ribosomal, and total membrane fractions from Y4 cells. Extracellular substances from other strains of A. actinomycetemcomitans (serotypes a and c) also reacted in the Western blots, but Haemophilus spp. or several periodontopathic bacteria did not. These results suggested that the 37-kDa protein is a cytosolic protein that is passed through the cell membrane, and its protein portion is specific for A. actinomycetemcomitans but common to serotypes. This protein induced Il-1 beta, Il-6, and TNF-alpha release from murine macrophages. The Il-6-inducing activity of the 37-kDa protein was higher than that of LPS. These findings suggested that the 37-kDa protein which is released from live cells plays a role in A. actinomycetemcomitans-associated periodontitis, as antigen inducing the release of inflammatory cytokines which are associated with alveolar bone loss.

Serum lipids modify periodontal infection – interleukin‐6 association

“…These observations are in accordance with animal studies, where periodontal pathogens have been reported to induce the production of proinflammatory cytokines (Tani et al. , Lalla et al. , Polak et al.…”

“…Non-experimental studies have shown that those who have periodontitis have higher levels of inflammatory markers such as IL-6 and TNF-a in serum (Loos et al 2000, Bretz et al 2005, Andrukhov et al 2011) and in gingival crevicular fluid than periodontally healthy persons (Geivelis et al 1993, Lee et al 1995, Becerik et al 2012). These observations are in accordance with animal studies, where periodontal pathogens have been reported to induce the production of proinflammatory cytokines (Tani et al 1997, Lalla et al 2003, Polak et al 2009), and with in vitro studies, where periodontal pathogens have been shown to be able to induce the production of pro-inflammatory cytokines from human gingival cells (Stathopoulou et al 2010, Peyyala et al 2013).…”

The highly leukotoxic JP2 clone of Aggregatibacter actinomycetemcomitans: evolutionary aspects, epidemiology and etiological role in aggressive periodontitis

“…Local formation of osteoclasts is suggested to occur via an enhanced expression of osteoclast‐stimulating cytokines in the inflamed tissues ( Taubman & Kawai , 2001). Stimulation of cytokine production can be induced by a variety of bacterial surface components and virulence factors, including those related to A. actinomycetemcomitans ( Reddi et al , 1995; Tani et al ., 1997; Schytte et al ., 1999; Nishida et al ., 2001; Uchida et al ., 2001; Lerner , 2004; Kelk et al ., 2005, 2008; Garlet et al ., 2006). The differentiation of osteoclasts is proposed to be caused by pro‐inflammatory cytokines, such as IL‐1, IL‐6, IL‐8, and TNF‐α.…”

Antigens of bacteria associated with periodontitis

“…Outer membrane and secreted proteins from A. actinomycetemcomitans have also been reported to be major inducers of in¯ammatory cytokines. Tani et al (348) isolated an extracellular 37 kDa antigenic glycoprotein from A. actinomycetemcomitans Y4 (serotype b) and evaluated its ability to induce the release of cytokines from murine macrophages. The 37-kDa glycoprotein was found to induce strong TNF-a, IL-1b, IL-6 responses and a weak granulocyte macrophage conlony stimulating factor (GM-CSF) response in macrophages (292).…”