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Volume 46, Number 3, June 2013

nvolvement of Toll-like receptor 2 in apoptosis of Aggregatibacter actinomycetemcomitans-infected THP-1 cells 


Satsuki Kato, Keisuke Nakashima, Toshiyuki Nagasawa, Yoshihiro Abiko, Yasushi Furuichi


Received: January 6, 2012    Revised: January 31, 2012    Accepted: February 9, 2012   

 

Corresponding author:

Division of Periodontology and Endodontology, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293, Japan
Corresponding author. Division of Periodontology and Endodontology, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293, Japan.Aggregatibacter (Actinobacillus) actinomycetemcomitans is a gram-negative bacterium that has been associated with aggressive periodontitis. A actinomycetemcomitans infection induces apoptosis in the human monocytic cell line THP-1, and p38 mitogen-activated protein kinase (p38) activity and tumor necrosis factor-α (TNF-α) production are enhanced by A actinomycetemcomitans infection. However, mechanisms governing the recognition of A actinomycetemcomitans by monocytes during apoptosis have not been elucidated. A actinomycetemcomitans cell wall components stimulate Toll-like receptor (TLR)2 and/or TLR4. The authors examined whether TLR2 and/or TLR4 were involved in the apoptosis of A actinomycetemcomitans-infected THP-1 cells. 



 

Background and purpose: 

A actinomycetemcomitans-infected THP-1 cells were transferred to six-well culture plates and incubated for 0 to 6 hours. In some experiments, THP-1 cells were incubated with anti-TLR2, anti-TLR4, or isotype control antibody (10 μg/mL) for 30 minutes prior to A actinomycetemcomitans infection. Expression of messenger RNA (mRNA) was examined by reverse transcription-polymerase chain reaction. Intracellular bacteria recovered from A actinomycetemcomitans-infected cells and apoptotic cells were detected by APOPercentage dye (Biocolor Ltd, Northern Ireland, UK) staining. Cellular p38 activity and TNF-α production were assessed with enzyme-linked immunosorbent assay. 



 

Results:

The expression of TLR2 mRNA was increased by A actinomycetemcomitans infection. Phagocytosis and apoptosis in A actinomycetemcomitans-infected THP-1 cells were inhibited by the addition of anti-TLR2 antibody. Also, anti-TLR2 antibody suppressed the activation of p38 and production of TNF-α in A actinomycetemcomitans-infected THP-1 cells.
 



 

Conclusion:

These results suggest that A actinomycetemcomitans induces increased expression of TLR2, leading to phagocytosis and apoptosis of THP-1 cells via p38 activation and TNF-α production. 



 

Key words:

Aggregatibacter (Actinobacillus) actinomycetemcomitans, Apoptosis, THP-1 cells, Toll-like receptor 2