Inflammation is a protective mechanism against pathogens, but if maintained continuously, it destroys tissue structures. Aggregatibacter actinomycetemcomitans is a gram-negative, facultative anaerobic bacterium often found in severe periodontitis. A. actinomycetemcomitans invades epithelial cells and triggers inflammatory response in the immune cells. In this study, we investigated the effect of water-soluble rosehip extract on A. actinomycetemcomitansinduced inflammatory responses. A human monocytic cell line (THP-1) was differentiated to macrophages by phorbol 12-mystristate 13-acetate treatment. The cytotoxic effect of extract was determined using the 3-(4,5-dimethylthiazol-2- yl)-2,5-diphenyltetrazolium bromide assay. The effects of extract on bacterial growth were examined by measuring the optical densities using a spectrophotometer. THP-1-derived macrophages were infected A. actinomycetemcomitans after extract treatment, and culture supernatants were analyzed for cytokine production using enzyme-linked immunosorbent assay. Protein expression was measured by western blotting. Extract was not toxic to THP-1- derived macrophages. A. actinomycetemcomitans growth was inhibited by 1% extract. The extract suppressed A. actinomycetemcomitans-induced tumor necrosis factor-α, interleukin (IL)-1β, and IL-8 production. It also decreased mitogen-activated protein kinase (MAP kinase) and nuclear factor-κB (NF-κB) phosphorylation. Moreover, the extract inhibited the expression of inflammasome components, including nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3, Absent in Melanoma 2, and apoptosis associated speck-like protein containing a CARD. And cysteine-aspartic proteases-1 and IL-1β expression were decreased by the extract. In summary, extract suppressed A. actinomycetemcomitans growth and decreased inflammatory cytokine production by inhibiting activation of MAP kinase, NF-κB, and inflammasome signaling. Rosehip extract could be effective in the treatment of periodontal inflammation induced by A. actinomycetemcomitans infection.

Introduction
Materials and Methods
    1. Reagents
    2. Bacterial and cell culture
    3. Cell cytotoxicity assay
    4. Growth curve
    5. Cytokine assay
Results
    1. Effect of rosehip extract on the viability ofTHP-1-derived macrophages and the growth ofA. actinomycetemcomitans
    2. Effect of rosehip extract on inflammatory cytokinesproduction induced by A. actinomycetemcomitansin THP-1-derived macrophages
    3. Effect of rosehip extract on MAP kinase and NF-κBactivation induced by A. actinomycetemcomitans inTHP-1-derived macrophages
    4. Effect of rosehip extract on inflammasomeactivation in THP-1-derived macrophages inducedby A. actinomycetemcomitans
Discussion
Acknowledgements
Conflicts of Interest
References

• Yuri Song(Department of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan 50162, Republic of Korea, Oral Genomics Research Center, Pusan National University, Yangsan 50162, Republic of Korea)
• Si young Kim(Department of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan 50162, Republic of Korea, Oral Genomics Research Center, Pusan National University, Yangsan 50162, Republic of Korea)
• Jin Chung(Department of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan 50162, Republic of Korea, Oral Genomics Research Center, Pusan National University, Yangsan 50162, Republic of Korea) Corresponding author