University Archives

Sulforaphane Inhibits Gingival Fibroblast IL-6 Production Induced by Periodontal Pathogens

Authors:
Arif Hussain¹, Laura Young2, Fatima Al Hakim1, Joseph Ferracciolo², David Fischer2, and Eric Krukonis²
¹University of Detroit Mercy School of Dentistry
²University of Detroit Mercy School of Dentistry, Division of Integrated Biomedical Sciences

Background: Sulforaphane (SFN) is a natural product obtained from leafy green vegetables like broccoli. SFN has been shown previously to inhibit inflammation and has been used to treat conditions like atopic dermatitis in animal models. SFN can act via inhibition of the NF-kB signalling pathway in host cells.

Objective:  We sought to determine whether SFN administered prior to infection of human gingival fibroblasts (HGFs) with various Gram-negative periodontal pathogens, can inhibit production of the proinflammatory cytokine IL-6. In addition, we investigated the effect of SFN on NF-kB nuclear localization (activation).

Methods: HGF-1 cells (ATCC) were grown overnight in 24-well tissue culture plates. Cells were washed with PBS and 0 or 5uM SFN was added to HGF-1 cells for 1 hour prior to the addition of ~3,000,000 bacteria. LPS-containing bacteria included: Escherichia coli (Ec), Fusobacterium nucleatum (Fn), Porphyromonas gingivalis (Pg), Treponema denticola (Td), and Aggregatibacter actinomycetemcomitans (Aa). After 3hrs, chamber slides were washed, fixed with paraformaldehyde, permeabilized with triton-X-100, and stained for NF-kB. After 24 hours, HGF-1 supernatants were harvested, and IL-6 was quantified by ELISA assay. The students’ T-test was used to assess statistical differences in Il-6 production.

Results: SFN exhibited a 70-90% reduction in IL-6 production in response to Ec, Fn, Td, and Aa (p<0.05). Similarly, SFN reduced the level of NF-kB nuclear localization in HGFs upon infection with these bacteria. Pg did not induce Il-6 production, likely due to Pg making an immunosuppressive tetra-acylated LPS at 37°C. In fact, addition of Pg along with Ec, Fn, or Aa led to complete inhibition of IL-6 production by these immunostimulatory bacteria.

Conclusions: SFN is potent suppressor of inflammation stimulated by several Gram-negative periodontal pathogens. SFN blocks IL-6 production, by inhibiting nuclear localization of NF-kB. When pre-grown at 37°C, Pg LPS blocks TLR4 activation by other periodontal pathogens.