Withaferin A inhibits inflammatory responses induced by Fusobacterium nucleatum and Aggregatibacter actinomycetemcomitans in macrophages

Abstract

Periodontitis is a progressive chronic inflammatory disease and a major cause of tooth loss in humans. As a withanolides, withaferin A (WA) is known to exhibit strong anti‑inflammatory activity. The present study examined whether WA inhibited inflammatory responses in macrophages in response to two representative periodontal pathogens, Fusobacterium nucleatum and Aggregatibacter actinomycetemcomitans. Murine bone marrow‑derived macrophages (BMDMs) were used in this study and cytokine production in culture supernatants was measured by enzyme‑linked immunosorbent assays. Western blot analysis was performed to determine the activation of nuclear factor‑κB and mitogen‑activated protein kinases (MAPKs) and the expression of inducible nitric oxide synthase (iNOS), toll‑like receptor (TLR) 2 and TLR4. The production of nitric oxide (NO) was determined by the Griess reaction. WA treatment was shown to decrease interleukin (IL)‑6 and tumor necrosis factor (TNF)‑α production in BMDMs in response to F. nucleatum and A. actinomycetemcomitans in a dose‑dependent manner. The phosphorylation of IκB‑α and MAPKs (p38, extracellular signal‑regulated kinases and c‑Jun N‑terminal kinases) induced by F. nucleatum and A. actinomycetemcomitans was also inhibited by WA. F. nucleatum and A. actinomycetemcomitans induced iNOS expression and NO production in BMDMs, which was inhibited by WA in a dose‑dependent manner. WA also reduced endogenous and induced expression of TLR2 and TLR4 in these cells. These results suggest that WA may be a potential therapeutic agent or preventive additive for periodontitis control.

Fusobacterium nucleatum Contributes to the Carcinogenesis of Colorectal Cancer by Inducing Inflammation and Suppressing Host Immunity

“…F. nucleatum infection increases the infiltration of inflammatory cells, such as macrophages, dendritic cells, and granulocytes, which create a pro-inflammatory microenvironment that is conducive to the occurrence of CRC [44]. Macrophages infected with F. nucleatum can induce the release of inflammatory cytokines [45], [46]. Natural cytotoxic receptor NKp46 of NK cell can directly recognize F. nucleatum through its surface ligand, secreting TNF-α to aggravate inflammation [20].…”

Ureaplasma Urealyticum Infection Contributes to the Development of Pelvic Endometriosis Through Toll-Like Receptor 2

“…PMCs were lysed in ice-cold RIPA buffer (Elpis Biotech, Daejeon, Korea) containing a protease inhibitor cocktail (Thermo Fisher Scientific, Waltham, Massachusetts, USA) and phosphatase inhibitor (Thermo Fisher Scientific). For cell lysates, standard western blotting was performed (33) as described using antibodies against MMP-2 and total and phospho-ERK, JNK, and p-38 (Cell Signaling Technology, Beverly, MA, USA). Immunofluorescent-labeled antibody to rabbit IgG (Cell Signaling Technology) was used to visualize western blots with the Fusion solo S chemidoc (Vilber, Collégien, France).…”

Withania somnifera (L.) Dunal: Opportunity for Clinical Repurposing in COVID-19 Management

“…WS has emerged as a potential modulator of the NF-κB pathway with withaferin A as an inhibitor of NF-κB activation ( Heyninck et al, 2014 ). Similarly, withaferin A also inhibited bacterial infection-induced NF-κB activation in macrophages and dendritic cells of mice ( Noh et al, 2016 ). Withaferin A ( Neog et al, 2018 ) ( Bale et al, 2018a ) and fatty acids of WS seeds ( Balkrishna et al, 2020 ) reduced NF-κB expression in monocytes, macrophages, and fibrotic lung tissues of mice.…”

“…WS aqueous extract along with fatty acids and withaferin A inhibited IL-6 and IL-1β released by monocytes, macrophages, and keratinocytes in vitro ( Balkrishna et al, 2020 ) ( Dubey et al, 2018 ) ( Neog et al, 2018 ) ( Sikandan et al, 2018 ). Withaferin A and WS aqueous extract significantly reduced serum levels of IL-1β and IL-6 secreted by bone marrow-derived dendritic cells and macrophages ( Morsy et al, 2019 ) ( Sultana etal., 2017 ) ( Noh et al, 2016 ) ( Kim et al, 2015 ) ( Khan et al, 2018 ). Withaferin A also suppressed IL-1β expression of the fibrotic lung of a mouse ( Bale et al, 2018a ).…”

“…Prx1 acts as DAMPs to bind TLR4 and stimulates the secretion of pro-inflammatory cytokines ( Riddell et al, 2010 ). A piece of strong mechanistic evidence revealed that withaferin A inhibits the chaperone activity of Prx1 and reduces the infection-induced TLR4 expression in macrophages ( Noh et al, 2016 ) ( Zhao Q. et al, 2015 ). Furthermore, cytoplasmic glucocorticoid receptors (GRs) contribute to the anti-inflammatory effect upon binding of glucocorticoids (class of steroids) through distinct molecular mechanisms ( Baschant and Tuckermann, 2010 ).…”