Filifactor alocis Promotes Neutrophil Degranulation and Chemotactic Activity

Abstract

Filifactor alocis is a recently recognized periodontal pathogen; however, little is known regarding its interactions with the immune system. As the first-responder phagocytic cells, neutrophils are recruited in large numbers to the periodontal pocket, where they play a crucial role in the innate defense of the periodontium. Thus, in order to colonize, successful periodontal pathogens must devise means to interfere with neutrophil chemotaxis and activation. In this study, we assessed major neutrophil functions, including degranulation and cell migration, associated with the p38 mitogen-activated protein kinase (MAPK) signaling pathway upon challenge with F. alocis. Under conditions lacking a chemotactic gradient, F. alocischallenged neutrophils had increased migration compared to uninfected cells, indicating that F. alocis increases chemokinesis in human neutrophils. In addition, neutrophil chemotaxis induced by interleukin-8 was significantly enhanced when cells were challenged with F. alocis, compared to noninfected cells. Similar to live bacteria, heat-killed F. alocis induced both random and directed migration of human neutrophils. The interaction of F. alocis with Toll-like receptor 2 induced granule exocytosis along with a transient ERK1/2 and sustained p38 MAPK activation. Moreover, F. alocis-induced secretory vesicle and specific granule exocytosis were p38 MAPK dependent. Blocking neutrophil degranulation with TAT-SNAP23 fusion protein significantly reduced the chemotactic and random migration induced by F. alocis. Therefore, we propose that induction of random migration by F. alocis will prolong neutrophil traffic time in the gingival tissue, and subsequent degranulation will contribute to tissue damage.

Filifactor alocismodulates human neutrophil antimicrobial functional responses

“…8A and B). Similar to the response we described with non-opsonized F. alocis (Armstrong, Miralda, Neff, Tian, Vashishta, Perez, … Uriarte, 2016); serum opsonized F. alocis significantly induced secretory vesicles and specific granule exocytosis (Fig. 8A and B).…”

“…We recently described that non-opsonized F. alocis , through TLR2 activation, promotes exocytosis of secretory vesicles and specific granules but not azurophil granules in human neutrophils; as well as enhanced random and directed migration (Armstrong, Miralda, Neff, Tian, Vashishta, Perez,… Uriarte, 2016). We next examined if opsonized HK F. alocis would induce a similar or distinct granule exocytosis pattern compared to the live organism.…”

“…These granules contain myeloperoxidase which is involved in the formation of hypochlorous acid (HOCl) within the phagosome; therefore, they are critical to the oxidative burst response and to phagosome maturation (Nauseef, 2007). Our laboratory recently reported that challenge of human neutrophils with non-opsonzied F. alocis did not induce azurophil granule exocytosis (Armstrong, Miralda, Neff, Tian, Vashishta, Perez, … Uriarte, 2016). Our confocal results from Fig.…”

Peptoanaerobacter stomatis Primes Human Neutrophils and Induces Granule Exocytosis

“…Similarly, F. nucleatum, P. gingivalis, and T. denticola can induce the release of elastase from neutrophils, and Aggregatibacter actinomycetemcomitans can induce MPO release (40,51,52). In contrast, we recently showed that the newly recognized periodontal pathogen Filifactor alocis did not induce azurophil granule release even at a high multiplicity of infection (53). Hence, various neutrophil responses may contribute to the pathogenic potential of different pathogenic bacteria.…”

Characterization of Filifactor alocis and its immune evasion strategies employed against human neutrophils.

“…Neutrophils recognize F. alocis through TLR2, which triggers degranulation and cell migration (325 saw that PAM3CSK4 induced priming in only certain neutrophil donor populations, however FSL-1 induced priming in neutrophils from all of our donor pool (Fig. 3-4 A-B).…”