Role of dendritic cell-mediated immune response in oral homeostasis: A new mechanism of osteonecrosis of the jaw.

Dendritic cells are an important link between innate and adaptive immune response. The role of dendritic cells in bone homeostasis, however, is not understood. Osteoporosis medications that inhibit osteoclasts have been associated with osteonecrosis, a condition limited to the jawbone, thus called medication-related osteonecrosis of the jaw. We propose that disruption of the local immune response renders the oral microenvironment conducive to osteonecrosis. We tested whether zoledronate (Zol) treatment impaired dendritic cell (DC) functions and increased bacterial load in alveolar bone in vivo and whether DC inhibition alone predisposed the animals to osteonecrosis. We also analyzed the role of Zol in impairment of differentiation and function of migratory and tissue-resident DCs, promoting disruption of T-cell activation in vitro. Results demonstrated a Zol induced impairment in DC functions and an increased bacterial load in the oral cavity. DC-deficient mice were predisposed to osteonecrosis following dental extraction. Zol treatment of DCs in vitro caused an impairment in immune functions including differentiation, maturation, migration, antigen presentation, and T-cell activation. We conclude that the mechanism of Zol-induced osteonecrosis of the jaw involves disruption of DC immune functions required to clear bacterial infection and activate T cell effector response.

Enterococcus faecalis Induces Differentiation of Immune-Aberrant Dendritic Cells from Murine Bone Marrow-Derived Stem Cells.

Enterococcus faecalis, long implicated in serious systemic infections and failure of root canal treatment, is a persistent inhabitant of oral periapical lesions. Dendritic cells (DCs) and other innate immune cells patrol the oral mucosa for infecting microbes. Dendritic cells are efficient at capturing microbes when immature, whereupon they can transform into potent antigen-presenting cells upon full maturation. Autophagy, a sophisticated intracellular process first described for elimination of damaged organelles, regulates DC maturation and other important immune functions of DCs. The present study examined how E. faecalis influences the differentiation of murine bone marrow-derived stem cells (BMSCs) into functional DCs in the presence of the cytokines granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). Although the viability and differentiation of DCs were not affected by E. faecalis, expression of the autophagy-related proteins ATG7, Beclin1, and LC3bI/II were significantly suppressed in an mTOR-dependent manner. Ultrastructurally, E. faecalis was identified in single-membrane vacuoles, some of which were in the process of binary fission. Bacterium-containing autophagosomes were absent within the cytoplasm. Accessory molecules (major histocompatibility complex class II [MHC-II], CD80, and CD86) and anti-inflammatory cytokine (transforming growth factor ß1 [TGF-ß1]) were suppressed in E. faecalis-induced DCs, while IL-1ß, tumor necrosis factor alpha (TNF-α), and IL-12 levels were upregulated. When pulsed with ovalbumin (OVA), the E. faecalis-induced DCs showed reduction in CD4+ OVA-specific OT-II T cell proliferation. It is concluded that E. faecalis promotes the differentiation of bone marrow stem cells into CD11c-positive DCs with aberrant immune functions while retaining the capability of proinflammatory cytokine induction.

Enterococcus faecalis shifts macrophage polarization toward M1-like phenotype with an altered cytokine profile.

Background: The macrophage is an innate immune defense cell involved in pathogen recognition and clearance. Aim: In view of the diversity of the macrophage phenotype and function, the present study investigated how Enterococcus faecalis infection affects the differentiation, phenotype and cytokine profile of macrophages. Methods: Murine bone marrow-derived stem cells were co-cultured with E. faecalis before and after differentiation. Macrophage M0 polarization towards M1 or M2 was initiated at day 6 by addition of LPS and INF-γ, or IL-4 and IL-13, respectively. Results: E. faecalis did not inhibit macrophage differentiation and were identified within macrophages. Viability of the macrophages infected with E. faecalis prior to differentiation was enhanced, evidenced by apoptosis inhibition, as was expression of CD38 and IRF5 proteins, indicators of M1-like polarization. These M1-like macrophages expressed an aberrant cytokine mRNA profile, with reduction in inflammatory cytokines IL-1ß and IL-12 and increase in regulatory cytokine IL-10. No changes in TNF-α or TGF-ß1 were detected, compared with the control groups. This atypical M1-like phenotype was retained even upon stimulation with growth factors that normally trigger their development into M2 macrophages. Conclusions: These findings suggested that E. faecalis infection of bone marrow-derived stem cells during differentiation into macrophages induces an atypical M1-like phenotype associated with intracellular bacterial survival.

Dendritic cell derived exosomes loaded with immunoregulatory cargo reprogram local immune responses and inhibit degenerative bone disease in vivo.

Chronic bone degenerative diseases represent a major threat to the health and well-being of the population, particularly those with advanced age. This study isolated exosomes (EXO), natural nano-particles, from dendritic cells, the "directors" of the immune response, to examine the immunobiology of DC EXO in mice, and their ability to reprogram immune cells responsible for experimental alveolar bone loss in vivo. Distinct DC EXO subtypes including immune-regulatory (regDC EXO), loaded with TGFB1 and IL10 after purification, along with immune stimulatory (stimDC EXO) and immune "null" immature (iDCs EXO) unmodified after purification, were delivered via I.V. route or locally into the soft tissues overlying the alveolar bone. Locally administrated regDC EXO showed high affinity for inflamed sites, and were taken up by both DCs and T cells in situ. RegDC EXO-encapsulated immunoregulatory cargo (TGFB1 and IL10) was protected from proteolytic degradation. Moreover, maturation of recipient DCs and induction of Th17 effectors was suppressed by regDC EXO, while T-regulatory cell recruitment was promoted, resulting in inhibition of bone resorptive cytokines and reduction in osteoclastic bone loss. This work is the first demonstration of DC exosome-based therapy for a degenerative alveolar bone disease and provides the basis for a novel treatment strategy.