Akkermansia muciniphila reduces Porphyromonas gingivalis-induced inflammation and periodontal bone destruction.

AIM: Akkermansia muciniphila is a beneficial gut commensal, whose anti-inflammatory properties have recently been demonstrated. This study aimed to evaluate the effect of A. muciniphila on Porphyromonas gingivalis elicited inflammation. MATERIAL AND METHODS: In lean and obese mice, A. muciniphila was administered in P. gingivalis-induced calvarial abscess and in experimental periodontitis model (EIP). Bone destruction and inflammation were evaluated by histomorphometric analysis. In vitro, A. muciniphila was co-cultured with P. gingivalis, growth and virulence factor expression was evaluated. Bone marrow macrophages (BMMϕ) and gingival epithelial cells (TIGK) were exposed to both bacterial strains, and the expression of inflammatory mediators, as well as tight junction markers, was analysed. RESULTS: In a model of calvarial infection, A. muciniphila decreased inflammatory cell infiltration and bone destruction. In EIP, treatment with A. muciniphila resulted in a decreased alveolar bone loss. In vitro, the addition of A. muciniphila to P. gingivalis-infected BMMϕ increased anti-inflammatory IL-10 and decreased IL-12. Additionally, A. muciniphila exposure increases the expression of junctional integrity markers such as integrin-ß1, E-cadherin and ZO-1 in TIGK cells. A. muciniphila co-culture with P. gingivalis reduced gingipains mRNA expression. DISCUSSION: This study demonstrated the protective effects of A. muciniphila administration and may open consideration to its use as an adjunctive therapeutic agent to periodontal treatment.

CC chemokine CCL1 receptor CCR8 mediates conversion of mesenchymal stem cells to embryoid bodies expressing FOXP3+CCR8+ regulatory T cells.

Embryoid bodies (EBs) are three dimensional aggregates of pluripotent stem cells primarily used to investigate morphogenesis and cell toxicity, are also attractive tools in regenerative medicine. While embryonic stem cells (ESCs) and induced pluripotent cells (IPSCs) have been shown to form EBs in mouse, primate and humans, EB formation have not been previously demonstrated in mesenchymal stem cells (MSCs). Here we show that rat MSCs form EBs; which express regulatory T cell (Treg) marker Foxp3 and CC chemokine CCL1 receptor CCR8. We show a novel method for formation of EBs from MSCs under stress and demonstrate that the induction of FoxP3+ CCR8+ EBs is dependent upon CCL1 gradients which mediate cell proliferation, migration and invasion of mTregs. The identification of EBs and novel FoxP3+ CCR8+ regulatory T cells (mTregs) for selective conversion and isolation of bone marrow derived MSCs offers novel avenues for research, diagnosis and treatment.

Isolation and characterization of a large soluble form of fibronectin that stimulates adhesion, spreading, and alignment of mouse erythroleukemia cells.

Fibronectin (FN) is a major component of the extracellular matrix which plays important roles in a variety of cellular processes including cell adhesion, and migration. The soluble cellular form of FN has a monomer molecular weight of approximately 250 kDa, and generally exists as a dimer of 500 kDa. We have isolated a different form of soluble FN from mouse breast cancer cell line SC115 conditioned medium (CM) and purified it to homogeneity as evidenced by both native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate PAGE. It still exhibits a monomeric form of about 250 kDa while its form in the CM is stable and soluble with an apparent tetrameric molecular weight in the range of 800-1000 kDa. This form of FN is a potent cell adhesion factor (AF) that induces adhesion to polystyrene, elongation, spreading, alignment or "track" formation, and migration of mouse erythroleukemia cells. Column fractions homogeneous for AF protein were able to stimulate 10% cell adhesion at concentrations of 23 ng/ml and 1.9 ng/cm(2). Purified AF induced 50% cell adhesion at 94 ng/ml and 7.5 ng/cm(2). AF also increased the migration of human aortic smooth muscle and vascular endothelial cells. However, this form of FN differs from other forms as it does not bind tightly to either gelatin or heparin. Studies of this AF should shed light on adhesion of cells to extracellular matrix molecules and on cell migration, both of which are critical in several biological processes such as wound healing, metastasis, matrix formation and structure, and organ development.

Role of CCR8 and other chemokine pathways in the migration of monocyte-derived dendritic cells to lymph nodes.

Studying the influence of chemokine receptors (CCRs) on monocyte fate may reveal information about which subpopulations of monocytes convert to dendritic cells (DCs) and the migration pathways that they use. First, we examined whether prominent CCRs on different monocyte subsets, CCR2 or CX3CR1, mediated migration events upstream of the accumulation of monocyte-derived DCs in lymph nodes (LNs). Monocytes were labeled and traced by uptake of latex microspheres in skin. Unexpectedly, neither CCR2 nor CX3CR1 were required. However, absence of CCR2 led to an increased labeling of the minor Gr-1int monocyte population, and the number of latex+ DCs that emigrated to LNs was correspondingly increased. Characterization of Gr-1int monocytes revealed that they selectively expressed CCR7 and CCR8 mRNA in blood. CCR7 and CCR8 pathways were used by monocyte-derived DCs during mobilization from skin to LNs. The role of CCR8 in emigration from tissues also applied to human monocyte-derived cells in a model of transendothelial trafficking. Collectively, the data suggest that Gr-1int monocytes may be most disposed to become a lymphatic-migrating DCs. When these monocyte-derived DCs exit skin to emigrate to LNs, they use not only CCR7 but also CCR8, which was not previously recognized to participate in migration to LNs.

Chemokine receptor-8 (CCR8) mediates human vascular smooth muscle cell chemotaxis and metalloproteinase-2 secretion.

The response of the arterial vascular wall to injury is characterized by vascular smooth muscle cell (VSMC) migration, a process requiring metalloproteinase production. This migration is induced by cytokines, however the agonists involved are not fully defined. The CC chemokine receptor 8 (CCR8) is expressed on monocytes and T lymphocytes and is the sole receptor for the human CC chemokine 1 (CCL1, I-309) and for the viral chemokine, vCCL1 (viral macrophage inflammatory protein 1 [vMIP-1]). We have reported that CCR8 is expressed on human umbilical vein endothelial cells (HUVECs) and mediates chemotaxis induced by CCL1. The conditioned medium from incubation mixtures of lipoprotein(a) (Lp(a)) and HUVECs (LCM) contained CCL1 and stimulated both monocyte and HUVEC chemotaxis, providing novel mechanisms for the atherogenicity of Lp(a). We now report that CCL1, vCCL1, and LCM stimulate chemotaxis of human VSMCs that is blocked by murine monoclonal antibody against CCR8 and by the G-protein inhibitor pertussis toxin. The effect of anti-CCR8 was specific, as this antibody failed to effect the chemotaxis of VSMCs in response to CCL3 or by platelet-derived growth factor BB (PDGF-BB). VSMCs contained CCR8 mRNA and CCR8 antigen coprecipitated with VSMC membranes. Antibodies against metalloproteinase-2 (MMP-2) inhibited the CCL1-induced chemotaxis of VSMCs, whereas anti-MMP-9 was less effective. CCL1 induced VSMC pro-MMP-2 mRNA and protein secretion. Poxvirus MC148 inhibited the increase in MMP-2 induced by CCL1, documenting that CCR8 was the receptor responsible. In mouse femoral arteries, CCR8 and TCA3 antigen colocalized with VSMCs and were up-regulated after injury. The induction of CCR8 and CCL1/TCA3 under conditions associated with VSMC proliferation and migration raises the possibility that CCR8 may play an important role in vessel wall pathology.

Chemokine receptor-8: potential role in atherogenesis.

Chemokines and their receptors play regulatory roles in inflammatory reactions. Lipoprotein(a) is an atherogenic lipoprotein, however the mechanisms of its actions are not defined. Our interest in chemokines and their receptors was stimulated by the finding that incubation of Lp(a) with human umbilical vein endothelial cells produced a conditioned medium that was chemotactic for human monocytes. Since infiltration of monocytes into the vessel wall is an early lesion in atherosclerosis, this finding provided a novel mechanism to explain the relationship between Lp(a) and atherosclerosis. The chemoattractant produced by HUVEC was identified as CCL1/I-309, a CC chemokine previously reported to be secreted by stimulated monocytes/macrophages and T lymphocytes. CCR8, the CCL1 receptor, was identified on endothelial cells, and CCL1 was found to be a chemoattractant for these cells. Most recently we demonstrated functional CCR8 on human vascular smooth muscle cells and found that the Lp(a)-HUVEC conditioned medium is a chemoattractant for these cells. CCL1 increased metalloproteinase-2 production by HUVEC, an activity that enables these cells to remodel the vascular matrix. These studies suggest that CCR8 may play an important role in arterial wall pathology.