Unfolding Role of a Danger Molecule Adenosine Signaling in Modulation of Microbial Infection and Host Cell Response.

Ectonucleotidases CD39 and CD73, specific nucleotide metabolizing enzymes located on the surface of the host, can convert a pro-inflammatory environment driven by a danger molecule extracellular-ATP to an adenosine-mediated anti-inflammatory milieu. Accordingly, CD39/CD73 signaling have has strongly implicated in modulating the intensity, duration, and composition of purinergic danger signals delivered to host. Recent studies have eluted potential roles for CD39 and CD73 in selective triggering of a variety of host immune cells and molecules in the presence of pathogenic microorganisms or microbial virulence molecules. Growing evidence also suggests that CD39 and CD73 present complimentary, but likely differential, actions against pathogens to shape the course and severity of microbial infection as well as the associated immune response. Similarly, adenosine receptors A2A and A2B have been proposed to be major immunomodulators of adenosine signaling during chronic inflammatory conditions induced by opportunistic pathogens, such as oral colonizer Porphyromonas gingivalis. Therefore, we here review the recent studies that demonstrate how complex network of molecules in the extracellular adenosine signaling machinery and their interactions can reshape immune responses and may also be targeted by opportunistic pathogens to establish successful colonization in human mucosal tissues and modulate the host immune response.

Host surface ectonucleotidase-CD73 and the opportunistic pathogen, Porphyromonas gingivalis, cross-modulation underlies a new homeostatic mechanism for chronic bacterial survival in human epithelial cells.

Cell surface nucleotide-metabolizing enzyme, ectonucleotidase-CD73, has emerged as a central component of the cellular homeostatic-machinery that counterbalances the danger-molecule (extracellular-ATP)-driven proinflammatory response in immune cells. While the importance of CD73 in microbial host fitness and symbiosis is gradually being unraveled, there remains a significant gap in knowledge of CD73 and its putative role in epithelial cells. Here, we depict a novel host-pathogen adaptation mechanism where CD73 takes a center role in the intracellular persistence of Porphyromonas gingivalis, a major colonizer of oral mucosa, using human primary gingival epithelial cell (GEC) system. Temporal analyses revealed, upon invasion into the GECs, P. gingivalis can significantly elevate the host-surface CD73 activity and expression. The enhanced and active CD73 significantly increases P. gingivalis intracellular growth in the presence of substrate-AMP and simultaneously acts as a negative regulator of reactive oxygen species (ROS) generation upon eATP treatment. The inhibition of CD73 by siRNA or by a specific inhibitor markedly increases ROS production. Moreover, CD73 and P. gingivalis cross-signaling significantly modulates pro-inflammatory interleukin-6 (IL-6) in the GECs. Conversely, exogenous treatment of the infected GECs with IL-6 suppresses the intracellular bacteria via amplified ROS generation. However, the decreased bacterial levels can be restored by overexpressing functionally active CD73. Together, these findings illuminate how the local extracellular-purine-metabolism, in which CD73 serves as a core molecular switch, can alter intracellular microbial colonization resistance. Further, host-adaptive pathogens such as P. gingivalis can target host ectonucleotidases to disarm specific innate defenses for successful intracellular persistence in mucosal epithelia.

In Situ Intraepithelial Localizations of Opportunistic Pathogens, Porphyromonas gingivalis and Filifactor alocis, in Human Gingiva.

Porphyromonas gingivalis and Filifactor alocis are fastidious oral pathogens and etiological agents associated with chronic periodontitis. Although previous studies showed increased levels of the two obligate anaerobic species in periodontitis patients, methodologies for this knowledge were primarily limited to sampling subgingival plaque, saliva, or gingival crevicular fluid. To evaluate the extent to which P. gingivalis and F. alocis may invade the periodontal tissues, an in situ cross-sectional study was comparatively conducted on the gingival biopsy specimens of patients diagnosed with periodontal health or chronic periodontitis. Immunostained tissue sections for each organism were imaged by Super-Resolution Confocal Scanning Microscopy to determine the relative presence and localization of target bacterial species. Fluorescence-in-situ-hybridization (FISH) coupled with species specific 16S rRNA method was utilized to confirm whether detected bacteria were live within the tissue. In periodontitis, P. gingivalis and F. alocis revealed similarly concentrated localization near the basement membrane or external basal lamina of the gingival epithelium. The presence of both bacteria was significantly increased in periodontitis vs. healthy tissue. However, P. gingivalis was still detected to an extent in health tissue, while only minimal levels of F. alocis were spotted in health. Additionally, the micrographic analyses displayed heightened formation of epithelial microvasculature containing significantly co-localized and metabolically active dual species within periodontitis tissue. Thus, this study demonstrates, for the first-time, spatial arrangements of P. gingivalis and F. alocis in both single and co-localized forms within the complex fabric of human gingiva during health and disease. It also exhibits critical visualizations of co-invaded microvascularized epithelial layer of the tissue by metabolically active P. gingivalis and F. alocis from patients with severe periodontitis. These findings collectively uncover novel visual evidence of a potential starting point for systemic spread of opportunistic bacteria during their chronic colonization in gingival epithelium.