Immunomodulatory Properties of Dental-Derived Mesenchymal Stem Cells

Mesenchymal stem cells are considered as an attractive tool for tissue regeneration. Almost all dental tissues contain a population of MSC-like cells, which were extensively studied within the last few years. Besides their ability to differentiate into different cell types, dental MSCs also possess strong immunomodulatory properties. Dental MSCs modulate both innate and adaptive immune response and influence the activity of almost all components of the immune system. The interaction between dental MSCs and the immune system is reciprocal because immunomodulatory activity of MSCs is strongly regulated by cytokines produced by immune cells. MSCs isolated from inflamed tissues might exhibit impaired immunomodulatory capacity, suggesting a potential role of these cells in inflammatory diseases and particularly periodontitis. Recent studies suggest that immunomodulatory properties of MSCs can also play an important role in their tissue regenerative capacity. The therapeutic effects of MSCs, including their immunomodulatory capacity, are largely explained by their tropic activity, including production of immunomodulatory proteins and growth factors. Summarizing, dental MSCs play an important role in tissue homeostasis under healthy and diseased conditions

Shut-down of type IX protein secretion alters the host immune response to Tannerella forsythia and Porphyromonas gingivalis

Tannerella forsythia and Porphyromonas gingivalis target distinct virulence factors bearing a structurally conserved C-terminal domain (CTD) to the type IX protein secretion system (T9SS). The T9SS comprises an outer membrane translocation complex which works in concert with a signal peptidase for CTD cleavage. Among prominent T9SS cargo linked to periodontal diseases are the TfsA and TfsB components of T. forsythia’s cell surface (S-) layer, the bacterium’s BspA surface antigen and a set of cysteine proteinases (gingipains) from P. gingivalis. To assess the overall role of the bacterial T9SS in the host response, human macrophages and human gingival fibroblasts were stimulated with T. forsythia and P. gingivalis wild-type bacteria and T9SS signal peptidase-deficient mutants defective in protein secretion, respectively. The immunostimulatory potential of these bacteria was compared by analyzing the mRNA expression levels of the pro-inflammatory mediators IL-6, IL-8, MCP-1 and TNF-$\alpha$ by qPCR and by measuring the production of the corresponding proteins by ELISA. Shot-gun proteomics analysis of T. forsythia and P. gingivalis outer membrane preparations confirmed that several CTD-bearing virulence factors which interact with the human immune system were depleted from the signal peptidase mutants, supportive of effective T9SS shut-down. Three and, more profoundly, 16 hours post stimulation, the T. forsythia T9SS mutant induced significantly less production of cytokines and the chemokine in human cells compared to the corresponding parent strain, while the opposite was observed for the P. gingivalis T9SS mutant. Our data indicate that T9SS shut-down translates into an altered inflammatory response in periodontal pathogens. Thus, the T9SS as a potential novel target for periodontal therapy needs further evaluation

Oral streptococci: modulators of health and disease

Streptococci are primary colonizers of the oral cavity where they are ubiquitously present and an integral part of the commensal oral biofilm microflora. The role oral streptococci play in the interaction with the host is ambivalent. On the one hand, they function as gatekeepers of homeostasis and are a prerequisite for the maintenance of oral health - they shape the oral microbiota, modulate the immune system to enable bacterial survival, and antagonize pathogenic species. On the other hand, also recognized pathogens, such as oral Streptococcus mutans and Streptococcus sobrinus, which trigger the onset of dental caries belong to the genus Streptococcus. In the context of periodontitis, oral streptococci as excellent initial biofilm formers have an accessory function, enabling late biofilm colonizers to inhabit gingival pockets and cause disease. The pathogenic potential of oral streptococci fully unfolds when their dissemination into the bloodstream occurs; streptococcal infection can cause extra-oral diseases, such as infective endocarditis and hemorrhagic stroke. In this review, the taxonomic diversity of oral streptococci, their role and prevalence in the oral cavity and their contribution to oral health and disease will be discussed, focusing on the virulence factors these species employ for interactions at the host interface

Multispecies biofilm behavior and host interaction support the association of Tannerella serpentiformis with periodontal health

The recently identified bacterium Tannerella serpentiformis is the closest phylogenetic relative of Tannerella forsythia, whose presence in oral biofilms is associated with periodontitis. Conversely, T. serpentiformis is considered health-associated. This discrepancy was investigated in a comparative study of the two Tannerella species. The biofilm behavior was analyzed upon their addition and of Porphyromonas gingivalis-each bacterium separately or in combinations-to an in vitro five-species oral model biofilm. Biofilm composition and architecture was analyzed quantitatively using real-time PCR and qualitatively by fluorescence in situ hybridization/confocal laser scanning microscopy, and by scanning electron microscopy. The presence of T. serpentiformis led to a decrease of the total cell number of biofilm bacteria, while P. gingivalis was growth-promoting. This effect was mitigated by T. serpentiformis when added to the biofilm together with P. gingivalis. Notably, T. serpentiformis outcompeted T. forsythia numbers when the two species were simultaneously added to the biofilm compared to biofilms containing T. forsythia alone. Tannerella serpentiformis appeared evenly distributed throughout the multispecies biofilm, while T. forsythia was surface-located. Adhesion and invasion assays revealed that T. serpentiformis was significantly less effective in invading human gingival epithelial cells than T. forsythia. Furthermore, compared to T. forsythia, a higher immunostimulatory potential of human gingival fibroblasts and macrophages was revealed for T. serpentiformis, based on mRNA expression levels of the inflammatory mediators interleukin 6 (IL-6), IL-8, monocyte chemoattractant protein-1 and tumor necrosis factor α, and production of the corresponding proteins. Collectively, these data support the potential of T. serpentiformis to interfere with biological processes relevant to the establishment of periodontitis

Investigating the role of a Tannerella forsythia HtrA protease in host protein degradation and inflammatory response

IntroductionDegradation of host proteins by bacterial proteases leads to the subversion of the host response and disruption of oral epithelial integrity, which is considered an essential factor in the progression of periodontitis. High-temperature requirement A (HtrA) protease, which is critical for bacterial survival and environmental adaptation, is found in several oral bacteria, including the periodontal pathogen Tannerella forsythia. This study investigated the proteolytic activity of HtrA from T. forsythia and its ability to modulate the host response.MethodsHtrA of T. forsythia was identified bioinformatically and produced as a recombinant protein. T. forsythia mutants with depleted and restored HtrA production were constructed. The effect of T. forsythia wild-type, mutants and recombinant HtrA on the degradation of casein and E-cadherin was tested in vitro. Additionally, the responses of human gingival fibroblasts and U937 macrophages to the different HtrA-stimuli were investigated and compared to those triggered by the HtrA-deficient mutant.ResultsT. forsythia wild-type producing HtrA as well as the recombinant enzyme exhibited proteolytic activity towards casein and E-cadherin. No cytotoxic effect of either the wild-type, T. forsythia mutants or rHtrA on the viability of host cells was found. In hGFB and U937 macrophages, both T. forsythia species induced an inflammatory response of similar magnitude, as indicated by gene and protein expression of interleukin (IL)-1β, IL-6, IL-8, tumour necrosis factor α and monocyte chemoattractant protein (MCP)-1. Recombinant HtrA had no significant effect on the inflammatory response in hGFBs, whereas in U937 macrophages, it induced a transient inflammatory response at the early stage of infection.ConclusionHtrA of T. forsythia exhibit proteolytic activity towards the host adhesion molecule E-cadherin and has the potential to influence the host response. Its role in the progression of periodontitis needs further clarification

Contraction Dynamics of Rod Microtissues of Gingiva-Derived and Periodontal Ligament-Derived Cells

Tissue engineering strategies using microtissues as “building blocks” have high potential in regenerative medicine. Cognition of contraction dynamics involved in the in vitro self-assembly of these microtissues can be conceived as the bedrock of an effective periodontal tissue regenerative therapy. Our study was directed at evaluating the shrinkage in the rod-shaped structure of a directed self-assembly of human gingiva-derived cells (GC) and periodontal ligament-derived cells (PDLC) and developing insights into the potential mechanisms responsible for the shrinkage. GC and PDLC were seeded in non-adherent agarose molds to form rod microtissues. Cells used for the experiments were characterized using fluorescence-activated cell sorting (FACS). To assess the viability, resazurin-based cytotoxicity assays, trypan blue dye exclusion assay, MTT and live/dead staining, and histological evaluation of rods based on hematoxylin and eosin staining were performed. Rod contraction was evaluated and measured at 0, 2, 6, and 24 h and compared to L-929 cells. The role of transforming growth factor (TGF)-β signaling, phosphoinositide 3-kinase (PI3K)/AKT, and mitogen activated protein kinase (MAPK) signaling was analyzed. Our results show that the rod microtissues were vital after 24 h. A reduction in the length of rods was seen in the 24 h period. While the recombinant TGF-β slightly reduced contraction, inhibition of TGF-β signaling did not interfere with the contraction of the rods. Interestingly, inhibition of phosphoinositide 3-kinase by LY294002 significantly delayed contraction in GC and PDLC rods. Overall, GC and PDLC have the ability to form rod microtissues which contract over time. Thus, approaches for application of these structures as “building blocks” for periodontal tissue regeneration should consider that rods have the capacity to contract substantially. Further investigation will be needed to unravel the mechanisms behind the dynamics of contraction

The Alterations in CD14 Expression in Periodontitis: A Systematic Review

Objective: Cluster of differentiation (CD14) is an important protein involved in activating toll-like receptors by bacterial components. It exists as either a transmembrane or soluble protein, called mCD14 or sCD14, respectively. Several studies show that CD14 regulates the inflammatory response to periodontal pathogens, and its expression is altered in periodontitis, an inflammatory disease of tooth-supporting tissues. It is the intent of this review to investigate the levels of expression of mCD14 and sCD14 in peripheral blood monocytes, saliva, gingival crevicular fluid, and gingival tissue biopsies in periodontitis patients. Methods: PubMed, Scopus, Ovid/Medline, Embase, and the Cochrane Library were consulted for the online literature search. To ensure methodical quality, titles and abstracts were reviewed in accordance to the PRISMA guidelines. Data extraction and evaluation of the full texts were executed in agreement with the GRADE approach. Results: This systematic review shows that mCD14 levels are decreased in peripheral blood monocytes of periodontitis patients in comparison to healthy patients, while sCD14 levels in sera, gingival crevicular fluid (GCF), and biopsies of periodontitis patients have a tendency to be increased in comparison to healthy controls. The evaluation of CD14 in gingival biopsies and periodontal tissues elucidated the fact that interpretation of the data obtained with qPCR, ELISA, and flow cytometry is questionable

Oxidative Stress and Antioxidant System in Periodontitis

Periodontitis is a common inflammatory disease, which is initiated by bacterial infection and subsequently progressed by aberrant host response. It can result in the destruction of teeth supporting tissues and have an influence on systemic health. When periodontitis occurs, reactive oxygen species, which are overproduced mostly by hyperactive neutrophils, could not be balanced by antioxidant defense system and cause tissues damage. This is characterized by increased metabolites of lipid peroxidation, DNA damage and protein damage. Local and systemic activities of antioxidants can also be influenced by periodontitis. Total antioxidant capacity, total oxidant status and oxidative stress index have been used to evaluate the oxidative stress associated with periodontitis. Studies have confirmed that inflammatory response in periodontitis is associated with an increased local and systemic oxidative stress and compromised antioxidant capacity. Our review focuses on increased oxidative stress in periodontal disease, specifically, on the relationship between the local and systemic biomarkers of oxidative stress and periodontitis and their association with the pathogenesis of periodontitis. Also, the relationship between periodontitis and systemic inflammation, and the effects of periodontal therapy on oxidative stress parameters will be discussed