Transcriptomic features of programmed and inflammatory cell death in gingival tissues.

The local gingival tissue environment with homeostasis and tissue-destructive events of periodontitis demonstrates major changes in histological features and biology of the oral/sulcular epithelium, fibroblasts, vascular cells, inflammatory cell infiltration, and alveolar bone. OBJECTIVE: This study used an experimental periodontitis model to detail the gingival transcriptome related to cell death processes of pyroptosis, necroptosis, ferroptosis, and cuproptosis. MATERIALS AND METHODS: Healthy Macaca mulatta primates stratified by age, ≤3 years (young), 7-12 years (adolescent), 12-15 years (adult), and 17-23 years (aged), provided gingival tissue biopsies for microarray analysis focused on 257 genes representative of the four cell death processes and bacterial plaque samples for 16S rRNA gene analysis. RESULTS: Age differences in the profiles of gene expression in healthy tissues were noted for cuproptosis, ferroptosis, necroptosis, and pyroptosis. Major differences were then observed with disease initiation, progression, and resolution also related to the age of the animals. Distinct bacterial families/consortia of species were significantly related to the gene expression differences for the cell death pathways. CONCLUSIONS: These results emphasized age-associated differences in the gingival tissue molecular response to changes in the quality and quantity of bacteria accumulating with the disease process reflected in regulated cell death pathways that are both physiological and pathophysiological.

Stereoselective synthesis of vinyl nitriles through a Ramberg-Bäcklund approach.

The applicability of vinyl nitriles in the preparation of pharmaceuticals, polymers, and other valuable materials benefits from robust preparative methodologies. In this work, we present a novel approach for the synthesis of vinyl nitriles based on the Ramberg-Bäcklund olefination reaction (RBR). While there are few examples for accessing functionalized olefins using the RBR, we believe that this methodology embodies useful means for installing privileged vinylnitrile building blocks, such as the acrylonitrile fragment of the US FDA-approved antiviral rilpivirine.

Sex effects on gingival transcriptomic patterns during initiation, progression, and resolution of periodontitis.

BACKGROUND: The prevalence and severity of periodontitis demonstrates altered population distribution with age, sex, and race and ethnicity. While males exhibit greater frequency of disease, particularly with aging, the underlying basis for this observation remains obscure. OBJECTIVE: This study used a nonhuman primate (Macaca mulatta) model of experimental ligature-induced periodontitis in adult animals to evaluate gingival transcriptomic differences stratified based upon sex of the animal. METHODS: The 18 animals represented humans ages 40-80 years, with gingival tissue samples obtained at baseline, 0.5 months (initiation), 1 and 3 months (progression), and at 5 months that were 60 days after ligature removal for clinical disease resolution. Microarray analysis was used to quantify gene expression profiles in the gingival tissues. RESULTS: The results demonstrated clear gene expression differences in healthy (baseline) tissues between the sexes, with elevations in females associated with immune responses and elevation in males related to tissue structural genes. With disease initiation, fewer genes differed between the sexes, while these differences were significantly increased in progressing disease and resolution, particularly in male animals. Overexpressed biological processes showed tissue structural/functional genes at initiation, with host response pathways altered during disease progression. Resolution samples generally demonstrated biological processes of cellular metabolism that differed from baseline healthy samples. CONCLUSION: The transcriptomic findings support sex as a biological variable in periodontitis using a nonhuman primate model of experimental periodontitis.

Gingival transcriptomic patterns of macrophage polarization during initiation, progression, and resolution of periodontitis.

Phenotypic and functional heterogeneity of macrophages is clearly a critical component of their effective functions in innate and adaptive immunity. This investigation hypothesized that altered profiles of gene expression in gingival tissues in health, disease, and resolution would reflect changes in macrophage phenotypes occurring in these tissues. The study used a nonhuman primate model to evaluate gene expression profiles as footprints of macrophage variation using a longitudinal experimental model of ligature-induced periodontitis in animals from 3 to 23 years of age to identify aging effects on the gingival environment. Significant differences were observed in distribution of expressed gene levels for M0, M1, and M2 macrophages in healthy tissues with the younger animals showing the least expression. M0 gene expression increased with disease in all but the aged group, while M1 was increased in adult and young animals, and M2 in all age groups, as early as disease initiation (within 0.5 months). Numerous histocompatibility genes were increased with disease, except in the aged samples. An array of cytokines/chemokines representing both M1 and M2 cells were increased with disease showing substantial increases with disease initiation (e.g. IL1A, CXCL8, CCL19, CCL2, CCL18), although the aged tissues showed a more limited magnitude of change across these macrophage genes. The analytics of macrophage genes at sites of gingival health, disease, and resolution demonstrated distinct profiles of host response interactions that may help model the disease mechanisms occurring with the formation of a periodontal lesion.

Mucosal circadian rhythm pathway genes altered by aging and periodontitis.

As circadian processes can impact the immune system and are affected by infections and inflammation, this study examined the expression of circadian rhythm genes in periodontitis. METHODS: Macaca mulatta were used with naturally-occurring and ligature-induced periodontitis. Gingival tissue samples were obtained from healthy, diseased, and resolved sites in four groups: young (≤3 years), adolescent (3-7 years), adult (12-26) and aged (18-23 years). Microarrays targeted circadian rhythm (n = 42), inflammation/tissue destruction (n = 11), bone biology (n = 8) and hypoxia pathway (n = 7) genes. RESULTS: The expression of many circadian rhythm genes, across functional components of the pathway, was decreased in healthy tissues from younger and aged animals, as well as showing significant decreases with periodontitis. Negative correlations of the circadian rhythm gene levels with inflammatory mediators and tissue destructive/remodeling genes were particularly accentuated in disease. A dominance of positive correlations with hypoxia genes was observed, except HIF1A, that was uniformly negatively correlated in health, disease and resolution. CONCLUSIONS: The chronic inflammation of periodontitis exhibits an alteration of the circadian rhythm pathway, predominantly via decreased gene expression. Thus, variation in disease expression and the underlying molecular mechanisms of disease may be altered due to changes in regulation of the circadian rhythm pathway functions.

Immunoglobulin gene expression profiles and microbiome characteristics in periodontitis in nonhuman primates.

Colonization of mucosal tissues throughout the body occurs by a wide array of bacteria in the microbiome that stimulate the cells and tissues, as well as respond to changes in the local milieu. A feature of periodontitis is the detection of adaptive immune responses to members of the oral microbiome that show specificity and changes with disease and treatment. Thus, variations in antibody responses are noted across the population and affected by aging, albeit, data are still unclear as to how these differences relate to disease risk and expression. This study used a nonhuman primate model of experimental periodontitis to track local microbiome changes as they related to the use and expression of a repertoire of immunoglobulin genes in gingival tissues. Gingival tissue biopsies from healthy tissues and following ligature-placement for disease initiation and progression provided gene expression analysis. Additionally, following removal of the ligatures, clinical healing occurs with gene expression in disease resolved tissues. Groups of 9 animals (young: <3 yrs., adolescent: 3-7 yrs., adult -12 to 15 yrs.; aged: 17-22 yrs) were used in the investigation. In healthy tissues, young and adolescent animals showed levels of expression of 78 Ig genes that were uniformly less than adults. In contrast, ⅔ of the Ig genes were elevated by > 2-fold in the aged samples. Specific increases in an array of the Ig gene transcripts were detected in adults at disease initiation and throughout progression, while increases in young and adolescent animals were observed only with disease progression, and in aged samples primarily late in disease progression. Resolved lesions continued to demonstrate elevated levels of Ig gene expression in only young, adolescent and adult animals. The array of Ig genes significantly correlated with inflammatory, tissue biology and hypoxia genes in the gingival tissues, with variations associated with age. In the young group of animals, specific members of the oral microbiome positively correlated with Ig gene expression, while in the older animals, many of these correlations were negative. Significant correlations were observed with a select assortment of bacterial OTUs and multiple Ig genes in both younger and older animal samples, albeit the genera/species showed little overlap. Incorporating this array of microbes and host responses clearly discriminated the various time points in transition from health to disease and resolution in both the young and adult animals. The results support a major importance of adaptive immune responses in the kinetics of periodontal lesion formation, and support aging effects on the repertoire of Ig genes that may relate to the increased prevalence and severity of periodontitis with age.

Comparative Analysis of Gene Expression Patterns for Oral Epithelial Cell Functions in Periodontitis.

The structure and function of epithelial cells are critical for the construction and maintenance of intact epithelial surfaces throughout the body. Beyond the mechanical barrier functions, epithelial cells have been identified as active participants in providing warning signals to the host immune and inflammatory cells and in communicating various detailed information on the noxious challenge to help drive specificity in the characteristics of the host response related to health or pathologic inflammation. Rhesus monkeys were used in these studies to evaluate the gingival transcriptome for naturally occurring disease samples (GeneChip® Rhesus Macaque Genome Array) or for ligature-induced disease (GeneChip® Rhesus Gene 1.0 ST Array) to explore up to 452 annotated genes related to epithelial cell structure and functions. Animals were distributed by age into four groups: ≤ 3 years (young), 3-7 years (adolescent), 12-16 years (adult), and 18-23 years (aged). For naturally occurring disease, adult and aged periodontitis animals were used, which comprised 34 animals (14 females and 20 males). Groups of nine animals in similar age groups were included in a ligature-induced periodontitis experiment. A buccal gingival sample from either healthy or periodontitis-affected tissues were collected, and microarray analysis performed. The overall results of this investigation suggested a substantial alteration in epithelial cell functions that occurs rapidly with disease initiation. Many of these changes were prolonged throughout disease progression and generally reflect a disruption of normal cellular functions that would presage the resulting tissue destruction and clinical disease measures. Finally, clinical resolution may not signify biological resolution and represent a continued risk for disease that may require considerations for additional biologically specific interventions to best manage further disease.

Gingival tissue antibody gene utilization in aging and periodontitis.

OBJECTIVE: This study used a nonhuman primate model of ligature-induced periodontitis to document the characteristics of immunoglobulin (Ig) gene usage in gingival tissues with disease and affected by age. BACKGROUND: Adaptive immune responses to an array of oral bacteria are routinely detected in local gingival tissues and the systemic circulation across the human population. The level and diversity of antibody increases with periodontitis, reflecting the increased quantity of B cells and plasmacytes in the tissues at sites of periodontal lesions. METHODS: Macaca mulatta (n = 36) in four groups (young - ≤3 years; adolescent >3-7 years; adult - 12-15 years; aged - 17-23 years) were used in this study. Gingival tissues were sampled at baseline (health), 2 weeks (initiation), 1 and 3 months (progression), and 5 months (resolution) of the lesion development and transcriptomic analysis included 78 Ig-related genes. RESULTS: The results demonstrated extensive variation in Ig gene usage patterns and changes with the disease process that was substantially affected by the age of the animal. Of note was that the aged animals generally demonstrated elevated expression on multiple Ig genes even in the baseline/healthy gingival tissues. The expression levels revealed 5 aggregates of Ig gene change profiles across the age groups. The number of gene changes were greatly increased in adult animals with the initiation of disease, while the young and adolescent animals showed extensive changes with disease progression. Elevated Ig gene transcripts remained with disease resolution except in the aged animals. The response profiles demonstrated selective heavy/light change gene transcripts that differed with age and clustering of the transcript expression was dominated by the age of the animals. CONCLUSIONS: The results suggested potential critical variations in the molecular aspects of Ig gene expression in gingival tissues that can contribute to understanding the kinetics of periodontal lesions, as well as the variation in episodes, rapidity of progression, and role in resolution that are impacted by age.

Gingival Transcriptome of Innate Antimicrobial Factors and the Oral Microbiome With Aging and Periodontitis.

The epithelial barrier at mucosal sites comprises an important mechanical protective feature of innate immunity, and is intimately involved in communicating signals of infection/tissue damage to inflammatory and immune cells in these local environments. A wide array of antimicrobial factors (AMF) exist at mucosal sites and in secretions that contribute to this innate immunity. A non-human primate model of ligature-induced periodontitis was used to explore characteristics of the antimicrobial factor transcriptome (n = 114 genes) of gingival biopsies in health, initiation and progression of periodontal lesions, and in samples with clinical resolution. Age effects and relationship of AMF to the dominant members of the oral microbiome were also evaluated. AMF could be stratified into 4 groups with high (n = 22), intermediate (n = 29), low (n = 18) and very low (n = 45) expression in healthy adult tissues. A subset of AMF were altered in healthy young, adolescent and aged samples compared with adults (e.g., APP, CCL28, DEFB113, DEFB126, FLG2, PRH1) and were affected across multiple age groups. With disease, a greater number of the AMF genes were affected in the adult and aged samples with skewing toward decreased expression, for example WDC12, PGLYRP3, FLG2, DEFB128, and DEF4A/B, with multiple age groups. Few of the AMF genes showed a >2-fold increase with disease in any age group. Selected AMF exhibited significant positive correlations across the array of AMF that varied in health and disease. In contrast, a rather limited number of the AMF significantly correlated with members of the microbiome; most prominent in healthy samples. These correlated microbes were different in younger and older samples and differed in health, disease and resolution samples. The findings supported effects of age on the expression of AMF genes in healthy gingival tissues showing a relationship to members of the oral microbiome. Furthermore, a dynamic expression of AMF genes was related to the disease process and showed similarities across the age groups, except for low/very low expressed genes that were unaffected in young samples. Targeted assessment of AMF members from this large array may provide insight into differences in disease risk and biomolecules that provide some discernment of early transition to disease.

Streptococcus gordonii-Induced miRNAs Regulate CCL20 Responses in Human Oral Epithelial Cells.

The mechanisms through which oral commensal bacteria mitigates uncontrolled inflammatory responses of the oral mucosa remain unknown. Here, we show that representative oral bacterial species normally associated with oral health [S. gordonii (Sg), V. parvula (Vp), A. naeslundii (An), C. sputigena (Cs), and N. mucosa (Nm)] enhanced differential chemokine responses in oral epithelial cells (OECs), with some bacteria (An, Vp, and Nm) inducing higher chemokine levels (CXCL1, CXCL8) than others (Sg, Cs). Although all bacterial species (except Cs) increased CCL20 mRNA levels consistent with protein elevations in cell lysates, only An, Vp, and Nm induced higher CCL20 secretion, similar to the effect of the oral pathogen F. nucleatum (Fn). In contrast, most CCL20 remained associated with OECs exposed to Sg and negligible amounts released into the cell supernatants. Consistently, Sg attenuated An-induced CCL20. MiR-4516 and miR-663a were identified as Sg-specifically induced miRNAs modulating validated targets of chemokine-associated pathways. Cell transfection with miR-4516 and miR-663a decreased An- and Fn-induced CCL20. MiRNA upregulation and attenuation of An-induced CCL20 by Sg were reversed by catalase. Up-regulation of both miRNAs was specifically enhanced by oral streptococci H2O2-producers. These findings suggest that CCL20 levels produced by OECs in response to bacterial challenge are regulated by Sg-induced miR-4516 and miR-663a in a mechanism that involves hydrogen peroxide. This type of molecular mechanism could partly explain the central role of specific oral streptococcal species in balancing inflammatory and antimicrobial responses given the critical role of CCL20 in innate (antimicrobial) and adaptive immunity (modulates Th17 responses).

Porphyromonas gingivalis: where do we stand in our battle against this oral pathogen?

Periodontal diseases, such as gingivitis and periodontitis, are inflammatory diseases triggered by pathogenic bacteria that lead to damage of the soft tissue and bone supporting the teeth. Amongst the identified oral periodontopathogenic bacteria, Porphyromonas gingivalis is able to enhance oral dysbiosis, which is an imbalance in the beneficial commensal and periodontal pathogenic bacteria that induces chronic inflammation. Given the critical role of oral pathogenic bacteria like P. gingivalis in the pathogenesis of periodontitis, local and/or systemic antibacterial therapy has been suggested to treat this disease, especially in its severe or refractory forms. Nevertheless, the majority of the antibacterial agents currently used for the treatment of periodontal diseases are broad-spectrum, which harms beneficial bacterial species that are critical in health, inhibit the growth of pathogenic bacteria, contribute in protecting the periodontal tissues to damage and aid in its healing. Thus, the development of more effective and specific antibacterial agents is needed to control oral pathogens in a polymicrobial environment. The strategies for the development of novel antibacterial agents include natural product isolation as well as synthetic and semi-synthetic methodologies. This review presents an overview of the periodontal diseases gingivitis and periodontitis along with current antibacterial treatment options (i.e., classes of antibacterial agents and the mechanism(s) of resistance that hinder their usage) used in periodontal diseases that specifically target oral pathogens such as P. gingivalis. In addition, to help medicinal chemists gain a better understanding of potentially promising scaffolds, this review provides an in-depth coverage of the various families of small molecules that have been investigated as potential anti-P. gingivalis agents, including novel families of compounds, repositioned drugs, as well as natural products.

Transcriptomic phases of periodontitis lesions using the nonhuman primate model.

We used a nonhuman primate model of ligature-induced periodontitis to identify patterns of gingival transcriptomic after changes demarcating phases of periodontitis lesions (initiation, progression, resolution). A total of 18 adult Macaca mulatta (12-22 years) had ligatures placed (premolar, 1st molar teeth) in all 4 quadrants. Gingival tissue samples were obtained (baseline, 2 weeks, 1 and 3 months during periodontitis and at 5 months resolution). Gene expression was analyzed by microarray [Rhesus Gene 1.0 ST Array (Affymetrix)]. Compared to baseline, a large array of genes were significantly altered at initiation (n = 6049), early progression (n = 4893), and late progression (n = 5078) of disease, with the preponderance being up-regulated. Additionally, 1918 genes were altered in expression with disease resolution, skewed towards down-regulation. Assessment of the genes demonstrated specific profiles of epithelial, bone/connective tissue, apoptosis/autophagy, metabolism, regulatory, immune, and inflammatory responses that were related to health, stages of disease, and tissues with resolved lesions. Unique transcriptomic profiles occured during the kinetics of the periodontitis lesion exacerbation and remission. We delineated phase specific gene expression profiles of the disease lesion. Detection of these gene products in gingival crevicular fluid samples from human disease may contribute to a better understanding of the biological dynamics of the disease to improve patient management.

Oral microbiome interactions with gingival gene expression patterns for apoptosis, autophagy and hypoxia pathways in progressing periodontitis.

Oral mucosal tissues must react with and respond to microbes comprising the oral microbiome ecology. This study examined the interaction of the microbiome with transcriptomic footprints of apoptosis, autophagy and hypoxia pathways during periodontitis. Adult Macaca mulatta (n = 18; 12-23 years of age) exhibiting a healthy periodontium at baseline were used to induce progressing periodontitis through ligature placement around premolar/molar teeth. Gingival tissue samples collected at baseline, 0·5, 1 and 3 months of disease and at 5 months for disease resolution were analysed via microarray. Bacterial samples were collected at identical sites to the host tissues and analysed using MiSeq. Significant changes in apoptosis and hypoxia gene expression occurred with initiation of disease, while autophagy gene changes generally emerged later in disease progression samples. These interlinked pathways contributing to cellular homeostasis showed significant correlations between altered gene expression profiles in apoptosis, autophagy and hypoxia with groups of genes correlated in different directions across health and disease samples. Bacterial complexes were identified that correlated significantly with profiles of host genes in health, disease and resolution for each pathway. These relationships were more robust in health and resolution samples, with less bacterial complex diversity during disease. Using these pathways as cellular responses to stress in the local periodontal environment, the data are consistent with the concept of dysbiosis at the functional genomics level. It appears that the same bacteria in a healthy microbiome may be interfacing with host cells differently than in a disease lesion site and contributing to the tissue destructive processes.

Oral Microbiome and Gingival Gene Expression of Inflammatory Biomolecules With Aging and Periodontitis.

Although data describe the presence and increase of inflammatory mediators in the local environment in periodontitis vs. health in humans, details regarding how these responses evolve in the transition from health to disease, changes during disease progression, and features of a resolved lesion remain unknown. This study used a nonhuman primate model of ligature-induced periodontitis in young, adolescent, adult, and aged animals to document features of inflammatory response affected by age. Rhesus monkeys had ligatures tied and provided gingival tissue biopsy specimens at baseline, 0.5, 1, and 3 months of disease and at 5 months of the study, which was 2 months post-ligature removal for clinically resolved tissues. The transcriptome was assessed using microarrays for chemokine (n = 41), cytokine (n = 45), chemokine receptor (n = 21), cytokine receptor (n = 37), and lipid mediator (n = 31) genes. Limited differences were noted in healthy tissues for chemokine expression with age; however, chemokine receptor genes were decreased in young but elevated in aged samples. IL1A, IL36A, and IL36G cytokines were decreased in the younger groups, with IL36A elevated in aged animals. IL10RA/IL10RB cytokine receptors were altered with age. Striking variation in the lipid mediator genes in health was observed with nearly 60% of these genes altered with age. A specific repertoire of chemokine and chemokine receptor genes was affected by the disease process, predominated by changes during disease initiation. Cytokine/cytokine receptor genes were also elevated with disease initiation, albeit IL36B, IL36G, and IL36RN were all significantly decreased throughout disease and resolution. Significant changes were observed in similar lipid mediator genes with disease and resolution across the age groups. Examination of the microbiome links to the inflammatory genes demonstrated that specific microbes, including Fusobacterium, P. gingivalis, F. alocis, Pasteurellaceae, and Prevotella are most frequently significantly correlated. These correlations were generally positive in older animals and negative in younger specimens. Gene expression and microbiome patterns from baseline were distinctly different from disease and resolution. These results demonstrate patterns of inflammatory gene expression throughout the phases of the induction of a periodontal disease lesion. The patterns show a very different relationship to specific members of the oral microbiome in younger compared with older animals.

Gingival tissue autophagy pathway gene expression profiles in periodontitis and aging.

OBJECTIVE: We hypothesized that autophagy-related genes will be differentially expressed in periodontitis, suggesting an impaired gingival autophagic response associated with disease. BACKGROUND: Autophagy is a cellular physiologic mechanism to maintain tissue homeostasis, while deficient autophagic responses increase inflammation and susceptibility to infection. METHODS: Rhesus monkeys [<3 years to 23 years of age (n = 34)] were examined for periodontal health and naturally occurring periodontitis. Gingival tissues samples were obtained from healthy or diseased sites, total RNA was isolated, and the Rhesus Gene Chip 1.0 ST (Affymetrix) was used for gene expression analysis of 150 autophagy-related genes. RESULTS: Comparison of expression levels with adult healthy tissues demonstrated a rather limited number of individual genes that were significantly different across the age-groups. In contrast, with periodontitis in the adults and aged animals, about 15% of the genes were significantly increased or decreased. The differences were reflected in the mTOR complex (5/12), ULK1/ATG1 complex (5/9), PI3K complex (5/21), ATG9 complex (2/7), ATG12 conjugation/LC3 lipidation (7/22), and lysosome fusion/vesicle degradation [LF/VD (5/10)] activities within the broader autophagic pathway. The genes most greatly altered in gingival tissues of naturally occurring periodontitis were identified in the ATG12 and LF/VD pathways that approximated 50% of the genes in each of those categories. While healthy gingival aging did not appear to reflect altered autophagy gene expression, substantial differences were noted with periodontitis irrespective of the age of the animals. Future studies into the role of autophagy in periodontitis and could offer potential new therapeutic strategies to prevent and/or treat periodontal disease.

Oral Microbiome and Gingival Tissue Apoptosis and Autophagy Transcriptomics.

Objective: This study focused on documenting characteristics of the gingival transcriptome during various stages of periodontitis targeting genes associated with apoptotic and autophagic pathways and changes that specifically associate with features of the oral microbiome. Methods:Macaca mulatta (n = 18; 12-23 years) were examined at baseline and 0.5, 1, and 3 months of disease progression, as well as 5 months with clinical disease resolution. 16S sequencing and microarray analyses examined changes in the microbiome and gingival transcriptome, respectively, at each time point from every animal. Results: Specific patterns of apoptotic and autophagic genes were identified related to the initiation and progression of disease. The analysis also provided insights on the principal bacteria within the complex microbiome whose abundance was significantly correlated with differences in apoptotic and autophagic gene expression. Bacteria were identified that formed associated complexes with similar effects on the host gene expression profiles. A complex of Leptotrichia_unclassifed, Capnocytophaga_unclassified, Prevotella sp. 317, and Veillonellaceae_[G-1] sp. 155 were significantly negatively correlated with both apoptosis and autophagy. Whereas, Veillonellaceae_[G-1], Porphyromonadaceae, and F. alocis 539 were significantly positively correlated with both pathways, albeit this relationship was primarily associated with pro-apoptotic genes. Conclusions: The findings provide evidence for specific bacteria/bacterial complexes within the oral microbiome that appear to have a more substantive effect on regulating apoptotic and autophagic pathways in the gingival tissues with periodontitis.

Second Generation of Zafirlukast Derivatives with Improved Activity against the Oral Pathogen Porphyromonas gingivalis.

Porphyromonas gingivalis is a Gram-negative anaerobic pathogen that can trigger oral dysbiosis as an early event in the pathogenesis of periodontal disease. The FDA-approved drug zafirlukast (ZAF) was recently shown to display antibacterial activity against P. gingivalis. Here, 15 novel ZAF derivatives were synthesized and evaluated for their antibacterial activity against P. gingivalis and for their cytotoxic effects. Most derivatives displayed superior antibacterial activity against P. gingivalis compared to ZAF and its first generation derivatives along with little to no growth inhibition of other oral bacterial species. The most active compounds displayed bactericidal activity against P. gingivalis and less cytotoxicity than ZAF. The superior and selective antibacterial activity of ZAF derivatives against P. gingivalis along with an increased safety profile compared to ZAF suggest these new compounds, especially 14b and 14e, show promise as antibacterials for future studies aimed to test their potential for preventing/treating P. gingivalis-induced periodontal disease.

Oral commensal bacteria differentially modulate epithelial cell death.

OBJECTIVE: Epithelial cell death is an important innate mechanism at mucosal surfaces, which enables the elimination of pathogens and modulates immunoinflammatory responses. Based on the antimicrobial and anti-inflammatory properties of cell death, we hypothesized that oral epithelial cell (OECs) death is differentially modulated by oral bacteria. MATERIAL AND METHODS: We evaluated the effect of oral commensals Streptococcus gordonii (Sg), Streptococcus sanguinis (Ss), and Veillonella parvula (Vp), and pathogens Porphyromonas gingivalis (Pg), Tannerella forsythia (Tf), and Fusobacterium nucleatum (Fn) on OEC death. Apoptosis and necrosis were evaluated by flow cytometry using FITC Annexin-V and Propidium Iodide staining. Caspase-3/7 and caspase-1 activities were determined as markers of apoptosis and pyroptosis, respectively. IL-1ß and IL-8 protein levels were determined in supernatants by ELISA. RESULTS: Significant increases in apoptosis and necrosis were induced by Sg and Ss. Pg also induced apoptosis, although at a substantially lower level than the commensals. Vp, Tf, and Fn showed negligible effects on cell viability. These results were consistent with Sg, Ss, and Pg activating caspase-3/7. Only Ss significantly increased the levels of activated caspase-1, which correlated to IL-1ß over-expression. CONCLUSIONS: OEC death processes were differentially induced by oral commensal and pathogenic bacteria, with Sg and Ss being more pro-apoptotic and pro-pyroptotic than pathogenic bacteria. Oral commensal-induced cell death may be a physiological mechanism to manage the extent of bacterial colonization of the outer layers of mucosal epithelial surfaces. Dysbiosis-related reduction or elimination of pro-apoptotic oral bacterial species could contribute to the risk for persistent inflammation and tissue destruction.

A Potential Role of Phospholipase 2 Group IIA (PLA2-IIA) in P. gingivalis-Induced Oral Dysbiosis.

Porphyromonas gingivalis is an oral pathogen with the ability to induce oral dysbiosis and periodontal disease. Nevertheless, the mechanisms by which P. gingivalis could abrogate the host-microbe symbiotic relationship leading to oral dysbiosis remain unclear. We have recently demonstrated that P. gingivalis specifically increased the antimicrobial properties of oral epithelial cells, through a strong induction of the expression of PLA2-IIA in a mechanism that involves activation of the Notch-1 receptor. Moreover, gingival expression of PLA2-IIA was significantly increased during initiation and progression of periodontal disease in non-human primates and interestingly, those PLA2-IIA expression changes were concurrent with oral dysbiosis. In this chapter, we present an innovative hypothesis of a potential mechanism involved in P. gingivalis-induced oral dysbiosis and inflammation based on our previous observations and a robust body of literature that supports the antimicrobial and proinflammatory properties of PLA2-IIA as well as its role in other chronic inflammatory diseases.

Novel zafirlukast derivatives exhibit selective antibacterial activity against Porphyromonas gingivalis.

Periodontal disease is an oral chronic immune-inflammatory disease highly prevalent worldwide that is initiated by specific oral bacterial species leading to local and systemic effects. The development of new preventive/therapeutic strategies to specifically target oral periodontopathogens without perturbing oral microbiome species normally colonizing the oral cavity is needed. The fast and affordable strategy of repositioning of already FDA-approved drugs can be an answer to the development of novel treatments against periodontal pathogens such as Porphyromonas gingivalis. Herein, we report the synthesis and antibacterial activity of novel zafirlukast derivatives, their bactericidal effect, and their cytotoxicity against oral epithelial cell lines. Many of these derivatives exhibited superior antibacterial activity against P. gingivalis compared to the parent drug zafirlukast. The most promising compounds were found to be selective against P. gingivalis and they were bactericidal in their activity. Finally, we demonstrated that these potent derivatives of zafirlukast provided a better safety profile against oral epithelial cells compared to zafirlukast.