Polymorphisms of pro-inflammatory cytokine genes and the risk for acute suppurative or chronic nonsuppurative apical periodontitis in a Colombian population.

AIM: To determine the association of functional single nucleotide polymorphisms in genes of the pro-inflammatory cytokines tumour necrosis factor-α, interleukin-1ß, interleukin-8 and interleukin-12B with the development of two clinical forms of apical periodontitis (AP): acute suppurative and chronic nonsuppurative. METHODOLOGY: The study included 120 patients from Bucaramanga City, Colombia, 63 diagnosed with acute suppurative AP (ASAP) and 57 diagnosed with chronic nonsuppurative AP (CNAP). Genotyping for IL1B +3954 (rs1143634), IL8 / CXCL8 -251 (rs4073), IL12B +1188 (rs3212227) and TNFA -308 (rs1800629) was performed by the PCR-restriction fragment length polymorphisms method. The statistical analysis was performed using STATA 10.0 and PLINK V1.07 software. RESULTS: Significant differences in the distribution of IL8 / CXCL8 -251 A allele (P adjusted = 0.041; OR adjusted = 0.41, CI adjusted = 0.31-0.97) and IL8 / CXCL -251 TT genotype (P adjusted = 0.04; OR adjusted = 2.24, CI adjusted = 1.04-4.84) were observed comparing patients diagnosed with ASAP and CNAP. No association was observed in genotype and allele distribution for other genetic polymorphisms analysed. CONCLUSION: This study provides molecular epidemiological evidence that suggests in the present cohort that IL8 / CXCL8 -251 T allele, which is associated with higher production of IL8/CXCL8, is also associated with a higher risk of developing acute suppurative form of AP, whereas IL8 / CXCL8 -251 A allele, which is associated with lower production of IL8/CXCL8, is associated with chronic nonsuppurative form of AP. This suggests a pivotal role for IL-8/CXCL8 in periapical disease because of its ability to induce chemotaxis and modulating the directed migration of neutrophils to the site of inflammation in response to microbial infection of pulp.

Comparative Metabolomics Reveals the Microenvironment of Common T-Helper Cells and Differential Immune Cells Linked to Unique Periapical Lesions.

Periapical abscesses, radicular cysts, and periapical granulomas are the most frequently identified pathological lesions in the alveolar bone. While little is known about the initiation and progression of these conditions, the metabolic environment and the related immunological behaviors were examined for the first time to model the development of each pathological condition. Metabolites were extracted from each lesion and profiled using gas chromatography-mass spectrometry in comparison with healthy pulp tissue. The metabolites were clustered and linked to their related immune cell fractions. Clusters I and J in the periapical abscess upregulated the expression of MMP-9, IL-8, CYP4F3, and VEGF, while clusters L and M were related to lipophagy and apoptosis in radicular cyst, and cluster P in periapical granuloma, which contains L-(+)-lactic acid and ethylene glycol, was related to granuloma formation. Oleic acid, 17-octadecynoic acid, 1-nonadecene, and L-(+)-lactic acid were significantly the highest unique metabolites in healthy pulp tissue, periapical abscess, radicular cyst, and periapical granuloma, respectively. The correlated enriched metabolic pathways were identified, and the related active genes were predicted. Glutamatergic synapse (16-20),-hydroxyeicosatetraenoic acids, lipophagy, and retinoid X receptor coupled with vitamin D receptor were the most significantly enriched pathways in healthy control, abscess, cyst, and granuloma, respectively. Compared with the healthy control, significant upregulation in the gene expression of CYP4F3, VEGF, IL-8, TLR2 (P < 0.0001), and MMP-9 (P < 0.001) was found in the abscesses. While IL-12A was significantly upregulated in cysts (P < 0.01), IL-17A represents the highest significantly upregulated gene in granulomas (P < 0.0001). From the predicted active genes, CIBERSORT suggested the presence of natural killer cells, dendritic cells, pro-inflammatory M1 macrophages, and anti-inflammatory M2 macrophages in different proportions. In addition, the single nucleotide polymorphisms related to IL-10, IL-12A, and IL-17D genes were shown to be associated with periapical lesions and other oral lesions. Collectively, the unique metabolism and related immune response shape up an environment that initiates and maintains the existence and progression of these oral lesions, suggesting an important role in diagnosis and effective targeted therapy.

Host and microbiological factors related to dental caries development.

Studies on dental caries suggest that in severe cases it may induce a systemic immune response. This occurs particularly when caries progresses into pulpal inflammation and results in abscess or fistula formation (AFF). We hypothesized that severe dental caries will affect the general health of children. The acute phase proteins alpha-1-acid glycoprotein (AGP), C-reactive protein (CRP) and the cytokine neopterin were chosen as parameters to monitor general health. Also, a polymorphism in the bacterial ligand CD14 (-260) was studied to investigate the relationship between genotype sensitivity for bacterial infections and AFF. In Suriname, children aged 6 years were recruited and enrolled into a dental care scheme, randomly assigned to 4 groups with different treatment strategies and monitored longitudinally. 348 children were included in the present study. Blood and saliva samples were taken at baseline and 1 year, and concentrations of serum AGP, CRP, neopterin, salivary Streptococcus mutans and CD14-260 C>T polymorphism were determined. There was no significant association between different treatment strategies and the serum parameters. Binary logistic regression analyses revealed a significant association between AFF as the outcome variable and the CD14 genotype and the concentrations of CRP and of neopterin as factors (p < 0.05). A significant negative association was found between the CD14-260 TT and AFF (p = 0.035, OR = 3.3) for the whole population. For children who had 4 or more carious lesions at baseline, the significance increased (p = 0.005, OR = 4.8), suggesting that the CD14-260 TT genotype was protective for AFF as a consequence of dental caries.

Histobacteriologic Conditions of the Apical Root Canal System and Periapical Tissues in Teeth Associated with Sinus Tracts.

INTRODUCTION: This histobacteriologic study described the pattern of intraradicular and extraradicular infections in teeth with sinus tracts and chronic apical abscesses. METHODS: The material comprised biopsy specimens from 24 (8 untreated and 16 treated) roots of teeth associated with apical periodontitis and a sinus tract. Specimens were obtained by periradicular surgery or extraction and were processed for histobacteriologic and histopathologic methods. RESULTS: Bacteria were found in the apical root canal system of all specimens, in the main root canal (22 teeth) and within ramifications (17 teeth). Four cases showed no extraradicular infection. Extraradicular bacteria occurred as a biofilm attached to the outer root surface in 17 teeth (5 untreated and 12 treated teeth), as actinomycotic colonies in 2 lesions, and as planktonic cells in 2 lesions. Extraradicular calculus formation (mineralized biofilm) was evident in 10 teeth. CONCLUSIONS: Teeth with chronic apical abscesses and sinus tracts showed a very complex infectious pattern in the apical root canal system and periapical lesion, with a predominance of biofilms.

Update on the adaptive immune responses of the dental pulp.

Recent advances in immunology have disclosed the enormous complexity of the immune regulatory system. The dental pulp is equipped to mount adaptive immune responses to caries, which include at least antigen-presenting cells, lymphocytes, mast cells and their cytokines, and chemokines. The purpose of this review is to summarize our current understanding of the roles of these cellular and molecular components in the irreversibly inflamed pulp. The immunopathology of abscess formation and the mechanisms for painless pulpitis are also discussed.

Human Exoproteome in Acute Apical Abscesses.

INTRODUCTION: An acute apical abscess is a severe response of the host to massive invasion of the periapical tissues by bacteria from infected root canals. Although many studies have investigated the microbiota involved in the process, information on the host factors released during abscess formation is scarce. The purpose of this study was to describe the human exoproteome in samples from acute apical abscesses. METHODS: Fourteen pus samples were obtained by aspiration from patients with an acute apical abscess. Samples were subjected to protein digestion, and the tryptic peptides were analyzed using a mass spectrometer and ion trap instrument. The human proteins identified in this analysis were classified into different functional categories. RESULTS: A total of 303 proteins were identified. Most of these proteins were involved in cellular and metabolic processes. Immune system proteins were also very frequent and included immunoglobulins, S100 proteins, complement proteins, and heat shock proteins. Polymorphonuclear neutrophil proteins were also commonly detected, including myeloperoxidases, defensins, elastases, and gelatinases. Iron-sequestering proteins including transferrin and lactoferrin/lactotransferrin were found in many samples. CONCLUSIONS: The human exoproteome included a wide variety of proteins related to cellular processes, metabolism, and immune response. Proteins involved in different mechanisms against infection, tissue damage, and protection against tissue damage were identified. Knowledge of the presence and function of these proteins using proteomics provides an insight into the complex host-pathogen relationship, the host antimicrobial strategies to fight infections, and the disease pathogenesis.

Phagocytosis of bacterial strains isolated from acute dentoalveolar abscess.

The phagocytosis by human polymorphonuclear leucocytes of 37 bacterial strains identified as Streptococcus milleri (10 strains), strictly anaerobic gram-positive cocci (10) Prevotella intermedia (6), Pr. oralis (5) and Fusobacterium nucleatum (6) was investigated in vitro. The ingestion of S. milleri and strictly anaerobic gram-positive cocci was significantly greater (p < 0.001) than that of strains of Prevotella spp. and F. nucleatum. The degree of uptake of capsulate and non-capsulate strains did not differ.

Ultrastructural analysis of MHC class II molecule-expressing cells in experimentally induced periapical lesions in the rat.

Periapical lesions were induced by making 28 days of unsealed pulp exposures in the lower first molars of Wistar rats. Major histocompatibility complex class II molecule-expressing cells were then demonstrated by means of immunoperoxidase staining using a monoclonal antibody OX6, and the ultrastructure of these cells was analyzed under electron microscopy. OX6+ cells were classified into two major populations, (i.e. macrophages and dendritic cell (DC)-like cells. DC-like cells had elongated cytoplasmic processes, contained a few lysosomal structures, lacked distinct phagosomes, and were the most predominant cell type in the established lesion. Some of lymphocytes and plasma cells also showed a positive immunoreactivity. Both OX6+ macrophages and DC-like cells often showed a cell-to-cell attachment with lymphocytes. These findings suggested that major histocompatibility complex class 11 molecule-expressing macrophages and DC-like cells may play a crucial role in periapical lesion development by acting as antigen-presenting cells to memory T lymphocytes.

Interleukin-1 alpha and interleukin-1 beta periapical exudates of infected root canals: correlations with the clinical findings of the involved teeth.

This study quantified the concentrations of interleukin-1 alpha (IL-1 alpha) and interleukin-1 beta (IL-1 beta) in the periapical exudates obtained from 69 single-rooted teeth using enzyme-linked immunosorbent assays, and examined their correlation with clinical findings of the involved teeth. Changes in the levels of these factors during root canal treatment were also investigated. The average levels of IL-1 beta (6.57 ng/ml) in periapical exudates were twice that of IL-1 alpha (3.25 ng/ml). The exudates containing pus showed significantly higher IL-1 alpha levels than those from the canals without pus (p < 0.01). The exudates from the canals with small radiolucent areas contained significantly higher IL-1 alpha levels than those from the canals with large radiolucent areas (p < 0.05). The tendency for there to be an increase in the levels of IL-1 alpha and a decrease in the levels of IL-1 beta was observed following root canal treatment. These observations suggest that IL-1 alpha and IL-1 beta are involved in the immunopathogenesis of periapical lesions and that IL-1 alpha and IL-1 beta may play different roles in the healing process of periapical lesions during root canal treatment.

Balance of interleukin-1 beta and interleukin-1 receptor antagonist in human periapical lesions.

Interleukin-1 beta (IL-1 beta) has been considered as a major potent mediator of bone resorption and implicated in the development of human periapical lesions. Among naturally occurring interleukin-1 (IL-1) inhibitors, IL-1 receptor antagonist (IL-1ra) is a 22 kDa protein that shares homology with IL-1 beta and IL-1 alpha, binds to IL-1 receptor with similar affinity to IL-1, and has no known agonist properties. In this study, we measured the periapical exudate (PE) levels of IL-1 beta and IL-1ra from human periapical lesions. PE samples were collected from root canals during routine endodontic treatments, and the enzyme-linked immunosorbent assay was used to measure PE-IL-1 beta and IL-1ra. Detectable levels of both IL-1 beta and IL-1ra were found in 25 of 29 clinical samples. Relatively high levels of IL-1ra compared with IL-1 beta (mean IL-1ra:IL-1 beta ratio = 128:7; range: 0.9 to 495.4), and significantly positive correlation between IL-1ra and IL-1 beta levels was found. The PE-IL-1ra:IL-1 beta ratios obtained from symptomatic lesions were significantly lower than those from asymptomatic lesions. These results suggest that IL-1ra-mediated IL-1 antagonism occurred to block locally produced IL-1 activity, and the balance of IL-1 to IL-1ra production may be crucial in the development of periapical lesions.

Immunohistochemical detection of prostaglandins E2, F2 alpha, and 6-keto-prostaglandin F1 alpha in experimentally induced periapical inflammatory lesions in rats.

The immunohistochemical localization of prostaglandin (PG) E2, PGF2 alpha, and 6-keto-PGF1 alpha (a stable metabolite of PGI2) was demonstrated in rat periapical inflammatory lesions induced by opening the pulp chamber. Two wk postoperatively, suppurative periapical lesions were formed, and active bone resorption was seen surrounding these lesions. Immunohistochemical examination showed that macrophages infiltrating in inflammatory tissue were positively stained for the examined PGs. In some lesions, wherein acute inflammatory changes subsided and proliferation of fibroblasts started, the fibroblasts were positively stained for 6-keto-PGF1 alpha. Osteocytes and osteoblasts were also positive for 6-keto-PGF1 alpha not only in experimental animals, but also in untreated animals. However the staining intensity of the PG in these cells was higher in periapical lesions than in normal condition. These findings suggested that the cellular sources of the PGs in the periapical lesions are mainly macrophages and fibroblasts, and that the PGs produced by these cells, and possibly osteoblast and osteocytes, may contribute to the osteolytic resorption of periapical lesions.

Host mediators in endodontic exudates. I. Indicators of inflammation and humoral immunity.

Exudate is often found in the root canal when entering the chamber and canal of teeth with periapical lesions. The aim of this study was to determine possible relationships between clinical or radiographic findings and the concentrations of different host mediators in endodontic exudates. Thirty-two nonvital teeth with periapical symptoms were included in the study. A Clinical Periapical Index was developed to quantify clinical findings. Endodontic exudates were collected with methylcellulose filter paper strips every 3 min, after opening of the pulp chamber. The concentrations of the lysosomal acid glycohydrolase beta-glucuronidase, IgG, IgA, IgM, and interleukin-1 beta in the endodontic exudates were analyzed. The results demonstrated that exudates collected from teeth with suppuration (cloudy exudates), and teeth with higher periapical index scores (Orstavik et al., 1986) contained higher concentrations of beta-glucuronidase and interleukin-1 beta. Furthermore, when the periapical index indicated severe involvement, higher IgG was observed in the first samples. The exudates from patients who presented with a sinus tract or swelling contained higher concentrations of IgM, compared with the patients with only periapical sensitivity. Data showed that endodontic exudates from patient with endodontic lesions can be analyzed for host mediators, and differences in the mediators were seen with different clinical and radiographic symptoms.

Enhanced expression of activation-associated molecules on macrophages of heterogeneous populations in expanding periapical lesions in rat molars.

Exudative macrophages are the most prevalent inflammatory cells during the entire pathogenetic process in experimentally induced rat periapical lesions. To clarify the significance of macrophages in the pathogenesis of periapical lesions, the way in which the phenotype of ED1 (a general marker for mononuclear phagocytes)-positive cells is modulated in actively expanding lesions was investigated, by immunoperoxidase staining with a panel of antibodies that recognize several activation-associated molecules on macrophages. Periapical lesions were induced experimentally by exposing the pulp in the lower first molars of Wistar rats. Active lesion expansion with morphological diversification of ED1-positive cells occurred between 14 and 28 days after the injury. Double immunoperoxidase staining revealed that ED1-positive cells coexpressing class II molecules of the major histocompatibility complex (MHC) molecules, inducible nitric oxide synthase (iNOS) and/or CD11a increased during the period of active lesion expansion. Increases of endothelial cells expressing intracellular adhesion molecule-1 and CD25 (interleukin-2 receptor)-expressing lymphocytes were also seen during the same period. Moreover, there existed two particular subpopulations of ED1 + cells in the established lesion at 28 days: (1) ED1++/class II MHC - /iNOS+ cells, located around the periapical abscess, and (2) ED1+/class II MHC+/ iNOS- cells with slender or dendritic morphology, distributed predominantly in the outer portion of the lesion where T lymphocytes were abundant. The first cell type could be a macrophage with potent phagocytic and antimicrobial actions, and the second might possess sufficient antigen-presenting capacity to cause the activation of T lymphocytes. It was concluded that macrophages, when activated, may participate in triggering lesion expansion. Functionally distinct subpopulations of macrophages may occupy different sites within the lesion where they can most effectively exert their specific functions.

Macrophage cell activation with acute apical abscess contents determined by interleukin-1 Beta and tumor necrosis factor alpha production.

INTRODUCTION: This clinical study has investigated the antigenic activity of bacterial contents from exudates of acute apical abscesses (AAAs) and their paired root canal contents regarding the stimulation capacity by levels of interleukin (IL)-1 beta and tumor necrosis factor alpha (TNF-α) throughout the root canal treatment against macrophage cells. METHODS: Paired samples of infected root canals and exudates of AAAs were collected from 10 subjects. Endodontic contents were sampled before (root canal sample [RCS] 1) and after chemomechanical preparation (RCS2) and after 30 days of intracanal medication with calcium hydroxide + chlorhexidine gel (Ca[OH]2 + CHX gel) (RCS3). Polymerase chain reaction (16S rDNA) was used for detection of the target bacteria, whereas limulus amebocyte lysate was used to measure endotoxin levels. Raw 264.7 macrophages were stimulated with AAA exudates from endodontic contents sampled in different moments of root canal treatment. Enzyme-linked immunosorbent assays were used to measure the levels of TNF-α and IL-1 beta. RESULTS: Parvimonas micra, Porphyromonas endodontalis, Dialister pneumosintes, and Prevotella nigrescens were the most frequently detected species. Higher levels of endotoxins were found in samples from periapical exudates at RCS1 (P < .005). In fact, samples collected from periapical exudates showed a higher stimulation capacity at RCS1 (P < .05). A positive correlation was found between endotoxins from exudates with IL-1 beta (r = 0.97) and TNF-α (r = 0.88) production (P < .01). The significant reduction of endotoxins and bacterial species achieved by chemomechanical procedures (RCS2) resulted in a lower capacity of root canal contents to stimulate the cells compared with that at RCS1 (P < .05). The use of Ca(OH)2 + CHX gel as an intracanal medication (RCS3) improved the removal of endotoxins and bacteria from infected root canals (P < .05) whose contents induced a lower stimulation capacity against macrophages cells at RCS1, RCS2, and RCS3 (P < .05). CONCLUSIONS: AAA exudates showed higher levels of endotoxins and showed a greater capacity of macrophage stimulation than the paired root canal samples. Moreover, the use of intracanal medication improved the removal of bacteria and endotoxins from infected root canals, which may have resulted in the reduction of the inflammatory potential of the root canal content.

Unrecognized odontogenic maxillary sinusitis: a cause of endoscopic sinus surgery failure.

BACKGROUND: Endoscopic sinus surgery (ESS) is reported to improve symptoms in approximately 85% of patients. Reasons for failure include misdiagnosis, technical inadequacies, underlying severe hyperplastic disease, biofilm, and immunodeficiency. Only one previous case of unrecognized odontogenic maxillary sinusitis has been cited in the literature as a reason for failure to improve with sinus surgery. This study was designed to characterize clinical and radiographic findings in patients who fail to improve with ESS because of an unrecognized dental etiology. METHODS: Five patients, with odontogenic maxillary sinusitis with prior unsuccessful ESS, were prospectively enrolled. Demographics and clinical aspects including duration of illness, prior sinus surgeries and therapies, and radiographic data were assessed. RESULTS: Five adults underwent an average of 2.8 sinus surgeries with persistence of disease and symptoms until their dental infection was treated. Duration of symptoms ranged from 3 to 15 years. In four of five patients, the periapical abscess was not noted on the original CT report but could be seen in retrospect. Three of five patients had been seen by their dentists and told they had no dental pathology. All five patients underwent dental extractions and one patient underwent an additional ESS after dental extraction. These procedures led to a resolution of sinusitis symptoms in all five patients. CONCLUSION: Unrecognized periapical abscess is a cause of ESS failure and the radiological report frequently will fail to note the periapical infection. Dentists are unable to recognize periapical abscesses reliably with dental x-rays and exam. In patients with maxillary sinus disease, the teeth should be specifically examined as part of the radiological workup.