Investigation of Bacterial Contents From Persistent Endodontic Infection and Evaluation of Their Inflammatory Potential

Abstract

This clinical study investigated and quantified cultivable bacteria and their levels of endotoxins in persistent endodontic infection, determining their antigenicity against macrophages and fibroblast cells by IL-1β and TNF-α secretion and evaluating their relationship with clinical and radiographic features. Samples from the root canals were obtained after root filling removal. Culture techniques were used to determine the bacterial count and the endotoxins were determined by LAL-assay. PCR analysis (16S rDNA) was used for bacterial detection. Raw 264.5 macrophages and V79 fibroblast were stimulated with endodontic contents. ELISA assay measured the amounts of IL-1ß/TNF-?#61537; secretion. Bacteria and endotoxin medians were 1.24x105 CFU/mL and 9.62 EU/mL, respectively. Porphyromonas endodontalis was the most frequently detected species. Higher levels of endotoxins were found in teeth with pain on palpation (23.56 EU/mL) rather than in its absence (8.21 EU/mL). Larger areas of bone destruction were related to higher levels of endotoxins and IL-1β and TNF-α secretion. The study findings revealed the presence of Gram-negative bacteria species in persistent endodontic infection, with their endotoxins related to both severity of bone destruction and development of symptomatology. Moreover, larger areas of bone destruction were related to higher levels of IL-1β and TNF-α secreted by macrophages and fibroblast cells.

Etiologic role of root canal infection in apical periodontitis and its relationship with clinical symptomatology

“…18 LPS concentrations found in infected root canals seem to promote a direct sensitization of receptors that activate the pain mechanism associated with bacterial infections, also sensitizing trigeminal sensory neurons. 131,132 The presence of endotoxin has been reported in samples taken from vital pulp, 13 irreversible pulpitis, 13 necrotic pulp, 11,12,13,15,16,17,20,25,28,29,30,113,131,133,134,135,136,137,138,139 root canals of teeth with endodontic failure, 19,20,26,132 and from root canals of teeth associated with periodontal disease. 140 Table 3 shows some of the endotoxin levels reported in initial samples collected from the root canals.…”

“…17 Macrophages are stimulated predominantly by the bacterial endotoxin. 12,132 Lymphocytes express different sets of inflammatory cells, proinflammatory and immunoregulatory cytokines and chemokines, which are considered important mediators in periapical immune response to infection. 132,143 Among the various host degradative pathways, considerable interest has been focused on the study of cytokines, not only because of their role in regulating the humoral immune system and in cellular responses against invasive bacteria, but also as mediators of periapical tissue destruction.…”

“…12,132 Lymphocytes express different sets of inflammatory cells, proinflammatory and immunoregulatory cytokines and chemokines, which are considered important mediators in periapical immune response to infection. 132,143 Among the various host degradative pathways, considerable interest has been focused on the study of cytokines, not only because of their role in regulating the humoral immune system and in cellular responses against invasive bacteria, but also as mediators of periapical tissue destruction. 145 Inflammatory cytokines produced from host cells (e.g., monocytes/ macrophages) reflect root canal conditions and determine the local immune process within the periapical environment.…”

Correlation Between Volume of Root Canal, Cultivable Bacteria, Bacterial Complexes and Endotoxins in Primary Infection

“…Additionally, the linear regression analysis has revealed a positive correlation between root canal volume and CFU count found in PEI with apical periodontitis. Over the years, clinical studies (14,20,21) have correlated the size of periapical lesion with the bacterial load, but no study established a correlation between root canal volume and bacterial load encountered in root canal infections.…”

“…It has long been known that endotoxins released from Gram-negative bacteria in infected root canals can contribute to increasing vasoactive and neurotransmitter substances at the nerve endings in inflamed periapical lesions (11). This study quantified the levels of endotoxins in root canal infections by using LAL-assay, which has been widely used in endodontics (21). Among the different LAL tests (25), the kinetic chromogenic limulus amebocyte assay was used in this study to evaluate the levels of endotoxins in our samples.…”