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Neuropeptide K-like immunoreactivity in human dental pulp.

Nerve fibres displaying such immunoreactivity were revealed by indirect immunofluorescence. Neuropeptide K-like immunoreactive fibres, entering the pulp within large nerve trunks, were distributed around blood vessels as well as in the stroma. Some immunoreactive fibres were also observed in the para-odontoblastic region. In view of the biological activity of neuropeptide K, it is tentatively proposed that it may act in the dental pulp as a regulatory peptide involved in neurogenic inflammation, blood flow regulation and sensory transmission.

Blood Vessels↗

A Novel piRNA-Mediated Epigenetic Axis: piR-36241 Exacerbates Pulpitis by Silencing the Protective Receptor ADGRG2 in Human Dental Pulp Stem Cells.

BACKGROUND: Pulpitis, a prevalent inflammatory dental disease, primarily results from bacterial infection, yet its epigenetic regulatory mechanisms remain poorly understood. PIWI-interacting RNAs (piRNAs), a class of small non-coding RNAs known to maintain genomic stability, have not been investigated in the context of pulpitis. OBJECTIVE: This study aimed to profile piRNA expression patterns in pulpitis and to elucidate the functional role and underlying mechanism of a specific piRNA, piR-36241 and its target gene ADGRG2 in disease progression. METHODS: piRNA and mRNA sequencing were performed on pulp tissues from patients with irreversible pulpitis and healthy controls. Potential piRNA targets were predicted bioinformatically. The interaction between piR-36241 and the 3'UTR of ADGRG2, an orphan adhesion G protein-coupled receptor (GPCR), was validated using dual-luciferase reporter assay, quantitative Real-Time PCR (qRT-PCR), fluorescence in situ hybridization (FISH), Western blot (WB), hematoxylin-eosin staining (HE), immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA). Functional analyses were conducted in LPS-stimulated human dental pulp cells (HDPCs) through gain-of-function and rescue experiments. RESULTS: We identified a distinct piRNA expression profile in pulpitis, with 21 piRNAs differentially expressed. piR-36241 was notably upregulated. It directly targeted the 3'UTR of ADGRG2 and suppressed its expression. ADGRG2, hypothesised to exert protective and homeostatic functions, was downregulated in pulpitis tissues and LPS-induced HDPCs. Overexpression of piR-36241 inhibited ADGRG2 expression and promoted the secretion of pro-inflammatory cytokines (IL-6, IL-8). Conversely, knockdown of ADGRG2 similarly enhanced inflammatory responses. Rescue experiments demonstrated that piR-36241 primarily regulates inflammation through ADGRG2 silencing. CONCLUSION: Our study reveals a novel pathogenic axis in pulpitis whereby upregulation of piR-36241 exacerbates inflammatory progression by repressing the protective receptor ADGRG2. These findings provide the first evidence of piRNA-mediated epigenetic regulation in pulpitis and highlight the piR-36241/ADGRG2 pathway as a potential therapeutic target for preserving pulp vitality.

Humans↗

Immunocytochemical distribution of human PMN elastase and cathepsin-G in dental pulp.

Components of primary (azurophilic) granules of polymorphonuclear leukocytes (PMNs) have been implicated as important mediators in pulpal inflammation. This anatomical study used ultracryoimmunocytochemical techniques and characterized and contrasted the subcellular distributions of human PMN elastase (PMN-E), PMN cathepsin-G (PMN-CG), and alpha-2 macroglobulin (alpha-2M) in healthy and inflamed dental pulps. Inflamed pulpal tissue sections revealed an intense distribution of PMN-E in the extracellular domain throughout the collagen matrix. PMN-E was also localized in the perinuclear cytoplasm of PMNs and distributed in a random fashion. PMN-CG was localized intensely in the intracellular granules of PMNs and observed moderately within the extracellular matrix. Healthy pulpal tissues exposed to PMN-E and PMN-CG antibodies revealed no evidence of PMN infiltration and no specific labeling. alpha-2M, a natural serum inhibitor of PMN-E and PMN-CG, was distributed in an intense fashion within the intravascular compartments of both inflamed and healthy pulpal samples. Immunogold-labeling for alpha-2M was observed in moderate amounts within the extravascular domain of inflamed pulpal samples but only in mild amounts within the same area of healthy tissues. These results suggest that PMN-E and PMN-CG are released to the extracellular matrix of irreversibly inflamed teeth, enabling them to facilitate pulpal connective tissue destruction. Conversely, moderate extravascular labeling for alpha-2M within inflamed samples suggests a physiological attempt at inhibiting the pulpal connective tissue destruction mediated by human PMN-E and PMN-CG.

Brain Stem↗

Stimulatory effect of interleukin-6 on plasminogen activator activity from human dental pulp cells.

The plasminogen activator (PA)-plasmin proteolytic system has recently received considerable attention because of its participation in a wide variety of biological activities and in pathological conditions involving tissue destruction. We examined the effects of interleukin-6 (IL-6) on PA activity and the gene expressions of tissue type (t) PA and PA inhibitor-1 (PAI-1) in human dental pulp (HDP) cells. IL-6 treatment induced significantly high PA activity in the HDP cells in a time- and dose-dependent manner, compared with nontreated controls. Western-blot analysis showed that tPA protein in the conditioned medium was stimulated by IL-6, compared with the control. The tPA and PAI-1 mRNA levels were increased in HDP cells treated with IL-6, as shown by reverse transcriptase-polymerase chain reaction. These results suggest that IL-6 stimulated PA activity through an enhancement of tPA gene expression and may be involved in extracellular matrix degradation through the stimulation of the PA-plasmin system of HDP cells.

Blotting, Western↗

Immunohistochemical localization of S-100beta in the dental pulp of the rat molar.

The present study was undertaken to reveal whether S-100alpha or S-100beta or both are present in the nerve fibers in the rat molar tooth pulp. No immunoreactivity for S-100alpha was observed in the molar pulp. In the root pulp, thick smooth-surfaced structures accompanying the blood vessel showed S-100beta-like immunoreactivity (-LI), and occasionally a very few thin beaded elements exhibited S-100beta-LI. In the coronal pulp, S-100beta-like immunoreactive (-IR) structures arborized repeatedly and extensively; they had a predominantly thick, smooth-surfaced appearance, though parts appeared thin and beaded. Numerous thin varicose S-100beta-IR structures ran through the odontoblast cell layer, and further penetrated into the predentin alongside the dentinal tubules. They could be traced for approximately 10-20 micrometers into the predentin from the pulp-predentin border. Immunoelectron microscopy revealed that the Schwann cells in the root pulp showed S-100beta-LI, and that S-100beta-LI was present in the axoplasm as well as Schwann cells in the coronal pulp. The S-100beta-IR axons were rarely surrounded by S-100beta-IR Schwann cells. In the predentin, S-100beta-IR nerve fibers terminated in a position close to the odontoblast processes. The present findings indicate that S-100beta, not S-100alpha, is present in the axon in the dental pulp and predentin as well as in the Schwann cells.

Animals↗

[Micro-plasma treatment of dental pulp in animal experiments].

The direct and indirect effect on the endodont after thermic treatment with the micro-plasma unit from EGIDE were examined in animal experiments. The EGIDE unit is considered by its producer to be an alternative for CO2 lasers. The histological evaluation showed that both the plasma-therapy of dentin and the exposed pulp lead to massive pathological changes of endodont and was inferior to traditional methods as the indirect pulpcapping.

Animals↗

IL-1 alpha and TNF-alpha expression in rat periapical lesions and dental pulp after unilateral sympathectomy.

OBJECTIVES: Apical periodontitis is an inflammatory disease characterized by bone resorption, and sympathetic nerves are known to modulate bone resorption and bone remodeling. Higher numbers of osteoclasts and larger periapical lesions have been observed after sympathectomy in rats, but the mechanisms underlying the inhibitory effect of sympathetic nerves on osteoclasts are unknown. This study aimed to test the hypothesis that sympathetic nerves inhibit the production of the bone-resorbing pro-inflammatory cytokines IL-1 alpha and TNF-alpha in rat periapical lesions. METHODS: Rats were unilaterally sympathectomized and apical lesions were induced by exposing the dental pulp of molar teeth to the oral microflora. We quantified the cytokines IL-1 alpha and TNF-alpha by enzyme-linked immunosorbent assay, and immunohistochemical analysis was done for qualitative localization. Pulp from intact incisor teeth was tested as a control. RESULTS: We showed that IL-1 alpha was increased, but not TNF-alpha, in the periapical lesions on the sympathectomized side. Both IL-1 alpha and TNF-alpha were expressed in unexposed pulp. TNF-alpha was significantly decreased in the denervated incisor pulp, whereas the level of IL-1 alpha remained unchanged. CONCLUSIONS: This study suggests that sympathetic nerves have an inhibitory effect on IL-1 alpha in periapical lesions and a stimulatory effect on TNF-alpha in the intact rat pulp.

Animals↗

Differential regulation of interleukin-6 and inducible cyclooxygenase gene expression by cytokines through prostaglandin-dependent and -independent mechanisms in human dental pulp fibroblasts.

Increased levels of interleukin-1 (IL)-1, tumor necrosis factor-alpha (TNF-alpha), IL-6, and prostaglandin E2 (PGE2) have been detected in inflamed pulp tissue. To gain further insight into the molecular pathogenesis of pulpitis, we investigated the effects of IL-1alpha or TNF-alpha and PGE2, either alone or in combination on IL-6 and cyclooxygenase (COX)-2 messenger RNA (mRNA) production in cultured human dental pulp (HDP) fibroblasts. Exposure of HDP fibroblasts to IL-1alpha or TNF-alpha resulted in elevated levels of IL-6 (approximately 3.4 to approximately 10.4-fold) and COX-2 (approximately 5 to approximately 6.2-fold) mRNA. Simultaneous addition of IL-1alpha and PGE2 or TNF-alpha and PGE2 to the cultures significantly reduced the cytokine-induced IL-6 mRNA synthesis ranging from 45% to 65%. However, indomethacin enhanced the cytokine-stimulated IL-6 mRNA synthesis by approximately 1.7 to approximately 3.4-fold. This action could be reversed by exogenous PGE2. In contrast, PGE2 or indomethacin failed to modify the stimulatory effect of IL-1alpha or TNF-alpha on COX-2 gene expression. Because excessive levels of IL-6 and prostaglandins have been connected with the pathogenesis of several inflammatory diseases, our results suggest the involvement of HDP fibroblasts in the development of pulpitis via producing IL-6 and COX-2. Furthermore, expression of IL-6 and COX-2 genes in this cell seems to be differentially regulated by cytokines through prostaglandin-dependent and -independent pathways.

Anti-Inflammatory Agents, Non-Steroidal↗

Stimulation of interleukin-6 production in human dental pulp cells by peptidoglycans from Lactobacillus casei.

Interleukin-6 (IL-6), which is a multifunctional cytokine, has an important role in acute and chronic inflammation. The peptidoglycan (PG) was purified from Lactobacillus casei, which was a Gram-positive bacteria frequently isolated from deep carious lesions and suspected to be a pathogen of pulpitis. In this study, the effects of PG on the production of IL-6 in human dental pulp cells were examined. PG stimulated IL-6 production in a time- and dose-dependent manner. Reverse transcriptase-polymerase chain reaction experiments showed that the increase was dependent on the enhancement of IL-6 mRNA levels. These findings suggest that Gram-positive bacteria, such as L. casei, from carious lesions, might be involved in developing pulpitis through the stimulation of IL-6 production.

Cells, Cultured↗

Detection of human IL-2 in normal and inflamed dental pulps.

Irreversible pulpitis has been associated with an increase in the number of pulpal T-cells. Interleukin-2 (IL-2) stimulates T-cell proliferation and signals the release of other proinflammatory mediators associated with connective tissue degradation. IL-2 has been suggested to be a useful marker of pathologic inflammatory activity in periodontal and systemic disease conditions. The purpose of this study was to analyze normal and inflamed dental pulps for the presence of immunoreactive IL-2 (iIL-2). Normal healthy pulpal tissue was obtained from 17 impacted third molars and inflamed samples were obtained from 12 symptomatic carious molars clinically diagnosed with irreversible pulpitis. Pulpal tissues were collected, prepared, and analyzed for histological status and iIL-2 concentration by a modified ELISA technique. iIL-2 was detected in all vital pulpal tissues. A t-test revealed significant differences in iIL-2 concentrations when inflamed pulpal tissues were compared to normal healthy samples (T = -2.75, p < 0.05). These results suggest that iIL-2 may serve as a marker of pathologic inflammatory activity in irreversible pulpitis.

Analysis of Variance↗

Rates of putrefaction of dental pulp in the Northwest coast environment.

Cytological stability is of interest to criminal investigators in instances where remnants of soft tissue have been preserved, since such tissue can aid in the identification of human remains, helping to determine either the sex of the individual or his or her identity. This study based on seven experiments shows that, in Northwest coast outdoor environments in both summer (three experiments) and winter (three experiments), the stability of dental pulp nuclei ranges from 4 days to 2 weeks. The seventh experiment serves to describe the morphological sequence observed in nuclear putrefaction. The specimens included human and pig extracted teeth and unextracted pig teeth. Deposition of the specimens was made both on the surface and in the subsurface (30-cm depth), and the environmental variables were recorded.

Animals↗

Bacterial contamination as a factor influencing the toxicity of materials to the exposed dental pulp.

Three materials, which had been previously found to be toxic when applied as pulp-capping agents in conventional rat molar pulps, were retested in germ-free rats. All produced much more favorable responses in the pulp, with a lack of inflammation and the presence of dentine bridges in the majority of teeth. It appears that much of the pulp damage previously attributed to the chemical toxicity of materials may be caused by the presence of bacteria. The design of studies intended to evaluate the response of the pulp to materials should include staining techniques that will detect the presence of bacteria on the floors of test cavities.

Aluminum Hydroxide↗

Dental pulp infarction in female rats following inhalation exposure to 2-butoxyethanol.

Female Fischer 344 (F344)/N rats (10 per exposure group) were exposed to 2-butoxyethanol (BE) vapors (0, 31, 62.5, 125, 250, or 500 ppm 6 h/d, 5 d/wk, for 13 weeks) to characterize its prechronic toxicity. Dental lesions consisting of bilateral multifocal dental pulp thrombosis, pulp infarction, and odontoblast infarction were noted in the maxillary incisors of 3 of 4 rats from the 500-ppm group that were sacrificed when moribund during the first week of exposure. In addition, 1 rat from the 500-ppm group that was sacrificed on day 32 had similar unilateral incisor lesions but with additional findings consistent with a unilateral maxillary incisor fracture. In contrast, rats sacrificed after 13 weeks of exposure lacked dental lesions. In conclusion, BE has the potential to cause pulp thrombosis and odontoblast infarction in female rats. The apparent variability in response to BE noted in moribund sacrificed vs terminally sacrificed rats was attributed to development of tolerance to BE-induced hemolysis and subsequent incisor regeneration.

Administration, Inhalation↗

Co-increase of nerve fibers and HLA-DR- and/or factor-XIIIa-expressing dendritic cells in dentinal caries-affected regions of the human dental pulp: an immunohistochemical study.

Neuro-immune interaction has been suggested to play some modulatory role in the immunodefense of the dentin/pulp complex. In this study, we performed a simultaneous immunohistochemical observation of neural elements and pulpal dendritic cells (PDCs) on human carious teeth, to obtain morphological evidence for neuro-immune interaction in response to dentinal tubule-derived carious stimuli. Human third molars bearing a pulp-exposure-free caries lesion were studied. Immunoperoxidase staining was performed with anti-HLA-DR, anti-coagulation factor XIIIa, and anti-CD14 as PDC markers, and anti-low-affinity nerve growth factor receptor (NGFR), anti-protein gene products 9.5, and anti-calcitonin gene-related peptide as nerve markers. The carious teeth usually exhibited localized accumulation of both PDCs and nerve fibers immunoreactive to each marker, in the para-odontoblastic region corresponding to the pulpal end of carious dentinal tubules. Semi-quantitative digital densitometry revealed that pixel numbers corresponding to factor-XIIIa- and NGFR-immunoreactivity were significantly higher in the carious regions than those in the non-carious regions of the same teeth as well as those in the corresponding regions of intact teeth. Classification of specimens with respect to caries depth showed that the co-increase was most apparent in teeth with superficial caries. The increase of PDCs was less pronounced in carious teeth with reparative dentin. These findings suggest that both pulpal nerves and PDCs respond promptly and actively to dentinal tubule-derived carious stimuli. The synchronized accumulation of the two structures suggests an increased opportunity for neuro-immune interaction that may be of significance in the modulation of pathological processes in the dental pulp.

Dendritic Cells↗

Establishment and characterization of a clonal cell line (RPC-C2A) from dental pulp of the rat incisor.

Maxillary incisor pulp cells from male Wistar rats (7 weeks old) were cultured. After the 52nd subculture, cloning was performed twice and a clonal cell line (RPC-C2A) with high alkaline phosphatase (ALP) activity was established. The population doubling time of RPC-C2A cells was 13.7 h, and the mode of the chromosome number was 42. The heat stability of ALP, the effect of the ALP inhibitors, and the ALP polyacrylamide gel electrophoresis pattern in RPC-C2A cells was the same as for the ALP of the isolated dental pulp.

Alkaline Phosphatase↗