Role of Lipopolysaccharide, Derived from Various Bacterial Species, in Pulpitis—A Systematic Review

Lipopolysaccharide (LPS) is widely used for induction of inflammation in various human tissues, including dental pulp. The purpose of this study was to summarize current medical literature focusing on (1) cell types used by researchers to simulate dental pulp inflammation, (2) LPS variants utilized in experimental settings and how these choices affect the findings. Our study was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). We searched for studies reporting outcomes of lipopolysaccharide application on dental pulp cells in vitro using electronic databases: MEDLINE, Web of Science and Scopus. Having gathered data from 115 papers, we aimed to present all known effects LPS has on different cell types present in dental pulp. We focused on specific receptors and particles that are involved in molecular pathways. Our review provides an essential foundation for further research using in vitro models of pulpitis.

Inhibition of the Nav1.7 Channel in the Trigeminal Ganglion Relieves Pulpitis Inflammatory Pain

Pulpitis causes significant changes in the peripheral nervous system, which induce hyperalgesia. However, the relationship between neuronal activity and Nav1.7 expression following pulpal noxious pain has not yet been investigated in the trigeminal ganglion (TG). The aim of our study was to verify whether experimentally induced pulpitis activates the expression of Nav1.7 peripherally and the neuronal activities of the TGs can be affected by Nav1.7 channel inhibition. Acute pulpitis was induced through allyl isothiocyanate (AITC) application to the rat maxillary molar tooth pulp. Three days after AITC application, abnormal pain behaviors were recorded, and the rats were euthanized to allow for immunohistochemical, optical imaging, and western blot analyses of the Nav1.7 expression in the TG. A significant increase in AITC-induced pain-like behaviors and histological evidence of pulpitis were observed. In addition, histological and western blot data showed that Nav1.7 expressions in the TGs were significantly higher in the AITC group than in the naive and saline group rats. Optical imaging showed that the AITC group showed higher neuronal activity after electrical stimulation of the TGs. Additionally, treatment of ProTxII, selective Nav1.7 blocker, on to the TGs in the AITC group effectively suppressed the hyperpolarized activity after electrical stimulation. These findings indicate that the inhibition of the Nav1.7 channel could modulate nociceptive signal processing in the TG following pulp inflammation.

Mitochondrial DNA leakage exacerbates odontoblast inflammation through gasdermin D-mediated pyroptosis

Alleviating odontoblast inflammation is crucial to control the progression of pulpitis. Mitochondrial DNA (mtDNA) is a vital driver of inflammation when it leaks from mitochondria of inflamed odontoblasts into the cytosol. Bacteria-induced inflammation leads to a novel type of cell death named pyroptosis. The canonical pyroptosis is a gasdermin (GSDM)-dependent cytolytic programmed cell death characterized by cell swelling and pore formation in the plasma membrane. To date, whether odontoblast cytosolic mtDNA regulates dental pulp inflammation through the canonical pyroptosis pathway remains to be elucidated. In this study, high gasdermin D (GSDMD) expression was detected in human pulpitis. We found that LPS stimulation of mDPC6T cells promoted BAX translocation from the cytosol to the mitochondrial membrane, leading to mtDNA release. Moreover, overexpression of isolated mtDNA induced death in a large number of mDPC6T cells, which had the typical appearance of pyroptotic cells. Secretion of the inflammatory cytokines CXCL10 and IFN-β was also induced by mtDNA. These results suggest that cytosolic mtDNA participates in the regulation of odontoblast inflammation through GSDMD-mediated pyroptosis in vitro. Interestingly, after overexpression of mtDNA, the expression of inflammatory cytokines CXCL10 and IFN-β was increased and not decreased in GSDMD knockdown mDPC6T cells. We further proposed a novel model in which STING-dependent inflammation in odontoblast-like cell is a compensatory mechanism to control GSDMD-mediated pyroptosis, jointly promoting the immune inflammatory response of odontoblasts. Collectively, these findings provide the first demonstration of the role of the mtDNA-GSDMD-STING in controlling odontoblast inflammation and a detailed description of the underlying interconnected relationship.

EZH2 Might Affect Macrophage Chemotaxis and Anti-Inflammatory Factors by Regulating CCL2 in Dental Pulp Inflammation

Objectives. We aimed to evaluate the effects of Enhancer of Zeste Homolog 2 (EZH2) on regulation of macrophage migration and expression of anti-inflammatory genes in pulpitis. Methods. Dental pulp inflammation was verified by histology in rat pulpitis model induced by lipopolysaccharide (LPS). Immunohistochemistry staining was used to detect changes of the expression of EZH2 and tumor necrosis factor alpha (TNF-α) in dental pulp inflammation. The expression of EZH2, CCL2, and cluster of differentiation 68 (CD68: macrophage surface marker) was measured by immunofluorescence staining. The effect of EZH2 on microphage migration was assessed by cell migration assay. The expressions of anti-inflammatory cytokine interleukins (IL-4 and IL-10) and transforming growth factor-β (TGF-β) in HDPCs which were treated by EZH2 complex, CCL2 complex, and CCL2 antibody were examined by quantitative real-time polymerase chain reaction (q-PCR). Results. The expression of TNF-α gradually increased in dental pulp inflammation. The expression of EZH2 in dental pulp decreased in 8 hours after LPS stimulation. However, the expression of EZH2 gradually increased in dental pulp after 1 day stimulation by LPS. The results of immunofluorescence staining showed that the expressions of EZH2, CCL2, and CD68 were significantly upregulated in dental pulp inflammation of rats. EZH2 could enhance macrophage migration. And the chemotactic activity of macrophages exposed to supernatants of EZH2-treated HDPCs could be inhibited by CCL2 inhibition. In addition, EZH2 suppressed the expression of anti-inflammatory genes, but CCL2 inhibition reversed the downregulation of anti-inflammatory factors, including IL-4 and TGF-β in HDPCs. Conclusions. EZH2 might affect chemotaxis of macrophages and the expression of anti-inflammatory factors by regulating CCL2. EZH2 plays an important role in the development of dental pulp inflammation, and it might be as a target for treatment of pulpitis.

Leukotriene B 4 Loaded in Microspheres Regulate the Expression of Genes Related to Odontoblastic Differentiation and Biomineralization by Dental Pulp Stem Cells

Background: Leukotriene B4 (LTB4) is a potent lipid mediator that stimulate the immune response. Because dental pulp inflammation and dentin repair are intrinsically related responses, the aim of this research was to investigate the potential of LTB4 in inducing differentiation of dental pulp stem cells. Methods: Microspheres (MS) loaded with LTB4 were prepared using an oil emulsion solvent extraction evaporation process and sterility, characterization, efficiency of LTB4 encapsulation and in vitro LTB4 release assay were investigated. Mouse dental pulp stem cells (OD-21) were stimulated with soluble LTB4 or MS loaded with LTB4 (0.01 and 0.1 μM). Cytotoxicity and cell viability was determined by lactate dehydrogenase (LDH) and MTT (methylthiazol tetrazolium) assays . Gene expression were measured by quantitative reverse transcription polymerase chain reaction (qRT-PCR) after 3, 6, 24, 48 and 72 h. Results: Mineralized nodule formation was assessed after 28 days of OD-21 cell stimulation with LTB4. Groups were compared using the one-way ANOVA test followed by Dunnett's post-test (α = 0.05). Treatment with LTB4 or MS loaded with LTB4 (0.01 and 0.1 micrometer- μM) were not cytotoxic to OD-21 cells. Treatment with LTB4 modulated the expression of the Ibsp (integrin binding sialoprotein) and Runx2 (runt-related transcription factor 2) genes differently depending on the experimental period analyzed. Interestingly LTB4 loaded in microspheres (0.1 μM) allowed long term dental pulp cell differentiation and biomineralization. LTB4 loaded in MS was not cytotoxic and induced an odontoblastic cell phenotype differentiation. Conclusion: These findings shed light on a novel pharmacological strategy to induce dental pulp cell differentiation.

THE EFFECT OF KARAMUNTING (Rhodomyrtus Tomentosa) LEAF EXTRACT ON THE NUMBER OF MACROPHAGES IN PULP INFLAMMATION

Background: Reversible pulpitis is an inflammation of dental pulp caused by the opening of the pulp due to cavities. One of the procedures in exposed pulp treatment is direct pulp capping using calcium hydroxide. However, this material has side effects, its high pH can cause necrosis, and due to that, a safer alternative material is needed. Karamunting leaf extract contains phenolic compounds, flavonoids, tannins, and saponins which have immunomodulatory properties that play an important role in healing exposed pulp. Objective: To determine the effect of karamunting leaf extract on the number of macrophages in pulp inflammation. Methods: This was a true experimental research with posttest-only group design, using simple random sampling that consist of 24 male Wistar rats which later be divided into 3 groups. The perforated rat dental pulp was then treated with karamunting leaf extract as a treatment group, calcium hydroxide as a positive control group, and not given any application (without drug) as a negative control group. The samples were analyzed histologically on the 3rd to 7th day after the application, inflammatory response occurred in all groups. Results: The two-way ANOVA results showed that there was a significant difference between the karamunting leaf extract group, the group that was not given drug, and the group given calcium hydroxide with a value p<0.05. Conclusion: Based on the research conducted, it is concluded that the administration of karamunting leaf extract can reduce the number of macrophages in pulp inflammation. Keywords: calcium hydroxid, exposed pulp, karamunting leaf, macrophages, pulp inflammation.

Diagnosis of Pulpitis from Dental Panoramic Radiograph Using Histogram of Gradients with Discrete Wavelet Transform and Multilevel Neural Network Techniques

Radiograph plays the major role of diagnosis, treatment and surgery in the Dental field. There are many types of Intra and extra oral radiographs in which Dental Panoramic Radiograph helps in visualising the full view of the oral cavity. Pulpitis is the dental diseases caused due to the inflammation of the dental pulp from untreated caries, trauma or multiple restorations which leads to Apical Periodontitis. To predict the severity of pulp vitality pulp inflammation has to be evaluated. Radiographs helps the dentist in diagnosing the extent of tooth decay and inflammation. An automatic diagnostic model is proposed using robust algorithms to diagnose pulpits. Dental Panoramic Radiograph is used in the proposed research to diagnose the pulpitis and to classify the normal teeth from the pulpitis. The collected images are pre-processed using Histogram Equalization and filtered using Gaussian and Median filters. Modified K-Means algorithm is applied to segment the bony and teeth area from the oral cavity area. Integral Histogram of Gradients with Discrete Wavelet Transform feature extraction techniques and Multi-Layer Neural Network Classifier is proposed to achieve the accuracy of 91.09% which can be used as an assistive tool by the dentist to diagnose pulpitis.

Differentially Expressed Salivary Proteins in Dental Caries Patients

Dental caries is a multifactorial disease mainly caused by cariogenic bacteria commonly found in the oral cavity. Dental caries may cause demineralization of the tooth, cavitation, hypersensitivity, pulp inflammation, and even tooth loss if left untreated. Saliva secreted in the oral cavity can serve as a tool for identification of biomarkers for early detection of diseases. In the present study, differential expression of salivary proteins from 33 dental caries patients was compared with 10 control subjects. The unstimulated saliva was analyzed by 12% SDS-PAGE and two-dimensional gel electrophoresis. Gelatin and casein zymography was performed to check for protease activity. Also, salivary IgAs from both groups were compared by sandwich ELISA technique. Dental caries patient’s saliva showed decreased caseinolytic and increased gelatinolytic activity probably due to metalloproteases and cathepsins. Mean salivary levels of sIgA were also significantly higher ( p < 0.018 ) in dental caries saliva samples. The 2D electrophoresis profile of both the groups showed regions on gel with visually detectable alterations in protein expression. The present study is among the few initial studies in the locality for identification of protein differences in saliva from dental caries patients and has demonstrated a good potential to identify alterations. However, a large population-based analysis is required to validate these findings to be translated as a tool for indicative applications.

The Role of microRNAs in Pulp Inflammation

The dental pulp can be affected by thermal, physical, chemical, and bacterial phenomena that stimulate the inflammatory response. The pulp tissue produces an immunological, cellular, and vascular reaction in an attempt to defend itself and resolve the affected tissue. The expression of different microRNAs during pulp inflammation has been previously documented. MicroRNAs (miRNAs) are endogenous small molecules involved in the transcription of genes that regulate the immune system and the inflammatory response. They are present in cellular and physiological functions, as well as in the pathogenesis of human diseases, becoming potential biomarkers for diagnosis, prognosis, monitoring, and safety. Previous studies have evidenced the different roles played by miRNAs in proinflammatory, anti-inflammatory, and immunological phenomena in the dental pulp, highlighting specific key functions of pulp pathology. This systematized review aims to provide an understanding of the role of the different microRNAs detected in the pulp and their effects on the expression of the different target genes that are involved during pulp inflammation.

The Effect of Lipoteichoic Acid from Lactobacillus plantarum on Dental Pulp Inflammation

Objective Lactobacillus plantarum, a bacterium located in deep caries, has a virulence factor in the form of lipoteichoic acid (LTA), which is found in the bacterial cell wall. LTA is able to trigger a neutrophils response in the dental pulp inflammation process within the first 6 to 24 hours. The quantity of bacteria is one factor influencing the increase in number of neutrophils in addition to the quality of the bacteria. This study seeks to analyze the effect of lipoteichoic acid of Lactobacillus plantarum (LTA-Lp) in the dental pulp inflammation by observing the number of neutrophil cells in a histopathological view. Materials and Methods The LTA was isolated from L. plantarum. The left upper molar of Rattus novergicus was mechanically perforated under anesthesia to induce dental pulp inflammation. The perforated tooth was then induced by 10 and 15 µg/mL of LTA-Lp and then restored by a temporary filling. The perforated tooth in the control group was only restored by a temporary filling. After 24, 48, and 72 hours, the tooth was extracted and then stained with hematoxylins and eosin to observe the neutrophils in the dental pulp via a light microscope. Result The number of neutrophils in the dental pulp after induction by 15 µg/mL of LTA-Lp is higher than 10 µg/mL of LTA-Lp and both controls. There were significant differences in the number of neutrophils in the dental pulp, in each group on 24, 48, and 72 hours after LTA-Lp inducing (p < 0.05). Conclusion The LTA-Lp dose of of 10 and 15 µg/mL affected the dental pulp inflammation by affecting the number of neutrophils.