Dataset Information

Chronic exposure to lipopolysaccharides as an in vitro model to simulate the impaired odontogenic potential of dental pulp cells under pulpitis conditions.

ABSTRACT:

Background

Simulating a bacterial-induced pulpitis environment in vitro may contribute to exploring mechanisms and bioactive molecules to counteract these adverse effects.

Objective

To investigate the chronic exposure of human dental pulp cells (HDPCs) to lipopolysaccharides (LPS) aiming to establish a cell culture protocol to simulate the impaired odontogenic potential under pulpitis conditions.

Methodology

HDPCs were isolated from four healthy molars of different donors and seeded in culture plates in a growth medium. After 24 h, the medium was changed to an odontogenic differentiation medium (DM) supplemented or not with E. coli LPS (0 - control, 0.1, 1, or 10 µg/mL) (n=8). The medium was renewed every two days for up to seven days, then replaced with LPS-free DM for up to 21 days. The activation of NF-κB and F-actin expression were assessed (immunofluorescence) after one and seven days. On day 7, cells were evaluated for both the gene expression (RT-qPCR) of odontogenic markers (COL1A1, ALPL, DSPP, and DMP1) and cytokines (TNF, IL1B, IL8, and IL6) and the production of reactive nitrogen (Griess) and oxygen species (Carboxy-H2DCFDA). Cell viability (alamarBlue) was evaluated weekly, and mineralization was assessed (Alizarin Red) at 14 and 21 days. Data were analyzed with ANOVA and post-hoc tests (α=5%).

Results

After one and seven days of exposure to LPS, NF-κB was activated in a dose-dependent fashion. LPS at 1 and 10 µg/mL concentrations down-regulated the gene expression of odontogenic markers and up-regulated cytokines. LPS at 10 µg/mL increased both the production of reactive nitrogen and oxygen species. LPS decreased cell viability seven days after the end of exposure. LPS at 1 and 10 µg/mL decreased hDPCs mineralization in a dose-dependent fashion.

Conclusion

The exposure to 10 µg/mL LPS for seven days creates an inflammatory environment that is able to impair by more than half the odontogenic potential of HDPCs in vitro, simulating a pulpitis-like condition.

SUBMITTER: Mendes Soares IP

PROVIDER: S-EPMC10382076 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Chronic exposure to lipopolysaccharides as an in vitro model to simulate the impaired odontogenic potential of dental pulp cells under pulpitis conditions.

Mendes Soares Igor Paulino IP Anselmi Caroline C Pires Maria Luiza Barucci Araujo MLBA Ribeiro Rafael Antonio de Oliveira RAO Leite Maria Luísa ML Soares Diana Gabriela DG DE Souza Costa Carlos Alberto CA Hebling Josimeri J

Journal of applied oral science : revista FOB 20230724

<h4>Background</h4>Simulating a bacterial-induced pulpitis environment in vitro may contribute to exploring mechanisms and bioactive molecules to counteract these adverse effects.<h4>Objective</h4>To investigate the chronic exposure of human dental pulp cells (HDPCs) to lipopolysaccharides (LPS) aiming to establish a cell culture protocol to simulate the impaired odontogenic potential under pulpitis conditions.<h4>Methodology</h4>HDPCs were isolated from four healthy molars of different donors a ...[more]

PMID: 37493701

Similar Datasets

Project description:Hyperglycemia, a major characteristic of diabetes, is considered to play a vital role in diabetic complications. High glucose levels have been found to inhibit the mineralization of dental pulp cells. However, gene expression associated with this phenomenon has not yet been reported. This is important for future dental therapeutic application.ObjectiveOur study aimed to investigate the effect of high glucose levels on mineralization of human dental pulp-derived cells (hDPCs) and identify the genes involved.MethodologyhDPCs were cultured in mineralizing medium containing 25 or 5.5 mM D-glucose. On days 1 and 14, RNA was extracted and expression microarray performed. Then, differentially expressed genes (DEGs) were selected for further validation using the reverse transcription quantitative polymerase chain reaction (RT-qPCR) method. Cells were fixed and stained with alizarin red on day 21 to detect the formation of mineralized nodules, which was further quantified by acetic acid extraction.ResultsComparisons between high-glucose and low-glucose conditions showed that on day 1, there were 72 significantly up-regulated and 75 down-regulated genes in the high-glucose condition. Moreover, 115 significantly up- and 292 down-regulated genes were identified in the high-glucose condition on day 14. DEGs were enriched in different GO terms and pathways, such as biological and cellular processes, metabolic pathways, cytokine-cytokine receptor interaction and AGE-RAGE signaling pathways. RT-qPCR results confirmed the significant expression of pyruvate dehydrogenase kinase 3 (PDK3), cyclin-dependent kinase 8 (CDK8), activating transcription factor 3 (ATF3), fibulin-7 (Fbln-7), hyaluronan synthase 1 (HAS1), interleukin 4 receptor (IL-4R) and apolipoprotein C1 (ApoC1).ConclusionsThe high-glucose condition significantly inhibited the mineralization of hDPCs. DEGs were identified, and interestingly, HAS1 and Fbln-7 genes may be involved in the glucose inhibitory effect on hDPC mineralization.

Project description:IntroductionOdontogenic differentiation of human dental pulp stem cells (hDPSCs) is a key step of pulp regeneration. Recent studies showed that circular RNAs (circRNAs) have many biological functions and that competing endogenous RNA (ceRNA) is their most common mechanism of action. However, the role of circRNAs in hDPSCs during odontogenesis is still unclear.MethodsIsolated hDPSCs were cultured in essential and odontogenic medium. Total RNA was extracted after 14 days of culture, and then, microarray analysis was performed to measure the differential expressions of circRNAs. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was then performed to validate the microarray results. Based on microarray data from this study and available in the database, a ceRNA network was constructed to investigate the potential function of circRNAs during odontogenesis. In addition, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to investigate the potential correlation between signaling pathways and circRNAs. In addition, qRT-PCR and Western blot analysis were used to explore the function of hsa_circRNA_104101.ResultsWe found 43 upregulated circRNAs and 144 downregulated circRNAs during the odontogenic differentiation process (fold change > 1.5 and <-1.5, respectively; P < 0.05). qRT-PCR results were in agreement with the microarray results. Bioinformatic analysis revealed that the Wnt signaling pathway and the TGF-β signaling pathway, as well as the other pathways associated with odontogenic differentiation, were correlated to the differentially expressed circRNAs. hsa_circRNA_104101 was proved to promote the odontogenic differentiation of hDPSCs.ConclusionThis study reported 187 circRNAs that were differentially expressed in hDPSCs during odontogenic differentiation. Bioinformatic analysis of the expression data suggested that circRNA-miRNA-mRNA networks might act as a crucial mechanism for hDPSC odontogenic differentiation, providing a theoretical foundation for the study of pulp regeneration regulation by circRNAs.

Dataset Information

Chronic exposure to lipopolysaccharides as an in vitro model to simulate the impaired odontogenic potential of dental pulp cells under pulpitis conditions.

Background

Objective

Methodology

Results

Conclusion

Publications

Chronic exposure to lipopolysaccharides as an in vitro model to simulate the impaired odontogenic potential of dental pulp cells under pulpitis conditions.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets