Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion
Objectives: The objective of the present study was to investigate whether NOD-like receptor family pyrin domain-containing 3 (NLRP3) and absent in melanoma 2 (AIM2) inflammasomes pathways were involved in an experimental model of fibroblast activation named nemosis, which was used to mimic circumsta...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2024-12-01
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| Series: | Regenerative Therapy |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2352320424000245 |
| _version_ | 1797268961586315264 |
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| author | Shafei Zhai Lihui Zhang Xue Li Qi Yu Changkui Liu |
| author_facet | Shafei Zhai Lihui Zhang Xue Li Qi Yu Changkui Liu |
| author_sort | Shafei Zhai |
| collection | DOAJ |
| description | Objectives: The objective of the present study was to investigate whether NOD-like receptor family pyrin domain-containing 3 (NLRP3) and absent in melanoma 2 (AIM2) inflammasomes pathways were involved in an experimental model of fibroblast activation named nemosis, which was used to mimic circumstances without bacteria stimulation. Methods: Nemosis of human dental pulp fibroblast (DPFs) was induced by three-dimensional culture in U-shaped 96-well plates and investigated by scanning electron microscopy (SEM). DPFs monolayers were used as control. Annexin V-FITC/7-AAD apoptosis assay was performed on the DPFs spheroids by flowcytometry. Caspase-1 activity detection assay was conducted on the DPFs spheroids. Quantitative real-time polymerase chain reaction (qRT-PCR), cytokine measurements, Western blot and the effect of COX-2 inhibitor on spheroids was studied. Results: SEM study observed human dental pulp fibroblast clusters and cell membranes damage on the surface of DPFs spheroids. The percentages of necrotic cells from DPFs spheroids gradually increased as the incubation time increased. A statistically significant increase in caspase-1 activity was observed after DPFs spheroids formation. DPFs spheroids displayed significant amounts of NLRP3, AIM2 mRNA and protein expression, caspase-1 mRNA expression and cleaved Caspase-1 protein expression and high IL-1β concentrations (P < 0.05) than DPFs monolayers. Specific COX-2 inhibitor (NS-398) decreased NLRP3 mRNA and protein expression, cleaved Caspase-1 protein expression, Caspase-1 activity and IL-1β mRNA expression and IL-1β concentrations (P < 0.05). However, Specific COX-2 inhibitor had no impact on AIM2 mRNA and protein expression, caspase-1 mRNA expression and pro-Caspase-1 protein expression. Conclusions: In conclusion, clustering human DPFs spontaneously activated NLRP3 and AIM2 inflammasomes and induced IL-1β secretion which could be partially attenuated by COX-2 inhibitor. Thus, nemosis could become a powerful model for studying mechanisms underlying aseptic pulpitis. |
| first_indexed | 2024-04-25T01:40:48Z |
| format | Article |
| id | doaj.art-2776d3d516794c9cac6376a73a6ffe47 |
| institution | Directory Open Access Journal |
| issn | 2352-3204 |
| language | English |
| last_indexed | 2024-04-25T01:40:48Z |
| publishDate | 2024-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Regenerative Therapy |
| spelling | doaj.art-2776d3d516794c9cac6376a73a6ffe472024-03-08T05:18:51ZengElsevierRegenerative Therapy2352-32042024-12-01271220Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretionShafei Zhai0Lihui Zhang1Xue Li2Qi Yu3Changkui Liu4Department of Stomatology, Xi'an Medical University, Xi'an, 710075, Shaanxi Province, ChinaDepartment of Stomatology, Xi'an Medical University, Xi'an, 710075, Shaanxi Province, ChinaDepartment of Stomatology, Xi'an Medical University, Xi'an, 710075, Shaanxi Province, ChinaShaanxi Key Laboratory of Ischemic Cardiovascular Disease, Institute of Basic and Translational Medicine, Xi'an Medical University, Xi'an, 710021, Shaanxi Province, ChinaDepartment of Stomatology, Xi'an Medical University, Xi'an, 710075, Shaanxi Province, China; Corresponding author.Objectives: The objective of the present study was to investigate whether NOD-like receptor family pyrin domain-containing 3 (NLRP3) and absent in melanoma 2 (AIM2) inflammasomes pathways were involved in an experimental model of fibroblast activation named nemosis, which was used to mimic circumstances without bacteria stimulation. Methods: Nemosis of human dental pulp fibroblast (DPFs) was induced by three-dimensional culture in U-shaped 96-well plates and investigated by scanning electron microscopy (SEM). DPFs monolayers were used as control. Annexin V-FITC/7-AAD apoptosis assay was performed on the DPFs spheroids by flowcytometry. Caspase-1 activity detection assay was conducted on the DPFs spheroids. Quantitative real-time polymerase chain reaction (qRT-PCR), cytokine measurements, Western blot and the effect of COX-2 inhibitor on spheroids was studied. Results: SEM study observed human dental pulp fibroblast clusters and cell membranes damage on the surface of DPFs spheroids. The percentages of necrotic cells from DPFs spheroids gradually increased as the incubation time increased. A statistically significant increase in caspase-1 activity was observed after DPFs spheroids formation. DPFs spheroids displayed significant amounts of NLRP3, AIM2 mRNA and protein expression, caspase-1 mRNA expression and cleaved Caspase-1 protein expression and high IL-1β concentrations (P < 0.05) than DPFs monolayers. Specific COX-2 inhibitor (NS-398) decreased NLRP3 mRNA and protein expression, cleaved Caspase-1 protein expression, Caspase-1 activity and IL-1β mRNA expression and IL-1β concentrations (P < 0.05). However, Specific COX-2 inhibitor had no impact on AIM2 mRNA and protein expression, caspase-1 mRNA expression and pro-Caspase-1 protein expression. Conclusions: In conclusion, clustering human DPFs spontaneously activated NLRP3 and AIM2 inflammasomes and induced IL-1β secretion which could be partially attenuated by COX-2 inhibitor. Thus, nemosis could become a powerful model for studying mechanisms underlying aseptic pulpitis.http://www.sciencedirect.com/science/article/pii/S2352320424000245Dental pulp fibroblastNemosisInflammasomePyrin domain-containing 3Absent in melanoma 2 |
| spellingShingle | Shafei Zhai Lihui Zhang Xue Li Qi Yu Changkui Liu Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion Regenerative Therapy Dental pulp fibroblast Nemosis Inflammasome Pyrin domain-containing 3 Absent in melanoma 2 |
| title | Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion |
| title_full | Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion |
| title_fullStr | Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion |
| title_full_unstemmed | Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion |
| title_short | Clustering human dental pulp fibroblasts spontaneously activate NLRP3 and AIM2 inflammasomes and induce IL-1β secretion |
| title_sort | clustering human dental pulp fibroblasts spontaneously activate nlrp3 and aim2 inflammasomes and induce il 1β secretion |
| topic | Dental pulp fibroblast Nemosis Inflammasome Pyrin domain-containing 3 Absent in melanoma 2 |
| url | http://www.sciencedirect.com/science/article/pii/S2352320424000245 |
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