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External Dentin Stimulation Induces ATP Release in Human Teeth.

ABSTRACT: ATP is involved in neurosensory processing, including nociceptive transduction. Thus, ATP signaling may participate in dentin hypersensitivity and dental pain. In this study, we investigated whether pannexins, which can form mechanosensitive ATP-permeable channels, are present in human dental pulp. We also assessed the existence and functional activity of ecto-ATPase for extracellular ATP degradation. We further tested if ATP is released from dental pulp upon dentin mechanical or thermal stimulation that induces dentin hypersensitivity and dental pain and if pannexin or pannexin/gap junction channel blockers reduce stimulation-dependent ATP release. Using immunofluorescence staining, we demonstrated immunoreactivity of pannexin 1 and 2 in odontoblasts and their processes extending into the dentin tubules. Using enzymatic histochemistry staining, we also demonstrated functional ecto-ATPase activity within the odontoblast layer, subodontoblast layer, dental pulp nerve bundles, and blood vessels. Using an ATP bioluminescence assay, we found that mechanical or cold stimulation to the exposed dentin induced ATP release in an in vitro human tooth perfusion model. We further demonstrated that blocking pannexin/gap junction channels with probenecid or carbenoxolone significantly reduced external dentin stimulation-induced ATP release. Our results provide evidence for the existence of functional machinery required for ATP release and degradation in human dental pulp and that pannexin channels are involved in external dentin stimulation-induced ATP release. These findings support a plausible role for ATP signaling in dentin hypersensitivity and dental pain.

SUBMITTER: Liu X

PROVIDER: S-EPMC4547316 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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External Dentin Stimulation Induces ATP Release in Human Teeth.

Liu X X Wang C C Fujita T T Malmstrom H S HS Nedergaard M M Ren Y F YF Dirksen R T RT

Journal of dental research 20150630 9

ATP is involved in neurosensory processing, including nociceptive transduction. Thus, ATP signaling may participate in dentin hypersensitivity and dental pain. In this study, we investigated whether pannexins, which can form mechanosensitive ATP-permeable channels, are present in human dental pulp. We also assessed the existence and functional activity of ecto-ATPase for extracellular ATP degradation. We further tested if ATP is released from dental pulp upon dentin mechanical or thermal stimula ...[more]

PMID: 26130258

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Project description:Exposure of dentin tubules due to loss of protective enamel (crown) and cementum (root) tissues as a result of erosion, mechanical wear, gingival recession, etc. has been the leading causes of dentin hypersensitivity. Despite being a widespread ailment, no permanent solution exists to address this oral condition. Current treatments are designed to alleviate the pain by either using desensitizers or blocking dentin tubules by deposition of minerals or solid precipitates, which often have short-lived effects. Reproducing an integrated mineral layer that occludes exposed dentin with concomitant peritubular mineralization is essential to reestablish the structural and mechanical integrity of the tooth with long-term durability. Here, we describe a biomimetic treatment that promotes dentin repair using a mineralization-directing peptide, sADP5, derived from amelogenin. The occlusion was achieved through a layer-by-layer peptide-guided remineralization process that forms an infiltrating mineral layer on dentin. The structure, composition, and nanomechanical properties of the remineralized dentin were analyzed by cross-sectional scanning electron microscopy imaging, energy dispersive X-ray spectroscopy, and nanomechanical testing. The elemental analysis provided calcium and phosphate compositions that are similar to those in hydroxyapatite. The measured average hardness and reduced elastic modulus values for the mineral layer were significantly higher than those of the demineralized and sound human dentin. The structural integration of the new mineral and underlying dentin was confirmed by thermal aging demonstrating no physical separation. These results suggest that a structurally robust and mechanically durable interface is formed between the interpenetrating mineral layer and underlying dentin that can withstand long-term mechanical and thermal stresses naturally experienced in the oral environment. The peptide-guided remineralization procedure described herein could provide a foundation for the development of highly effective oral care products leading to novel biomimetic treatments for a wide range of demineralization-related ailments and, in particular, offers a potent long-term solution for dentin hypersensitivity.

Project description:The mechanism of pain in dentine hypersensitivity is poorly understood but proposed to result from the activation of dental sensory neurons in response to dentinal fluid movements. Odontoblasts have been suggested to contribute to thermal and mechanosensation in the tooth via expression of transient receptor potential (TRP) channels. However, a mechanism by which odontoblasts could modulate neuronal activity has not been demonstrated. In this study, we investigated functional TRP channel expression in human odontoblast-like cells and measured ATP release in response to TRP channel activation. Human immortalized dental pulp cells were driven toward an odontoblast phenotype by culture in conditioned media. Functional expression of TRP channels was determined with reverse transcription polymerase chain reaction and ratiometric calcium imaging with Fura-2. ATP release was measured using a luciferin-luciferase assay. Expression of mRNA for TRPA1, TRPV1, and TRPV4 but not TRPM8 was detected in odontoblasts by reverse transcription polymerase chain reaction. Expression of TRPV4 protein was detected by Western blotting and immunocytochemistry. The TRPA1 agonists allyl isothiocyanate and cinnamaldehyde and the TRPV4 agonist GSK1016790A caused a concentration-dependent increase in intracellular Ca(2+) concentration that was inhibited by the selective antagonists HC030031, AP18, and HC067047, respectively. In contrast, exposure to the TRPV1 agonist capsaicin or the TRPM8 agonist icilin had no effect on intracellular Ca(2+) concentration. Treatment with allyl isothiocyanate, cinnamaldehyde, or GSK1016790A caused an increase in ATP concentration in culture medium that was abolished by preincubation with TRP channel antagonists. These data demonstrate that activation of TRPA1 and TRPV4 channels in human odontoblast-like cells can stimulate ATP release. We were unable to confirm the presence of thermosensitive TRPV1 and TRPM8 that has previously been reported in odontoblasts.

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External Dentin Stimulation Induces ATP Release in Human Teeth.

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External Dentin Stimulation Induces ATP Release in Human Teeth.

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