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Nine rats (18\u2009M) were allocated for histological evaluation and immunofluorescence imaging (STED), six rats (12\u2009M) for FIB\u2010SEM observation, and three rats (6\u2009M) served as untreated controls; control molars were used for both immunofluorescence imaging and FIB\u2010SEM. For experimental allocation, two molars from each rat were randomly assigned to one of three groups: NEX MTA (NMTA), Dycal, or no\u2010capping material (NC). After 4\u2009weeks, teeth were collected for histological analysis, FIB\u2010SEM observation and immunofluorescence imaging. Tubulin\u2010b3 and nestin expression were examined by immunofluorescence staining and imaged with STED microscopy. The Kruskal\u2013Wallis test followed by Dunn's multiple comparisons test and one\u2010way ANOVA followed by Tukey's multiple comparisons test were performed to assess significant differences.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      All experimental groups exhibited complete mineralised tissue formation near the exposure site. The NMTA group showed a significantly more regular odontoblastic layer than the Dycal and NC groups (\n                      <jats:italic>p<\/jats:italic>\n                      \u2009&lt;\u20090.05). FIB\u2010SEM demonstrated that NMTA induced a well\u2010organised and continuous tubular structure, whereas Dycal and NC produced irregular, discontinuous, and poorly organised mineralised barriers. STED imaging of the NMTA group revealed strong and organised co\u2010localization of tubulin\u2010\u03b23 and nestin resembling primary dentine, whereas the Dycal and NC groups showed weak, disorganised, or absent signals and lacked co\u2010localised processes.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>NMTA showed superior performance compared with Dycal, consistently promoting well\u2010organised dentine bridge formation, a continuous odontoblastic layer, and coordinated neural marker\u2010positive processes. This indicates that NMTA more effectively supports structural formation of the pulp\u2010dentine complex after DPC.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1111\/iej.70144","type":"journal-article","created":{"date-parts":[[2026,3,18]],"date-time":"2026-03-18T04:11:16Z","timestamp":1773807076000},"page":"1632-1644","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Ultrastructural Evaluation of Newly Formed Dentine After Direct Pulp Capping With Mineral Trioxide Aggregate and Calcium Hydroxide Using Focused Ion Beam Scanning Electron Microscopy and Stimulated Emission Depletion"],"prefix":"10.1111","volume":"59","author":[{"ORCID":"https:\/\/orcid.org\/0009-0008-5168-6774","authenticated-orcid":false,"given":"Bu'er","family":"Qi","sequence":"first","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Graduate School of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu","family":"Toida","sequence":"additional","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rafiqul","family":"Islam","sequence":"additional","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4696-5751","authenticated-orcid":false,"given":"Md Refat Readul","family":"Islam","sequence":"additional","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-3920-3448","authenticated-orcid":false,"given":"Kenta","family":"Tsuchiya","sequence":"additional","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuan","family":"Yuan","sequence":"additional","affiliation":[{"name":"School\/Hospital of Stomatology Zhejiang Chinese Medical University  Hangzhou Zhejiang China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hidehiko","family":"Sano","sequence":"additional","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tomoka","family":"Hasegawa","sequence":"additional","affiliation":[{"name":"Ultrastructure of Hard Tissue, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0886-4407","authenticated-orcid":false,"given":"Monica","family":"Yamauti","sequence":"additional","affiliation":[{"name":"Department of Biomedical and Applied Science Indiana University School of Dentistry  Indianapolis Indiana USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1638-4008","authenticated-orcid":false,"given":"Atsushi","family":"Tomokiyo","sequence":"additional","affiliation":[{"name":"Department of Restorative Dentistry and Endodontology, Faculty of Dental Medicine Hokkaido University  Sapporo Hokkaido Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2026,3,17]]},"reference":[{"key":"e_1_2_12_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.joen.2010.11.001"},{"key":"e_1_2_12_3_1","doi-asserted-by":"publisher","DOI":"10.3390\/molecules26092725"},{"key":"e_1_2_12_4_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.joen.2017.06.031"},{"key":"e_1_2_12_5_1","doi-asserted-by":"publisher","DOI":"10.1186\/s13287-021-02141-y"},{"key":"e_1_2_12_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.joen.2016.04.019"},{"key":"e_1_2_12_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.dental.2016.06.013"},{"key":"e_1_2_12_8_1","doi-asserted-by":"publisher","DOI":"10.1111\/iej.13947"},{"key":"e_1_2_12_9_1","first-page":"4","article-title":"Tunnel Defects in Dentin Bridges: Their Formation Following Direct Pulp Capping","volume":"21","author":"Cox C. 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