{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,12,1]],"date-time":"2025-12-01T06:40:41Z","timestamp":1764571241641,"version":"3.44.0"},"reference-count":45,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,8,20]],"date-time":"2025-08-20T00:00:00Z","timestamp":1755648000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100005893","name":"China Academy of Traditional Chinese Medicine","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100005893","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:sec><jats:title>Introduction<\/jats:title><jats:p>Autoimmune thyroiditis (AIT) is a chronic autoimmune disease characterized by lymphocytic infiltration of the thyroid gland and elevated specific antibodies. Its incidence rises annually, yet no standardized animal model fully mimics human AIT. Given unclear pathogenesis and lack of targeted immunotherapies, researchers invest significant time in developing suitable models. This study systematically compares pathological and immunological effects of different immunization conditions (antigen dose, frequency, administration methods) in NOD\/LtJ mice to establish an optimal model for elucidating AIT pathogenesis and therapies.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>Eighty female NOD\/LtJ mice were divided into subcutaneous (SC) and tail vein intravenous (IV) injection groups. SC groups received porcine thyroglobulin (pTg) emulsified in CFA (primary) and IFA (booster), with doses of 50\/100\/200 \u03bcg and frequencies of 2 or 3 immunizations. IV groups received pTg in PBS followed by LPS (3 immunizations: weeks 1, 3, 4). All model groups drank 0.05% NaI water. Thyroid histopathology (HE staining, infiltration scoring), serum TPO-Ab\/TG-Ab (ELISA), cytokines (multiplex assay), Th17\/Treg cells (multiplex immunofluorescence), and thyroid IL-17A\/NLRP3\/Caspase-1 (immunohistochemistry) were analyzed 2 weeks post-last immunization.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>High-dose antigen (200 \u03bcg pTg) with high-frequency immunization (three times) via SC or IV routes induced severe thyroid lymphocyte infiltration (scores: SC 3.4\u00b10.55, IV 3.2\u00b10.45; p&amp;lt;0.01 vs. controls), follicular destruction, and elevated serum antibodies (TPO-Ab: IV 438.8\u00b113.15 &amp;gt; SC 406.2\u00b17.46; TG-Ab: IV 158.4\u00b15.32 &amp;gt; SC 141.9\u00b12.36). This protocol activated Th1\/Th17 cytokines (IL-17A, IL-6, TNF-\u03b1), increased Treg cells (p&amp;lt;0.001), and specifically enhanced NLRP3 (p&amp;lt;0.001) and Caspase-1 in thyroid tissue, with IV injection showing superior antibody production and inflammasome activation.<\/jats:p><\/jats:sec><jats:sec><jats:title>Discussion<\/jats:title><jats:p>The combination of high-dose pTg (200 \u03bcg) and three immunizations maximally induced AIT pathology and immune responses in NOD\/LtJ mice. Tail vein injection excelled in stimulating antibody production and NLRP3 activation, while subcutaneous injection promoted stronger histological inflammation. After balancing operational feasibility, pathological reproducibility, and immunological specificity, three subcutaneous and intravenous tail injection of 200 \u03bcg pTg are recommended as the optimal modeling protocol. This approach accelerates model selection, improves experimental efficiency, and reduces animal use, providing a robust foundation for AIT research.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fimmu.2025.1591196","type":"journal-article","created":{"date-parts":[[2025,8,20]],"date-time":"2025-08-20T05:34:01Z","timestamp":1755668041000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":1,"title":["Constructing the optimal experimental autoimmune thyroiditis mouse model using porcine thyroglobulin"],"prefix":"10.3389","volume":"16","author":[{"given":"Ke","family":"Liu","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Pei","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Zi-Shan","family":"Jin","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiang-Kun","family":"Meng","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jin-Li","family":"Luo","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lin","family":"Han","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiao-Tong","family":"Yu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1965","published-online":{"date-parts":[[2025,8,20]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.1016\/j.beem.2019.101367","article-title":"Hashimotos\u2019 Thyroiditis: epidemiology, pathogenesis, clinic and therapy","volume":"33","author":"Ragusa","year":"2019","journal-title":"Best Pract Res Clin Endocrinol Metab"},{"key":"B2","doi-asserted-by":"publisher","first-page":"1788","DOI":"10.3390\/biomedicines12081788","article-title":"The impact of environmental factors on the development of autoimmune thyroiditis-review","volume":"12","author":"Cyna","year":"2024","journal-title":"Biomedicines"},{"key":"B3","doi-asserted-by":"publisher","first-page":"918","DOI":"10.3390\/cells12060918","article-title":"Molecular mechanisms in autoimmune thyroid disease","volume":"12","author":"Vargas-Uricoechea","year":"2023","journal-title":"Cells"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1016\/j.beem.2022.101661","article-title":"Genetics and epigenetics of autoimmune thyroid diseases: translational implications","volume":"37","author":"Lee","year":"2023","journal-title":"Best Pract Res Clin Endocrinol Metab"},{"key":"B5","doi-asserted-by":"publisher","DOI":"10.3389\/fpubh.2022.1020709","article-title":"Global prevalence and epidemiological trends of hashimoto\u2019s thyroiditis in adults: A systematic review and meta-analysis","volume":"10","author":"Hu","year":"2022","journal-title":"Front Public Health"},{"key":"B6","doi-asserted-by":"publisher","DOI":"10.1089\/thy.2019.0067","article-title":"Efficacy and safety of long-term universal salt iodization on thyroid disorders: epidemiological evidence from 31 provinces of mainland China","volume":"30","author":"Li","year":"2020","journal-title":"Thyroid"},{"key":"B7","doi-asserted-by":"publisher","DOI":"10.1016\/j.autrev.2014.01.007","article-title":"Hashimoto thyroiditis: clinical and diagnostic criteria","volume":"13","author":"Caturegli","year":"2014","journal-title":"Autoimmun Rev"},{"key":"B8","doi-asserted-by":"publisher","DOI":"10.1515\/jpem-2013-0370","article-title":"Multiple autoimmunity, type 1 diabetes (T1dm), autoimmune thyroiditis and thyroid cancer: is there an association? 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