{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,8,5]],"date-time":"2026-08-05T01:50:18Z","timestamp":1785894618073,"version":"3.56.0"},"posted":{"date-parts":[[2025,12,2]]},"group-title":"Neuroscience","reference-count":78,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,12,2]],"date-time":"2025-12-02T00:00:00Z","timestamp":1764633600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"funder":[{"award":["R01HD082314"],"award-info":[{"award-number":["R01HD082314"]}],"id":[{"id":"https:\/\/ror.org\/04byxyr05","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R37HD019938"],"award-info":[{"award-number":["R37HD019938"]}],"id":[{"id":"https:\/\/ror.org\/04byxyr05","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000062","name":"National Institute of Diabetes and Digestive and Kidney Diseases","doi-asserted-by":"crossref","award":["R01DK106193"],"award-info":[{"award-number":["R01DK106193"]}],"id":[{"id":"10.13039\/100000062","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,12,2]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:sec>\n                  <jats:title>Background<\/jats:title>\n                  <jats:p>Obesity and type 2 diabetes mellitus (T2D) are major risk factors for male hypogonadism, a disorder with multi-system impacts on health. However, the mechanisms underlying obesity-associated male secondary hypogonadism remain poorly understood. Here, we aimed to dissect dysfunction of the hypothalamic\u2013pituitary\u2013testicular (HPT) axis and elucidate underlying epigenetic mechanisms, using a high-fat diet mouse model.<\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Methods<\/jats:title>\n                  <jats:p>Male C57BL6 mice were fed standard chow or high-fat diet (HFD, 60% fat) for 16 weeks starting at age 6 weeks. Plasma testosterone levels, sperm counts, and gonadotropin responses to senktide and kisspeptin stimulation were assessed. HFD-induced transcription changes in the hypothalamic arcuate nucleus (ARC) were evaluted using bulk and single-cell RNA-sequencing. Genome-wide changes in 5-hydroxymethylcytosine (5hmC) were analyzed by hydroxymethyl-DNA immunoprecipitation sequencing (hMeDIP-seq). Functional relevance of 5hmC changes was evaluated by ectopic TET expression in an immortalized ARC Kiss1 neuron cell line and RT-qPCR.<\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Results<\/jats:title>\n                  <jats:p>\n                    HFD-fed mice developed obesity, hyperglycemia, glucose intolerance, and insulin resistance, indicative of the development of a type 2 diabetes-like metabolic disorder. This was accompanied by low testosterone, reduced sperm counts, and unchanged basal luteinizing hormone (LH), consistent with obesity\/T2D-associated male secondary hypogonadism, as observed clinically in humans. Impaired LH responses to senktide, a Kiss1 neuron activator, but not to kisspeptin itself, identified suppressed Kiss1 neuron function as a key mechanism underlying secondary hypogonadism. RNA-seq analysis revealed dysregulation of metabolic and neural pathways in Kiss1 neurons. hMeDIP-seq demonstrated widespread 5hmC alterations in the ARC of HFD-induced obese\/diabetic mice, correlated with dysregulation of fatty acid metabolism, neuronal activity, and synapse function pathways. Ectopic TET expression\n                    <jats:italic>ex vivo<\/jats:italic>\n                    in Kiss1 neuronal cell lines restored 5hmC levels and upregulated key metabolic and neuronal genes that were repressed in the ARC of DIO mice\n                  <\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Conclusion<\/jats:title>\n                  <jats:p>Our findings demonstrate that Kiss1 neurons are highly sensitive to diet and metabolic changes, and that obesity\/diabetes-induced 5hmC modifications play a key role in dysregulating metabolic and neuronal pathways in Kiss1 neurons. These findings reveal a novel mechanism linking metabolic disturbances to reproductive dysfunction, through direct effects on Kiss1 neurons.<\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Graphic Abstract<\/jats:title>\n                  <jats:fig id=\"ufig1\" position=\"float\" orientation=\"portrait\" fig-type=\"figure\">\n                    <jats:graphic xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"691258v1_ufig1\" position=\"float\" orientation=\"portrait\"\/>\n                  <\/jats:fig>\n                <\/jats:sec>","DOI":"10.64898\/2025.11.30.691258","type":"posted-content","created":{"date-parts":[[2025,12,8]],"date-time":"2025-12-08T12:45:56Z","timestamp":1765197956000},"source":"Crossref","is-referenced-by-count":1,"title":["High-Fat Diet Induces Epigenetic and Metabolic Changes in Kisspeptin Neurons in Association with Obesity and Male Secondary Hypogonadism"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3040-2903","authenticated-orcid":false,"given":"Rui","family":"Fang","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rona S.","family":"Carroll","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8237-0704","authenticated-orcid":false,"given":"Ursula B.","family":"Kaiser","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2025120800373034000_2025.11.30.691258v1.1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1742-1241.2006.00992.x"},{"key":"2025120800373034000_2025.11.30.691258v1.2","doi-asserted-by":"publisher","DOI":"10.1530\/EJE-12-0890"},{"key":"2025120800373034000_2025.11.30.691258v1.3","doi-asserted-by":"publisher","DOI":"10.1210\/jc.2011-0118"},{"key":"2025120800373034000_2025.11.30.691258v1.4","doi-asserted-by":"publisher","DOI":"10.1016\/S0140-6736(20)32667-2"},{"key":"2025120800373034000_2025.11.30.691258v1.5","doi-asserted-by":"publisher","DOI":"10.1016\/s0140-6736(25)00355-1"},{"issue":"8","key":"2025120800373034000_2025.11.30.691258v1.6","doi-asserted-by":"crossref","first-page":"407","DOI":"10.1038\/s41574-020-0363-7","article-title":"Metabolic regulation of kisspeptin - 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