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Membranes from small antral follicles (2\u20139\u00a0mm) were enzymatically digested and cultured for 48\u00a0h. A mixed cell sample was obtained before MACS separation into ANPEP-positive and ANPEP-negative cell fractions, followed by an additional 48-h culture. Cell purity and viability were assessed by flow cytometry (FACS) using ANPEP and FSHR antibodies. Gene expression (\n                      <jats:italic>ANPEP, CYP17A1, LHCGR, FSHR, CYP19A1<\/jats:italic>\n                      ) was analyzed by RT-qPCR. Steroid secretion (pregnenolone, progesterone, 17-OH-pregnenolone, 17-OH-progesterone, DHEA, androstenedione) was quantified using LC\u2013MS\/MS.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      Before cell sorting, 29.6\u201382.9% of viable cells expressed ANPEP. After MACS,\u2009~\u200968% of cells in the ANPEP-positive fraction expressed ANPEP, compared with\u2009~\u20093% in the negative fraction (\n                      <jats:italic>P<\/jats:italic>\n                      \u2009&lt;\u20090.05).\n                      <jats:italic>CYP17A1<\/jats:italic>\n                      expression was higher in pre-MACS and ANPEP-positive populations than in the negative fraction. ANPEP-positive cells secreted androgen precursors (17-OH-pregnenolone, 17-OH-progesterone, DHEA, androstenedione), whereas the negative fraction produced relatively more progesterone, consistent with theca cell enrichment.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>ANPEP-based MACS enables enrichment of viable, steroidogenic human theca interna cells. The ANPEP-positive fraction exhibits molecular and functional characteristics consistent with theca lineage and provides a useful model for studying ovarian steroidogenesis.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1007\/s10815-026-03983-2","type":"journal-article","created":{"date-parts":[[2026,7,23]],"date-time":"2026-07-23T14:54:11Z","timestamp":1784818451000},"page":"3007-3019","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Isolation of theca cells from human antral follicle membranes using ANPEP-based magnetic cell sorting"],"prefix":"10.1007","volume":"43","author":[{"ORCID":"https:\/\/orcid.org\/0009-0001-3231-435X","authenticated-orcid":false,"given":"Lea Bejstrup","family":"Jensen","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-4552-3147","authenticated-orcid":false,"given":"Malene Louise","family":"Johannsen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jane Alr\u00f8","family":"B\u00f8tkj\u00e6r","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna","family":"Ronnefeld Veng","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kirsten","family":"Tryde Macklon","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anette","family":"T\u00f8nnes Pedersen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bjarne","family":"Styrishave","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stine Gry","family":"Kristensen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,7,23]]},"reference":[{"key":"3983_CR1","doi-asserted-by":"publisher","first-page":"121","DOI":"10.1210\/edrv-17-2-121","volume":"17","author":"A Gougeon","year":"1996","unstructured":"Gougeon A. 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