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This study investigated the anti-obesity effects and molecular mechanisms of Antrodia cinnamomea alcohol extract (ACE) in 3T3-L1 preadipocytes and a high-fat diet (HFD)-induced obesity mouse model. In vitro, Antrodia cinnamomea alcohol extract significantly inhibited adipocyte differentiation and lipid accumulation in 3T3-L1 cells by downregulating PPAR\u03b3 and C\/EBP\u03b1, while activating the AMPK pathway and suppressing MAPK signaling. In vivo, Antrodia cinnamomea alcohol extract administration reduced body weight, adipose tissue mass, and liver lipid accumulation in high-fat diet-fed mice, ameliorating non-alcoholic fatty liver disease (NAFLD) symptoms. Transcriptomic analysis of adipose tissue revealed that Antrodia cinnamomea alcohol extract modulated key gene expression profiles related to fatty acid metabolism and adipogenesis, suppressing lipid synthesis while enhancing \u03b2-oxidation. Furthermore, Antrodia cinnamomea alcohol extract rebalanced gut microbiota, increasing beneficial bacterial populations such as Akkermansia and Bifidobacterium, while reducing pro-inflammatory Escherichia-Shigella species. These findings demonstrate that Antrodia cinnamomea alcohol extract exerts multifaceted anti-obesity effects by regulating lipid metabolism, adipogenesis pathways, and gut microbiota composition, highlighting its potential as a natural therapeutic agent for obesity management.<\/jats:p>","DOI":"10.3390\/ijms26125856","type":"journal-article","created":{"date-parts":[[2025,6,19]],"date-time":"2025-06-19T06:11:07Z","timestamp":1750313467000},"page":"5856","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Antrodia cinnamomea Extract Attenuates Obesity by Targeting Adipogenic Pathways and Gut Dysbiosis in High-Fat Diet-Fed Mice"],"prefix":"10.3390","volume":"26","author":[{"given":"Kuen-Tze","family":"Lin","sequence":"first","affiliation":[{"name":"Department of Radiation Oncology, Cardinal Tien Hospital, New Taipei City 23148, Taiwan"},{"name":"School of Medicine, College of Medicine, Fu-Jen Catholic University, New Taipei City 24205, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4713-6410","authenticated-orcid":false,"given":"Shih-Yu","family":"Lee","sequence":"additional","affiliation":[{"name":"Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center, Taipei 11490, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lee","family":"Ya-Jy","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, National Defense Medical Center, Taipei 11490, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Po-Jui","family":"Wu","sequence":"additional","affiliation":[{"name":"School of Pharmacy, National Defense Medical Center, Taipei 11490, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tsu-Chung","family":"Chang","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, National Defense Medical Center, Taipei 11490, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wen-Liang","family":"Chang","sequence":"additional","affiliation":[{"name":"School of Pharmacy, National Defense Medical Center, Taipei 11490, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"I-Chuan","family":"Yen","sequence":"additional","affiliation":[{"name":"School of Pharmacy, National Defense Medical Center, Taipei 11490, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,6,18]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"113","DOI":"10.1111\/j.1559-4572.2008.00044.x","article-title":"The obesity, metabolic syndrome, and type 2 diabetes mellitus pandemic: Part I. 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