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However, methodologies that can be applied to achieve this can vary greatly, leading to a limited type of information. Thus, the accuracy, robustness, reliability and potential (human) translation are highly reliant on the experimental design and selected methodological models. This review presents an update exploring the main features, advantages and disadvantages of most important pre-clinical models used at the present time to study the effects of bioactive compounds on metabolic diseases. Moreover, future challenges in developing new methods are also depicted. In vitro models (enzyme assays and standard two-dimensional cultures of adipocytes, skeletal muscle cells) are intrinsically well established and constitute the first choice and most widely used methods to study bioactive compounds in metabolic diseases. However, novel models such as three-dimensional cultures (spheroids, organoids) are also starting to emerge and complement traditional culture systems. Models of small organisms (\n                    <jats:italic>C. elegans<\/jats:italic>\n                    ,\n                    <jats:italic>D. melanogaster<\/jats:italic>\n                    ) and non-mammal vertebrates (\n                    <jats:italic>D. rerio<\/jats:italic>\n                    ) represent a scientific advantage and a middle-step before traditional mammalian models (rats and mice). This article provides extensive information and a critical overview of a wide range of methods that represent present and future avenues towards a further understanding of metabolic diseases. Combining and developing new methods will be key for future progression on the effects of bioactive compounds on metabolic diseases, as well as to minimize the use of mammalian models due to ethical reasons.\n                  <\/jats:p>","DOI":"10.1007\/s13105-025-01130-6","type":"journal-article","created":{"date-parts":[[2025,10,24]],"date-time":"2025-10-24T09:39:04Z","timestamp":1761298744000},"page":"1225-1255","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Preclinical research in obesity-associated metabolic diseases using in vitro, multicellular, and non-mammalian models"],"prefix":"10.1007","volume":"81","author":[{"given":"Paula","family":"Aranaz","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marina","family":"Clavel-Millan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katherine","family":"Gil-Cardoso","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maitane","family":"Gonz\u00e1lez-Arceo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"Hern\u00e1ndez-Gonz\u00e1lez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Francisco","family":"Les","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"J\u00e9r\u00f4me","family":"Salles","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ez-Zoubir","family":"Amri","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jos\u00e9 M.","family":"Arbones-Mainar","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Claude","family":"Atgi\u00e9","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fr\u00e9d\u00e9ric","family":"Capel","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Arnaud","family":"Courtois","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xavier","family":"Escot\u00e9","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar\u00eda Jos\u00e9","family":"Garc\u00eda-Barrado","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"St\u00e9phanie","family":"Krisa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"V\u00edctor","family":"L\u00f3pez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ferm\u00edn I.","family":"Milagro","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar\u00eda P.","family":"Portillo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Silvia","family":"Lorente-Cebri\u00e1n","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,10,24]]},"reference":[{"key":"1130_CR1","doi-asserted-by":"publisher","first-page":"671","DOI":"10.1007\/s00441-013-1655-1","volume":"354","author":"V Aas","year":"2013","unstructured":"Aas V, Bakke SS, Feng YZ, Kase ET, Jensen J, Bajpeyi S, Thoresen GH, Rustan AC (2013) Are cultured human myotubes far from home? 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