Domingues, MarcoGomes, BárbaraHollmann, AxelSantos, Nuno C.2022-09-062022-09-062021Int J Mol Sci. 2021 Mar 4;22(5):25741661-6596http://hdl.handle.net/10451/54333© 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).Cholesterol is responsible for the plasticity of plasma membranes and is involved in physiological and pathophysiological responses. Cholesterol homeostasis is regulated by oxysterols, such as 25-hydroxycholesterol. The presence of 25-hydroxycholesterol at the membrane level has been shown to interfere with several viruses' entry into their target cells. We used atomic force microscopy to assess the effect of 25-hydroxycholesterol on different properties of supported lipid bilayers with controlled lipid compositions. In particular, we showed that 25-hydroxycholesterol inhibits the lipid-condensing effects of cholesterol, rendering the bilayers less rigid. This study indicates that the inclusion of 25-hydroxycholesterol in plasma membranes or the conversion of part of their cholesterol content into 25-hydroxycholesterol leads to morphological alterations of the sphingomyelin (SM)-enriched domains and promotes lipid packing inhomogeneities. These changes culminate in membrane stiffness variations.eng25-hydroxycholesterolAtomic force microscopyCholesterolForce spectroscopySupported lipid bilayersjournal article10.3390/ijms220525741422-0067