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. 2022 Dec 27;24(1):447. doi: 10.3390/ijms24010447

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© 2022 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 (https://creativecommons.org/licenses/by/4.0/).

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(A) MSC were treated for 21 days with canonical differentiation medium (DM) to achieve a complete osteogenic (upper panel) and adipogenic (lower panel) differentiation evaluated by alkaline phosphatase (ALP) and oil red staining, respectively. This condition served as positive control (Ctrl+) of the experiment. In parallel, MSC were cultured for 21 days in a proliferation medium (PM) and treated every 3 days with EVs collected from children with normal weight (NW EVs), overweight (Ow EVs) and obesity (Ob EVs), and compared to control condition (Ctrl-) with no EV treatment. These samples were also stained for ALP and oil red to assess osteogenic (upper panel) and adipogenic (lower panel) differentiation, respectively. Scale bar: 200 μm. Quantification of the percentage of ALP (B) and oil red (C) staining in MSCs treated with EVs in PM for 21 days. RT-PCR for the expression of osteoblastic master-gene Runx2 (D) and the adipogenic master-gene PPAR-γ (E) in MSCs treated with NW, Ow and Ob EVs for 21 days, normalized versus the housekeeping GAPDH. * p < 0.05; ** p < 0.01.