In vitro modeling of adipogenesis using MSCs from subcutaneous adipose tissue to explore the cellular and molecular mechanisms underlying pediatric obesity and obesogenic memory.

Objective Pediatric obesity is associated with early metabolic complications and increased risk of persistent obesity in adulthood. Beyond fat accumulation, obesity may induce long-lasting molecular alterations in adipose tissue, described as "obesogenic memory." However, whether such intrinsic alterations are already present in pediatric adipose progenitor cells remains poorly understood. This study aimed to develop an in vitro model of adipogenesis using pediatric adipose-derived mesenchymal stem cells (hMSCs) and to investigate whether obesity is associated with intrinsic metabolic reprogramming of adipose progenitors. Methods hMSCs were isolated from periumbilical subcutaneous adipose tissue obtained from pediatric subjects with normal weight (NW), overweight (OW) and obesity (OB). Cells were expanded and induced to undergo adipogenic differentiation for up to 14 days using defined adipogenic media. Differentiation was evaluated through morphological analysis, lipid accumulation (BODIPY staining), gene expression profiling by RT-qPCR, and protein analysis by Western blot. Results Two differentiation protocols efficiently induced adipocyte maturation without cytotoxicity. hMSCs derived from NW and OB subjects showed comparable adipogenic differentiation capacity, with similar lipid droplet accumulation and expression of canonical adipogenic markers including C/EBPα, PPARγ, and FABP4. Despite this comparable differentiation efficiency, OB-derived adipocytes exhibited altered transcriptional regulation of genes involved in lipid metabolism, including pathways associated with lipogenesis, lipolysis, and fatty acid β-oxidation. Protein analyses further revealed dysregulated expression of key metabolic regulators such as SCD1, SREBP1, PNPLA2, and FADS1, suggesting altered lipid metabolic programming. Conclusions Pediatric obesity does not impair adipogenic differentiation but is associated with intrinsic metabolic alterations in adipose progenitor cells. These findings support the presence of an early obesogenic memory in adipose tissue that may contribute to long-term metabolic dysfunction.