487-P: Lipocalin-2 Regulates the Function of Phosphatidic Acid in Senescence and mTOR Signaling in Adipose Stromal-Vascular Cells

Introduction and Objective: Phosphatidic acid (PA), a key intermediate in phospholipid synthesis, regulates mTOR, a nutrient-sensing pathway critical for metabolism, growth and development. However, PA's role in aging and cellular senescence, and its regulatory mechanisms, remain unclear. Lipocalin 2 (LCN2), an adipocyte secreted protein, was identified as a PA-binding protein. Lcn2 deficiency is shown to increase the risk of obesity, diabetes, and reduces adipose tissue PA species, including PA 16:0 18:1 (PA1) and PA 16:0 18:2 (PA2). This study explores LCN2's role in PA-mediated regulation of cellular senescence and mTOR signaling. Methods: Stromal-vascular (SV) cells from inguinal white adipose tissue (Ing-WAT) of wild-type (WT) and Lcn2 knockout (KO) mice were treated with PA1, mouse recombinant LCN2 (apoLCN2), or PA1+LCN2 (holoLCN2) for two days. Additionally, WT SV cells were treated with PA1 and PA2. Cellular senescence and mTOR signaling were assessed via β-galactosidase staining, and gene/protein expression analyses. Results: In WT SV cells, PA1 or PA2 significantly reduced β-galactosidase-positive senescent cells as well as downregulated of senescence and senescence-associated secretory phenotype (SASP) marker genes, including p16, Cebpb, Il1b, Il18, and Tnfa. Lcn2 deficiency in Ing SV cells resulted in increased senescence shown in β-galactosidase staining. Gene expression analysis revealed a significant upregulation of senescence markers in Lcn2 KO SV cells. While neither PA1 alone nor apo-LCN2 (LCN2 alone) had a significant effect on mTOR signaling activation, holo-LCN2 markedly enhanced mTOR signaling pathway activation in both WT and Lcn2 KO SV cells. Conclusion: Our findings show PA is protective against cellular senescence, while Lcn2 deficiency exacerbates it by reducing PA levels. LCN2 is essential for the PA-mediated mTOR activation, highlighting the functional interplay between PA and LCN2 in regulating adipose tissue metabolism and aging. Disclosure X. Jin: None. H. Su: None. X. Chen: None. Funding NIDDK Grant (R01 DK123042)