(P18-027-24) Phosphatidic Acid (16:0 - 18:1) Mediates Lipocalin 2 Regulation of mTOR Signaling Activity in Adipose Stromal Vascular Cells

Objectives: Phosphatidic acid (PA) is known as the key intermediate in the synthesis pathway of phospholipids, it also has a notable role in the regulation of mTOR, the nutrient-sensing signaling pathway that is important for cellular metabolism and tissue growth and development, while PA’s function in aging and cellular senescence is not well defined. In previous studies, we discovered that lipocalin 2 (LCN2), a protein secreted by adipocytes, functions as the PA binding protein; Lcn2 deficiency was found to increase the risk for diet-induced obesity and diabetes and to reduce adipose tissue levels of certain PA species, including PA (16:0 - 18:1). In this study, we aimed to investigate the role of PA in mediating LCN2 regulation of mTOR signaling pathway.

Methods: Stromal vascular (SV) cells were isolated from inguinal white adipose tissue of wild type (WT) and Lcn2 knockout (KO) mice. SV cells were allocated to control, PA, mouse recombinant (mr) LCN2, and PA+LCN2 groups and they were treated with their corresponding intervention for 2 days. RNA and protein were extracted from cells and culture medium, both gene and protein expressions were analyzed.

Results: We observed the Lcn2 deficiency increased senescence in inguinal SV cells, as β-galactosidase staining exhibited an increased number of senescent cells in the KO group. Gene expression of several senescence markers, p16, Mcp1, and C-ebpb, had significantly upregulated in Lcn2 deficiency, while Igfbp7 had a trend of upregulation. TGFβ, a protein marker of senescence-associated secretory phenotype (SASP), and the mTOR signaling components were also upregulated in Lcn2 KO cells. We found that PA-treated cells showed reduced protein expression of mTOR signaling pathway and TGFβ secretion; the supplementation of mrLCN2 alone has no effect, while the combination of PA and mrLCN2 together induced a significant reversal effect on these markers in Lcn2 KO SV cells.

Conclusions: We demonstrated Lcn2 deficiency led to increased cellular senescence and mTOR signaling pathway activation, while PA countered these effects. This further links PA and LCN2 in the regulation of metabolism and aging in adipose tissue.

Funding Sources: This work was supported by NIDDK Grant (R01 DK123042) awarded to X.C.