Lipocalin 2 (Lcn2) has previously been characterized as an adipokine/cytokine playing a role in glucose and lipid homeostasis. In this study, we investigate the role of Lcn2 in adipose tissue remodeling during high-fat diet (HFD)-induced obesity. We find that Lcn2 protein is highly abundant selectively in inguinal adipose tissue. During 16 weeks of HFD feeding, the inguinal fat depot expanded continuously, whereas the expansion of the epididymal fat depot was reduced in both wild-type (WT) and Lcn2(-/-) mice. Interestingly, the depot-specific effect of HFD on fat mass was exacerbated and appeared more pronounced and faster in Lcn2(-/-) mice than in WT mice. In Lcn2(-/-) mice, adipocyte hypertrophy in both inguinal and epididymal adipose tissue was more profoundly induced by age and HFD when compared with WT mice. The expression of peroxisome proliferator-activated receptor-γ protein was significantly down-regulated, whereas the gene expression of extracellular matrix proteins was up-regulated selectively in epididymal adipocytes of Lcn2(-/-) mice. Consistent with these observations, collagen deposition was selectively higher in the epididymal, but not in the inguinal adipose depot of Lcn2(-/-) mice. Administration of the peroxisome proliferator-activated receptor-γ agonist rosiglitazone (Rosi) restored adipogenic gene expression. However, Lcn2 deficiency did not alter the responsiveness of adipose tissue to Rosi effects on the extracellular matrix expression. Rosi treatment led to the further enlargement of adipocytes with improved metabolic activity in Lcn2(-/-) mice, which may be associated with a more pronounced effect of Rosi treatment in reducing TGF-β in Lcn2(-/-) adipose tissue. Consistent with these in vivo observations, Lcn2 deficiency reduces the adipocyte differentiation capacity of stromal-vascular cells isolated from HFD-fed mice in these cells. Herein Rosi treatment was again able to stimulate adipocyte differentiation to a similar extent in WT and Lcn2(-/-) inguinal and epididymal stromal-vascular cells. Thus, combined, our data indicate that Lcn2 has a depot-specific role in HFD-induced adipose tissue remodeling.