Regulatory T cells (Tregs) mediate critical anti-inflammatory actions in the skin, potentially via a wide range of mechanisms. In contact hypersensitivity (CHS)-mediated skin inflammation, Tregs undergo prolonged contact with the endothelium within dermal postcapillary venules and inhibition of Treg/endothelial cell interactions during select phases of CHS leads to increased intravascular adhesion of pro-inflammatory leukocytes and exacerbation of skin inflammation. These observations suggest that the microvascular endothelium is a key site of the anti-inflammatory actions of Tregs. Here we tested this hypothesis by measuring adhesion molecule expression in resting and inflamed skin in the presence and absence of Tregs. In a two-challenge model of oxazolone-induced CHS, imaging of cleared skin revealed that CHS induced upregulation of E-selectin and ICAM-1 but not P-selectin and VCAM-1. These alterations were particularly prominent in venules extending from the hypodermis into the superficial dermis. These changes were associated with upregulation of expression of mRNA for a host of chemokines and cytokines. Acute in vivo ablation of Tregs during the hapten challenge phase of CHS resulted in exacerbated skin inflammation, associated with enhanced expression of E-selectin and ICAM-1 in the dermal microvasculature. Moreover, in skin that was not undergoing hapten challenge, absence of Tregs also resulted in E-selectin upregulation in postcapillary venules. Together these observations demonstrate that the microvascular endothelium is a target of the anti-inflammatory actions of Tregs in the skin, both in hapten-induced CHS and in steady-state skin.