Processing of POMC peptides by dermal microvascular endo‐thelial cells: implication for EC biologic functions and skin inflammation

Abstract

Dermal microvascular endothelial cells (ECs) are both source and target of the pro‐opiomelanocortin (POMC) peptides ACTH and α‐melanocyte‐stimulating hormone (α‐MSH). The availability of neuropeptides as important modulators of innate and adaptive immune responses is controlled by neuropeptide‐specific peptidases such as neutral endopeptidase (NEP) or angiotensin‐converting enzyme (ACE). In this study, we have tested the possibility that NEP or ACE expressed by ECs may influence the local bioavailability of POMC peptides. Incubation of ACTH1−39 with cell membranes prepared from the high NEP‐/low ACE‐expressing microvascular EC line 1 (HMEC‐1) or from low NEP‐/high ACE‐expressing primary human dermal ECs (HDMECs) for 30–480 min resulted in a decrease in ACTH immunoreactivity (IR) over time in membrane supernatants that could be partially blocked with NEP inhibitors as detected by radioimmunoassay. In parallel, α‐MSH IR increased peaking after 60 min. Fragments generated by incubation of HMEC‐1 or HDMEC membranes with ACTH1−39, ACHT1−24 or α‐MSH for 1–120 min were further analysed by mass spectroscopy. HMEC‐1 membranes generated peptide products which could be altered by inhibition of NEP, but not ACE. Likewise, HDMEC membranes fragmented ACTH similar to HMEC‐1 membranes in the presence of NEP inhibitors. Some of the proteins can be assigned to regular proteolytic cleavage, while others seem to be modified. Importantly, HMEC‐1 and HDMEC membranes also slowly degraded α‐MSH, suggesting that EC proteolytic peptidases locally control ACTH/α‐MSH bioavailability, which may be important in controlling cutaneous inflammation.