Abstract
Recent reports have indicated prolidase (PEPD) as a ligand of the epidermal growth factor receptor (EGFR). Since this receptor is involved in the promotion of cell proliferation, growth, and migration, we aimed to investigate whether prolidase may participate in wound healing in vitro. All experiments were performed in prolidase-treated human keratinocytes assessing cell vitality, proliferation, and migration. The expression of downstream signaling proteins induced by EGFR, insulin-like growth factor 1 (IGF-1), transforming growth factor β1 (TGF-β1), and β1-integrin receptors were evaluated by Western immunoblotting and immunocytochemical staining. To determine collagen biosynthesis and prolidase activity radiometric and colorimetric methods were used, respectively. Proline content was determined by applying the liquid chromatography coupled with mass spectrometry. We found that prolidase promoted the proliferation and migration of keratinocytes through stimulation of EGFR-downstream signaling pathways in which the PI3K/Akt/mTOR axis was involved. Moreover, PEPD upregulated the expression of β1-integrin and IGF-1 receptors and their downstream proteins. Proline concentration and collagen biosynthesis were increased in HaCaT cells under prolidase treatment. Since extracellular prolidase as a ligand of EGFR induced cell growth, migration, and collagen biosynthesis in keratinocytes, it may represent a potential therapeutic approach for the treatment of skin wounds.
Highlights
Wound healing is a physiological process restoring skin functionality
We found that phosphorylation of epidermal growth factor receptor (EGFR) instantly entailed phosphorylation of Akt and extracellular signal-regulated kinase (ERK) in response to PEPD treatment
Whereas these signaling proteins were activated rapidly, activation of mammalian target of rapamycin (mTOR) was slower, as p-mTOR reached a maximal level at 1 h after treatment with PEPD. These results demonstrated that prolidase could elicit EGFR transactivation leading to sustained phosphorylation of Akt/mTOR and MAPK signaling pathways in keratinocytes
Summary
Wound healing is a physiological process restoring skin functionality. It consists of four phases occurring in proper time and order. This process starts from hemostasis, inflammation followed by proliferation and eventually tissue remodeling. The sequence of events during wound healing is strictly programmed and any disturbances may impair normal tissue repair. In the microenvironment of the damaged tissue, there are numerous biological factors and cell types involved in this process. One of them is keratinocytes, which proliferate and migrate to the wounded area during the proliferative phase induced by growth factors [1]
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