Towards therapeutic control in cutaneous wound healing

Scheper, R.J. [Promotor]Gibbs, S. [Copromotor

Autocrine Regulation of Re-Epithelialization After Wounding by Chemokine Receptors CCR1, CCR10, CXCR1, CXCR2, and CXCR3.

This study identifies chemokine receptors involved in an autocrine regulation of re-epithelialization after skin tissue damage. We determined which receptors, from a panel of 13, are expressed in healthy human epidermis and which monospecific chemokine ligands, secreted by keratinocytes, were able to stimulate migration and proliferation. A reconstructed epidermis cryo(freeze)-wound model was used to assess chemokine secretion after wounding and the effect of pertussis toxin (chemokine receptor blocker) on re-epithelialization and differentiation. Chemokine receptors CCR1, CCR3, CCR4, CCR6, CCR10, CXCR1, CXCR2, CXCR3, and CXCR4 were expressed in epidermis. No expression of CCR2, CCR5, CCR7, and CCR8 was observed by either immunostaining or flow cytometry. Five chemokine receptors (CCR1, CCR10, CXCR1, CXCR2, and CXCR3) were identified, the corresponding monospecific ligands (CCL14, CCL27, CXCL8, CXCL1, CXCL10, respectively) of which were not only able to stimulate keratinocyte migration and/or proliferation but were also secreted by keratinocytes after introducing cryo-wounds into epidermal equivalents. Blocking of receptor–ligand interactions with pertussis toxin delayed re-epithelialization, but did not influence differentiation (as assessed by formation of basal layer, spinous layer, granular layer, and stratum corneum) after cryo-wounding. Taken together, these results confirm that an autocrine positive-feedback loop of epithelialization exists in order to stimulate wound closure after skin injury

Histatins enhance wound closure with oral and non-oral cells

The role of human saliva in oral wound-healing has never been fully elucidated. We previously demonstrated that parotid-salivary histatins enhance in vitro wound closure. The question remains whether other salivary-gland secretions enhance wound closure, and also the effects of histatins on primary and non-oral cells. Since the presence of histatins is not limited to parotid saliva, we expected to observe wound-closure activity of other salivary-gland secretions. However, here we show that non-parotid saliva does not stimulate wound closure, most probably due to the presence of mucins, since the addition of MUC5B to parotid saliva abolished its effect. Furthermore, we found that histatins stimulated wound closure of (primary) cells of both oral and non-oral origin. This suggests that the cellular receptor of histatins is widely expressed and not confined to cells derived from the oral cavity. These findings encourage the future therapeutic application of histatins in the treatment of all kinds of wounds

Structure-activity analysis of histatin, a potent wound healing peptide from human saliva: cyclization of histatin potentiates molar activity 1000-fold

Wounds in the mouth heal faster and with less scarification and inflammation than those in the skin. Saliva is thought to be essential for the superior oral wound healing, but the involved mechanism is still unclear. We have previously discovered that a human-specific peptide, histatin, might be implicated in the wound-healing properties of saliva. Here we report that histatin enhances reepithelialization in a human full-skin wound model closely resembling normal skin. The peptide does not stimulate proliferation but induces cell spreading and migration, two key initiating steps in reepithelialization. Activation of cells by histatin requires a G-protein-coupled receptor that activates the ERK1/2 pathway. Using a stepwise-truncation method, we determined the minimal domain (SHREFPFYGDYGS) of the 38-mer-parent peptide that is required for activity. Strikingly, N- to C-terminal cyclization of histatin-1 potentiates the molar activity similar to 1000-fold, indicating that the recognition of histatin by its cognate receptor requires a specific spatial conformation of the peptide. Our results emphasize the importance of histatin in human saliva for tissue protection and recovery and establish the experimental basis for the development of synthetic histatins as novel skin wound-healing agents.-Oudhoff, M. J., Kroeze, K. L., Nazmi, K., van den Keijbus, P. A. M., van 't Hof, W., Fernandez-Borja, M., Hordijk, P. L., Gibbs, S., Bolscher, J. G. M., Veerman, E. C. I. Structure-activity analysis of histatin, a potent wound healing peptide from human saliva: cyclization of histatin potentiates molar activity 1000-fold. FASEB J. 23, 3928-3935 (2009). www.fasebj.or