Previous studies have shown that inflammation is a key factor in determining scar formation in cutaneous fetal wounds. In a fetal mouse model, sterile wounds generated on embryonic day 15 (E15) have little to no inflammation and heal scarlessly, while wounds generated on embryonic day 18 (E18) have inflammation and heal with a scar. The mechanisms leading to age-related differences in inflammation and healing are not fully understood.
Alarmins, which comprise a group of endogenous danger signals, have been implicated in promoting inflammation in various organ systems and disease processes. This study evaluates the role of a specific alarmin, high mobility group box 1 (HMGB-1), in cutaneous fetal wound healing. HMGB-1 is a non-histone architectural DNA binding protein localized within the nucleus of the cell. However, it can be released by both passive and active mechanisms. Passive release into the extracellular matrix occurs as a result of cell injury or necrosis. While active secretion is described as a capability of several cell types, including immune cells. Once in the extracellular space, HMGB-1 stimulates inflammation by binding to RAGE (receptor for advanced glycation end products) or toll-like receptors present on inflammatory cells.
This is the first study to evaluate the role of HMGB-1 in the process of fetal wound healing. We hypothesized that alterations in the expression or release of HMGB-1 between early and late gestation fetal wounds contribute to the differences seen in healing outcomes. To begin, comparisons of HMGB-1 expression in fetal wounds that heal scarlessly and wounds that heal by scar formation were made. Later experiments tested the ability of HMGB-1 to induce scar formation in E15 wounds that have previously been shown to heal scarlessly. Finally, mechanisms of action were explored to decipher how HMGB-1 affects fibroblasts, the cells ultimately responsible for scar formation.
Immunohistochemical staining of E15 and E18 skin revealed age-related differences in HMGB-1 expression patterns in both unwounded and wounded skin. As compared to E15 basal keratinocytes, basal keratinocytes of E18 skin expressed higher nuclear HMGB-1 staining and demonstrated a more substantial loss of nuclear staining after injury, suggesting that HMGB-1 is released to a greater extent in E18 wounds. Furthermore, E15 wounds treated with HMGB-1 could be induced to heal by fibrosis with differences seen in both the amount and quality of collagen present within the scar.
Age-dependent differential release of HMGB-1 from fetal keratinocytes during wound healing and the capability of HMGB-1 to induce scar formation in E15 wounds that naturally heal scarlessly were found. These results support the hypothesis that alterations in expression or release of HMGB-1 contribute to the difference seen in healing outcome between early and late fetal wounds. This study suggests both the timing and degree of extracellular HMGB-1 release are potential factors in determining whether scarless or fibrotic wound healing will result. In light of these findings, HMGB-1 remains a viable therapeutic target for optimizing wound repair.