In the present study, we introduced a new group of small cells, Dot cells that were detected within both of fetal and adult mouse blood. We believe that Dot cells are a previously unidentified group of cells since no other reports have described a similar cell morphology or marker profile. Dot cells strongly express to E-cadherin, integrin β1, CD184. They have weaker to CD34, CD13 and low to Sca1 expression. Because E-cadherin is expressed mainly by epithelial cells, and integrin β1 is a well-known epidermal stem cell marker, the strong expression of both E-cadherin and integrin β1 by Dot cells in unwounded E16.5 mouse skin are likely targeted to epidermal development. We believe that Dot cells are relatively primitive cells or stem cells due to their unusually small size and the fact that they express stem cell markers such as E-cadherin, integrin β1 and CD34.
Our data show that Dot cells have a strong homing effect to sites of injury for tissue regeneration. They specifically migrate to wounds and differentiate into dermal cells, which release less interstitial collagen and reduce scar. We believe that the mechanism for Dot cell migration to the damaged area could be due to the presence of CD184, a seven-transmembrane G-protein coupled receptor, which functions as a receptor for stromal cell-derived factor-1 (SDF-1), an important chemokine for heart development [36
] and stem cell migration [37
]. The release of SDF-1 is upregulated at sites of tissue injury. The increased SDF-1, in turn, leads to the homing of circulating stem cells [38
]. The fibroblast-transplanted group did not show the same scarless repair effect as seen in the Dot cell-transplanted group. This indicates that transplanted fibroblasts do not home to damaged tissue. This phenomenon has also been described by other reporters [39
Our data also show one mechanism of Dot cell tissue repair occurs by differentiation of Dot cells within wounds to dermal and endothelial cells. The differentiation of Dot cells is a rapid process that results the repairing tissues derived all from Dot cells differentiation (). We used Dot cell-transplantation method instead of topic application of Dot cells to the wound bed in our experiments. Because Dot cells are blood-derived and very small, so it is easier and more physiological to apply them through circulation. In addition, blood-transplantation is easier to control the cell numbers without loosing cells that could be happened in wound topic application. One mechanism of Dot cell-induced scarless healing could be their lack of some cytokine expression. For example, the differentiated Dot cells strongly express FGF-2, which function to increase proliferation of the Dot cell-derived fibroblasts. Their rapid proliferation may have anti-differentiation effects, such as reduction of latent local TGF-β, since TGF-β is pro-fibrotic. FGF-2 may function to reduce fibroblasts differentiation into myofibroblasts, the cells that release large amounts of interstitial collagens and induce strong tissue contraction, i.e. resulting scarring. Because all wounds in the present study were made in 6 mm diameter, we only detected only a small of accelerated wound healing by Dot cells. However, we believe that Dot cells can also increase healing speed, and my help in the healing of diabetic wounds. These hypothesis need to be investigated in the future studies.
Dot cells function during early stages of repair, which may be requisite to scarless healing. We observed that the scar on saline control wounds remodels after day 20 when there were no significant differences of scaring between Dot cells and saline injected wounds. The recovery of saline-injected group by 20 days could be explained by host stem cells in saline-injected mice also migrate to the wounds and repair tissue, however, in a slower way, due to the smaller number of stem cells in saline-injected mice compared to the Dot cell-transplanted wounds.
The ratio of Dot cells in E16.5 fetal mouse blood is more than twenty times higher than that of adult mice, suggesting Dot cells may be the key for scarless healing. To determine if the higher Dot cell number enhances tissue repair, we transplanted 500,000 isolated Dot cells into each wounded adult mouse. The Dot cell number for transplantation was calculated as 40% of the total number Dot cells in one recipient animal, i.e. 1.5 (ml of blood in one mouse) × 4 ×109 (total blood cells per ml) × 0.02% (the ratio of adult Dot cells within total blood cells) = 500,000. Our data suggest that a 40% increase in the number of Dot cells can induce scarless healing. The ratio of Dot cells in E16.5 fetal mice is twenty times higher than in adult mice, the high number of Dot cells in fetus maybe the key for scarless wound healing.
In our transplantation model, we found that GFP-labeled Dot cells lose their GFP expression in wounds after the dermal structure is fully restored. This suggests that the self-renew of Dot cells population is well regulated, which effectively limit their proliferation rate in vivo
. Dys-regulation or other damage during self-renewal could cause hyperproliferation of the transplanted stem cells, and thus cause them to become ‘cancer stem cells’ in recipients [40
]. We believe that after wound has healed, Dot cells stay in the circulation in the number as same as the normal unwounded mouse.
Our data provide evidence that Dot cells have a major role in repairing wounded dermis to reduce scar formation. We believe the high number of circulating Dot cells in fetal blood is the key for fetal scarless wound healing. In contrast, scarring in adults could be due to the lower number of Dot cells in blood and BM in adults. However, the mechanism of tissue repair by Dot cells is still unclear and further studies on the skin progenitor effects of Dot cells need to be performed.