The Effect Of Substance P On Differentiation And Migration Of Epidermal Stem Cells In Wound Healing

Wound healing is a complex process in which growth factors secreted by repair cells plays an important role on promoting the repair process. Mean while neuropeptide released from nerve ending, such as SP (substance P), CGRP (calcitonin gene-related peptide), has a closed relationship with wound healing. The mechanism of neuropeptide promoting wound healing has been considered as switching effect on neutrogena inflammation in wound and promoting the DNA synthesis by repair cells as yet. It is revealed now that the ESC (epidermal stem cells) is the source of skin wound healing and appendage regeneration. The niche of ESC is very important in modulating ESC directed differentiation and skin appendages reformation in wound healing. Neuropeptide is an important factor in ESC niche. We speculate that neuropeptide can guide the differentiation and migration of ESC in wound healing. If the relationship between neuropeptide and ESC is found, a new therapeutic method, applying neuropeptide to influence ESC differentiation, will be established for promoting wound healing.In the present study, we observed the expression of NK1-receptor (specific receptor for SP) on ESC membrane in rat's skin at different development stage. The rats with full thickness skin injury on back were used to observe the differentiation and migration of ESC in wound healing. All rats were random divided into following three groups in present experiment: ①SP group, rats in this group were injected SP in the middle of the wound postinjury. ②capsaicin group, rats in this group were subcutaneously injected capsaicin pre-injury, in order to damage sensory neuron and block the secretion of neuropeptides. ③Control group, rats in this group were not given SP or capsaicin. Based on slow cell cycling and label restraining of ESC, we use BrdU, a nuclei maker, as the trace maker to observe ESC migration in wound healing. In order to find the effect of SP on the differentiation of ESC, the experiment in vitro was carried out. The cultured ESC was stimulated by SP and the differentiation was observed by using flow cytomete. ESC marker (K19 and β1 integrin) and TAC marker (K14 and K5) was used to identify ESC and TAC (transient amplifying cell).Our studies in vivo show that there is positive staining of NK1-receptor on ESCmembrane in rat's skin at different development stage (fetal rat, new born rat and mature rat). Compared with control group, the healing speed in SP group is faster, and that in capsaicin group is much lower. BrdU positive cells, which was proved to be ESC with double immunohistochemistry staining (K19 antibody andβ1integrin antibody), existed in the skin border near the wound and granulation tissue in SP group after skin injury, but the BrdU positive cell were found only in the wound border in control group and capsaicin group. The BrdU positive cells can be found on the 3rd day postinjury in the SP group and control group, but in capsaicin group, they can be found until the 12th day postinjury. The proportion of BrdU positive cells to all observed cells in SP group is obviously higher than that in control group and capsaicin group. The culture method for ESC was improved in the present study and the effect of SP on differentiation of cultured ESC was found. There are positive K14-K5 double immunohistochemistry staining cells appeared in the cultured ESC 24 hours after SP was added into culture media. This means ESC develop into TAC under the stimulation by SP in our experiment. The results suggested that: 1st, there is NK1-receptor on the membrane of ESC, this is the molecular foundation of effect of SP on the stem cells. 2nd, SP obviously promote the wound healing and shorten the healing duration, this may be contribute to the effect of SP on ESC migration to a definite location after injury. 3rd, ESC can develop into the differentiation state under the stimulation by SP.In summary, the neuropeptide SP induce ESC migration and come into differentiation state, this may be the molecular mechanisms for SP promoting wound he...