| Mesenchymal stem cells(MSCs)have a significant promise for regenerative medicine for the self-renewal and multipotent capacity.MSCs can be isolated from bone marrow,adipose tissue,umbilical cord tissue.Fetal MSCs are a new potential source of MSCs.Compared with adult MSCs,fetal MSCs exhibit lower immunogenicity,higher proliferation and differentiation potential.FDMSCs are derived from accidental aborted fetus,and they are thought as the main functional cells involved in scarless wound healing.Furthermore,owing to the histological origin of FDMSCs,they may deserve unique properties on skin regeneration.In summary,FDMSCs are better candidates than adult MSCs in wound healing.Recent literature suggests that the regenerative effect of MSCs is mainly mediated through paracrine signaling to regulate recipient cells.MSCs are high secretive capacity cells which can secrete lots of biologically active paracrine molecules including cytokines,chemokines,growth factors and exosomes.Accumulating evidences showed that the tissue repairing effect of MSCs is mediated by exosomes.In the last decades,researchers have shown increased interests in exosomes.Exosomes are 30-150nm small membranous vesicles secreted by most cell types.There are nuclear acids,lipids,and proteins in exosomes,and their main function is to transfer bioactive molecular in cell-cell communication.Exosomes participate in multiple physiological and pathological processes by regulating cell migration,invasion,gene expression,angiogenesis and immune response.Moreover,recent studies have shown the role of exosomes in disease diagnosis and drug delivery.Exosomes derived from MSCs(FDMSC-Ex)are involved in the acceleration of wound healing by activating endogenous repairing process,regulating inflammation,promoting angiogenesis and altering the target cell status.However,the molecular mechanism of promoting wound healing by MSC exosomes is still not clear.Therefore,we used a promising MSCs type,FDMSCs,to investigate the effect of their exosomes on cutaneous wound healing in vivo.Then we applied the FDMSC-Ex on adult dermal fibroblasts in vitro,and analyzed the proliferation,migration and secreting capacity of adult dermal fibroblasts.Then the Notch signal pathway associated to wound healing process was detected to illustrate the underlying mechanism of the FDMSC-Ex in promoting wound healing.We found that Notch signaling is activated after incubating with FDMCS-Ex;and in FDMSC-Ex,the ligands of the Notch pathway were undetectable except for Jagged 1,and the results of Jagged 1 mimic by peptide and knockdown by siRNA suggested that Jagged 1 may lead the activation of the Notch signal in ADFs.Our results suggest that FDMSC-Ex can provide a useful tool to design treatment strategies for unhealed wound and large area burn wound.Research purposesTo isolate FDMSC-Ex and investigate the molecular mechanism of the wound-healing promoting effect of FDMSC-Ex.Research methods1)Isolation and identification of FDMSC-Ex.FDMSC-Ex were isolated using ExoQuick-TC kit(SBI,USA)following the instruction.The morphology of FDMSC-Ex was identified by transmission electron microscope and the marker protein(CD63,Alix,and Tsg101)and the diameter of FDMSC-Ex were detected.2)Animal assay.Full thickness 10mm×10mm dermal wounds were created in the skin on the back of the mouse.The wounds were treated with 200?l FDMSC-Ex(1mg/ml)at the edge of the wound.The wounds were evaluated at day 7/14 after surgery and the wound tissues were collected for histopathological study.3)Exosome labeling.Exosomes were labeled with PKH26 and incubated with ADFs for 24 hours.Then the cells were observed by fluorescence microscope after cell nucleus dyeing by DAPI.4)Cell proliferation.After incubating with FDMSC-Ex,CCK-8 assay was performed to evaluate the AFDs proliferation.5)Cell migration.After incubating with FDMSC-Ex,the migration of ADFs was observed by wound healing and transwell assay.6)ECM protein synthesis.The ECM protein synthesis ability was evaluated by RT-PCR after incubating with FDMSC-Ex.7)Notch signal pathway.The protein expression levels of Notch signal were evaluated by Western blot.To identify the function of Notch pathway in the wound healing process,Notch was blocked by the specific inhibitor,DAPT,and then the migration and proliferation of ADFs were detected again.8)The biological function analysis of Jagged 1 in FDMSC-Ex.Jagged 1 mimic by peptide and knockdown by siRNA were performed to detect the functional role of Jagged 1 in FDMSC-Ex.Research results1.FDMSC-Ex were isolated and identified.2.FDMSC-Ex can promote cutaneous wound healing in vivo.3.FDMSC-Ex internalization by ADFs.4.FDMSC-Ex enhanced proliferation,migration,and ECM synthesis capacity of ADFs.5.FDMSC-Ex activated Notch signaling pathway and DAPT can partly block the promoting effect of FDMSC-Ex on ADFs proliferation and migration.6.Jagged 1 in FDMSC-Ex can promote wound healing by activating Notch signaling.Conclusion1.FDMSC-Ex can promote cutaneous wound healing in vivo.2.FDMSC-Ex enhanced proliferation,migration,and ECM synthesis capacity of ADFs.3.FDMSC-Ex activated Notch signaling pathway and DAPT can partly block the promoting effect of FDMSC-Ex on ADFs proliferation and migration.4.Jagged 1 in FDMSC-Ex can activate Notch signaling,enhance proliferation and migration of ADFs to promote wound healing. |
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