Taken together, these data suggest that AMMs possess considerable therapeutic potential for chronic wounds through the secretion of angiogenic factors and enhanced engraftment/differentiation capabilities.Ĭitation: Kim S-W, Zhang H-Z, Guo L, Kim J-M, Kim MH (2012) Amniotic Mesenchymal Stem Cells Enhance Wound Healing in Diabetic NOD/SCID Mice through High Angiogenic and Engraftment Capabilities. Notably, transplanted AMMs exhibited high engraftment rates and expressed keratinocyte-specific proteins and cytokeratin in the wound area, indicating a direct contribution to cutaneous closure. AMM transplantation significantly promoted wound healing and increased re-epithelialization and cellularity.
NOD SCID MICE SKIN
Diabetic mice were generated using streptozotocin and wounds were created by skin excision, followed by AMM transplantation. In vitro scratch wound assays also showed that AMM-derived conditioned media (CM) significantly accelerated wound closure. Quantitative real-time PCR and ELISA results revealed that the angiogenic factors, IGF-1, EGF and IL-8 were markedly upregulated in AMMs when compared with adipose-derived mesenchymal stem cells (ADMs) and dermal fibroblasts. In this study, we evaluated the therapeutic potential of AMMs using a diabetic mouse wound model. Cells were cultured either in serum-free media as previously described (6) or in media supplemented with 10% fetal calf serum.Although human amniotic mesenchymal stem cells (AMMs) have been recognised as a promising stem cell resource, their therapeutic potential for wound healing has not been widely investigated. The levels of engraftment were dependent on the injected cell dose, the duration of the experiment, and the source of human CD34 + cells. Percent engraftment always indicates the percent of either human DNA or of human CD45 cells in the mouse bone marrow. Southern (DNA) blot analysis with a human-specific α satellite probe and human-specific progenitor assays were done as previously described (5, 6). NOD/SCID, and NOD/SCID β 2-microglobulin knockout (20) mice were bred and maintained under defined flora in intraventilated cages and transplanted by injection into the tail vein after sublethal (375R) irradiation according to established protocols (5, 6) approved by the Weizmann animal ethics committee. Natural killer cells differentiated into mature CD56 + cells after incubation with human SCF (100 ng/ml) and human IL-15 (100 ng/ml, R&D Systems) for 10 days. Human lymphoid and myeloid cells were immunostained with anti-CD45 (Immuno Quality Products, Groningen, Netherlands), anti-CD19, and anti-CD56 (Coulter). Control cells were incubated with anti-CD34. Polyclonal anti–SDF-1 (10 μg per mouse, R&D Systems) was injected intravenously with the cells (2 × 10 5 cells per mouse) and 24 hours later injected again intraperitoneally. CXCR4 expression was always analyzed by double staining with anti-CD34. The sources for the reagents are as follows: PMA (100 ng/ml), was purchased from Sigma, stem cell factor (SCF) and IL-6 (50 ng/ml) from R&D Systems, and antibodies to CXCR4 from Pharmingen or R&D Systems (10 μg per 2 × 10 5 cells). Percents in the results represent percent of initial 2 × 10 5 cells in the migrating and nonmigrating cell fractions. SDF-1 (125 ng/ml, R&D Systems) transmigration assays were done as previously described (13) with 2 × 10 5 CD34 + cells. CD34 + enrichment, flow cytometry, and fluorescence-activated cell sorting (FACS) were performed as previously described (5, 6). Differences in the results are due to the different CD34 + cell sources (cord blood, bone marrow, and mobilized peripheral blood). In all experiments, samples of the same initial cell pool were compared. Human cells were obtained after informed consent according to procedures approved by the Weizmann Committee.
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