Noninvasive Assessment of Progenitor Cell Persistence in Wound Beds of Immunocompetent Mice
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This suggested that while they may have a functional role in the initial stages of wound healing, it does not presumably involve proliferation because the signal is decreased rather than propagated through the wound. Alternatively, the bioluminescent cells that were grafted directly into the wound bed may have migrated to other organ systems and locations. To evaluate this possibility, the grafted animals were sacrificed by cervical ligation and the organs imaged individually ex vivo. We did not observe any differences in the bioluminescence of the internal organs between control and transplanted animals (data not shown). Because the experimental approach measures the expression of luciferase activity as a surrogate for transplanted cells, it is possible that the transplanted BMSC lose their ability to express the transgene due to terminal differentiation or gene loss. Finally, cell fusion between host and transplanted cells may abrogate the luciferase signal. PCR and more invasive detection techniques like immunostaining can evaluate the fate and characteristics of the transplanted cells that persist, but they do not provide kinetic information. Together with quantitative noninvasive imaging, it may be possible to manipulate BMSCs to facilitate wound healing, and optimize their contribution to the kinetics of repair and regeneration.
Most previous research evaluating the fate of stem and progenitor cells transplanted into the wound bed was performed in model systems like chimeric mice, immunodeficient hosts, or undifferentiated stem cells that were specifically designed to minimize donor cell rejection.2,4,7–11 This is the first study demonstrating a specific and quantifiable example of the persistence of progenitor cells in the wound bed in immunocompetent hosts. While we show that only a small portion of the BMSCs remain in the wound bed, they are indeed relatively long-lasting and point to the need for further assessments of their long term contribution to repair and regeneration in the wound bed. The specificity of the model can also be extended by deploying more cell-type specific promoters other than actin to monitor specific responses in the wound bed. Because the only significant bioluminescence signal detected was emitted from the transplanted cells, there is an excellent signal: noise ratio compared to non-transplantation models and noninvasive imaging enables detection, quantification and localization of discrete changes in the expression of the luciferase reporter.
In the case of chronic wounds, large acute wounds and burns, there is a significant unmet need to develop alternative strategies for synthetic grafts that incorporate growth factors, genes and progenitor cells so as to yield a clinically significant improvement in wound healing.12–15 In the experiments here, we focused on the question of stem cell persistence and the deployment of its noninvasive quantitation in the wound bed. Several years ago, Falanga and Sabolinski16 demonstrated that the presence of fetal allogenic skin cells within a synthetic skin-substitute matrix could accelerate wound healing. Indeed, using traditional methods such as in situ hybridization or fluorescent cell labeling to track donor cells in skin wound healing, these models have shown that donor cells persist in the wound bed in some cases for up to 2.5 years.1,3 Similarly, tagging of BMSC with a number of different reporters (ie, green fluorescent protein, lac Z, and firefly luciferase) can be used to monitor their incorporation and differentiation in the wound bed.7,17–19 While these studies demonstrated that dermal fibroblasts and bone marrow-derived cells can be tracked as they incorporated into the wound bed, they all required ex vivo tissue analyses at multiple time-points over the duration of the study.