(WHS-P3.01) INNATE IMMUNE INTERACTIONS DEFINE KEY MECHANISMS OF EXOSOME-INDUCED HEALING OF CHRONIC WOUNDS
Friday, May 17, 2024
10:30 AM – 11:30 AM East Coast USA Time
Chronic wounds, thus far, are managed clinically but are not treated for underlying pathology. Specifically, in the population with diabetes, chronic wounds do not manifest the repair or regenerative properties of skin and thus, are one of the leading causes of non-traumatic lower limp amputations. Major hallmarks of these chronic wounds include neuropathy, peripheral artery defects, and immune dysfunction. We found that local administration of secreted nanoscale vesicles, called exosomes, harvested from cultured primary human bone marrow multipotent stromal cells, accelerate wound closure in a genetic animal model of type 2 diabetic delayed wound healing. The cellular and molecular sequelae of exosome-based treatment, however, remain to be investigated and is at the heart of our study. Cutaneous administration of exosomes at post-operative day 1 (POD1) significantly decreased time to closure and wound burden of type 2 diabetic wounds, in a dose dependent manner compared to PBS-treated control diabetic wounds. In order to characterize exosome-induced healing mechanisms, we employed multiplexed immunofluorescence imaging and single cell RNA sequencing. Our analysis revealed a pro-healing phenotype, likely modulated through interactions between innate immunity and cutaneous vasculature. Immunofluorescence imaging of exosome-treated wounds displayed extensive CD31+ neovascularization in expanded areas of granulation tissue in the diabetic wound bed by POD10, phenocopying typical healing wounds of wildtype mice. We also observed increased presence of undifferentiated monocytes/macrophages and macrophages in the diabetic wound tissue by 1.6-fold and 6.2-fold of that in control wounds, respectively. In vitro studies confirmed that exosomes were capable of modulating primary murine macrophages to a pro-healing-like phenotype. Further confirming our hypothesis of innate immune activation, exosome treatment failed to rescue the delayed healing in clodronate liposome-treated, macrophage-depleted mice. These macrophage-depleted but exosome-treated wounds also lack the development of highly vascularized granulation tissue indicating that macrophage induction is paramount for exosome-induced wound healing. Interestingly, even the endogenous control of macrophage plasticity throughout the sequential phases of skin healing remains elusive even in the context of non-pathological or typical healing wounds. Our results demonstrate that macrophage modulation is at least one avenue through which exosomes mediate wound closure benefit. Our results begin to address the obvious need of studies of cellular events engaged following exosome-induced wound healing, a critical barrier in the safe clinical translation of this tissue engineering approach.