Xenotransplantation of Human Cord Blood Derived Unrestricted Somatic Stem Cells Rescue the Blistering Phenotype in a Murine Model of Recessive Dystrophic Epidermolysis Bullosa

Background: RDEB is a severe inherited skin blistering disease caused by mutations in the COL7A1 gene that encodes type VII collagen (C7), a major component in the anchoring fibrils at the basement membrane. Allogeneic stem cell transplantation in both experimental animals and human subjects with RDEB resulted in increased C7 deposition and alleviation of the blistering phenotype in some recipients (Kiuru/Cairo/Christiano et al. 2011). However, there was no distinct anchoring fibril formation in the recipient skin and the effective cell types that contribute to the regeneration in RDEB skin remain to be identified. Our previous studies demonstrated that CB-derived USSCs express C7 and promote epithelialization and wound closure in a murine wound healing model (Liao/Cairo et al. 2013).

Methods: CB-derived USSCs were labeled with GFP/Luc reporter gene and injected in newborn col7a1-/- (RDEB) mice without immunosuppression. The migration of USSCs was determined by bioluminescent imaging (BLI) of RDEB mice as well as excised internal organs.

Results: A single intra-hepatic injection of USSCs (0.2 x 10⁶) in newborn RDEB mice arrested the blistering phenotype and significantly enhanced the median survival to 8 days (n=21), as compared to 2 days in PBS injected animals (n=18) (P<0.0001)). About 30% of the USSC- treated mice lived beyond 2 weeks of life span (Fig 1). A second USSC intra- hepatic injection further elongated the median life span (16 days, n = 18, P<0.0001). Remarkably, two mice that received repeated injections survived more than 12 weeks. As compared to intra-hepatic injection of USSCs, other methods, i.e., a single intradermal injection of USSCs (n=15) and multiple facial vein injections of USSC-conditioned medium (n=17) did not result in a longterm benefit in the recipient RDEB mice, although the median life span was significantly increased to 4 days and 3 days respectively (P<0.0001).  The data suggest that circulation of the USSC cell population was necessary for the longterm survival of RDEB mice. Histological analysis on the footpad skin of the RDEB mice after a single intra-hepatic USSC injection demonstrated that the percentage of the skin with intact epidermis-dermis attachment was significantly improved. USSC injection also enhanced the expression of C7 in the recipient skin and electron microscopy revealed a partial restoration of anchoring fibrils in skin biopsies from the USSC-treated RDEB mice. BLI of the RDEB mice following USSC injection further demonstrated that USSCs rapidly went into circulation after intra-hepatic injection and did not result in detectable retention in organs such as heart, kidney, spleen etc (Fig 2). Indeed, specific localization of USSCs was observed in the blistered paws and GI tracts, suggesting a specific migration of USSCs toward the sites of blisters. 

Conclusion: CB-derived USSCs represent a novel stem cell source for the treatment of RDEB.