Contribution of cytokeratin 19-expressing cells towards islet regeneration induced by multipotent stromal cell secreted proteins.

Abstract

Residual beta cell function has been documented in "medalist" patients who have lived with Type 1 diabetes (T1D) for >50 years. In addition, endocrine cell neogenesis first occurs in the developing human embryo from progenitor cells derived from pancreatic ductal epithelial structure. Thus, beta cell conversion from a dormant epithelial precursor remains a promising approach to regenerate islets during T1D. We have previously shown that intra-pancreatic (iPan) injection of Wnt pathway-stimulated conditioned media (Wnt+ CdM) generated from human bone marrow-derived multipotent stromal cells (MSC) contained islet regenerative factors that reduced hyperglycemia and recovered beta cell mass in streptozotocin-treated mice. However, the endogenous source of regenerated beta cells remains unknown. Herein, we employed cytokeratin 19 (CK19)-CreERT Rosa26-mTomato lineage-tracing mice to assess the endocrine conversion of CK19+ cells during MSC CdM-induced islet regeneration. Mice iPan-injected with Wnt+ CdM demonstrated reduced blood glucose levels and improved glucose tolerance compared to mice injected with unconditioned basal media. CdM-injected mice also showed increased islet number and beta cell mass, as well as CK19+ cells within regenerating islets. The frequency of insulin + cells that co-expressed tdTomato within dissociated pancreas samples observed via flow cytometry was 5-fold higher in Wnt+ CdM-injected mice (~5%) compared to basal media-injected controls (~1%). Collectively, in vivo lineage tracing revealed conversion of CK19+ cells to functional beta cells partially contributed to islet regeneration induced by Wnt-activated MSC CdM. Future studies are required to delineate alternate cell types and mechanisms participating in islet regeneration induced by direct delivery of MSC-CdM.