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In utero intracardial injection of a transducible MafA protein accelerates pancreatic islet maturation

Nancy Vargas, Silvia Alvarez-Cubela, Margarita Nieto, Nicholas Fort, Jaime Giraldo, Sirlene Cechin, Enrique Garcia, Camillo Ricordi, Luca Inverardi, Ricardo Pastori, Juan Dominguez-Bendala

Stem Cell Laboratory, Diabetes Research Institute, University of Miami, Miami, FL, United States

In concert with Pdx1 and Beta2/NeuroD, the nuclear protein MafA is essential for the maintenance of the adult beta cell phenotype by contributing to the proper regulation of the insulin promoter. When ectopically expressed together, MafA, Pdx1 and Ngn3 (an upstream regulator of Beta2/NeuroD) were recently shown to reprogram acinar exocrine cells into functional, insulin-producing beta cells. This novel approach has opened the door to potential clinical interventions aimed at restoring beta cell mass in the context of pancreatic disease. For this to happen, however, alternatives to the viral delivery of these factors need to be defined. Protein transduction is one such possible alternative. Using this technology, any protein of interest can be made cell-permeable by the mere addition of a short membrane-penetrating peptide. Recombinant purified proteins can then be added in vitro to the culture medium or delivered locally or systemically in vivo, where they are expected to exert their function in the same manner as their native counterparts. This DNA-free system is deemed to be safer than viral-based approaches for future medical use. We have already described transducible versions of Ngn3 and Pdx1, and here we present a recombinant transducible version of the MafA protein (TAT-MafA) that penetrates across cell membranes with an extremely high efficiency and binds to the insulin promoter in vitro. When injected in utero into the heart of live murine embryos, TAT-MafA reaches the developing pancreas and increases the expression of key target genes, enhances insulin production and causes cytoarchitectural changes that are consistent with faster islet maturation. The purification and characterization of a functional TAT-MafA protein sets the stage for prospective therapeutic applications that circumvent the use of viruses. To our knowledge, this is also the first report on the use of protein transduction in utero.