Vav1 Sustains the Expression of Insulin, PDX1 and miR-375 During Differentiation of hiPSCs to β Cells: A Potential Target to Improve the In Vitro Generation of Insulin-Producing Cells

Backround: Human-induced pluripotent stem cells (hiPSCs) have emerged as a promising source of transplantable insulinproducing cells (IPCs) to restore insulin levels in Type 1 Diabetes (T1D) patients. Despite progress, obtaining fully functional β cells from hiPSCs remains challenging, underscoring the need to better understand the intracellular mechanisms involved. We investigated here the potential role of Vav1, a multidomain protein that we identified as crucial for the maturation of human biliary stem cells (hBTSCs) into β-like cells and in the trans-differentiation of pancreatic adenocarcinoma (PDAC) cells into IPCs; Methods: Levels and subcellular localization of Vav1 were investigated throughout a seven-step differentiation process of hiPSCs to β cells. Vav1expression was forcedly modulated in pancreatic progenitors, and the potential effects were evaluated on insulin production and on PDX1, miR-375, and Akt, key regulators of β cells generation; RESULTS. Vav1 showed dynamic modulation, with pancreatic precursor cells requiring adequate levels of the protein to generate IPCs. Results: Vav1 sustains the expression of PDX1, a primary regulator of insulin expression, and of its target miR-375, essential for determining β cell mass. Furthermore, Vav1 reduction correlated with increased activation of Akt, which regulates cell survival and insulin secretion in β cells and is down-regulated by miR- 375. Conclusion: Our findings suggest the existence of a Vav1/PDX1/miR-375/Akt axis as part of the complex network orchestrating the generation of functional β cells. These insights indicate that strategies aimed at specifically modulating Vav1 levels may positively impact the generation of IPCs in vitro and, ultimately, β cell replacement therapy for T1D.