Pregnancy is characterized by the development of maternal insulin resistance and increased hepatic glucose production to provide sufficient glucose to be used as nutrient by the developing fetus. The maternal insulin-producing beta-cells expand to counterbalance the increased glucose levels and failure of this process can lead to development of gestational diabetes, which has great health implications for both the mother and the developing fetus. Yet, our knowledge on the mechanisms behind the pregnancy-induced beta-cell expansion in humans is limited. To address this knowledge gap, in the betaPPreg project, I will use the pig model as an ideal surrogate large mammal to study the mechanisms and identify physiological drivers of pregnancy-induced beta-cell expansion. First, I will identify the extent and the different cellular mechanisms of beta-cell adaptation in the porcine pancreas over the three trimesters of gestational time (~114 days) by histological analysis. To reveal the physiological candidates of beta-cell adaptation, I will perform untargeted metabolomics coupled with proteomics analysis of serum and pancreas at sequential temporal windows during gestation. This analysis will point to the significantly changed metabolites and growth factors that can be potentially involved in beta-cell expansion. I will perform a small-scale metabolite/growth factor screen in isolated neonatal porcine islets with the most regulated factors, to causatively link the metabolites/growth factors stimulating beta-cell expansion. The most promising candidates will be then tested in isolated human islets and human-derived ductal organoids to assess their translational potential as beta-cell proliferation/differentiation inducers and human therapeutics. Overall, the proposed project will greatly advance our understanding of beta-cell adaptation during pregnancy in large mammals and provide novel therapeutic candidates for gestational diabetes treatment.