Type 1 diabetes Mellitus (T1DM) is an autoimmune disease characterized by the destruction of the insulin-producing β-cells of the pancreatic islets of Langerhans. Allogeneic intraportal islet transplantation offers a promising treatment to restore insulin independence, yet post-transplant survival and function remain critically limited by hypoxia, inflammation, and poor vascularization. Therefore, addressing these challenges is crucial to improve long-term transplantation outcomes. Among various strategies, the use of accessory cells has shown promise in protecting and supporting islets and their function. In particular, human amniotic epithelial cells (AECs) have demonstrated cytoprotective effects on pancreatic islets under normal and stressful conditions such as hypoxia and inflammation, due to their anti-inflammatory, immunomodulatory, and regenerative properties. Recently, cell-free derivatives of stem cells such as secretome (conditioned medium, CM) and extracellular vesicles (EVs) have emerged as safer and scalable alternatives to live cell therapies. These derivatives harness the paracrine effects of stem cells by delivering bioactive molecules, including proteins, lipids, and nucleic acids, to target cells, while minimizing risks linked to cell transplantation such as immunogenicity and tumorigenicity. Notably, AEC-derived secretome and EVs (AEC-CM and AEC-EVs) have shown therapeutic potential in various regenerative contexts but remain unexplored in islet transplantation. Therefore, this thesis investigates the therapeutic potential of AEC derivatives on pancreatic islets.
The first aim was to assess the effect of AEC derivatives on the function and survival of pancreatic islets in normal, hypoxic, and inflammatory environments. We addressed this aim by dividing it into two parts. First, on an analytical level, by isolating and characterizing AEC-EVs from AEC-CM. We successfully isolated and characterized AEC-EVs. We also demonstrated that their cargo is rich in proteins related to the extracellular matrix, cell growth, and metabolic regulation, all of which may support islet survival and function. The second part was on a functional level, assessing the effects of AEC derivatives on islets. Our results demonstrated that AEC-CM significantly enhanced insulin secretion in islets under normoxic conditions. In contrast, the EV fraction confers a cytoprotective effect primarily under hypoxic stress, promoting enhanced islet function and mildly ameliorated survival, while showing limited benefit under inflammatory conditions.
The second aim was to elucidate the underlying mechanisms mediating beneficial effects on islets, focusing on AEC-EV treatment, which showed the most benefit under hypoxia following the first aim. We employed mass spectrometry to uncover proteomic changes in islets treated with or without AEC-EVs. We found that AEC-EVs mediate their cytoprotective effects on hypoxic islets by promoting the establishment of rich extracellular matrix protein levels and the regulation of different mechanisms, including pathways related to autophagy and insulin receptor signaling.
In summary, this work supports AEC derivatives as promising cell-free therapeutic strategies to enhance pancreatic islet survival and function. These insights open new avenues for improving graft survival and long-term clinical outcomes in T1DM patients undergoing islet transplantation.
