Bio-based hydrogel for beta cell islet transplantation in type 1 diabetes therapy

Objectives: Type 1 Diabetes Mellitus (T1DM) is a chronic autoimmune disease that destroys insulin-producing pancreatic cells, affecting over 9 million people globally. The primary treatment involves insulin therapy, but improper delivery can lead to poor glucose control or hypoglycemia. Pancreatic islet transplantation offers a potential alternative, but long-term graft survival remains a challenge. This study aims to enhance cell-based therapies by developing a permeable, pro-angiogenic immune-isolation core- shell hydrogel using decellularized pancreatic extracellular matrix (dECM) for the encapsulation of beta cells. The hydrogel will be formed using Riboflavin (RB) a bio crosslinker also known as Vitamin B2, which is reported to ameliorate oxidative stress and inflammation, key factors in the pathophysiology of insulin resistance. Methods: Porcine pancreas was decellularized using a supercritical carbon dioxide (scCO2) protocol developed by our group, to effectively preserve the biochemical structure of the ECM, while removing the cellular components. After five cycles of decellularization (110-120 bar, 36-38ºC for 1h cycle), the tissue was thoroughly analyzed. The residual DNA content, a key indicator of the efficiency of decellularization, and the ECM components (glycosaminoglycans, collagen, and elastin) were quantified using Blyscan, Sircol and Fasin assay kits. The hydrogel was developed by digesting the dECM using pepsin/HCl and crosslinked with Riboflavin (RB) in combination with Horseradish peroxidase (HRP). Pancreatic islets from a pancreatic cell line were seeded into the hydrogel, by placing a beta cell line spheroid into the core of the dECM RB system. Also, cell viability will be assessed by live/dead assay, and insulin and PDX 1 expression will be characterized by Elisa and RT PCR. Results: The DNA quantification revealed a significant reduction in DNA content (≈12.7 ng/mg) after five cycles, indicating effective removal of cellular material. Moreover, the results from the quantification of ECM components (GAGs, elastin and collagen) demonstrated that the decellularization process effectively preserved its ECM components. The crosslinked dECM-RB hydrogel exhibited mechanical stability, making it ideal for cell culture. The crosslinking process also helped to preserve the native structure of the ECM, which is crucial for maintaining the biological activity of the seeded cells. Ongoing studies are focused on regulating insulin secretion by stimulating the cells with glucose. The preliminary results revealed that beta cells within the hydrogel could produce insulin in response to glucose, showcasing its functional capability. Conclusion: The findings highlight the potential of the use of riboflavin as a bio crosslinker with an added activity to improve the stability and performance of pancreatic decellularized ECM. The dECM-RB hydrogel provided a favorable microenvironment for beta cell activity. Overall, a robust and functional cell delivery platform was created, and studies are presently ongoing to monitor cell longevity within the hydrogel and insulin production upon glucose stimulation.