Establishment of decellularized extracellular matrix scaffold derived from caprine pancreas as a novel alternative template over porcine pancreatic scaffold for prospective biomedical application

Abstract

In this study, the caprine pancreas has been presented as an alternative to the porcine organ for pancreatic xenotransplantation with lesser risk factors. The obtained caprine pancreas underwent a systematic cycle of detergent perfusion for decellularization. It was perfused using anionic (0.5% w/v sodium dodecyl sulfate) as well as non‐ionic (0.1% v/v triton X‐100, t‐octyl phenoxy polyethoxy ethanol) detergents and washed intermittently with 1XPBS supplemented with 0.1% v/v antibiotic and nucleases in a gravitation‐driven set‐up. After 48 h, a white decellularized pancreas was obtained, and its extracellular matrix (ECM) content was examined for scaffold‐like properties. The ECM content was assessed for removal of cellular content, and nuclear material was evaluated with temporal H&E staining. Quantified DNA was found to be present in a negligible amount in the resultant decellularized pancreas tissue (DPT), thus prohibiting it from triggering any immunogenicity. Collagen and fibronectin were confirmed to be preserved upon trichrome and immunohistochemical staining, respectively. SEM and AFM images reveal interconnected collagen fibril networks in the DPT, confirming that collagen was unaffected. sGAG was visualized using Prussian blue staining and quantified with DMMB assay, where DPT has effectively retained this ECM component. Uniaxial tensile analysis revealed that DPT possesses better elasticity than NPT (native pancreatic tissue). Physical parameters like tensile strength, stiffness, biodegradation, and swelling index were retained in the DPT with negligible loss. The cytocompatibility analysis of DPT has shown no cytotoxic effect for up to 72 h on normal insulin‐producing cells (MIN‐6) and cancerous glioblastoma (LN229) cells in vitro. The scaffold was recellularized using isolated mouse islets, which have established in vitro cell proliferation for up to 9 days. The scaffold received at the end of the decellularization cycle was found to be non‐toxic to the cells, retained biological and physical properties of the native ECM, suitable for recellularization, and can be used as a safer and better alternative as a transplantable organ from a xenogeneic source.

Impact of various detergent-based immersion and perfusion decellularization strategies on the novel caprine pancreas derived extracellular matrix scaffold

“…Apart from collagen, the cell population in the MTT analysis was also seen to be increased. We have previously established that caprine pancreatic ECM promotes cell migration, adhesion, and proliferation under in vitro culture conditions; therefore, this increase in cell population can be attributed to the abundance of ECM-associated molecules persisting in the scaffold ( Singh et al, 2022 ). M1 macrophages can contribute to the initial inflammatory response to such scaffolds, which can lead to tissue damage if not appropriately controlled.…”

In-Depth Analysis of the Pancreatic Extracellular Matrix during Development for Next-Generation Tissue Engineering

“…The power of our approach was demonstrated by the detection of factors only expressed at one developmental stage and that could support pancreatic differentiation/development. While viscoelasticity studies have previously been conducted on mature pancreatic tissues [ 44 ], our study presents the first-ever assessment of the embryonic pancreas and its ECM, revealing a relatively soft and low-viscosity matrix during development. We believe that this comprehensive list of proteins and the viscoelasticity properties at the E14.5 and P1 stages are important sources of information that should guide or influence the design of scaffolds for an artificial pancreas, but also help develop protocols for obtaining mature pancreatic cells from embryonic stem cells.…”