Optimization of Liver Decellularization Maintains Extracellular Matrix Micro-Architecture and Composition Predisposing to Effective Cell Seeding

Abstract

Hepatic tissue engineering using decellularized scaffolds is a potential therapeutic alternative to conventional transplantation. However, scaffolds are usually obtained using decellularization protocols that destroy the extracellular matrix (ECM) and hamper clinical translation. We aim to develop a decellularization technique that reliably maintains hepatic microarchitecture and ECM components. Isolated rat livers were decellularized by detergent-enzymatic technique with (EDTA-DET) or without EDTA (DET). Histology, DNA quantification and proteomics confirmed decellularization with further DNA reduction with the addition of EDTA. Quantification, histology, immunostaining, and proteomics demonstrated preservation of extracellular matrix components in both scaffolds with a higher amount of collagen and glycosaminoglycans in the EDTA-DET scaffold. Scanning electron microscopy and X-ray phase contrast imaging showed microarchitecture preservation, with EDTA-DET scaffolds more tightly packed. DET scaffold seeding with a hepatocellular cell line demonstrated complete repopulation in 14 days, with cells proliferating at that time. Decellularization using DET preserves microarchitecture and extracellular matrix components whilst allowing for cell growth for up to 14 days. Addition of EDTA creates a denser, more compact matrix. Transplantation of the scaffolds and scaling up of the methodology are the next steps for successful hepatic tissue engineering.

Biocompatibility and hemocompatibility of efficiently decellularized whole porcine kidney for tissue engineering

“…Morphologically, our decellularized kidney preserved the glomerulus and its vascular network upon examination with H&E and SEM. It was reported that high concentrations of SDS (1%) and Triton‐x cause destruction of the vascular network of rat livers during decellularization process, which makes the scaffolds unsuitable for potential transplantation . Importantly, we have shown the maintenance of the vascular network.…”

Decellularized human liver scaffold‐based three‐dimensional culture system facilitate hepatitis B virus infection

“…PHH is the best cell model for HBV studies. [22][23][24] The advantages of decellularization liver scaffold are obviously. Advanced 3D liver models aimed at recreating hepatocyte physiological function have been established and believed to be promising in field such as regeneration and drug screening.…”

Decellularized liver matrix as substrates for rescue of acute hepatocytes toxicity

“…Liver cirrhosis is accompanied by abnormal changes in ECM structure and composition (Klaas et al, ), remodeling in the ECM composition may alter cell signaling and facilitate liver regeneration for liver diseases (Klaas et al, ; Williams, Clouston, & Forbes, ). On the other hand, the decellularization procedure without using harsh detergents may result in structural integrity of ECM (Maghsoudlou et al, ); moreover, Triton X‐100, a mild detergent and typically used for liver decellularization (Badylak, Taylor, & Uygun, ), can only reduce cell component/DNA content without depleting any glycosaminoglycans (GAGs) during the washing process (Bruyneel & Carr, ). The effect could be attributed that GAGs and other components (collagen, proteoglycans) constitute the liver extracellular microenvironment, and also play an important role in hepatic functions and viability (Damania et al, ; Lee et al, ); furthermore, the wound ECM of GaIN‐damaged cells can be subsequently replaced by mature ECM and return to the preinjury state (Jonker, Dijkhuis, Boes, Hardonk, & Grond, ).…”