Method for perfusion decellularization of porcine whole liver and kidney for use as a scaffold for clinical-scale bioengineering engrafts.

BACKGROUND

Whole-organ engineering provides a new alternative source of donor organs for xenotransplantation. Utilization of decellularized whole-organ scaffolds, which can be created by detergent perfusion, is a strategy for tissue engineering. In this article, our aim is to scale up the decellularization process to human-sized liver and kidney to generate a decellularized matrix with optimal and stable characteristics on a clinically relevant scale.

METHODS

Whole porcine liver and kidney were decellularized by perfusion using different detergents (1% SDS, 1% Triton X-100, 1% peracetic acid (PAA), and 1% NaDOC) via the portal vein and renal artery of the liver and kidney, respectively. After rinsing with PBS to remove the detergents, the obtained liver and kidney extracellular matrix (ECM) were processed for histology, residual cellular content analysis, and ECM components evaluation to investigate decellularization efficiency, xenoantigens removal, and ECM preservation.

RESULTS

The resulting liver and kidney scaffolds in the SDS-treated group showed the most efficient clearance of cellular components and xenoantigens, including DNA and protein, and preservation of the extracellular matrix composition. In comparison, cell debris was observed in the other decellularized groups that were generated using Triton X-100, PAA, and NaDOC. Special staining and immunochemistry of the porcine liver and kidney ECMs further confirmed the disrupted three-dimension ultrastructure of the ECM in the Triton X-100 and NaDOC groups. Additionally, Triton X-100 effectively eliminated the residual SDS in the SDS-treated group, which ensured the scaffolds were not cytotoxic to cells. Thus, we have developed an optimal method that can be scaled up for use with other solid whole organs.

CONCLUSIONS

Our SDS-perfusion protocol can be used for porcine liver and kidney decellularization to obtain organ scaffolds cleared of cellular material, xenoimmunogens, and preserved vital ECM components.