Hemocompatibility improvement of perfusion-decellularized clinical-scale liver scaffold through heparin immobilization.

Ji Bao,Xin Jiang,Yongjie Zhou,Yujia Wang,Hong Bu,Qiong Wu,Jiu Sun,Yujun Shi,Li Li

doi:10.1038/srep10756

Abstract

Whole-liver perfusion-decellularization is an attractive scaffold–preparation technique for producing clinical transplantable liver tissue. However, the scaffold’s poor hemocompatibility poses a major obstacle. This study was intended to improve the hemocompatibility of perfusion-decellularized porcine liver scaffold via immobilization of heparin. Heparin was immobilized on decellularized liver scaffolds (DLSs) by electrostatic binding using a layer-by-layer self-assembly technique (/h-LBL scaffold), covalent binding via multi-point attachment (/h-MPA scaffold), or end-point attachment (/h-EPA scaffold). The effect of heparinization on anticoagulant ability and cytocompatibility were investigated. The result of heparin content and release tests revealed EPA technique performed higher efficiency of heparin immobilization than other two methods. Then, systematic in vitro investigation of prothrombin time (PT), thrombin time (TT), activated partial thromboplastin time (APTT), platelet adhesion and human platelet factor 4 (PF4, indicates platelet activation) confirmed the heparinized scaffolds, especially the /h-EPA counterparts, exhibited ultralow blood component activations and excellent hemocompatibility. Furthermore, heparin treatments prevented thrombosis successfully in DLSs with blood perfusion after implanted in vivo. Meanwhile, after heparin processes, both primary hepatocyte and endothelial cell viability were also well-maintained, which indicated that heparin treatments with improved biocompatibility might extend to various hemoperfusable whole-organ scaffolds’ preparation.

Highlights

Whole-liver perfusion-decellularization is an attractive scaffold– preparation technique for producing clinical transplantable liver tissue
The remnant DNA extract was subjected to agarose gel electrophoresis to reveal the length of DNA fragments in the decellularized liver scaffold (DLS), which can barely be seen by visual inspection
Combined with the numbers of the adhered platelets, these results indicated that platelet activation on the heparin treated DLSs was greatly suppressed

Summary

Introduction

Whole-liver perfusion-decellularization is an attractive scaffold– preparation technique for producing clinical transplantable liver tissue. A portal implantable functional tissue-engineered liver using a decellularized liver scaffold (DLS) and hepatocytes in rats[8] This process has been scaled up, generating biocompatible scaffolds at a clinically relevant scale[9]. We evaluated the performance of the heparinized scaffolds by measuring immobilized heparin contents and release rates, coagulation time, platelet adhesion and activation, cell culture in vitro and implantation in vivo (Fig. 1B). These hemocompatibility improvement studies are the first step toward generating an engineered transplantable functional liver decellularized whole organ scaffolds

Methods

Results

Conclusion