Successful establishment of in-vivo selective liver lobe decellularization : the starting point for In-vivo liver engineering

In-vivo liver decellularization has become a promising strategy to study in-vivo liver engineering. However, long term survival after in-vivo liver decellularization has not yet been achieved due to anatomical and technical challenges. The aim of this study is to generate a survival model of in-vivo selective liver lobe decellularization in rat. Prior to perform in-vivo decellularization, we generated an in-vivo selective liver lobe perfusion model using the left lateral lobe. The selected liver lobe was perfused with heparin saline for either 20 min, 2h, 3 h or 4 h respectively (n=3/group). The procedure was well tolerated by all animals (1w survival rate 100%, 12/12 animals). We compared three different protocols to identify the optimal protocol for in-vivo liver lobe decellularization. We observed that only using 2 hours of 1% SDS rather than 1% Triton X100 or 1% Triton X100 followed by 1% SDS resulted in the complete removal of cellular components as confirmed by histological assessment. The integrity of the vascular structure was confirmed by CT, histology and SEM, respectively. The main matrix proteins including collagen IV, elastin, fibronectin, and laminin were preserved as detected with strong signals in immunohistochemistry. Additionally, we compared two methods to drain the waste fluid. The drainage method of polyvinylidene chloride (PVDC) film + dry gauze + aspiration tube resulted in complete protection from the corrosive detergent. All animals reached survived more than one week (n=6/6), in contrast to only 16.7% (n=1/6) using the other drainage method with only PVDC film + dry gauze. We concluded that in-vivo selected liver perfusion is technically feasible. We also concluded that using 1% SDS as detergent and using PVDC film+ gauze +suction tube is optimal for in-vivo partial liver lobe decellularization.



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