Improving hemocompatibility of decellularized liver scaffold using Custodiol solution.

Organ decellularization is one of the most promising approaches of tissue engineering to overcome the shortage of organs available for transplantation. However, there are key hurdles that still hinder its clinical application, and the lack of hemocompatibility of decellularized materials is a central one. In this work, we demonstrate that Custodiol (HTK solution), a common solution used in organ transplantation, increased the hemocompatibility of acellular scaffolds obtained from rat livers. We showed that Custodiol inhibited ex vivo, in vitro, and in vivo blood coagulation to such extent that allowed successful transplantation of whole-liver scaffolds into recipient animals. Scaffolds previously perfused with Custodiol showed no signs of platelet aggregation and maintained in vitro and in vivo cellular compatibility. Proteomic analysis revealed that proteins related to platelet aggregation were reduced in Custodiol samples while control samples were enriched with thrombogenicity-related proteins. We also identified distinct components that could potentially be involved with this anti-thrombogenic effect and thus require further investigation. Therefore, Custodiol perfusion emerge as a promising strategy to reduce the thrombogenicity of decellularized biomaterials and could benefit several applications of whole-organ tissue engineering.

Bone marrow progenitor cells do not contribute to liver fibrogenic cells.

AIM: To investigate the contribution of bone marrow (BM) cells to hepatic fibrosis. METHODS: To establish a model of chimerism, C57Bl/6 female mice were subjected to full-body irradiation (7 Gy) resulting in BM myeloablation. BM mononuclear cells obtained from male transgenic mice expressing enhanced green fluorescent protein (GFP) were used for reconstitution. Engraftment was confirmed by flow cytometry. To induce liver injury, chimeric animals received carbon tetrachloride (CCl(4)) 0.5 mL/kg intraperitoneally twice a week for 30 d (CCl(4) 30 d) and age-matched controls received saline (Saline 30 d). At the end of this period, animals were sacrificed for post mortem analysis. Liver samples were stained with hematoxylin and eosin to observe liver architectural changes and with Sirius red for collagen quantification by morphometric analysis. α-smooth muscle actin (α-SMA) was analyzed by confocal microscopy to identify GFP+ cells with myofibroblast (MF) characteristics. Liver tissue, BM and peripheral blood were collected and prepared for flow cytometric analysis using specific markers for detection of hepatic stellate cells (HSCs) and precursors from the BM. RESULTS: Injury to the liver induced changes in the hepatic parenchymal architecture, as reflected by the presence of inflammatory infiltrate and an increase in collagen deposition (Saline 30 d = 11.10% ± 1.12% vs CCl(4) 30 d = 12.60% ± 0.73%, P = 0.0329). Confocal microscopy revealed increased reactivity against α-SMA in CCl(4) 30 d compared to Saline 30 d, but there was no co-localization with GFP+ cells, suggesting that cells from BM do not differentiate to MFs. Liver flow cytometric analysis showed a significant increase of CD45+/GFP+ cells in liver tissue (Saline 30 d = 3.2% ± 2.2% vs CCl(4) 30 d = 5.8% ± 1.3%, P = 0.0458), suggesting that this increase was due to inflammatory cell infiltration (neutrophils and monocytes). There was also a significant increase of common myeloid progenitor cells (CD117+/CD45+) in the livers of CCl(4)-treated animals (Saline 30 d = 2.16% ± 1.80% vs CCl(4) 30 d = 5.60% ± 1.30%, P = 0.0142). In addition the GFP-/CD38+/CD45- subpopulation was significantly increased in the CCl(4) 30 d group compared to the Saline 30 d group (17.5% ± 3.9% vs 9.3% ± 2.4%, P = 0.004), indicating that the increase in the activated HSC subpopulation was not of BM origin. CONCLUSION: BM progenitor cells do not contribute to fibrosis, but there is a high recruitment of inflammatory cells that stimulates HSCs and MFs of liver origin.