Hepatocyte Culture in Autologous Decellularized Spleen Matrix

Abstract

ABSTRACT. Background and Aims: Using decellularized scaffold to reengineer liver tissue is a promising alternative therapy for end-stage liver diseases. Though the decellularized human liver matrix is the ideal scaffold for reconstruction of the liver theoretically, the shortage of liver donors is still an obstacle for potential clinical application. Therefore, an appropriate alternative scaffold is needed. In the present study, we used a tissue engineering approach to prepare a rat decellularized spleen matrix (DSM) and evaluate the effectiveness of this DSM for primary rat hepatocytes culture. Methods: Rat decellularized spleen matrix (DSM) was prepared by perfusion of a series of detergents through spleen vasculature. DSM was characterized by residual DNA and specific extracellular matrix distribution. Thereafter, primary rat hepatocytes were cultured in the DSM in a 3-dimensional dynamic culture system, and liver cell survival and biological functions were evaluated by comparison with 3-dimensional sandwich culture and also with cultured in decellularized liver matrix (DLM). Results: Our research found that DSM did not exhibit any cellular components, but preserved the main extracellular matrix and the intact vasculature evaluated by DNA detection, histology, immunohistochemical staining, vessel corrosion cast and upright metallurgical microscope. *Correspondence to: Yi Lv; Email: luyi169@126.com Received July 7, 2014; Revised September 10, 2014; Accepted January 18, 2015.16 Organogenesis, 11:16-29, 2015 Ó 2015 Taylor & Francis Group, LLC ISSN: 1547-6278 print / 1555-8592 online DOI: 10.1080/15476278.2015 Moreover, the method of DSM preparation procedure was relatively simple with high success rate (100%). After seeding primary hepatocytes in DSM, the cultured hepatocytes survived inside DSM with albumin synthesis and urea secretion within 10 d. Additionally, hepatocytes in dynamic culture medium had better biological functions at day 10 than that in sandwich culture. Albumin synthesis was 85.67 § 6.34 mg/10 7 cell/24h in dynamic culture in DSM compared to 62.43 § 4.59 mg/10 7 cell/ 24h in sandwich culture (P < 0.01) and to 87.54 § 5.25 mg/10 7 cell/24h in DLM culture (P > 0.05); urea release was 32.14 § 8.62 mg/10 7 cell/24h in dynamic culture in DSM compared to 20.47 § 4.98 mg/10 7 cell/24h in sandwich culture (P < 0.05) and to 37.38 § 7.29 mg/10 7 cell/24h cultured in DLM (P > 0.05). Conclusion: The present study demonstrates that DSM can be prepared successfully using a tissue engineering approach. The DSM is an appropriate scaffold for primary hepatocytes culture.

Decellularized spleen matrix for reengineering functional hepatic-like tissue based on bone marrow mesenchymal stem cells

“…Remarkably, an abundant blood sinus structure was observed in DSM, which was favorable to satisfy the large amount of cell metabolism and oxygen consumption. It has been concluded in our published reports 15,16 that the hepatocyte vitality when cultured in DSM as well as the synthesis and secretion function were comparable with DLM-based HLT.…”

“…Herein, in order to expand the scaffold donor pool, we have established the decellularized spleen matrix (DSM), which had homogeneous ECM with DLM, and presented comparable condition for hepatocyte cultivation. 15,16 DSM scaffolds may be more promising for clinical application due to their extensive sources such as traumatic rupture, hypersplenism caused by various reasons, idiopathic thrombocytopenic purpura, floating spleen, and donation after cardiac/ brain death.…”

Decellularized amniotic membrane Scaffolds improve differentiation of iPSCs to functional hepatocyte‐like cells

“…In another studies, rat decellularized spleen matrix have used to maintain viability and functions of primary hepatocyte or stem cell-derived HLCs. 15,52 In attempt to resolve the human liver shortage and provide a replacement for xenogeneic tissues, extra-embryonic tissues have attracted great attention in research and clinical practice due to their easy availability, low probability of immune rejection and lack of ethical problems. 36,45,46 In this study, we investigate the effects of natural spongy scaffold derived from dAM on hepatic differentiation and function of hiPSCs.…”

“…As an option, animal derived-decellularized organs is widely applied as a promising alternative for generating transplantable and functional livers, but there are concerns about the relevant differences between human and animal liver architecture, in addition to immunological reactions and pathogen transmission. [15][16][17] Recently, the human amniotic membrane (hAM), the innermost layer of the placenta, has been used as a scaffold in tissue engineering. [18][19][20] In a majority of tissue engineering applications, the basement membrane part of the amniotic membrane after isolation from other layers, become decellularized to produce a biological tissue material for scaffold fabrication.…”

Strategies based on organ decellularization and recellularization

“…However, an organ might be reconstructed by cultivating organ‐specific cells in an ECM derived from another organ (e.g. spleen and hepatocytes or hepatocyte‐like cells) . Although this concept promises to be very practical, it only appears to be suitable for organs possessing an endocrine function (e.g.…”