Zeisberg Michael, Kramer Kyle, Sindhi Nazia, Sarkar Pradip, Upton Melissa, Kalluri Raghu
Center for Matrix Biology, Department of Medicine, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA 02215, USA.
Mol Cell Biochem. 2006 Feb;283(1-2):181-9. doi: 10.1007/s11010-006-2677-8.
Basement membrane (BM) is a highly specialized extracellular matrix (ECM), which is associated with epithelia and endothelia. BM provide epithelia with structural support and also regulate cell behavior. The liver contains a unique ECM within the space of Disse, which consists of basement membrane constituents as well as fibrillar ECM molecules. Changes in composition of this ECM are considered detrimental for viability of hepatocytes during progression of liver disease. Mouse tumor-derived BM preparations, such as Matrigel, which are commonly used as a model for BM in vitro, differ significantly in their composition from liver BM present in vivo. In order to gain further insights into the role of BM in the regulation of hepatocyte behavior in health and disease, we generated a liver-derived basement membrane matrix (LBLM). LBLM allowed investigation of BM-hepatocyte interactions in vitro. Here we report a novel approach of generating a liver-derived basement membrane matrix by separate isolation of type IV collagen, laminin, nidogen, and heparan sulfate proteoglycans, and subsequent reconstitution into a matrix-like gel. Adhesion of primary human hepatocytes to LBLM was increased and the rate of de-differentiation was decreased compared to hepatocyte cultivation on Matrigel or type I collagen matrix. Primary human hepatocytes maintained their differentiated epithelial phenotype on LBLM isolated from normal human livers for more than 21 days, whereas they de-differentiated rapidly on LBLM isolated from cirrhotic human livers. Normal human LBLM contains a unique isoform composition of type IV collagen, namely alpha1 (IV), alpha2(IV), alpha4(IV), and alpha6(IV) chains, whereas cirrhotic LBLM contains only alpha1(IV) and alpha2(IV) isoforms, albeit present in increased amounts. These findings suggest that the composition of liver basement membrane is important for the maintenance of hepatocyte viability and provide anti-de-differentiation clues.
基底膜（BM）是一种高度特化的细胞外基质（ECM），与上皮细胞和内皮细胞相关。基底膜为上皮细胞提供结构支持并调节细胞行为。肝脏在狄氏间隙内含有独特的细胞外基质，其由基底膜成分以及纤维状细胞外基质分子组成。在肝脏疾病进展过程中，这种细胞外基质组成的变化被认为对肝细胞的生存能力有害。小鼠肿瘤来源的基底膜制剂，如基质胶，通常用作体外基底膜模型，其组成与体内存在的肝脏基底膜有显著差异。为了进一步深入了解基底膜在健康和疾病状态下对肝细胞行为调节中的作用，我们制备了肝脏来源的基底膜基质（LBLM）。LBLM使得体外研究基底膜与肝细胞的相互作用成为可能。在此我们报告一种通过分别分离IV型胶原、层粘连蛋白、巢蛋白和硫酸乙酰肝素蛋白聚糖，随后重构为类似基质的凝胶来制备肝脏来源基底膜基质的新方法。与在基质胶或I型胶原基质上培养肝细胞相比，原代人肝细胞对LBLM的黏附增加且去分化速率降低。原代人肝细胞在从正常人肝脏分离的LBLM上可维持其分化的上皮表型超过21天，而在从肝硬化人肝脏分离的LBLM上则迅速去分化。正常人LBLM含有IV型胶原的独特异构体组成，即α1（IV）、α2（IV）、α4（IV）和α6（IV）链，而肝硬化LBLM仅含有α1（IV）和α2（IV）异构体，尽管其含量增加。这些发现表明肝脏基底膜的组成对于维持肝细胞生存能力很重要，并提供了抗去分化线索。
