Molecular Characterization of a Precision-Cut Rat Liver Slice Model for the Evaluation of Anti-Fibrotic Compounds. Molecular Characterization of a Precision-Cut Rat Liver Slice Model for the Evaluation of Anti-Fibrotic Compounds

Precision-cut liver tissue slice (PCLS) contains all major cell types of the liver parenchyma and preserves the original cell-cell and cell-matrix contacts. It represents a promising ex vivo model to study liver fibrosis and test the anti-fibrotic effect of experimental compounds in a physiologic environment. In this study using RNAquencing we demonstrated that various pathways functionally related to fibrotic mechanisms were dysregulated in PCLSs derived from rats subjected to bile duct ligation. The Alk5 inhibitor SB525334, nintedanib and sorafenib each reversed a subset of genes dysregulated in fibrotic PCLSs and of those genes we identified 608 genes whose expression was reversed by all three compounds. These genes define a molecular signature characterizing many aspects of liver fibrosis pathology and its attenuation in the model. A panel of 12 genes and 4 secreted biomarkers including procollagen I, HA, IGFBP5 and WISP1, were further validated as efficacy endpoints for the evaluation of anti-fibrotic activity of experimental compounds. Finally, we showed that blockade of αV integrins with a small molecule inhibitor attenuated the fibrotic phenotype in the model. Overall, our results suggest that the rat fibrotic PCLS model may represent a valuable system for target validation and to determine the efficacy of experimental compounds. Overall design: Precision-cut liver tissue slices (PCLS) from BDL rats were treated with drug and profiled with RNA-Seq

精准切割肝组织切片（Precision-cut liver tissue slice, PCLS）包含肝实质的所有主要细胞类型，并保留了原始的细胞-细胞及细胞-基质连接。该切片是一种极具应用前景的离体模型，可用于研究肝纤维化，并在生理环境中检测实验性化合物的抗纤维化活性。本研究通过RNA测序（RNA Sequencing）证实，在胆管结扎（bile duct ligation, BDL）大鼠来源的纤维化PCLS中，多种与纤维化机制功能相关的通路出现表达失调。ALK5抑制剂SB525334、尼达尼布（nintedanib）与索拉非尼（sorafenib）均可逆转纤维化PCLS中失调基因的部分表达谱；其中，我们鉴定出608个基因可被这三种化合物共同逆转其表达水平。这些基因构成了可表征肝纤维化病理特征及其在该模型中缓解过程的分子特征谱。本研究进一步验证了包含Ⅰ型前胶原蛋白、透明质酸（HA）、IGFBP5及WISP1在内的12个基因与4种分泌型生物标志物组成的检测组合，可作为评估实验性化合物抗纤维化活性的药效学终点。此外，本研究证实，使用小分子抑制剂阻断αV整合素可缓解该模型中的纤维化表型。综上，本研究结果表明，大鼠纤维化PCLS模型可作为用于靶点验证及评估实验性化合物疗效的可靠系统。整体实验设计：对胆管结扎大鼠来源的精准切割肝组织切片施以药物处理，并通过RNA-Seq进行转录组表达谱分析。