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.精确切割肝组织切片（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-基质连接关系。该模型是极具应用前景的离体研究模型，可在生理环境下用于肝纤维化研究以及实验化合物的抗纤维化活性评估。本研究通过RNA测序（RNA sequencing）证实，胆管结扎大鼠来源的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）均可分别逆转纤维化PCLS中部分失调基因的表达；在上述基因中，我们鉴定出608个可被这三种化合物共同逆转其表达水平的基因。这些基因构成了一组分子特征标签，可表征该模型中肝纤维化病理的多个层面及其缓解状态。本研究进一步验证了一套包含12个基因及4种分泌型生物标志物的检测组合，其中分泌型生物标志物包括I型前胶原（procollagen I）、透明质酸（HA）、胰岛素样生长因子结合蛋白5（IGFBP5）及WNT1诱导信号通路蛋白1（WISP1），该组合可作为评估实验化合物抗纤维化活性的疗效终点指标。最后，本研究证实，通过小分子抑制剂阻断αV整合素（αV integrins）可减轻该模型的纤维化表型。综上，本研究结果表明，纤维化大鼠PCLS模型可作为极具价值的研究体系，用于靶点验证及实验化合物的疗效评估。