Background and Aims: During liver fibrosis, proteolytic activity is timely regulated in infiltrating and liverresident cells depending on the cellular demands and controls not only matrix turnover but also, the activation and repression of growth factors and chemokines. Despite our growing understanding of the roles played by lysosomal proteases in liver fibrosis, our knowledge of their specific targets and signaling networks remains limited. Thus, the aim of this study was to analyse cathepsin D (CtsD) cell-specific role in hepatocytes during liver fibrosis. Method: We generated a novel knock-out mouse strain by breeding Albumin-Cre (hepatocytes) mice with CtsD floxed mice. Fibrosis was established using bile duct ligation (BDL) for 14 days or chronic administration of CCl4 twice a week for 8 weeks in CtsDF/F and CtsDDHep mice. Liver damage was determined by serum ALT. CtsD deletion was assessed by enzymatic activity assay, WB, IF and RTPCR. Fibrosis was analysed by Sirius Red staining, α-SMA IHP and α-SMA, Col1A1 and TGF-beta RTPCR. Liver inflammation was determined by NIMP and F4/80 IHP and TNF-α, CCL2 and CCL3 RTPCR. Results:CtsD cell-specific deletion in hepatocytes was validated by CtsD WB in primary mouse hepatocytes and dual IF (F4/80-CtsD) in liver section from CtsDF/F and CtsDDHep mice.Of note, CtsD expression remained unaffected in liver non-parenchymal cells. Next, fibrosis was established for 14 days using BDL. CtsD deletion in CtsDDHep livers was confirmed by CtsD activity assay and gene expression. CtsD deletion in hepatocytes did not affect liver damaged (ALT) and liver fibrosis as determined by Sirius red staining, αSMA IHP and hepatic α-SMA, Col1A1 and TGF-beta gene expression. Furthermore, liver inflammation was also not significantly affected in CtsDDHep mice after BDL as assessed by NIMP and F4/80 IHP and TNF-α, CCL2 and CCL3 gene expression. In agreement, no significant changes in liver damage, fibrosis and inflammation were observed after chronic CCl4 administration between CtsDF/F and CtsDDHep mice supporting our results in the BDL model. Conclusion: CtsD expressed in hepatocytes does not play an essential role in liver fibrosis development.

Resumen del trabajo presentado en el 17º Congreso de la Societat Catalana de Transplantament, celebrado en Barcelona, del 22 al 24 de marzo de 2023