4A). The increase of HSPC-related markers is more prominent at 4 weeks of age than at 1 year. Histologically, A6-positive cells appear as early as 3-4 weeks of age in parenchyma and the hyperplastic ductal region of albNScko livers, whereas no A6+ signals are detected in NSflx/flx livers (Fig. 4B). The majority of A6+ cells also coexpress the CK19 antigen on serial sections of 4 um in thickness (Fig. 4C). To determine the role of NS in liver regeneration, Cabozantinib datasheet we analyzed responses of albNScko and NSflx/flx livers to CCl4 treatment at 2 weeks of age when Cre expression and DNA damage were

maximal and histological changes and serum measurement of hepatocellular injury were mild in albNScko livers. Liver samples were collected in pairs from CCl4- and oil-treated mice at the first, second, or fourth day of the injection. NSflx/flx livers show acute pericentral necrosis with infiltrating leukocytes during the first 2 days after injection

(Fig. 5A1, white arrows). Without CCl4 exposure, albNScko livers contained regenerative nodules and nonregenerative regions (Fig. 5A2, perinodule). In response to CCl4 treatment, albNScko livers began to show not only the same acute pericentral necrosis and leukocyte infiltration as observed in NSflx/flx livers, but also severe hydropic degeneration (black arrows) in the perinodular areas (Fig. 5A3). Notably, the regenerative nodules were relatively resistant AZD6738 to the acute necrotic effect of CCl4 treatment (Fig. 5A4), which is consistent with their lower expression of the key enzyme, CYP2E1, that metabolizes CCl4 and forms free radicals (Supporting Fig. 4A). Mice recovered from CCl4-triggered pericentral necrosis after 4 days (Supporting Fig. 4B). Unlike NSflx/flx livers, which show no increase of CK19+ cells after CCl4-induced damage, albNScko

livers displayed a significant increase of CK19+ bile ductules and small CK19+ progenitor-like cells located in the periportal region in response to CCl4 treatment (Fig. 5B and Supporting Fig. 4C). Immunostaining on serial sections showed that the numbers of A6- and CK19 double-positive progenitor cells were increased by CCl4 treatment in albNScko ifoxetine livers (Fig. 5C). Consistently, the number of Sox9 and CK19 double-positive cells was also increased in CCl4-treated albNScko livers (Supporting Fig. 4D). Supporting the idea that HSPCs may be expanded in CCl4-treated albNScko livers, qRT-PCR assays demonstrated that the messenger RNA levels of two HSPC-related markers, EpCAM and AFP, were both up-regulated in albNScko livers after CCl4 treatment (Fig. 5D). The increase of EpCAM occurred within 1 day, peaked on the second day, and dropped after 4 days, whereas the increase of AFP was found primarily on the second day of the injection.