The mRNA level of c-myc was also higher in the ILK/liver−/− mice as compared to the WT mice at day 7. Because c-myc is regulated posttranscriptionally,24 we analyzed the protein expression of c-myc after TCPOBOP

administration. In the WT mice c-myc expression was induced as early as day 1 (Fig. 5A). Its expression, however, was maximum at day 2. By days 5 and 7 its expression had started to go down. In the ILK/liver−/− mice the expression levels were higher as compared to the WT mice at all timepoints except for day 2, suggesting a sustained induction of c-myc. Overall, there seemed to be a higher and more sustained induction of c-myc in the ILK/liver−/− mice. Transcriptional buy CHIR-99021 activity of c-myc was also higher in the ILK/liver−/− mice as compared to the WT mice at day 1 after TCPOBOP administration (Fig. 5C). To further corroborate the findings that there was a sustained induction of c-myc in the ILK/liver−/− mice, we performed c-myc immunohistochemistry at days 1 and 7 after TCPOBOP administration. At day 1

both WT and ILK/liver−/− mice showed nuclear staining. By day 7 the WT mice showed minimal nuclear staining, whereas the ILK/liver−/− still had plenty see more of hepatocytes that showed nuclear staining, suggesting a sustained induction of c-myc (Fig. 5D). Studies have shown FoxM1 to be a key target of c-Myc,1 which in turn is involved in promoting hepatocyte proliferation.25 We looked

at the levels of FoxM1 mRNA in the WT and ILK/liver−/− mice after TCPOBOP administration. FoxM1 mRNA levels, even though lower in the ILK/liver−/− mice at day 1 after TCPOBOP administration, were more sustained as compared to the WT mice (Fig. 5E). Despite the dramatic effects of specific chemical mitogens such as TCPOBOP on the liver, the signaling pathways responsible for limiting the chemically induced hypertrophic and hyperplastic responses remain largely unknown. Studies in our laboratory have shown the role of ECM in inhibiting hepatocyte proliferation.16, 18, 26 Because it is practically impossible to eliminate ECM from an intact organ, elimination of the proteins responsible for transmission of the ECM signals to hepatocytes became a feasible alternative when ILKloxP/loxP mice became available. FAK (focal MCE adhesion kinase), Mig2 and ILK are three proteins, primarily involved with transmission of the integrin signals. Our results demonstrate that the final size of the liver due to the TCPOBOP-induced hypertrophic and hyperplastic response is to a significant level dependent on ILK. Livers deficient in ILK show prolonged and sustained proliferative response to TCPOBOP. They also show higher liver/body weight ratios as compared to the WT counterpart given the same treatment. TCBOPOP is the strongest chemical mitogen5-7 for the liver.