Given the early increase in cell size as well as a more rapid and enhanced growth factor expression in CO-treated mice, we next evaluated cell cycle progression in liver homogenates after PHTx. CO-treated find protocol mice showed a more rapid and greater induction of cyclin D1 and cyclin E over that observed with air-treated controls after PHTx appearing as early as 3 hours in CO-treated mice versus 24 hours in air controls (Fig. 4B). Corroborating

the effects on the cyclins, assessment of the cyclin-dependent kinase (cdk) inhibitor p21, which controls cell cycle progression at G1 phase of the cell cycle, was decreased in livers from CO-treated mice beginning at 3 hours and remained unchanged and partially increased in air-treated mice through 24 hours (Fig. 4B), suggesting some degree of growth arrest which might be related to injury and loss of function (Fig. 2A). Finally, we assessed retinoblastoma (Rb), which regulates differentiation, apoptosis, cell cycle, and coordinates G1 to S phase transition. During G1 phase of the cell cycle, Rb converts from a transcriptionally repressive state to an inactive phosphorylated state by differential phosphorylation of serine and threonine residues. Similar to the cyclins, CO-treated this website mice showed strong phospho-Rb as early as 3 hours, versus 12 hours in controls (Fig. 4B). Taken together, these data support the proliferative index measured

in Fig. 1. Given this result, we next searched for a potential signaling mechanism and target of CO in enhancing liver regeneration and HC proliferation after PHTx. We first measured activation of Akt medchemexpress after PHTx, which is known to be a key regulator of hepatocyte size enlargement and proliferation.31 As expected, PHTx led to a modest increase in phospho-Akt over naïve animals, which was substantially augmented at each timepoint in CO-treated mice at 3, 6, and 24 hours after PHTx. Upstream of Akt, c-Met is also known to be activated during liver regeneration

after PHTx as the receptor for HGF. Air-treated mice showed a time-dependent increase in Met which peaked at 12 and 24 hours. This kinetic remained similar in CO-treated mice; however, the activation of Met was significantly greater and peaked as early as 3 hours (Fig. 4C). We also measured phospho-signal transducer and activator of transcription 3 (STAT-3), which is also known as a very important transcription factor in the mitotic response after PHTx. As with Akt and Met, PHTx induced STAT-3 phosphorylation, and surprisingly, CO blocked STAT-3 activation in the liver in response to PHTx (Fig. 4C), suggesting that STAT-3 was not involved in the ability of CO to enhance regeneration. Collectively these findings suggest that CO induces cell cycle entry more rapidly and extensively, which involves, in part, the induction of an Akt-HGF axis as the mechanism of action and presumably a blockade of STAT-3 activation.