(A) Immunostainings showed that hepatocytes were normally quiescent but proliferated at 18 hours after 2/3 PH in control mice. Hepatocytes of mice injected with miR-21–ASO at 6 hours, but not at 10 hours, after 2/3 PH failed to express markers of progression through G1 (cyclin D1 and Ki67, brown) and into S (Ki67 and PCNA, brown) phase of the cell cycle. Original magnification, ×200. (B) Quantification of hepatocytes expressing markers of cell cycle progression. For each immunostaining, approximately 1,500 hepatocytes (250 per frame) were analyzed per time point and treatment. (C) Immunoblotting showed failure to increase cyclin D1 protein levels in livers of mice injected with miR-21–ASO at 6 hours, but not at 10 hours, after 2/3 PH. Numbers indicate protein levels relative to controls. Gapdh was analyzed as a loading control. (D) qRT-PCR showed that miR-21–ASO injection did not interfere with induction of liver Ccnd1 transcription after 2/3 PH. At least 3 mice were analyzed for each time point and treatment. No differences were observed between control mice injected with carrier at 6 versus 10 hours after 2/3 PH. Data represent mean ± SEM. *P < 0.05, **P < 0.01.