mRNA and protein expression of Mcl-1L during liver regeneration.

<p>Remnant liver tissue from the indicated time points was used to determine (A) mcl-1 mRNA expression by q-RT-PCR; *P<0.05 and (B) protein levels of Mcl-1L as detected by Western blotting. (C) Parafilm sections of the remnant liver were used for Mcl-1 detection by IHC. Magnification, 400×.</p

IL-6 induced angiotensinogen expression in primary cultures of mouse hepatocytes.

<p>(A) Time course of IL-6-induced (10 ng/ml) angiotensinogen protein expression as detected by enzyme immunoassay (n = 3/group). (B) Dose response of IL-6-induced angiotensinogen protein expression after 24 hours of culture, as detected by enzyme immunoassay. Comparisons are between the indicated groups. *<i>P</i><0.05, n = 3.</p

JAK/PI3K/Akt/CREB signaling is involved in IL-6-induced Mcl-1L expression in hepatocytes.

<p>(A) Rat hepatocytes were pretreated with chemical inhibitors, JAK Inhibitor InSolution™ 420097 (20 nM), for 1 h prior to IL-6 (10 ng/ml) treatment. After 4 h, the Mcl-1L protein levels were determined by Western blotting. Between-group comparisons are as indicated with *p<0.05. (B) Rat hepatocytes were pretreated with chemical inhibitors, LY294002 (50 µg/mL), PD98059 (50 µg/mL), and staurosporine (20 nM) for 1 h prior to IL-6 (10 ng/ml) treatment. After 4 h, Mcl-1L expression was analyzed by Western blotting. Between-group comparisons are as indicated. *p<0.05. (C) Rat hepatocytes were pretreated with the NF-κB and CREB decoy ODN (10 µM) for 24 h prior to IL-6 treatment. After 4 h, mcl-1 mRNA expression was analyzed by q-RT-PCR. Data are fold-induction relative to the control (Lane 1). Between-group comparisons are as indicated. *p<0.05. (D) Rat hepatocytes were pretreated with JAK Inhibitor InSolution™ 420097 (20 nM), or LY294002 (50 µg/mL) for 1 h prior to IL-6 treatment. After 30 minutes, p-Akt, Akt, p-CREB and CREB were determined by Western blotting. Between-group comparisons are as indicated. *p<0.05.</p

Signal transduction pathways involved in IL-6-induced angiotensinogen expression in primary cultures of mouse hepatocytes.

<p>(A) Inhibition effects of chemical inhibitors 1 to 6 on IL-6-activated signaling mediators detected by Western blotting and quantified by calculating the ratios of phosphorylated/non-phosphorylated protein forms. The ratio in lane 1 is defined as 1. Comparison is between lanes 2 and 3 in each group. *<i>P</i><0.05, n = 3. (B) The effects of different chemical inhibitors on IL-6-induced angiotensinogen protein expression as detected by enzyme immunoassay. Comparison is between the indicated groups. *<i>P</i><0.05, n = 3.</p

IL-6 induced Mcl-1L expression in rat hepatocytes.

<p>(A) Serum levels of IL-6 in PH rats or flavopiridol (2.5 mg/kg) treated for 24 hours before PH group or control group were determined by EIA. Data are provided as mean +/− S.D. *P<0.05. (B) Rat hepatocytes were serum-starved for 24 hours prior treated with recombinant rat IL-6 (0.1, 1 and 10 ng/ml), under serum-starved conditions, and after 4 hours, Mcl-1L protein levels were determined by Western blot analysis. (C) Rat hepatocytes were serum-starved for 24 hours prior treated with recombinant rat IL-6 ( 1 ng/ml), under serum-starved conditions, at the indicated time periods, Mcl-1L protein levels were determined by Western blot analysis. (D) Rat hepatocytes were treated with rat recombinant IL-6 (10 ng/ml) at the indicated time periods. Mcl-1 mRNA expression in rat hepatocytes was determined by q-RT-PCR. *P<0.05.</p

Mcl-1L chemical inhibitor flavopiridol inhibits liver regeneration in rats.

<p>(A) Rats were pretreated with flavopiridol (2.5 mg/kg) or vehicle (control) 24 hours before partial hepatectomy (PH). (A) The remnant liver weight (RLW) of each group was measured at the indicated time periods after PH and expressed as a percentage of the original liver weight (OLW). Data are compared between the vehicle and flavopiridol-treated groups. *P<0.05. (B) The expression of Mcl-1L in remnant liver tissue was determined by IHC. (C) ki67 staining. (D) TUNEL staining. Magnification, 400×.</p

Effect of IL-6-related signaling inhibition on angiotensinogen protein expression and liver regeneration in remnant livers after 70% partial hepatectomy in mice.

<p>Mice were pretreated with chemical inhibitors of JAK2 (AG490, 10 mg/kg subcutaneously), p38 (intraperitoneal SB203580, 15 mg/kg), and STAT3 (intraperitoneal 5,15-DPP, 15 mg/kg) for 4 hours prior to partial hepatectomy. (A) Serum angiotensinogen levels of mice pretreated with different chemical inhibitors (AG490, SB203580, and 5,15-DPP) as detected by enzyme immunoassay. (B) Changes in the ratio of remnant to original liver weight after 70% partial hepatectomy. Remnant liver weight was estimated retrospectively from the excised liver weight after 70% PH. Data are presented as mean ± S.D., and comparisons were made between groups as indicated. *<i>P</i><0.05. (C) Ki-67 staining of regenerated liver tissue sections of the indicated group. Magnification, 400x. (D) Quantification of Ki-67 staining. Data presented here are the quantification of Ki-67-positive nuclei per high-power field. Data are presented as mean percentage of positive nuclei ± S.D., and comparisons were made between groups as indicated. *<i>P</i><0.05.</p

Changes in the ratio of remnant liver weight to original liver weight (RLW/OLW) after 70% partial hepatectomy (PH).

<p>(A) OLW was estimated retrospectively from the excised liver weight after 70% PH. Data are presented as mean ± S.D., and comparisons were made between groups as indicated. *P<0.05. (B) ki67 staining of remnant liver tissue. Magnification, 400×.</p

Schematic of the proposed signal transduction pathway of Mcl-1L induction in hepatocytes during liver regeneration.

<p>IL-6 regulates Mcl-1L expression through the IL-6 dependent JAK/PI3K/Akt/CREB activation pathway in rat hepatocytes.</p

Serum IL-6 and angiotensinogen levels, angiotenisogen mRNA and protein expression in remnant livers after 70% partial hepatectomy in mice.

<p>(A) Serum angiotensinogen levels detected by enzyme immunoassay (n = 5/group). (B) Angiotensinogen mRNA detected by reverse transcription-polymerase chain reaction quantified by calculating the ratios of angiotensinogen/GAPDH. (C) Angiotensinogen protein expression detected by Western blot and quantified by calculating the ratios of angiotensinogen/ß-actin. The ratio in lane 1 is defined as 1. Comparison is between the time 0 group and specified time periods. *<i>P</i><0.05, n = 5.</p