1. Volume 44, Issue 2, Pages 391-399 (February 2006)
Iron overload promotes Cyclin D1 expression and alters cell cycle in mouse hepatocytes 
Marie-Bérengère Troadec, Brice Courselaud, Lénaïck Détivaud, Christelle Haziza-Pigeon, Patricia Leroyer, Pierre Brissot, Olivier Loréal 
Journal of Hepatology 
Volume 44, Issue 2, Pages (February 2006)
DOI: /j.jhep
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

2. Fig. 1
Iron exposure induces Liver Iron Concentration increase and hepatomegaly. Iron overload obtained using carbonyl–iron supplementation (37 weeks) or iron–dextran injection (1 week or 8 weeks) was associated with an increase of (A) Liver Iron Concentrations and (C) Ratio of liver weight/total animal weight. (A) Student's t-test appreciates the significance of results (*P<0.05; **P<0.01). (B) These models of iron-overload showed distinct histological features of iron deposition, as visualised in blue by Perls'staining. In carbonyl–iron and 8 weeks iron–dextran models, hepatocytes were iron overloaded (arrow head), whereas in 8 weeks iron–dextran livers, iron was mainly stored in Kupffer cells (arrow). Original magnification ×200.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

3. Fig. 2
Iron exposure induces nuclear ploidy. Typical analysis of nuclear ploidy profiles by flow cytometry of (A) mouse lymphocytes, and hepatocytes isolated from (B) 1-week iron–dextran and (C) 2-month iron–dextran. (Red line=iron overloaded mouse; blue=control mouse of identical age). We performed t-test (*P<0.05, **P<0.005) comparing to respective controls, on three independent mice (see Table 1).
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

4. Fig. 3
Expression of Cyclin D1 protein in liver and hepatocytes. Western blots were performed using polyclonal anti-Cyclin D1 antibody on (A) BALB/cJ mice total livers and on rat hepatocytes stimulated or not with Epithelial Growth Factor (EGF) used as positive and negative controls, (B) C57BL/6 mouse total livers, and (C) freshly isolated C57BL/6 mouse hepatocytes. Iron exposure duration and treatment are indicated upon western blot figure. Corresponding densitometries are presented on the right panel. A Student's t-test appreciated the significance of results (*, P<0.05). (D) Relationship between Cyclin D1 protein level quantified by densitometry, and Cyclin D1 mRNA level quantified by quantitative RT-PCR (log 2(Cyclin D1/Actin)) in the 1-week/2 months iron–dextran/dextran C57BL/6 mice. Correlation appreciated by a Spearman test.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

5. Fig. 4
Expression of Cyclin D1 protein in cultured iron overloaded hepatocytes. Western blot were performed using polyclonal anti-Cyclin D1 antibody, on hepatocytes isolated from (A) 1-week iron–dextran or (B) 2-month iron–dextran C57BL/6 mice, and controls. Four hours after cell seeding, cultures were stimulated (+) or not (−) by 50ng/mL Epithelial Growth Factor (EGF). Each experiment has been reproduced twice. Corresponding densitometries are presented below.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

6. Fig. 5
In vitro hepatocytes DNA synthesis differs from controls depending of iron exposure duration. [3H]-methyl-thymidine was incubated 24h prior measurements, on isolated hepatocytes from (A) 1-week iron–dextran, (B) 2 months iron–dextran C57BL/6 mice or respective controls. Cultures were stimulated or not by 50ng/mL Epithelial Growth Factor (EGF). [3H]-methyl-thymidine incorporation is expressed in cpm/mg proteins. Each curve has been reproduced twice by independent experiments.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

7. Fig. 6
Mitotic index of iron overloaded in vitro hepatocytes. Mitotic index was calculated on 1000 nuclei in isolated hepatocytes from (A) 1-week iron–dextran, (B) 2 months iron–dextran C57BL/6 mice and respective control dextran C57BL/6 mice, stimulated (+) or not (−) by 50ng/mL Epithelial Growth Factor (EGF).
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions