The Role And Possible Mechanism Of Iron Element In Liver Fibrosis

Objective : Experiments in vivo, the ma le Sprague–Dawley rat mo dule of iron deposition in liver. Experiments in vitro, the possible mechanism of iron tranportation in mononuclear phagocyte with the HIF- 1α pathway were observed in Raw264.7 cells.Methods: 60 male Sprague–Dawley rats were randomly divided into three groups by weight: normal group and iron dextran group with 30 rats respectively. Iron dextran(50mg/kg) was administrated as intramuscular injection to each rat for three periods as four months, five months and six months. During each period, we had 20 rats sacrificed with 10 rats in each group. We recorded the body weight, liver weight, biochemical and fibrotic indicators of each rat. The m RNA expression and protein expression of HIF-1α、FPN1、DMT1 after the administration of AFC on Raw264.7 for 24 hours in three different doses(25 μM、50 μM、100 μM) was measured by RT-PCR and Western Blot. Experiments in vitro contain four groups: control group, AFC low dose group, medium dose group and high dose group. The m RNA expression and protein expression of HIF-1α、FPN1、DMT1 after the administration of YC-1(100 μM) and the interaction of YC-1(100 μM) and AFC, in three different doses(25 μM、50 μM、100 μM), on Raw264.7 for 24 hours was measured by RT-PCR and Western Blot. Containing five groups: control group, YC-1 group, YC-1-AFC low dose group, medium dose group and high dose group.Results: Comparing to the control group, the liver weight of Iron Dextran group is significantly rised(P<0.05). By the way, the biochemical index shows that liver injury is appeared, considering with fibrotic data, so we come to the conclusion that the intramuscular injection of Iron Dextran causes liver injury and gradual liver fibrosis with longer injection period. Experiments in vitro,after the administration of distinctive doses of AFC on Raw264.7, RT-PCR results show that the different amount of HIF-1α m RNA expression between each group have no significant variance(P>0.05), while the amount of FPN1 m RNA expression in high dose group(100 μM) is significant higher and DMT1 m RNA expression was decreased significantlyboth comparing to control group(P<0.05). Western Blot show the same result in HIF-1α protein expression(P>0.05), but comparing to other groups, the high dose group(100 μM) show the significant higher protein expression in FPN1(P<0.05). When the high dose group(100 μM) shows the significant lower protein expression in DMT1 compared with control group(P<0.05), the medium dose group(50 μM) and high dose group(100 μM) also have a significant lower DMT1 protein expression compared with low dose group(25 μM)(P<0.05). Under the interaction of YC-1(100 μM),there are different results came up. After the administration of YC-1(100 μM) and distinctive doses of AFC on Raw264.7, RT-PCR results show that the different amount of HIF-1α m RNA expression between each group has significant higher variance. Comparing to control group, YC-1 group(100 μM) and YC-1(100 μM)-AFC medium dose(50 μM) group both show significant higher diffence in HIF-1α m RNA expression(P>0.05). Except the YC-1(100 μM)-AFC low dose(25 μM) group, other groups show significant higher FPN1 m RNA expression than control group(P<0.05) and those four experimental groups all show higher DMT1 m RNA expression than control group as well(P<0.05). Western Blot results show the decrease of HIF-1α、FPN1、DMT1 protein expression in each experimental group comparing to control group. The YC-1(100 μM)-AFC low dose group(25 μM) and YC-1(100 μM)-AFC high dose group(100 μM) both show the significant lower protein expression than control group in HIF-1α(P<0.05). Also YC-1(100 μM)-AFC high dose group(100 μM) shows the significant lower protein expression in FPN1 compared with other groups, except the low dose group(P<0.05); comparing to the control group, other groups, except the YC-1(100 μM)-AFC medium dose group(50 μM) as well, have a significant decrease in DMT1 protein expression(P<0.05).Conclusions: Intramuscular injection of iron dextran four to six months turns to be proved that it causes the genesis of liver fibrosis and grows serious as time goes. After the analysis of results of AFC administraion, the function of HIF-IRP-FPN1/DMT1 pathway in iron transportation of liver macrophage cells is not certainly founded. Due to the insignificant variance of the expression of HIF-1α, the evidence of the changed expression of FPN1 and DMT1 is not enough to make a conclusion that confirming the pathwat’s value. The YC-1 depresses the expression of HIF-1αand, to some extent, causes the changed expression of FPN1 and DMT1, which means the lower iron transportion rate. Therefore, this further experiment proves the HIF-IRP-FPN1/DMT1 pathway’s value in liver macrophage cells. The HIF-1α protein could also control iron transportation in liver macrophae cells with the help of FPN1 and DMT1 protein and maybe other unknow pathways.