Distribution Analysis of Iron and Copper by STEM-EDX Spectroscopy of Hemosiderin Particles in the Liver of Rats Overloaded With Iron.

BACKGROUND/AIM: Our recent studies have indicated that trace copper co-existed with iron in hemosiderin particles of human genetic iron overload. To understand this phenomenon, we analyzed hemosiderin particles in iron-overloaded rat liver by using scanning transmission electron microscopy - energy-dispersive X-ray (STEM-EDX) spectroscopy. MATERIALS AND METHODS: Samples for STEM-EDX spectroscopy were prepared from the liver of rats administered an intraperitoneal injection of dextran iron. RESULTS: The micro-domain analysis with STEM-EDX spectroscopy showed that dense bodies contained high levels of iron and trace copper. Quantitative analysis of copper levels in the liver specimen using atomic spectrophotometry showed that copper concentration in the liver was not increased by iron overload. These findings suggest that the overload of iron induced distribution of trace copper to hemosiderin particles without changing cellular copper levels. CONCLUSION: Co-existence of copper with iron was observed in hemosiderin particles of the liver of an experimental model of iron overload, suggesting that iron overload induced distribution of trace copper into hemosiderin particles.

(-)-Epigallocatechin-3-gallate Down-regulates Doxorubicin-induced Overexpression of P-glycoprotein Through the Coordinate Inhibition of PI3K/Akt and MEK/ERK Signaling Pathways.

BACKGROUND/AIM: (-)-Epigallocatechin-3-gallate (EGCG) has been indicated to regulate the function of P-glycoprotein (P-gp), which is a drug transporter encoded by the MDR1 (ABCB1) gene. P-gp expression is induced by doxorubicin (DOX). We aimed to clarify the mechanisms and inhibitory effects of EGCG on DOX-induced P-gp expression in HepG2 cells. MATERIALS AND METHODS: Rhodamine 123 (Rho123) was used for P-gp substrate. Western blotting and polymerase chain reactions (PCRs) were conducted using specific antibodies and primer sets. RESULTS: The DOX-pretreated cells accumulated a significantly decreased amount of Rho123), than control cells; however, the cells pretreated with EGCG and DOX, in combination, accumulated Rho123 more than DOX-pretreated cells. DOX induced the overexpression of MDR1 mRNA and increased the phosphorylation of Akt, ERK1/2, p38 MAPK and JNK. EGCG significantly inhibited the phosphorylation of Akt and ERK. The DOX-induced P-gp overexpression was partially suppressed by an inhibitor of MEK1/2 (U0126), but not by a PI3K inhibitor (LY294002). Interestingly, the expression of P-gp was synergistically inhibited by combined treatment of U0126 with LY294002 and also inhibited by an mTORC1 inhibitor, rapamycin. CONCLUSION: EGCG inhibited DOX-induced overexpression of P-gp through the coordinate inhibitory action on MEK/ERK and PI3K/Akt signaling pathways.

Inclusion bodies of aggregated hemosiderins in liver macrophages.

Hemosiderin formation is a structural indication of iron overload. We investigated further adaptations of the liver to excess iron. Five patients with livers showing iron-rich inclusions larger than 2 µm were selected from our database. The clinical features of patients and structures of the inclusions were compared with those of 2 controls with mild iron overload. All patients had severe iron overload with more than 5000 ng/mL of serum ferritin. Etiologies were variable, from hemochromatosis to iatrogenic iron overload. Their histological stages were either portal fibrosis or cirrhosis. Inclusion bodies were ultra-structurally visualized as aggregated hemosiderins in the periportal macrophages. X-ray analysis always identified, in addition to a large amount of iron complexes including oxygen and phosphorus, a small amount of copper and sulfur in the mosaic matrixes of inclusions. There were no inclusions in the control livers. Inclusion bodies, when the liver is loaded with excess iron, may appear in the macrophages as isolated organella of aggregated hemosiderins. Trace amounts of copper-sulfur complexes were always identified in the mosaic matrices of the inclusions, suggesting cuproprotein induction against excess iron. In conclusion, inclusion formation in macrophages may be an adaptation of the liver loaded with excess iron.

Coexistence of Copper in the Iron-Rich Particles of Aceruloplasminemia Brain.

The interaction between iron and copper has been discussed in association with human health and diseases for many years. Ceruloplasmin, a multi-copper oxidase, is mainly involved in iron metabolism and its genetic defect, aceruloplasminemia (ACP), shows neurological disorders and diabetes associated with excessive iron accumulation, but little is known about the state of copper in the brain. Here, we investigated localization of these metals in the brains of three patients with ACP using electron microscopes equipped with an energy-dispersive x-ray analyzer. Histochemically, iron deposition was observed mainly in the basal ganglia and dentate nucleus, and to lesser degree in the cerebral cortex of the patients, whereas copper grains were not detected. X-ray microanalysis identified two types of iron-rich particles in their brains: dense bodies, namely hemosiderins, and their aggregated inclusions. A small number of hemosiderins and most inclusions contained a significant amount of copper which was enough for distinct Cu x-ray images. These copper-containing particles were observed more frequently in the putamen and dentate nucleus than the cerebral cortex. Coexistence of iron and copper was supported by good correlations in the molecular ratios between these two metals in iron-rich particles with Cu x-ray image. Iron-dependent copper accumulation in iron-rich particles may suggest that copper recycling is enhanced to meet the increased requirement of cuproproteins in iron overload brain. In conclusion, the iron-rich particles with Cu x-ray image were found in the ACP brain.