Iron homeostasis is altered in response to hypoxia and hypothermic preconditioning in brain glial cells

Arzu L. ARAL,Alp Özgün BÖRCEK,Mehmet Ali ERGÜN,Ayşe Başak ENGİN,Hayrunnisa BOLAY

doi:10.3906/sag-2003-41

Abstract

Background/aimAltered iron metabolism is one of the pathophysiological mechanisms occurring during hypoxic injuries in the central nervous system. Proper homeostasis of cellular iron is regulated by iron import, storage, and export proteins that prevent excess iron overload or iron starvation in cells. Therapeutic hypothermia is an approved treatment for hypoxic ischemia in newborns, but the underlying molecular mechanism is still unknown. We studied the effects of hypoxia, preceded with preconditioning, on the iron homeostasis of glial cells, known as a major actor in the inflammatory process during perinatal brain injury.Materials and methodsPrimary microglia and astrocytes in culture were exposed to 12 h of hypoxia with or without mild hypothermic preconditioning. The mRNA expression was assessed using qPCR. Iron accumulation was visualized via modified Perl’s histochemistry. Cytokine levels in cell cultures were measured using ELISA.ResultsHypothermic preconditioning enhanced microglial viability, which previously was decreased in both cell types due to hypoxia. Hypoxia increased iron accumulation in the mixed glial cells and in ferritin expression in both microglia and astrocytes. Hypotermic preconditioning decreased the elevated ferritin-light chain expression significantly in microglia. Iron importer proteins, DMT1 and TfR1, both increased their mRNA expression after hypoxia, and hypothermic preconditioning continued to support the elevation of DMT1 in both glial cell types. Ferroportin expression increased as a survival factor of the glial cell following hypoxia. Hypothermic preconditioning supported this increase in both cell types and was especially significant in astrocytes. IL-10 levels were prominently increased in cell culture after hypothermic preconditioning.ConclusionThe data suggest that hypothermic preconditioning affects cellular iron homeostasis by regulating the storage and transfer proteins of iron. Regulation of the cellular iron traffic may prevent glial cells from experiencing the detrimental effects of hypoxia-related inflammation.

Highlights

Motor or cognitive neurological disorders originating from the perinatal period pose an increasingly serious health problem in developing countries [1]
Hypothermic preconditioning enhanced microglial viability, which previously was decreased in both cell types due to hypoxia
DMT1 and TfR1, both increased their mRNA expression after hypoxia, and hypothermic preconditioning continued to support the elevation of DMT1 in both glial cell types

Summary

Introduction

Motor or cognitive neurological disorders originating from the perinatal period pose an increasingly serious health problem in developing countries [1]. Hypoxic ischemic (HI) conditions are the main cause of brain damage in newborns. Excessive accumulation of iron causes iron-mediated free radical damage within the cell. Studies in infants and neonatal rats have shown that hypoxic-ischemic damage increases free iron levels in serum and cerebrospinal fluid [7]. Accumulated iron, which is associated with hypoxia at the cellular level, especially in microglia, stimulates reactive oxygen products, cytokine production, and the resultant apoptosis of oligondendrocytes. Microglia prevents excessive iron accumulation in the environment by binding it following hypoxic damage [8]. It has been suggested that excessive iron retained in microglia may be a regulator for oligodendrocyte cytotoxicity seen in periventricular white matter following hypoxia [9]. Transferrin receptor (TfR) and divalent methyl carrier (DMT), proteins involved in cellular iron transport are regulated with expression of HIF-1a in neurons, astrocytes, [11] and microglia [12]

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