Studies on brain ferritin

Ferritin was isolated from the livers and brains of two groups of rats, one of which was fed AICI₃ (100/μM) for one year in the drinking water. Brain contained about a third of the ferritin found in the liver. While brain ferritin from normal rats contained 42.1 ± 14.3 g atoms of aluminum (Al⁺³), that from the Al⁺³-fed group contained 115.4 ± 48.3 g atoms per mole of ferritin. Liver ferritin from both groups contained less Al⁺³ and and more iron (Fe) compared with brain ferritin.

Ferritin isolated from the brains of patients who died of Alzheimer's disease (AD) contained more Al⁺³ and more iron than that from age-matched controls. Human brain ferritin (HBF) is composed of two types of subunits, about 70% heavy (Mr 22,000) and 30% light (Mr 19,500). The isoelectric focusing pattern of HBF was considerably different from that of human liver ferritin (HLF). Only five of the twenty brain ferritin bands migrated similarly to the acidic isoferritins from the liver and the major component of human brain ferritin, representing 30% of the total ferritin, had a pi of 8.

We have further characterized rat brain ferritin and human brain ferritin isolated form both AD and normal brain tissue with respect to the effect of Al⁺³ on iron loading and release and subunit composition. Al⁺³ causes a concentration dependent decrease in the initial rate of iron loading into demetallo-apo human brain ferritin and the rates were similar for ferritin isolated from both AD and normal tissue. The rates of iron release were: horsespleen ferritin (HSF) >> HLF > RBF > HBF = rat liver ferritin. The rates of iron release from AD and Normal ferritin were similar and were unaffected by preloading with Al⁺³. Several mammalian ferritins were compared for their total iron uptake; HSF > HLF > HBF. Comparison of human brain ferritin from AD and N brain tissue showed no statistical difference in their iron uptake. Consistent with our earlier observations, holoferritin is more resistant to precipitation by Al⁺³ than apoferritin. Human brain ferritin displays two subunits on SDS-PAGE: Mr 22,000(H) and 19,000(L) which show immunological reactivity with antihuman H chain and anti-human liver ferritin (L) antibodies, respectively. Rat brain ferritin displays three subunits on SDS-PAGE Mr 23,000(H), 22,000(M) and 20,000(L). Only the L chain reacted with anti-human L chain antibodies. The H and M chains did not react with anti-human H chain or anti-human L chain antibodies.

Concavalin A (Con A) is reported to be a ferritin. However this protein had no amino acid sequence homology with any of the known ferritins and did not react specifically with either anti-human brain or anti-soybean V ferritin antibodies.

Reverse phase HPLC resolved human brain ferritin into clusters of H (7 peaks) and L(5 peaks) components. The two major H peaks represented 24% and 41% of the total H and one major L peak represented 74% of the total L protein. The cluster of H-chain peaks migrated with an R_f on SDS-PAGE similar to that of the human brain ferritin H chain. Amino acid analysis of two homogeneous H chain peaks were very similar. Comparison of the amino acid composition of human brain ferritin H and L chains with the respective human liver ferritin subunits shows that while they are similar, the human brain ferritin H chain has fewer lys residues and the human brain ferritin L chain has more glx residues. Human brain homogenates in sucrose were fractionated by centrifugation. The ferritin distribution was: cellular debris 19.3%, nuclear 1.7%, mitochondrial 5.4%, microsomal 15.6% and cytosolic 58.0%. Most of the iron was found in the cellular debris (60%) and microsomal fractions (26%). While serum ferritin is a glycoprotein, SDS-PAGE gels of human brain and rat brain ferritin did not stain for carbohydrates. These data suggest that the L chain is more conserved between species and H chain heterogeneity implicates the H chain in differing isoferritin functions.