T1 and T2 relaxation times and iron concentrations were measured in 24 specimens of gray matter from fresh human and monkey brains at magnetic fields from 0.05 to 1.5 Tesla. Three different effects were found that correlate with iron content: a T1-shortening that falls off somewhat at high fields, a T2-shortening that is field-independent and thus important at low fields, and a contribution to 1/T2 that increases linearly with field strength. This linear field dependence has been seen only in ferritin and other ferric oxyhydroxide particles. Our results are in agreement with in vivo MRI studies and are generally consistent with values for ferritin solution, except for differences such as clustering of ferritin in tissue. A cerebral cavernous hemangioma specimen showed similar T2-shortening, but with a 2.7 times larger magnitude, attributed to larger clusters of hemosiderin in macrophages. The dependence on interecho time 2 tau was measured in three brains; 1/T2 increased significantly for tau up to 32 ms, as expected from the size of the ferritin clusters. These findings support the theory that ferritin iron is the primary determinant of MRI contrast in normal gray matter.

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This paper aims to define the role of the threefold intersubunit channels in iron uptake and sequestration processes in the iron-storage protein, ferritin. Iron uptake, measured as loss of availability of Fe(II) to ferrozine (due to oxidation), has been studied in recombinant human H-chain ferritins bearing amino acid substitutions in the threefold channels or ferroxidase centres. Similar measurements with recombinant horse L-chain ferritin are compared. It is concluded that significant Fe(II) oxidation occurs only at the H-chain ferroxidase centres and not in the threefold channels, although this route is used by Fe(II) for entry. Investigations by Mössbauer and u.v.-difference spectroscopy show that part of the iron oxidized by H-chain ferritin returns to the threefold channels as Fe(III). This monomeric Fe(III) can be displaced by addition of Tb(III). Fe(III) also moves into the cavity for formation of the iron-core mineral, ferrihydrite. Iron incorporated into ferrihydrite becomes kinetically inert.

The paper describes a study of Fe(II) oxidation and the formation of Fe(III)-apoferritin complexes in recombinant human H-chain ferritin and its variants. The effects of site-directed changes in the conserved residues associated with a proposed ferroxidase centre have been investigated. A change in any of these residues is shown to reduce the rate of Fe(II) oxidation, confirming the importance of the ferroxidase centre in the catalysis of Fe(II) oxidation. Mössbauer and u.v.-difference spectroscopy show that in the wild-type protein Fe(II) oxidation gives rise to Fe(III) monomers, dimers and larger clusters. The formation of Fe(III) mu-oxo-bridged dimers occurs at the ferroxidase centre and is associated with fast oxidation: in three variants in which Fe(II) oxidation is especially slow, no Fe(III) dimers are seen. Within the time scale 0.5-20 min in wild-type human H-chain ferritin, dimer formation precedes that of the monomer and the progression dimer-->monomer-->cluster is observed, although not to completion. In a preliminary investigation of oxidation intermediates using a stopped-flow instrument, an Fe(III)-tyrosine complex reported by Waldo et al. (1993), is attributed to Tyr-34, a residue at the ferroxidase centre. The Fe(III)-Tyr-34 complex, forms in 0.5 s and then decays, as dimer absorbance increases. The relationship between Fe(III)-tyrosinate and the formation of Fe(III) dimers is uncertain.