D.J. Macey

Iron Mineralization in the Radula Teeth of the Chiton Acanthopleura hirtosa

The structure, morphology and organization of the radula teeth of the chiton Acanthopleura hirtosa, together with their surrounding superior epithelium tissue, have been examined by using transmission electron microscopy and electron diffraction. Considerable amounts of iron are first seen in inclusions resembling ferritin aggregates and haemosiderin within the cells of the superior epithelium just before the onset of tooth-cusp mineralization. Iron appears to be delivered to the mineralizing surface of the cusp (presumably as FeII) via a series of microvilli, which extend out from the cells of the superior epithelium and which are contiguous with the surface of the tooth cusp. Before mineralization, the tooth cusp consists of a series of fine organic fibrils arranged perpendicular to the posterior edge but parallel to the anterior edge of the cusp. Mineralization within the cusp begins with the deposition of small spherules of ferrihydrite, which appear to have a close spatial relation to the organic matrix. Depending on their location within the cusp these spherules are rapidly replaced by crystals of either goethite, lepidocrocite and/or magnetite as mineralization proceeds. Mineralization is preferentially concentrated towards the posterior surface of the tooth cusp with a greater deposition of magnetite than is found on the anterior surface. We suggest that the initial control of mineralization in the tooth cusp is mediated by the organic matrix. However, the phase transformations among the several mineral phases found in the cusp may well be due to differential competition among reactions leading to these phases and this, in turn, may be controlled by the generation of differential redox and related potentials within the cusp.

The form of iron oxide deposits in thalassemic tissues varies between different groups of patients: a comparison between Thai β-thalassemia/hemoglobin E patients and Australian β-thalassemia patients

Mössbauer spectra of 12 beta-thalassemia/hemoglobin E spleen samples from Thai patients who had not received multiple blood transfusions and chelation therapy and seven beta-thalassemia spleen samples from Australian patients who had received multiple blood transfusions and chelation therapy were recorded with sample temperatures of 78 K. Each spectrum was found to consist of a superposition of a relatively intense central doublet characteristic of high-spin Fe(III), a low intensity sextet of peaks due to magnetic hyperfine-field splitting, and occasionally a doublet that could be attributed to heme iron. A significant (P=0.01) difference (Kolmogorov-Smirnov statistic of 0.71) between the distributions of sextet signal intensity as a fraction (Fs) of the total non-heme iron Mössbauer spectral signal for the two groups of patients was detected. The distribution of Fs for the Thai beta-thalassemia/hemoglobin E spleens had a mean value of 0.128 (S.D. 0.035) while that for the Australian beta-thalassemia spleens had a mean of 0.27 (S.D. 0.12). No significant difference between the distributions of non-heme iron concentrations in the tissues for the two groups of patients was detected by atomic absorption spectrometry. This study shows that the Australian beta-thalassemia patients had a higher fraction of their non-heme spleen iron in a goethite-like form than the Thai beta-thalassemia/Hb E patients.

Changes in serum thyroxine and triiodothyronine concentrations during metamorphosis of the Southern Hemisphere Lamprey Geotria australis, and the effect of propylthiouracil, triiodothyronine and environmental temperature on serum thyroid hormone concentrations of ammocoetes.

Serum thyroid hormone concentrations were measured during the seven stages of metamorphosis (1–7) of the southern hemisphere lamprey, Geotria australis. The respective mean concentrations ± SEM of serum thyroxine (T4) and triiodothyronine (T3) fell from 31.73 ± 4.09 and 5.06 ± 0.70 nM in large ammocoetes sampled in February, at the time when metamorphosis was initiated, to 4.54 ± 0.36 and 1.03 ± 0.12 nM at stage 5. Although there was a small, but significant, recovery of serum T4 concentrations during stages 6 and 7, no such corresponding statistically significant rise occurred in serum T3 concentrations.Serum thyroid hormone concentrations in ammocoetes sampled during the period when metamorphosis was taking place, exhibited a marked seasonal increase between February and May–June (late autumn/early winter); serum T3 and T4 concentrations peaked in May–June and were, respectively, > 2 fold and > 8 fold higher than those recorded for samples in late February (mid summer). By mid-July the serum T4 and T3 levels had declined from the peak values.Ammocoetes taken from streams at 16°C in June and acclimated to aquaria water at 25°C or 6°C had significantly lower serum T3 and T4 concentrations at the higher temperature, and also a lower serum T4, but not T3 concentration, at the lower temperature.Treatment of separate groups of ammocoetes with either propylthiouracil or T3 for 70 days significantly depressed and raised respectively, the serum thyroid hormone and hepatic T3 concentrations and caused significant changes in the body weight, but did not induce the onset of metamorphosis.

Characterization of biominerals in the radula teeth of the chiton, Acanthopleura hirtosa

Understanding biomineralization processes provides a route to the formation of novel biomimetic materials with potential applications in fields from medicine to materials engineering. The teeth of chitons (marine molluscs) represent an excellent example of a composite biomineralized structure, comprising variable layers of iron oxide, iron oxyhydroxide and apatite. Previous studies of fully mineralized teeth using X-ray diffraction, Raman spectroscopy and scanning electron microscopy (SEM) have hinted at the underlying microstructure, but have lacked the resolution to provide vital information on fine scale structure, particularly at interfaces. While transmission electron microscopy (TEM) is capable of providing this information, difficulties in producing suitable samples from the hard, complex biocomposite have hindered progress. To overcome this problem we have used focused ion beam (FIB) processing to prepare precisely oriented sections across interfaces in fully mineralized teeth. In particular, the composite structure is found to be more complex than previously reported, with additional phases (goethite and amorphous apatite) and interface detail observed. This combination of FIB processing and TEM analysis has enabled us to investigate the structural and compositional properties of this complex biocomposite at higher resolution than previously reported and has the potential to significantly enhance future studies of biomineralization in these animals.

Properties and role of ferritin in the hemolymph of the chiton Clavarizona hirtosa

The major iron-binding protein found in the hemolymph of the chiton Clavarizona hirtosa has been purified for the first time and identified as ferritin. This ferritin, which is present at a concentration of approx. 400 μg·ml−1, has a Mr of 28 000 and 25 500, exhibits microheterogeneity with isoelectric values in the range 5.3–6.0, binds 1500–2500 Fe atoms·mol−1 and is immunologically distinct from horse spleen ferritin. The initial rate of iron accumulation by ferritin molecules was determined to be markedly higher than that exhibited by horse spleen ferritin. Taken together, these data suggest that ferritin found in the hemolymph serves as a key component of the high-capacity transport system necessary to deliver iron to the rapidly mineralizing tissue of the radula in these molluscs.

Iron levels and major iron binding proteins in the plasma of ammocoetes and adults of the southern hemisphere lamprey geotria australis gray

1. The major plasma iron binding proteins (IBP) of both larval and adult Geotria australis differ from those of other vertebrates. 2. Larval IBP has a M.W. of approximately 354,000 and is composed of 20 subunits of equal M.W. 3. Adult IBP is a tetramer (M.W. = c. 296,000) which readily dissociates into subunits of approximately 78,000. 4. Confirmation that plasma iron binding proteins were similar to ferritin in larvae and transferrin in adults was provided by pI values, Fe/protein ratios and electron microscopy. 5. Total plasma iron was 19,760 micrograms/100 ml in larvae and 34 micrograms/100 ml in adults.

In situ Raman spectroscopic studies of the teeth of the chiton Acanthopleura hirtosa

Abstract In situ Raman spectroscopy, in combination with energy dispersive spectroscopy, has been used for the first time to determine the identities and locations, at the micron level, of mineral phases present in single chiton teeth that have been extensively mineralized. At the later stages of development the major lateral teeth of the chiton Acanthopleura hirtosa show characteristic spectroscopic evidence for the presence of lepidocrocite (γ-FeOOH), magnetite (Fe3O4), and an apatitic calcium phosphate. Goethite (α-FeOOH) and ferrihydrite (5 Fe2O3·9 H2O), which have been detected previously in teeth at the early stages of mineralization, were not detected in this mature tooth. The spatial distribution of these phases was determined, providing evidence for the presence of a discrete layer of lepidocrocite between the magnetite and apatite regions, illustrating the complexity of the biomineralization process. The technique of laser Raman microscopy is shown to be ideal for the examination of small biomineralized structures in situ, such as chiton teeth.

Organ-specific crystalline structures of ferritin cores in β-thalassemia/hemoglobin E

SummaryThe cores of ferritins isolated from different organs of human subjects withβ-thalassemia/hemoglobin E (β-thal/HbE) disease have different size distributions and crystallinities depending on the source organ. These patients have not been treated by hypertransfusion regimen or iron chelation therapy.β-Thal/HbE spleens and livers yield ferritin cores which are less crystalline than those isolated from normal spleens and livers, reflecting the more rapid deposition of iron in the diseased state. Ferritins isolated from the hearts and pancreases ofβ-thal/HbE subjects were found to have larger, more crystalline cores than those from theβ-thal/HbE livers and spleens, possibly as a consequence of the role of the heart and pancreas as long-term iron deposition sites in this iron overload pathology.

Calcium biomineralization in the radular teeth of the chiton, Acanthopleura hirtosa

SummaryA method has been devised for isolating the calcium biomineral from the iron biominerals and organic components present in the major lateral teeth of the chiton Acanthopleura hirtosa. Fourier-transform infrared spectroscopy of the calcium biomineral indicated that it was an apatite material containing carbonate and fluoride ions. Carbonate was not found to be present as a separate phase. The apatite was further separated into low and high density fractions, both of which showed crystallinity intermediate between that of bovine tibia cortical bone and human tooth enamel, as indicated by powder X-ray diffraction analysis. The calcified region of the major lateral teeth was also studied in situ using transmission electron microscopy and electron diffraction analysis, revealing a close spatial relationship between the mineral apatite phase and underlying organic matrix. It is suggested that the architectural arrangement of apatite biomineral and fibrous organic constituents imparts specialized mechanical properties to the tooth making it ideally suited for the task of obtaining food from hard surfaces.

Identification of ferritin as a major high molecular weight zinc-binding protein in the tropical rock oyster,Saccostrea cuccullata

The metal-binding proteins of the body mass of the tropical rock oysterSaccostrea cuccullata from the iron-ore shipping terminals near Dampier, Western Australia, have been separated by gel chromatography and analyzed for Zn, Fe, and Cu. Appreciable amounts of Zn (40%) and Cu (25%) were eluted in association with a high molecular weight fraction (>60,000 daltons), the major component of which was identified as the heat stable Fe-binding protein, ferritin (approx. mol. wt. 550,000), which also contains 95% of the Fe detected. Ferritin thus accounts for significant amounts of protein-bound Zn in this species and most probably in other bivalves containing elevated tissue levels of Zn.