Fine structure of in situ microbial iron deposits

The fine structure of iron deposits produced by Leptothrix spp. from two caverns and a surface spring were observed. In the case of Leptothrix pseudovacuolata, the iron occurred as an amorphous deposit within the cell sheath and the cells contained gas vesicles. In the case of the other Leptothrix spp., the iron was deposited within the cell sheath either as a hexagonal or fibrillar matrix. In the case of fibrillar iron deposits, fibrils resembling those of the sheath were found within the cytoplasm of the cells. This suggests that the deposition of iron could have occurred within the cells as well as at the cell surface. In some cases, the fine structure of microbial iron deposits could provide a means of distinguishing biological from abiological iron deposition.     

Clinical, anatomical and biophysical data on a case of encephalopathy with aluminum deposits]

A man presented at 27 years of age, the first signs of a progressive neurological and mental deterioration which eventually lead to his death at 37, from an intercurrent pulmonary infection. He was a severe alcoholic. The pathological examination of the brain revealed patchy demyelination and deposits of crystalline material, resembling so called calcifications. The biophysical examination showed the presence in the brain and other organs, of aluminum and phosphorus in all deposits, sometimes associated with iron, sulfur and calcium. Nothing in his past history suggests any contact with aluminum. Presumably this is a case of "generalized aluminosis", with the main clinical and pathological features manifesting in the central nervous system.     

Topographical localization of iron in brains of the aged fat-tailed dwarf lemur (Cheirogaleus medius) and gray lesser mouse lemur (Microcebus murinus)

Iron deposits in the human brain are characteristic of normal aging but have also been implicated in various neurodegenerative diseases. Among nonhuman primates, strepsirhines are of particular interest because hemosiderosis has been consistently observed in captive aged animals. In particular, the cheirogaleids, because of their small size, rapid maturity, fecundity, and relatively short life expectancy, are a useful model system for the study of normal and pathological cerebral aging. This study was therefore undertaken to explore iron localization in the brain of aged cheirogaleids (mouse and dwarf lemurs) with histochemistry and magnetic resonance microscopy. Results obtained with both techniques were comparable. There was no difference between old animals in the two species. The young animals (3 years old) showed no iron deposits. In the old animals (8–15 years old), iron pigments were mainly localized in the globus pallidus, the substantia nigra, the neocortical and cerebellar white matter, and anterior forebrain structures, including the nucleus basalis of Meynert. This distribution agrees with previous findings in monkeys and humans. In addition, we observed iron in the thalamus of these aged nonhuman primates. Microscopic NMR images clearly reveal many features seen with the histochemical procedure, and magnetic resonance microscopy is a powerful method for visualizing age-related changes in brain iron. Am. J. Primatol. 45:291–299, 1998. © 1998 Wiley-Liss, Inc.     

Electron-microscopic demonstration of the distribution of calcium deposits in the exocrine pancreas of the rat after application of carbachol,atropine, cholecystokinin,and procaine

Summary In an attempt to identify a cellular Ca²⁺-pool, from which calcium is released when secretagogues are applied, tissue fragments of the rat exocrine pancreas were incubated and fixed with glutaraldehyde in the presence of calcium. By means of this procedure electron-dense deposits were found on plasma membranes. X-ray microanalysis showed that these deposits contain calcium. Stimulation of tissue fragments with the use of the secretagogues carbachol or cholecystokinin reduced the number of deposits by about 80%. When the antagonist atropine was applied after carbachol stimulation, deposits reappeared on cell membranes, which then disappeared again after a second stimulation with cholecystokinin. In the presence of procaine, carbachol was inhibited and only slightly reduced the Ca²⁺-deposits on the plasma membranes.These results suggest that a calcium pool, from which calcium is released to induce enzyme secretion on stimulation, is located in the cell membrane     

Bleomycin-Detectable Iron in Brain Tissue

The normal brain contains regions with high concentrations of iron, part of which appears to be in a low molecular mass chelatable form. Iron complexes with a molecular mass of below 10,000, were measured in ultrafiltrates of homogenized gerbil brains using the bleomycin assay, and were found to average 20.5 ± 3.5 μM (n = 8). As expected, no bleomycin detectable iron was found in the plasma of these animals.

No obvious difference in the tissue levels of bleomycin-detectable iron was recorded following ischaemia and reperfusion. This is probably due to the already abundant presence of iron in the brain and the likely release of iron from protected sites due to structural damage inherent in the preparative procedures used.     

Insulin immunohistochemistry of rodent CNS: Apparent species differences but good correlation with radioimmunological data

Summary Using an indirect immunofluorescence technique the distribution of insulin was mapped in brains of Wistar strain rats and mice. Insulin immunoreactivity was found to be widely distributed throughout mouse CNS, whereas in rat brain a restriction of immunoreactive material to cerebral blood vessels and ependymal cells and/or tanycytes of the brain ventricles was observed. In radioimmunological studies the amount of insulin (IRI) was estimated for different brain areas (cerebral cortex, brain stem, cerebellum, hippocampus, thalamus and hypothalamus). In the case of Wistar rats very low levels of IRI were found. On the contrary, the same regions in mouse brain contained considerably greater amounts of IRI. The comparison between histochemical and biochemical data revealed a good correlation. It is concluded that part of the insulin measured by radioimmunoassay is associated with neuronal structures.Research supported by the Ministerium für Hoch- und Fachschulwesen der DDR and Ministerium für Gesundheitswesen der DDR     

Haemopexin affects iron distribution and ferritin expression in mouse brain

Haemopexin (Hx) is an acute phase plasma glycoprotein, mainly produced by the liver and released into plasma where it binds heme with high affinity and delivers it to the liver. This system provides protection against free heme‐mediated oxidative stress, limits access by pathogens to heme and contributes to iron homeostasis by recycling heme iron. Hx protein has been found in the sciatic nerve, skeletal muscle, retina, brain and cerebrospinal fluid (CSF). Recently, a comparative proteomic analysis has shown an increase of Hx in CSF from patients with Alzheimer’s disease, thus suggesting its involvement in heme detoxification in brain. Here, we report that Hx is synthesised in brain by the ventricular ependymal cells. To verify whether Hx is involved in heme scavenging in brain, and consequently, in the control of iron level, iron deposits and ferritin expression were analysed in cerebral regions known for iron accumulation. We show a twofold increase in the number of iron‐loaded oligodendrocytes in the basal ganglia and thalamus of Hx‐null mice compared to wild‐type controls. Interestingly, there was no increase in H‐ and L‐ferritin expression in these regions. This condition is common to several human neurological disorders such as Alzheimer’s disease and Parkinson’s disease in which iron loading is not associated with an adequate increase in ferritin expression. However, a strong reduction in the number of ferritin‐positive cells was observed in the cerebral cortex of Hx‐null animals. Consistent with increased iron deposits and inadequate ferritin expression, malondialdehyde level and Cu–Zn superoxide dismutase‐1 expression were higher in the brain of Hx‐null mice than in that of wild‐type controls. These data demonstrate that Hx plays an important role in controlling iron distribution within brain, thus suggesting its involvement in iron‐related neurodegenerative diseases.     

Determination of Zinc,Copper, Iron,and Manganese in Different Regions of Lamb Brain

The levels of zinc, copper, iron, and manganese were measured in the hippocampus, thalamus, gyrus cinguli, hypothalamus, and in the prefrontal, parietal, temporal, and occipital cortexes of lamb brain by flame atomic absorption spectrometry. Standard addition method was also applied to samples and it was observed that there was no matrix interference affecting the determination of elements interested. In the determination of Cu, a slotted tube atom trap was used to improve sensitivity. One-way analysis of variance was used for the statistical evaluation of the results for each element. Limit of detection results for Zn, Cu, Fe, and Mn were found to be 24, 4.5, 60, and 25 ng/mL, respectively. Region-specific differences were found for all elements interested. Mn concentration ranging from 0.29 ± 0.04 to 0.61 ± 0.04 mg/kg was found to be the lowest in all brain regions among the all elements interested.     

Alpha-melanocyte-stimulating hormone (alpha-MSH) in the brain of the cartilagenous fish. Immunohistochemical localization and biochemical characterization

The distribution of immunoreactive alpha-melanocyte-stimulating hormone (alpha-MSH) in the central nervous system and pituitary of the elasmobranch fish Scyliorhinus canicula was determined by the indirect immunofluorescence and the peroxidase-antiperoxidase methods using a highly specific antiserum. Perikarya containing alpha-MSH-like immunoreactivity were localized in the dorsal portion of the posterior hypothalamus, mainly in the tuberculus posterioris and sacci vasculosus nuclei. Immunoreactive alpha-MSH cell bodies were found in the dorsal wall and ventral region of the caudal part of the tuberculum posterioris. These structures were densely innervated by fine beaded immunoreactive fibers. Some alpha-MSH immunoreactive cells were occasionally detected in the ventral part of the nucleus periventricularis. Scattered cell bodies and fibers were also observed in the dorsal wall of the posterior recess. Outside the hypothalamus very few fibers were detected in the dorsal thalamus and mesencephalon. No immunoreactivity was found in any other parts of the brain. The alpha-MSH immunoreactive material localized in the brain was characterized by combining high-performance liquid chromatography (HPLC) analysis and radioimmunological detection. Brain and pituitary extracts exhibited displacement curves which were parallel to that obtained with synthetic alpha-MSH. The concentrations of alpha-MSH immunoreactive material were determined in 5 different regions of the brain. The highest concentration was found in the hypothalamus. HPLC analysis resolved two major forms of immunoreactive alpha-MSH in the hypothalamus, which had been same retention times as des-N alpha-acetyl-alpha-MSH and its sulfoxide derivative. These results provide the first evidence for the presence of alpha-MSH-like peptides in the fish brain.(ABSTRACT TRUNCATED AT 250 WORDS)     

Resistance of peripheral and sub-cortical somatosensory pathway to electrical noise

The somatosensory system is vulnerable to large amounts of noise distortion. But how does the central nervous system distinguish the peripheral inputs which carry information to the brain from that which does not possess information? To address this question we studied the effect of electrical stimulation of the median nerve on tactile spatial frequency perception in healthy subjects and Parkinson's disease (PD) patients. Subjects were categorized in two groups (healthy and PD patients) and were asked to report if a test tactile frequency pattern (TFP) was the same as the reference TFP given to the other hand. In each case stimulation was either present or absent on the median nerve of the hand holding the test pattern. We observed no impairment of tactile performance in the presence of electrical stimulation of the median nerve. This result together with previous work on direct stimulation of the somatosensory relay nucleus of the thalamus (Abbassian et al., Stereotact Funct Neurosurg 76: 19–28, 2001) in which the same result of no impairment of the tactile discrimination task was observed suggest a high degree of noise tolerance exists in the somatosensory pathway.     

Absence of an orphan mitochondrial protein, c19orf12, causes a distinct clinical subtype of neurodegeneration with brain iron accumulation

The disease classification neurodegeneration with brain iron accumulation (NBIA) comprises a clinically and genetically heterogeneous group of progressive neurodegenerative disorders characterized by brain iron deposits in the basal ganglia. For about half of the cases, the molecular basis is currently unknown. We used homozygosity mapping followed by candidate gene sequencing to identify a homozygous 11 bp deletion in the orphan gene C19orf12. Mutation screening of 23 ideopathic NBIA index cases revealed two mutated alleles in 18 of them, and one loss-of-function mutation is the most prevalent. We also identified compound heterozygous missense mutations in a case initially diagnosed with Parkinson disease at age 49. Psychiatric signs, optic atrophy, and motor axonal neuropathy were common findings. Compared to the most prevalent NBIA subtype, pantothenate kinase associated neurodegeneration (PKAN), individuals with two C19orf12 mutations were older at age of onset and the disease progressed more slowly. A polyclonal antibody against the predicted membrane spanning protein showed a mitochondrial localization. A histopathological examination in a single autopsy case detected Lewy bodies, tangles, spheroids, and tau pathology. The mitochondrial localization together with the immunohistopathological findings suggests a pathomechanistic overlap with common forms of neurodegenerative disorders.     

Ultrastructure of CaCO3 deposits of terrestrial isopods (Crustacea, Oniscidea)

 The ultrastructure of the sternal CaCO₃ deposits of 3 species of the Diplochaeta and 15 of the Crinochaeta was investigated by means of scanning electron microscopy of fractured surfaces. In the Diplochaeta Li-gia italica and L. oceanica, the deposits consist exclusively of individual spherules with diameters between 0.2 and 1.4 μm. No material was observed within the spaces between the spherules. In Ligidium hypnorum, two structurally distinct regions exist. A proximal layer resembling the deposit of Ligia italica and L. oceanica and a distal layer in which the spherules appear to be fused with each other. In the species of the Crinochaeta, the CaCO₃ deposits comprise a spherular region which resembles the deposits of Ligidium hypnorum, and a homogeneous layer located between the spherular part of the deposit and the hypodermal cell layer. In some species the diameters of the spherules may be up to 3.1 μm. In the homogeneous layer and the distal spherular layer more calcium per volume can be stored than in the proximal spherular layer in which the spaces between the spherules are devoid of CaCO₃. This suggests that the multiple layered deposits are an adaptation to terrestrial life, as a consequence of the need for increased resorption of cuticular calcium. Accepted: 7 January 1997     

Enhanced intestinal uptake of iron,zinc and calcium in rats fed pungent spice principles – Piperine,capsaicin and ginger (Zingiber officinale)

In view of the wide-spread deficiency of iron and zinc in populations dependent on plant foods, it is desirable to improve the bioavailability of the same. Specific dietary spices may alter the ultrastructure and permeability characteristics of intestines. Groups of Wistar rats were fed piperine, capsaicin and ginger containing diets for 8 weeks in order to examine their possible influence on intestinal absorption of iron, zinc and calcium. Everted segments of duodenum, jejunum and ileum portions of small intestines isolated from these rats were examined for ex vivo uptake of iron, zinc and calcium from incubations containing digesta of finger millet. Higher uptake of iron, zinc and calcium by the intestinal segments from spice-fed animals was observed. The increase in the mineral uptake was the highest for calcium with >100% in some cases. The positive influence of dietary capsaicin was more pronounced on zinc uptake as compared to that of iron. Uptake of the glutamic acid standard was 87% and 62% higher in the case of jejunal segments of rats fed piperine and ginger. The higher intestinal uptake of iron and zinc as a result of consumption of pungent spices could encourage a strategy to reduce deficiency of these trace elements prevalent in population dependent on plant based foods.     

Proopiomelanocortin (POMC)-related peptides in the brain of the rainbow trout, Salmo gairdneri

We have investigated the presence of ACTH, -MSH and β-endorphin, three peptides which derive from the multifunctional precursor protein proopiomelanocortin (POMC) in the brain of the rainbow trout Salmo gairdneri. Using both the indirect immunofluorescence and peroxidase-antiperoxidase techniques, a discrete group of positive cells was identified in the hypothalamus, within the anterior part of the nucleus lateralis tuberis. -MSH-containing neurons represented the most abundant immunoreactive subpopulation. Coexistence of -MSH, ACTH and β-endorphin was observed in the lateral part of the nucleus. ACTH- and β-endorphin-containing cells were mainly distributed in the rostral and caudal regions of the nucleus. In the medial portion of the nucleus lateralis tuberis, numerous cells were only stained for -MSH. Moderate to dense plexuses of immunoreactive fibers were observed in the ventral thalamus and the floor of the hypothalamus. Some of these fibers projected towards the pituitary. The concentrations of ACTH, -MSH and β-endorphin-like immunoreactivities were measured in microdissected brain regions by means of specific radioimmunoassays. Diencephalon, mesencephalon and medulla oblongata extracts gave dilution curves which were parallel to standard curves. The highest concentrations of POMC-derived peptides were found in the diencephalon (-MSH: 4.28±0.43 ng/mg prot.; ACTH: 1.08±0.09 ng/mg prot.; β-endorphin: 1.02±0.1 ng/mg prot.), while lower concentrations were detected in the mesencephalon, medulla oblongata and telencephalon. The present results demonstrate that various peptides derived from POMC coexist within the same cell bodies of the fish hypothalamus. Taken together, these data suggest that expression and processing of POMC in the fish brain is similar to that occurring in pituitary melanotrophs.     

The occurrence and fine structural characterization of microbodies inEuglena gracilis

Summary The fine structure of an organelle morphologically similar to microbodies found in higher plants and animals was studied in cells ofEuglena gracilis fixed simultaneously in glutaraldehyde and osmium tetroxide. These organelles were 0.4 to 0.8 microns in diameter, bounded by a single membrane, and frequently observed in close spatial association with both endoplasmic reticulum and mitochondria. Their finely granular matrices frequently contained membranous cores. Though these organelles were relatively abundant in acetate- and ethanolgrown cells, they were rarely observed in glucose-grown cells, an indication that they play the same role in the metabolism of 2-carbon substrates as do glyoxysomes in higher plants. The presence of these organelles, assumed to be microbodies, is also of considerable interest since catalase, an enzyme characteristic of microbodies from a variety of sources, was not detected.This work was supported in part by grant GY 3804 from the National Science Foundation to L.B.G.     

Ceruloplasmin and β-amyloid precursor protein confer neuroprotection in traumatic brain injury and lower neuronal iron

Traumatic brain injury (TBI) is in part complicated by pro-oxidant iron elevation independent of brain hemorrhage. Ceruloplasmin (CP) and β-amyloid protein precursor (APP) are known neuroprotective proteins that reduce oxidative damage through iron regulation. We surveyed iron, CP, and APP in brain tissue from control and TBI-affected patients who were stratified according to time of death following injury. We observed CP and APP induction after TBI accompanying iron accumulation. Elevated APP and CP expression was also observed in a mouse model of focal cortical contusion injury concomitant with iron elevation. To determine if changes in APP or CP were neuroprotective we employed the same TBI model on APP^−/− and CP^−/− mice and found that both exhibited exaggerated infarct volume and iron accumulation postinjury. Evidence supports a regulatory role of both proteins in defence against iron-induced oxidative damage after TBI, which presents as a tractable therapeutic target.     

Subcellular distribution of calcium during spermatogenesis of zebrafish,Danio rerio

Calcium plays a variety of vital regulatory functions in many physiological and biochemical events in the cell. The aim of this study was to describe the ultrastructural distribution of calcium during different developmental stages of spermatogenesis in a model organism, the zebrafish (Danio rerio ), using a combined oxalate–pyroantimonate technique. Samples were treated by potassium oxalate and potassium pyroantimonate during two fixation stages and examined using transmission electron microscopy to detect electron dense intracellular calcium. The subcellular distribution of intracellular calcium was characterized in spermatogonium, spermatocyte, spermatid, and spermatozoon stages. The area which is covered by intracellular calcium in different stages was quantified and compared using software. Isolated calcium deposits were mainly detectable in the cytoplasm and the nucleus of the spermatogonium and spermatocyte. In the spermatid, calcium was partially localized in the cytoplasm as isolated deposits. However, most calcium was transformed from isolated deposits into an unbound pool (free calcium) within the nucleus of the spermatid and the spermatozoon. Interestingly, in the spermatozoon, calcium was mainly localized in a form of an unbound pool which was detectable as an electron‐dense mass within the nucleus. Also, sporadic calcium deposits were scattered in the midpiece and flagellum. The proportional area which was covered by intracellular calcium increased significantly from early to late stages of spermatogenesis. The extent of the area which was covered by intracellular calcium in the spermatozoon was the highest compared to earlier stages. Calcium deposits were also observed in the somatic cells (Sertoli, myoid, Leydig) of zebrafish testis. The notable changes in the distribution of intracellular calcium of germ cells during different developmental stages of zebrafish spermatogenesis suggest its different homeostasis and physiological functions during the process of male gamete development.     

Microvascular anatomy of the brain of the adult pipid frog,Xenopus laevis (Daudin): A scanning electron microscopic study of vascular corrosion casts

To demonstrate the 3D microvascular anatomy of the brain of the model organism Xenopus laevis Daudin scanning electron microscopy of vascular corrosion casts was correlated with light microscopy of stained 7 µm thick serial tissues sections. Results showed that supplying arteries descended from the leptomeningeal surface without remarkable branchings straight to the subventricular zone where they branched and capillarized. Capillaries showed few H‐ and/or Y‐shaped anastomoses during their centrifugal course toward the leptomeningeal surface where they drained into cerebral venules and veins. Apart from the accessory olfactory bulb and the vestibule‐cochlear nucleus where capillaries were densely packed, capillaries formed a wide‐meshed 3D network throughout the brain parenchyma and thus contrasted to urodelian brains where hairpin‐shaped capillaries descend from the leptomeningeal vessels into varying depths of the brain parenchyma. In about two‐third of specimens, a closed arterial circle of Willis was found at the base of the brain. If this circle in Xenopus might serve the same two functions as in men is briefly discussed. Choroid plexuses of third and fourth ventricle were found to have a high venous, but a low arterial inflow via one small choroidal artery only. Findings are compared with previous studies on the vascularization of the anuran brain and discrepancies in respect to presence or absence of particular arteries and/or veins in Ranids, Bufonids, and Pipids studied so far are discussed with particular emphasis on the techniques used in the various studies published so far.     

Clotting Problems with the Teflon-Silastic Arteriovenous Shunt in Patients on Regular Haemodialysis

Episodes of clotting that occurred in 22 patients on regular haemodialysis were studied over a six-month period. The venous pressure during dialysis and the radiology of the Teflon-Silastic arteriovenous shunt were found to be satisfactory guides for the management of the shunt. The failure of the shunt during the early stage was mainly due to technical reasons. Histological study of the excised vessels in removed long-term shunts showed that these had failed because of rigidity and thickening of the vessel wall due to calcium and iron deposits or chronic inflammation, or both.     

Immunohistochemical evaluation of natural cases of encephalitic listeriosis in sheep

ABSTRACT

Listeriosis is an important public health problem in the world. It can cause abortion, encephalitis, septicemia, conjunctivitis and mastitis in ruminants. The development of central nervous system lesions is not fully understood in encephalitic listeriosis. We performed a retrospective analysis of 15 sheep with encephalitic listeriosis. Hyperemia and opacification of the meninges were common necropsy findings. Lesions generally were localized in the caudal part of the brain including the pons, medulla oblongata, thalamus and cerebellum. Microabscesses usually were found in the caudal brain and cerebellum, while perivascular infiltrates were found most often in other parts of the brain. Evidence of Listeria monocytogenes was detected immunohistochemically in the medulla oblongata, pons, thalamus and cerebellum. Prominent reactions for glial fibrillary acidic protein (GFAP), S100 protein, N-methyl-D-aspartate receptor-1 (NMDAR1) and inducible co-stimulatory protein (ICOS) were detected in the caudal brain, which indicates that these proteins may play roles in the pathogenesis of encephalitic listeriosis.