Storage Bodies Research Articles

IMAGING DIAGNOSIS-NEURONAL CEROID LIPOFUSCINOSIS WITH A CHRONIC SUBDURAL HEMATOMA

A subdural hematoma was found to accompany neuronal ceroid lipofuscinosis in an 11-month-old Dachshund. Results from clinical, magnetic resonance (MR) imaging, histopathologic, ultrastructural, and molecular assessments are described. The dog had a 3-month history of progressive neurologic signs. In MR images, there was severe asymmetric cerebral atrophy with a subdural hematoma. Histopathologically, there was autofluorescent, periodic acid-Schiff-positive lipopigment in neurons and transmission electron microscopy confirmed a typical curvilinear profile of the storage bodies. We hypothesize that rapid brain atrophy contributed to the subdural hematoma formation, a complication not described previously in dogs with neuronal lipofuscinosis.

Geometrical Aspects of the Incorporation of Free Space in Magnetomechanics at Finite Strains

AbstractThis paper addresses the modeling of finite magnetoelasticity with a particular focus on the incorporation of free space energy storage and magnetic body forces. First, a coordinate‐invariant formulation of an extended Neo‐Hookean model with magnetostrictive coupling is presented. Then, a finite element model for finite magnetomechanics based on a variational stationarity principle is formulated. The influence of magnetic body forces is studied in an exemplary boundary value problem. (© 2009 Wiley‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Development and maturation of invariant NKT cells in the presence of lysosomal engulfment

A defect in invariant NKT (iNKT) cell selection was hypothesized in lysosomal storage disorders (LSD). Accumulation of glycosphingolipids (GSL) in LSD could influence lipid loading and/or presentation causing entrapment of endogenous ligand(s) within storage bodies or competition of the selecting ligand(s) by stored lipids for CD1d binding. However, when we analyzed the iNKT cell compartment in newly tested LSD animal models that accumulate GSL, glycoaminoglycans or both, we observed a defective iNKT cell selection only in animals affected by multiple sulfatase deficiency, in which a generalized aberrant T-cell development, rather than a pure iNKT defect, was present. Mice with single lysosomal enzyme deficiencies had normal iNKT cell development. Thus, GSL/glycoaminoglycans storage and lysosomal engulfment are not sufficient for affecting iNKT cell development. Rather, lipid ligand(s) or storage compounds, which are affected in those LSD lacking mature iNKT cells, might indeed be relevant for iNKT cell selection.

An Investigation of Bacillus thuringiensis in Rectal-Collected Fecal Samples of Cows

In order to better understand the range and role of Bacillus thuringiensis (Bt) and its toxins in nature, we have undertaken a study of Bt taken directly from the rectum of 117 cows from 37 farms on the Caribbean island of Trinidad. Thirty-seven fecal samples (32%) were found to contain at least one Bt. Generally only one or two isolates with a particular crystal morphology were isolated from any one sample, however, a few samples contained more, up to 11 isolates, suggesting post-ingestion amplification. Bioassays using larvae of Musca domestica, Caenorhabditis elegans and Tetrahymena pyriformis showed no observable toxicity in gross bioassays. Very small dot-like parasporal bodies, not generally characteristic of Bt, were isolated from 44% of the samples, which in many instances appeared unstable and whose relation to Bt Cry protein-containing parasporal bodies is unknown. In conclusion, we find little evidence for a host adapted strain of Bt in the cows examined, nor toxicity to organisms that might logically be associated with either the cow or its feces. The presence of a large number of isolates containing small dot-like parasporal bodies, possibly either poly-beta-hydroxybutyrate storage bodies or Cry proteins, was unexpected and merits further investigation.

Effect Of Sports Beverage Temperature On Body Temperature And Cycling Performance In The Heat

Rising body temperature may influence the evolution of fatigue during exercise in the heat. Cooler beverage temperature may attenuate the rise in body temperature in the heat by generating a heat sink or via oral sensory mechanisms. PURPOSE: To determine the effect of a cold (4°C; C) or thermoneutral (37°C; N) sports beverage on evolution of body temperature and exercise performance in the heat and explore the influence of sensory factors to ingesting a cold stimulus (ice puree). METHODS: Seven non-acclimatised males (age 32.8±6.1y, mass 81.1±11.1kg) were recruited to complete C, N, and N plus ice (NI) trials in randomised order. Subjects cycled for 90 min at 65% VO2peak (SSE) followed by a 15 min time trial (TT) in 28°C, 70% relative humidity and ingested 3.5 ml.kg-1 of 7.4% carbohydrate-electrolyte drink every 10 mins including 30ml ice puree every 5 mins in NI during SSE. Rectal (Tr) and skin temperature (Tsk) were measured, and mean body temperature (), heat storage (HS), and total body heat content (TBHC) calculated from these. Total work (TW), distance (D), average and peak power output (AP, PP) were measured during TT. One and two factor repeated-measures ANOVAs were used to evaluate differences in performance and physiological variables across time and within trials respectively. Statistical significance was set at p<0.05. RESULTS: Differences in Tr, Tsk or between C, N, and NI were not significant across SSE however for C versus N, rise pre to post in Tsk (0.22±1.10°C v 1.14±0.90°C; p=0.02) and (1.2±0.3 v 1.6±0.3°C; p=0.03), and HS (45.1±8.8J. °C-1.kg-1.min-1 v 60.1±1.0J. °C-1.kg-1.min-1) were lower. There was a significant improvement in TW (263±60kJ v 252±63kJ; p<0.01), AP (296.0±66.3W v 287.0±71.3; p=0.03) and D (10.1±0.09km v 9.8±0.9km; p=0.01) over the TT on C compared to the N trials. Difference in TW and AP for C compared to the N trial was 4.9% and 3.9% respectively. Although most measures tended to improve with I, results were not significant when compared to N. CONCLUSIONS: Consumption of cold sports beverage during prolonged exercise in the heat improves body temperature measures and performance. Provision of ice did not reveal a sensory response, but requires further study as frequency of ingestion may have masked the sesnory effect by generating a minor heat sink. Supported by a NESC Discretionary Research Grant

Fixation and fate of C and N in the cyanobacterium Trichodesmium using nanometer-scale secondary ion mass spectrometry

The marine cyanobacterium Trichodesmium is ubiquitous in tropical and subtropical seas and is an important contributor to global N and C cycling. We sought to characterize metabolic uptake patterns in individual Trichodesmium IMS-101 cells by quantitatively imaging (13)C and (15)N uptake with high-resolution secondary ion mass spectrometry (NanoSIMS). Trichodesmium fix both CO(2) and N(2) concurrently during the day and are, thus, faced with a balancing act: the O(2) evolved during photosynthesis inhibits nitrogenase, the key enzyme in N(2) fixation. After performing correlated transmission electron microscopy (TEM) and NanoSIMS analysis on trichome thin-sections, we observed transient inclusion of (15)N and (13)C into discrete subcellular bodies identified as cyanophycin granules. We speculate that Trichodesmium uses these dynamic storage bodies to uncouple CO(2) and N(2) fixation from overall growth dynamics. We also directly quantified both CO(2) and N(2) fixation at the single cell level using NanoSIMS imaging of whole cells in multiple trichomes. Our results indicate maximal CO(2) fixation rates in the morning, compared with maximal N(2) fixation rates in the afternoon, bolstering the argument that segregation of CO(2) and N(2) fixation in Trichodesmium is regulated in part by temporal factors. Spatial separation of N(2) and CO(2) fixation may also have a role in metabolic segregation in Trichodesmium. Our approach in combining stable isotope labeling with NanoSIMS and TEM imaging can be extended to other physiologically relevant elements and processes in other important microbial systems.

A Variant Form of Mucolipidosis IV: Report on 4 Patients From the Indian Subcontinent

The clinical manifestations and histopathologic and neuroimaging findings in 4 Indian patients with a variant form of mucolipidosis IV are described. The presenting symptoms were psychomotor delay, spastic paraplegia, and mild mental retardation. One patient also had visual deterioration due to optic atrophy. None had corneal or retinal abnormalities. Magnetic resonance imaging in 3 patients showed a uniformly thin corpus callosum in all patients and white matter changes in 2 patients. Electron microscopic examination of skin biopsy specimens revealed storage bodies characteristic of mucolipidosis IV. These patients differ from previously described patients with this disorder in the absence of corneal abnormalities and in their presentation with spastic paraplegia during the second decade of life. Correct diagnosis is needed for genetic counseling, prognostication. and reduction of additional familial burden of this rare disease.

Alte und neue Eisenparameter – Bedeutung im Eisenstoffwechsel und in der Diagnostik

Iron is an element which is essential to life but also potentially toxic. Therefore, clever mechanisms exist in the human body for uptake, transport and storage of iron. Hepcidin, which seems to be the master protein for regulation of intestinal iron absorption, is known for a short time. The expression of hepcidin is not only influenced by iron levels but also by mediators of inflammation and growth factors of erythropoiesis. Hence hepcidin plays also a crucial role in the development of anemia of chronic disease and iron overload due to ineffective erythropoiesis. Serum ferritin is a reliable parameter to estimate the storage iron. It is an acute phase protein which is elevated during infections and inflammations, though. In these situations, measurement of soluble transferrin receptors is a useful tool to differentiate between iron deficiency and anemia of chronic disease. Newer parameters as erythrocyte zink protoporphyrin or percentage of hypochromic erythrocytes (%HYPO) are suited to detect a functional iron deficiency. Early diagnosis of iron overload is essential to prevent organ damage. Serum ferritin and transferrin are useful parameters to screen for iron overload. If no clear reason for a secondary iron overload can be found, the search for a hereditary haemochromatosis is recommended. Most of these hereditary haemochromatoses are a result of mutations in the HFE gene (homozygous state for Cys282Tyr or compound heterozygosity for Cys282Tyr/ His63Asp) which can be detected by PCR technique. Liver biopsy is still the gold standard for quantification of storage iron. However, a method of increasing importance for quantification of iron overload is magnetic resonance imaging with new approaches as for example T2*.

Protection of specific maternal messenger RNAs by the P body protein CGH-1 (Dhh1/RCK) during Caenorhabditis elegans oogenesis.

During oogenesis, numerous messenger RNAs (mRNAs) are maintained in a translationally silenced state. In eukaryotic cells, various translation inhibition and mRNA degradation mechanisms congregate in cytoplasmic processing bodies (P bodies). The P body protein Dhh1 inhibits translation and promotes decapping-mediated mRNA decay together with Pat1 in yeast, and has been implicated in mRNA storage in metazoan oocytes. Here, we have investigated in Caenorhabditis elegans whether Dhh1 and Pat1 generally function together, and how they influence mRNA sequestration during oogenesis. We show that in somatic tissues, the Dhh1 orthologue (CGH-1) forms Pat1 (patr-1)-dependent P bodies that are involved in mRNA decapping. In contrast, during oogenesis, CGH-1 forms patr-1–independent mRNA storage bodies. CGH-1 then associates with translational regulators and a specific set of maternal mRNAs, and prevents those mRNAs from being degraded. Our results identify somatic and germ cell CGH-1 functions that are distinguished by the involvement of PATR-1, and reveal that during oogenesis, numerous translationally regulated mRNAs are specifically protected by a CGH-1–dependent mechanism.

B-4 躯体蓄熱方式の蓄・放熱特性の改善 : その5:PCM混入ジェル状水蓄熱体の提案

B-4 躯体蓄熱方式の蓄・放熱特性の改善 : その5:PCM混入ジェル状水蓄熱体の提案

Retinal Pathology in a Canine Model of Late Infantile Neuronal Ceroid Lipofuscinosis

Late infantile neuronal ceroid lipofuscinosis (NCL) is an inherited disorder characterized by progressive vision loss. The disease results from mutations in the TPP1 (CLN2) gene. Studies were undertaken to characterize the effects of a TPP1 frameshift mutation on the retina in Dachshunds. A litter of four puppies consisting of one homozygous affected dog, two heterozygotes, and one homozygous normal dog were monitored for neurologic and retinal changes through 10 months of age. The affected and homozygous normal dogs, as well as one of the heterozygotes, were then euthanatized, and the retinas were examined morphologically. The affected dog exhibited normal visual behavior and retinal function at 3 months of age, but vision was clearly impaired by 7 months, with markedly reduced ERG b-wave amplitudes. Beyond 7 months of age, the affected dog was functionally blind, and pupillary light reflexes and ERG response amplitudes continued to decline through 10 months of age. Both rod and cone system functions were severely impaired. The retina exhibited accumulation of autofluorescent storage bodies with distinctive curvilinear contents. Substantial cell loss occurred in the inner nuclear layer, with a smaller reduction in photoreceptor cell density. The canine TPP1 mutation results in progressive vision loss and retinal degeneration similar to that which occurs in human late infantile NCL. With the canine model, the natural history of disease progression in the retina provides a better understanding of the pathologic course of the disease and provides objective markers that can be used to assess the efficacy of therapeutic interventions.

Mice deficient in Neu4 sialidase exhibit abnormal ganglioside catabolism and lysosomal storage

Mammalian sialidase Neu4, ubiquitously expressed in human tissues, is located in the lysosomal and mitochondrial lumen and has broad substrate specificity against sialylated glycoconjugates. To investigate whether Neu4 is involved in ganglioside catabolism, we transfected beta-hexosaminidase-deficient neuroglia cells from a Tay-Sachs patient with a Neu4-expressing plasmid and demonstrated the correction of storage due to the clearance of accumulated GM2 ganglioside. To further clarify the biological role of Neu4, we have generated a stable loss-of-function phenotype in cultured HeLa cells and in mice with targeted disruption of the Neu4 gene. The silenced HeLa cells showed reduced activity against gangliosides and had large heterogeneous lysosomes containing lamellar structures. Neu4(-/-) mice were viable, fertile and lacked gross morphological abnormalities, but showed a marked vacuolization and lysosomal storage in lung and spleen cells. Lysosomal storage bodies were also present in cultured macrophages preloaded with gangliosides. Thin-layer chromatography showed increased relative level of GD1a ganglioside and a markedly decreased level of GM1 ganglioside in brain of Neu4(-/-) mice suggesting that Neu4 may be important for desialylation of brain gangliosides and consistent with the in situ hybridization data. Increased levels of cholesterol, ceramide and polyunsaturated fatty acids were also detected in the lungs and spleen of Neu4(-/-) mice by high-resolution NMR spectroscopy. Together, our data suggest that Neu4 is a functional component of the ganglioside-metabolizing system, contributing to the postnatal development of the brain and other vital organs.

Accumulation of glial fibrillary acidic protein and histone H4 in brain storage bodies of Tibetan terriers with hereditary neuronal ceroid lipofuscinosis

The neuronal ceroid lipofuscinoses (NCLs) are inherited neurodegenerative diseases characterized by massive accumulation of autofluorescent storage bodies in neurons and other cells. A late-onset form of NCL occurs in Tibetan terrier dogs. Gel electrophoretic analyses of isolated storage body proteins from brains of affected dogs indicated that a protein of approximately 50 kDa was consistently prominent and a 16 kDa component was present in some brain storage body preparations. Mass spectral analysis identified the 50 kDa protein as glial fibrillary acidic protein (GFAP), isoform 2. GFAP identification was supported by immunoblot and immunohistochemical analyses. Histone H4 was the major protein in the 16 kDa component. Specific accumulation of GFAP and histone H4 in storage bodies has not been previously reported for any of the NCLs. Tibetan terrier NCL may be the canine correlate of one of the human adult-onset NCLs for which the genetic bases and storage body compositions have not yet been determined.

A new large animal model of CLN5 neuronal ceroid lipofuscinosis in Borderdale sheep is caused by a nucleotide substitution at a consensus splice site (c.571 + 1G >>> A) leading to excision of exon 3

Batten disease (neuronal ceroid lipofuscinoses, NCLs) are a group of inherited childhood diseases that result in severe brain atrophy, blindness and seizures, leading to premature death. To date, eight different genes have been identified, each associated with a different form. Linkage analysis indicated a CLN5 form in a colony of affected New Zealand Borderdale sheep. Sequencing studies established the disease-causing mutation to be a substitution at a consensus splice site (c.571 + 1G > A), leading to the excision of exon 3 and a truncated putative protein. A molecular diagnostic test has been developed based on the excision of exon 3. Sequence alignments support the gene product being a soluble lysosomal protein. Western blotting of isolated storage bodies indicates the specific storage of subunit c of mitochondrial ATP synthase. This flock is being expanded as a large animal model for mechanistic studies and trial therapies.

Phenotypic characterization of a mouse model of juvenile neuronal ceroid lipofuscinosis

Juvenile neuronal ceroid lipofuscinosis (JNCL) is an autosomal recessively inherited neurodegenerative disorder that results from mutations in the CLN3 gene. JNCL is characterized by accumulation of autofluorescent lysosomal storage bodies, vision loss, seizures, progressive cognitive and motor decline, and premature death. Studies were undertaken to characterize the neuronal ceroid lipofuscinosis phenotype in a Cln3 knockout mouse model. Progressive accumulation of autofluorescent storage material was observed in brain and retina of affected mice. The Cln3 −/− mice exhibited progressively impaired inner retinal function, altered pupillary light reflexes, losses of inner retinal neurons, and reduced brain mass. Behavioral changes included reduced spontaneous activity levels and impaired learning and memory. In addition, Cln3 −/− mice had significantly shortened life spans. These phenotypic features indicate that the mouse model will be useful for investigating the mechanisms underlying the disease pathology in JNCL and provide quantitative markers of disease pathology that can be used for evaluating the efficacies of therapeutic interventions.

Immunological Consequences of Apoptotic Cell Phagocytosis

Cells undergo apoptosis in development, tissue homeostasis, and disease and are subsequently cleared by professional and nonprofessional phagocytes. There is now overwhelming evidence that phagocyte function is profoundly altered following apoptotic cell uptake, with consequences for the ensuing innate and adaptive immune response. Pathogens and tumors exploit the changes in macrophage function following apoptotic cell uptake. Here, we will outline the consequences of apoptotic cell phagocytosis and illustrate how apoptotic cells could be used to manipulate the immune response for therapeutic gain.

Metabolomic investigation of CLN6 neuronal ceroid lipofuscinosis in affected South Hampshire sheep

The neuronal ceroid lipofuscinoses (NCLs; Batten disease) are a group of fatal inherited neurodegenerative diseases in humans and animals distinguished by a common clinical pathology, characteristic storage body accumulation in cells, and gross brain atrophy. An (1)H NMR spectroscopy- and GC-MS-based metabolomic investigation of changes in the cerebellum, frontal and occipital lobes, and cerebrospinal fluid (CSF) of CLN6 NCL affected South Hampshire sheep charted changes from the preclinical state to advanced disease. Glutamine and succinate concentrations increased in all brain regions in affected sheep relative to controls, whereas concentrations of aspartate, acetate, glutamate, N-acetyl aspartate (NAA), and gamma-aminobutyric acid (GABA) decreased. Changes in the concentrations of inositols, NAA, and GABA were consistent with glial cell activation and neurodegeneration beginning in the frontal and occipital lobes, in agreement with previous histopathological data. Further metabolic deficits were defined in all regions at earlier time points, including the cerebellum, where very little neurological degeneration has been reported. Biochemical abnormalities in the CSF of affected sheep at 18-31 months include relative increases in lactate, acetate, tyrosine, and creatine/creatinine concentrations and decreases in myo- and scyllo-inositol and citrate concentrations. The changes detected in the CSF and brain tissue mirrored those previously apparent in NCL mouse models, suggesting that they are common to all NCLs. However, the changes in glutamate and glutamine concentrations in CSF occurred after clinical disease, indicating that any changes in glutamate/glutamine cycling occur as a consequence of the primary deficits associated with the NCLs.

A cryptic intracellular green alga in Ginkgo biloba: ribosomal DNA markers reveal worldwide distribution

Intracellular symbioses involving eukaryotic microalgae and a variety of heterotrophic protists and invertebrates are widespread, but are unknown in higher plants. Recently, we reported the isolation and molecular identification of a Coccomyxa-like green alga from in vitro cell cultures of Ginkgo biloba L. This alga resides intracellularly in an immature "precursor" form with a nonfunctional chloroplast, implying that algal photosynthetic activity has no role in this endosymbiosis. In necrotizing Ginkgo cells, precursors evolved into mature algae, proliferated, and were liberated into the culture medium after host cell bursting. In the present paper we demonstrate by molecular methods a worldwide distribution of the alga in planta. Endosymbiont-specific sequences of ribosomal DNA could be traced in Ginkgo tissues of each specimen examined from different geographic locations in Europe, North America, and Asia. The Ginkgo/Coccomyca association represents a new kind of intracellular, vertically inherited symbiosis. Storage bodies, probably of lipid nature, present in the cytoplasm of each partner suggest a possible involvement of the endosymbiont in metabolic pathways of its host.

Distinct roles of free leptin, bound leptin and soluble leptin receptor during the metabolic‐inflammatory response in patients with liver cirrhosis

Alteration of the leptin system appears to play a role in the inflammatory-metabolic response in catabolic diseases such as chronic liver diseases. To investigate the association between leptin components, inflammatory markers and hepatic energy and substrate metabolism. We investigated in vivo hepatic substrate and leptin metabolism in 40 patients employing a combination of arterial and hepatic vein catheterization techniques and hepatic blood flow measurements. In addition to metabolic, inflammatory and neuroendocrine parameters, circulating levels of free leptin, bound leptin and soluble leptin receptor were determined. Compared with controls, bound leptin and soluble leptin receptor levels were significantly elevated in cirrhosis, while free leptin did not increase. In cirrhosis bound leptin was correlated with soluble leptin receptor (r = 0.70, P < 0.001). Free leptin was positively correlated with metabolic parameters such as energy storage (body fat mass; r = 0.36, P < 0.05), insulin and insulin resistance (r = 0.48; r = 0.46, P < 0.01) as well as with hepatic glucose and energy release (r = 0.35 and r = 0.40, P < 0.05). In contrast, bound leptin and soluble leptin receptor were linked to proinflammatory cytokines and sympathetic activity (r = 0.61 and r = 0.56, P < 0.01). The components of the leptin system (free leptin, bound leptin and soluble leptin receptor) have distinct roles in metabolic and inflammatory processes in patients with liver cirrhosis. The better understanding of this metabolic and inflammatory tissue-repair response may lead to innovative new therapeutic strategies in liver disease as well as in various other catabolic diseases.

Pathogenic mechanisms in lysosomal disease: a reappraisal of the role of the lysosome

The view that lysosomes simply represent end organelles in the serial degradation of polymeric molecules derived from the cell surface and its interior has led to major misconceptions about the nature of lysosomal storage diseases and the pathogenic cascades that characterize them. Accordingly, lysosomal storage bodies are often considered 'inert', inducing cell dysfunction and death primarily through mechanical overcrowding of normal organelles or by other non-specific means leading to generalized cytotoxicity. However, modern studies of lysosomes and their component proteins provide evidence to support a far greater role for these organelles in cell metabolism. In intimate association with endosomal, autophagosomal and related vesicular systems, the greater lysosomal system can be conceptualized as a vital recycling centre that serves as a central metabolic coordinator, influencing literally every aspect of the cell, from signal transduction to regulation of gene expression. This broader view of the role of lysosomes in cells not only provides insight into how single gene defects impacting on lysosomal function can result in the plethora of complex cellular transformations characteristic of these diseases, but also suggests new and innovative therapies that may hold considerable promise for ameliorating disease progression.