Ground-glass hepatocellular inclusions are associated with polypharmacy.

Ground-glass (GG) hepatocytes are classically associated with chronic hepatitis B (HBV) infection, storage disorders, or cyanamide therapy. In a subset of cases, an exact etiology cannot be identified. In this study, we sought to characterize the clinical, histological, and ultrastructural findings associated with HBV-negative GG hepatocytes. Our institutional laboratory information system was searched from 2000 to 2019 for all cases of ground-glass hepatocytes. Ten liver biopsies with GG hepatocellular inclusions and negative HBV serology, no known history of storage disorders, or cyanamide therapy were reviewed. Half of the patients had history of organ transplantation and/or malignancy. These patients took on average 8.1 medications (range: 3-14) with the most common medications being immunosuppressive and health supplements. Histologically, GG hepatocytes show either peri-portal or centrizonal distribution. The inclusions are PAS-positive and diastase sensitive. Electron microscopy showed intracytoplasmic granular inclusions with low electron density, consistent with unstructured glycogen. In summary, GG hepatocytes are a rare finding in liver biopsies, but are more common in patients with hepatitis B. They can also be seen in HBV-negative patients who have polypharmacy. In these cases, they are the result of unstructured glycogen accumulation putatively due to altered cell metabolism.

Arginine vasopressin neuronal loss results from autophagy-associated cell death in a mouse model for familial neurohypophysial diabetes insipidus.

Familial neurohypophysial diabetes insipidus (FNDI) characterized by progressive polyuria is mostly caused by mutations in the gene encoding neurophysin II (NPII), which is the carrier protein of the antidiuretic hormone, arginine vasopressin (AVP). Although accumulation of mutant NPII in the endoplasmic reticulum (ER) could be toxic for AVP neurons, the precise mechanisms of cell death of AVP neurons, reported in autopsy studies, remain unclear. Here, we subjected FNDI model mice to intermittent water deprivation (WD) in order to promote the phenotypes. Electron microscopic analyses demonstrated that, while aggregates are confined to a certain compartment of the ER in the AVP neurons of FNDI mice with water access ad libitum, they were scattered throughout the dilated ER lumen in the FNDI mice subjected to WD for 4 weeks. It is also demonstrated that phagophores, the autophagosome precursors, emerged in the vicinity of aggregates and engulfed the ER containing scattered aggregates. Immunohistochemical analyses revealed that expression of p62, an adapter protein between ubiquitin and autophagosome, was elicited on autophagosomal membranes in the AVP neurons, suggesting selective autophagy induction at this time point. Treatment of hypothalamic explants of green fluorescent protein (GFP)-microtubule-associated protein 1 light chain 3 (LC3) transgenic mice with an ER stressor thapsigargin increased the number of GFP-LC3 puncta, suggesting that ER stress could induce autophagosome formation in the hypothalamus of wild-type mice as well. The cytoplasm of AVP neurons in FNDI mice was occupied with vacuoles in the mice subjected to WD for 12 weeks, when 30-40% of AVP neurons are lost. Our data thus demonstrated that autophagy was induced in the AVP neurons subjected to ER stress in FNDI mice. Although autophagy should primarily be protective for neurons, it is suggested that the organelles including ER were lost over time through autophagy, leading to autophagy-associated cell death of AVP neurons.

Effect of Yisui Shengxue Granule () on the oxidative damage of erythrocytes from patients with hemoglobin H disease.

OBJECTIVE: To investigate the effect of Yisui Shengxue Granule (, YSSXG), a complex Chinese medicine, on the oxidative damage of erythrocytes from patients with hemoglobin H (HbH) disease. METHODS: Twenty-two patients with HbH disease and 22 healthy volunteers were observed. YSSXG was given to patients with HbH disease for 3 months. Before and after the 3-month treatment, blood parameters [hemoglobin (Hb), red blood cells (RBCs), and reticulocyte percent (Ret)] were examined; inclusion bodies in erythrocytes were observed by transmission electron microscopy (TEM); activities of antioxidant defense enzymes [superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (Cat)] and erythrocyte membrane malondialdehyde (MDA) concentrations were determined. RESULTS: In patients with HbH disease, measured values of RBC and Hb obtained from the first to the third months after treatment with YSSXG were significantly higher than before treatment (P<0.01). Measured values of Ret from the second to the third months after treatment were significantly lower than before treatment (P<0.05 and P<0.01, respectively). Prior to treatment with YSSXG, TEM images of RBCs showed the presence of numerous inclusion bodies. After treatment with YSSXG, the amount and volume of inclusion bodies decreased. Treatment with YSSXG also led to a significant increase in SOD activity (P<0.01), a decrease in Cat activity (P<0.01), and no significant differences in GSHPx activity (P>0.05) or MDA concentration (P>0.05). However, compared with the healthy counterparts, SOD, GSH-Px, and Cat activities presented at high levels (P<0.01) both before and after treatment. CONCLUSIONS: YSSXG could improve the degree of hemolysis and anemia in patients with HbH disease. The mechanism may be related to its antioxidative effects, which could elevate the activity of total SOD in erythrocytes and efficiently inhibit the oxidative precipitation of ß-globin chains.

Oil body biogenesis during Brassica napus embryogenesis.

Although the oil body is known to be an important membrane enclosed compartment for oil storage in seeds, we have little understanding about its biogenesis during embryogenesis. In the present study we investigated the oil body emergence and variations in Brassica napus cv. Topas. The results demonstrate that the oil bodies could be detected already at the heart stage, at the same time as the embryos began to turn green, and the starch grains accumulated in the chloroplast stroma. In comparison, we have studied the development of oil bodies between Arabidopsis thaliana wild type (Col) and the low-seed-oil mutant wrinkled1-3. We observed that the oil body development in the embryos of Col is similar to that of B. napus cv. Topas, and that the size of the oil bodies was obviously smaller in the embryos of wrinkled1-3. Our results suggest that the oil body biogenesis might be coupled with the embryo chloroplast.

Substrate reduction therapy in juvenile GM2 gangliosidosis.

Substrate reduction therapy (SRT) is considered to be a potential therapeutic option for juvenile GM2 gangliosidosis (jGM2g). We evaluated the efficacy of SRT in jGM2g, assessing neurological, neuropsychological and brain magnetic resonance imaging (MRI) outcomes over a 24-month period of treatment. In an open-label and single-center study, five jGM2g patients (mean age 14.6+/-4.5 years) received oral miglustat at doses of 100-200mg t.i.d. adjusted to body surface area. Patients underwent general and neurological examinations, neuropsychological, electrophysiological, and brain MRI studies. All patients showed neurological deterioration over the period of the study, with particularly notable worsening of gait, speech and coordination. One patient experienced acute psychosis, and another showed worsening of pre-existing epilepsy. Some neuropsychological tests showed no evidence of deterioration in the three patients with high enough cognitive functioning for reliable assessment. Profound cognitive impairment in two children precluded neuropsychological evaluation. In four patients, evaluation of brain MRI showed no changes in white matter signal abnormalities and cerebellar atrophy noted at baseline, while one patient showed progression of cerebellar and supratentorial brain atrophy. Transmission electron microscopy analysis of peripheral mononuclear cells showed reduction of intracytoplasmatic inclusions with treatment. SRT with miglustat of patients with jGM2g failed to ameliorate progressive neurological deterioration, but apparently no worsening of some areas of cognitive function tested and brain MRI lesions was noted over 24 months of treatment. The results must be interpreted with care owing to the small sample of patients and the lack of a control-arm.

Long-term effects of natural amino acids on infection with Chlamydia trachomatis.

Supplementation of culture media with leucine, isoleucine, methionine, or phenylalanine was previously found to inhibit Chlamydia trachomatis growth in HEp-2 cells. Here, we investigated the long-term effects of these additives on C. trachomatis infection in the same cell model. Amino acid addition 30h post-infection (pi) effectively suppressed the generation of infectious progeny monitored for 10 days pi. With the exception of phenylalanine, amino acid treatment beginning at 2h pi for up to 15 days led to a complete lack of infectious progeny. Phenylalanine treatment resulted in residual minimal infectivity. In extended supplementation experiments, very small aberrant chlamydial inclusions formed, whose numbers decreased considerably over time, and the production of infectious chlamydiae could not be rescued even upon amino acid withdrawal. Interestingly, a state of chlamydial persistence was induced under these conditions, as 16S rRNA transcripts were detected throughout treatment. However, expression of several key chlamydial genes including omp1, groEL, omcB, and those functioning for chlamydial DNA replication and cytokinesis was generally very low or even undetected, particularly in monolayers treated with Leu, Ile, or Met. These data revealed a capacity of certain amino acids to eliminate infectious chlamydial progeny. Additionally, supplementation of certain amino acids resulted in the formation of a small persistent population. Extrapolating from these findings may help formulate an anti-chlamydial treatment based on nutritional elements.

Sorption of hydrophobic organic compounds (HOC) in rapeseed oil bodies.

Oil-bodies are minute plant organelles (0.5-2.0microm diameter) consisting of an oil core surrounded by a phospholipid monolayer/proteinaceous membrane. Oil-bodies have been isolated from rapeseed seeds and demonstrated to constitute a novel type of micro-capsule suitable for the extraction of hydrophobic organic compounds from aqueous environments. Three hydrophobic pesticides: atrazine (2-chlor-4-ethyl-amino-6-isopropylamino-1,3,5-triazine), carbaryl (1-naphthyl methylcarbamate) and parathion (O,O-diethyl O-(4-nitrophenyl) phosphorothioate), as well as naphthalene and 2-phenylethanol were successfully extracted from aqueous solutions, with absorption in the inner oily core of OB as sorption mechanism. The OB membrane does not represent a barrier for the mass transfer of the compound towards the inner oily core of OB. Moreover, due to very high surface area to volume ratio, oil-bodies exhibit very good mass transfer properties compared with larger synthetic microcapsules or two-phase liquid-liquid extraction (LLE) techniques, which diminishes the need for strong agitation and avoids the formation of difficult to separate stable emulsions.

Suppression of soybean oleosin produces micro-oil bodies that aggregate into oil body/ER complexes.

Using RNAi, the seed oil body protein 24-kDa oleosin has been suppressed in transgenic soybeans. The endoplasmic reticulum (ER) forms micro-oil bodies about 50 nm in diameter that coalesce with adjacent oil bodies forming a hierarchy of oil body sizes. The oil bodies in the oleosin knockdown form large oil body-ER complexes with the interior dominated by micro-oil bodies and intermediate-sized oil bodies, while the peripheral areas of the complex are dominated by large oil bodies. The complex merges to form giant oil bodies with onset of seed dormancy that disrupts cell structure. The transcriptome of the oleosin knockdown shows few changes compared to wild-type. Proteomic analysis of the isolated oil bodies of the 24-kDa oleosin knockdown shows the absence of the 24-kDa oleosin and the presence of abundant caleosin and lipoxygenase. The formation of the micro-oil bodies in the oleosin knockdown is interpreted to indicate a function of the oleosin as a surfactant.

Bundles of hexagonally arranged tubules in timothy grass pollen: detection of a novel pollen component using anhydrous fixation and image analysis techniques in transmission electron microscopy.

Pollen from timothy grass (Phleum pratense L.) was subjected to various aqueous and non-aqueous fixation and preparation protocols for transmission electron microscopy. Only in the cytoplasm of anhydrously prepared pollen grains were conspicuous inclusions observed that range in size from less than 1 mum up to 8 or 10 mum. These bodies have so far not been described in the literature. Higher magnifications show that these inclusions consist of bundles of hexagonally arranged small tubules. In order to obtain details of the ultrastructure of this novel pollen component, TEM micrographs of ultrathin sections of hexagonally arranged tubules were analyzed using Fourier transform techniques of image analysis. It was found that the tubules form groups with quasi-periodic hexagonal arrangement, with an average centre-to-centre spacing between the neighbouring tubules of approximately 42 nm. Individual tubules are formed by 12 or 13 particles. The outer diameter of the tubules ranges between 22 and 24 nm. From our experiments, we conclude that the quasi-periodic hexagonally arranged tubules forming conspicuous cytoplasmic inclusions in dry timothy grass pollen grains are structurally similar to microtubules.

Lafora-like ground-glass inclusions in hepatocytes of pediatric patients: a report of two cases.

We report 2 cases of ground-glass hepatocyte inclusions occurring in pediatric patients. Case 1 had alpha-thalassaemia major and was receiving iron chelation therapy, whereas case 2 had trisomy 21 with a history of bone marrow transplantation for acute myeloid leukemia. The liver sections in both cases showed eosinophilic, periodic acid-Schiff diastase-positive intracytoplasmic inclusions that were negative for hepatitis B surface antigen. Immunohistochemically the inclusions showed positive staining with KM279, a monoclonal antibody against polyglucosan derived from Lafora inclusions. On electron microscopy, in case 1, intracytoplasmic inclusions were composed of degenerate organelles, glycogen, and irregular fibrillar structures; in case 2, they were composed of vesicular structures containing granular material. Ultrastructural changes in both cases differed from classical Lafora inclusions and ruled out hepatitis B surface antigen, glycogenosis type IV, and fibrinogen storage disease. Genetic analysis of the Lafora's disease genes performed in case 2 revealed no mutations. The development of hepatocyte cytoplasmic inclusions in both our cases could be related to medication effects, because similar inclusions were reported in patients using cyanamide. Drug-induced inclusions, mimicking Lafora's disease, should be included in the differential diagnosis of hepatocyte ground-glass inclusions.

Cell-based fluorescence assay for evaluation of new-drugs potential for phospholipidosis in an early stage of drug development.

To evaluate new-drugs potential for phospholipidosis (PL), we developed a cell-based fluorescence assay using a fluorescent-labeled phospholipid analogue (NBD-PE). CHL/IU cells derived from newborn hamster lung were exposed to positive reference compounds (amiodarone, imipramine, chloroquine, propranolol, chlorpromazine and amantadine) in the presence of NBD-PE, and the level of PL, as indicated by accumulation of fluorescent inclusions in the cytoplasm, was evaluated using fluorescence microscopy and fluorometry. All positive reference compounds induced accumulation of fluorescent inclusions in a concentration-dependent manner with an increase in fluorescence intensity. Fluorescence microscopically, the positive dose of test compound was determined as the concentration with a grade equivalent to or above that of 3.13 microM of amiodarone. Based on this criterion, 8 of 20 test compounds including PL-positive or -negative compounds were judged positive that were concurrent with the pathological results from rat toxicity studies. Furthermore, a positive criterion for fluorometry was decided as equivalent to or above 25% of maximum intensity induced by 1.56-25.0 microM amiodarone. In comparison of fluorometry methods with fluorescence microscopy method, 19 of 20 compounds were judged same. From these findings, we concluded that the assay developed in this study is a rapid and reliable method to predict new-drugs potential for PL at an early stage of drug development.

Intracellular inclusions of uncultured magnetotactic bacteria.

Magnetotactic bacteria produce magnetic crystals in organelles called magnetosomes. The bacterial cells may also have phosphorus-containing granules, sulfur globules, or polyhydroxyalkanoate inclusions. In the present study, the ultrastructure and elemental composition of intracellular inclusions from uncultured magnetotactic bacteria collected in a marine environment are described. Magnetosomes contained mainly defect-free, single magnetite crystals with prismatic morphologies. Two types of phosphorus-containing granules were found in magnetotactic cocci. The most common consisted of phosphorus-rich granules containing P, O, and Mg; and sometimes also C, Na, Al, K, Ca, Mn, Fe, Zn, and small amounts of S and Cl were also found. In phosphorus-sulfur-iron granules, P, O, S, Na, Mg, Ca, Fe, and frequently Cl, K, and Zn, were detected. Most cells had two phosphorus-rich granules, which were very similar in elemental composition. In rod-shaped bacteria, these granules were positioned at a specific location in the cell, suggesting a high level of intracellular organization. Polyhydroxyalkanoate granules and sulfur globules were less commonly seen in the cells and had no fixed number or specific location. The presence and composition of these intracellular structures provide clues regarding the physiology of the bacteria that harbor them and the characteristics of the microenvironments where they thrive.

Acute pulmonary inflammation induced by lung overloading with selenium particles: leukocyte response and in situ detection of selenium at high resolution.

The kinetics of the acute inflammatory response of the lung was triggered in CD-1 mice by a single intratracheal instillation of a large amount of Se (10 mg); it was studied by quantitative cytology of bronchoalveolar lavage samples, light microscopy, and scanning electron microscopy coupled with x-ray elemental microanalysis. Bronchoalveolar lavage leukocytes were mostly neutrophils and increased from 12 to 24 h of Se treatment and decreased at 72 h. Only less than half of the granulocytes showed ingested Se particles; in contrast, virtually all BAL macrophages contained Se particles. Scanning electron microscopy coupled with X-ray elemental microanalysis revealed that the intracellular Se particles were heterogeneous in size (diameters from 0.4 and up to 14 microm), and that Se inclusions were sometimes accumulated at a pole of the cell. At 72 h after instillation of the particles, Se-loaded alveolar macrophages were migrated in the interstitial space of the alveoli. Se-positive regions had a focal distribution in the lung; accumulation of inflammatory cells erased the alveolar architecture of these areas of the deep lung. Our data indicates that Se overloading of the lung results in: (1) an acute inflammatory response that is dominated by neutrophils; (2) early removal of Se done mostly by alveolar macrophages, and (3) formation of focal areas of invasion of the lung parenchyma by inflammatory infiltrates.

Size and stability of reconstituted sesame oil bodies.

Oil bodies of sesame seeds comprise a triacylglycerol matrix, which is surrounded by a monolayer of phospholipids embedded with unique proteins, mainly structural proteins termed oleosins. Artificial oil bodies were successfully reconstituted with various compositions of triacylglycerols, phospholipids, and oil-body proteins. The sizes of reconstituted oil bodies displayed a normal distribution with an average size proportional to the ratio of triacylglycerols to oil-body proteins. Both thermostability and structural stability of reconstituted oil bodies decreased as their sizes increased, and vice versa. Proteinase K digestion indicated that oleosins anchored both native and reconstituted oil bodies via their central hydrophobic domains. The stability of reconstituted oil bodies, as well as the purified ones from sesame seeds, could be substantially enhanced after their surface proteins were cross-linked by glutaraldehyde or genipin.

Correlation between in vivo and in vitro pulmonary responses to jet propulsion fuel-8 using precision-cut lung slices and a dynamic organ culture system.

In tissue slice models, interactions between the heterogeneous cell types comprising the lung parenchyma are maintained thus providing a controlled system for the study of pulmonary toxicology in vitro. However, validation of the model in vitro system must be affirmed. Previous reports, in in vivo systems, have demonstrated that Clara cells and alveolar type II cells are the targets following inhalation of JP-8 jet fuel. We have utilized the lung slice model to determine if cellular targets are similar following in vitro exposure to JP-8. Agar-filled adult rat lung explants were cored and precision cut, using the Brende/Vitron tissue slicer. Slices were cultured on titanium screens located as half-cylinders in cylindrical Teflon cradles that were loaded into standard scintillation vials and incubated at 37 degrees C. Slices were exposed to JP-8 jet fuel (0.5 mg/ml, 1.0 mg/ml, and 1.5 mg/ml in medium) for up to 24 hours. We determined ATP content using a luciferin-luciferase bioluminescent assay. No significant difference was found between the JP-8 jet fuel doses or time points, when compared to controls. Results were correlated with structural alterations following aerosol inhalation of JP-8. As a general observation, ultrastructural evaluation of alveolar type cells revealed an apparent increase in the number and size of surfactant secreting lamellar bodies that was JP-8 jet fuel-dose dependent. These results are similar to those observed following aerosol inhalation exposure. Thus, the lung tissue slice model appears to mimic in vivo effects of JP-8 and therefore is a useful model system for studying the mechanisms of lunginjury following JP-8 exposure.

Coffee ringspot virus vectored by Brevipalpus phoenicis (Acari: Tenuipalpidae) in coffee.

Coffee ringspot is characterized by conspicuous ringspot symptoms on leaves, berries, and less frequently on twigs. It is caused by coffee ringspot virus (CoRSV), a short, bacilliform virus (40 nm x 100-110 nm). The virus is not seed borne and is transmitted by Brevipalpus phoenicis (Geijskes). Transovarial transmission within the mite does not occur. CoRSV has been mechanically transmitted to Chenopodium amaranticolor Coste and Reynaud, C. quinoa Wildenow, Beta vulgaris L., and Alternanthera tenella Colla resulting in local lesions. Systemic infection within both C. amaranticolor and C. quinoa occurs. Virions are found in the nucleus or cytoplasm of infected cells, commonly associated with membranes. Occasionally, membrane bounded particles are found within the cisternae of the endoplasmic reticulum. A characteristic electron lucent, nuclear inclusion is commonly found in many infected cells. These cytopathic effects place CoRSV among the nuclear type of Brevipalpus-borne viruses. The disease has been reported in several Brazilian states (São Paulo, Paraná, Minas Gerais, and Federal District) and recently found in Costa Rica. A similar disease is known in the Philippines, but no information exists about its relationship to CoRSV. Coffee ringspot had no economical significance until recently when a large scale infection was reported in Minas Gerais that resulted in yield loss.

Intracellular lipophilic inclusions of mycobacteria in vitro and in sputum.

Although most mycobacterial lipids are thought to be associated with the cell envelope, the authors previously observed substantial deposits of intracellular lipophilic material. A Nile-red-based cytological assay was used to determine factors which affect the presence and natural history of intracellular lipophilic inclusions (ILIs) in Mycobacterium smegmatis. Development of ILIs was associated with stationary-phase cultures in broth and with aged (6 days) colonies on agar. Using variants of Youmans' defined medium, the frequency and size of ILIs was observed to be minimal in carbon-poor medium. ILIs were observed to form within 15 min after provision of fatty acids to the medium and after a period of several days in nitrogen-poor medium. Analysis of the non-polar lipid extracts of ILI-rich and -poor preparations indicated that the triacylglycerols (TAGs) were a major component of the inclusions. The acyl substituents of the TAGs varied according to whether they were formed in Middlebrook 7H9 broth, in low-nitrogen Youmans' broth or rapidly after oleic acid supplementation of Youmans' broth. These studies support a storage function for TAGs in mycobacteria in addition to their previously suggested occurrence as components of the cell envelope. To assess a possible role for ILIs in Mycobacterium tuberculosis, a combined acid-fast (Auramine)/Nile red assay was applied to heavily positive sputum samples from patients with tuberculosis. Strong intracellular Nile red signals were obtained from acid-fast cells, indicating that ILI occur in M. tuberculosis in vivo. This may reflect a distinct physiological state of these cells, which it has not been possible to reproduce in vitro. These findings indicate that the uptake of long-chain fatty acids and TAG biosynthetic and degradative pathways are important aspects of mycobacterial lipid metabolism, meriting further investigation.

Fatal herpes simplex infection in a pygmy African hedgehog (Atelerix albiventris).

An adult pygmy African hedgehog developed acute posterior paresis attributed to a prolapsed intervertebral disc diagnosed by C-T scan. Corticosteroid therapy resulted in prompt resolution of the ataxia, but 2 weeks later the animal became anorexic and died. Macroscopically, the liver was stippled with punctate off-white foci which were confirmed microscopically to be foci of necrosis. Numerous hepatocytes contained intranuclear inclusions and syncytial cell formation was also present. A herpes virus was isolated and identified by fluorescent antibody and polymerase chain reaction studies as herpesvirus simplex type 1. To our knowledge, this is the first report of herpes infection in the African hedgehog and the first time herpes simplex has been identified as a cause of disease in insectivores.

Ultrastructure and composition of asteroid bodies.

PURPOSE: Asteroid hyalosis is a disease of the vitreous, characterized by brilliant reflecting particles, termed asteroid bodies, which are surrounded by a tightly adhering network of fibrils. The composition and mode of formation of asteroid bodies is not yet understood in detail. The purpose of this study was to investigate the ultrastructure of asteroid bodies and to identify the intrinsic inorganic and organic components that contribute to the nature and development of asteroid bodies. METHODS: Electron energy loss spectroscopy and energy-filtered transmission electron microscopy were used for the elemental analysis of asteroid bodies. The ultrastructural localization of glycosaminoglycans was investigated, using lectin and antibody conjugates in conjunction with transmission electron microscopy and epifluorescence microscopy. Anionic sites of glycosaminoglycans were detected with 15 nm cationic colloidal gold at low pH, applied as a postembedding technique. Ultrastructural details of asteroid bodies were documented using fast Fourier transform analysis of zero-loss filtered images. RESULTS: Element mapping of asteroid bodies by electron spectroscopic imaging revealed a homogeneous distribution of calcium, phosphorus, and oxygen. The electron energy loss spectra of these elements showed details similar to those found for hydroxyapatite. Additionally, high contrast and sensitivity against a calcium-specific chelator highlighted the crystalline, apatite-like nature of asteroid bodies. Immunofluorescence microscopy revealed the presence of chondroitin-6-sulfate at the periphery of asteroid bodies, which is in agreement with the ultrastructural colocalization of anionic sites. Fast Fourier transform analysis revealed that each 7-nm periodicity of asteroid lamellar stacks is divided by a fine, parallel-oriented line, separating each 7-nm layer into two halves of 3.5-nm thickness. Carbohydrates specific for hyaluronic acid were observed by lectin-gold labeling to be part of the inner matrix of asteroid bodies. CONCLUSIONS: The results of this study demonstrate the structural and elemental similarity of asteroid bodies with hydroxyapatite. Proteoglycans and their glycosaminoglycan side chains are implicated in playing a role in regulating the biomineralization process.

Ultrastructure of two oil-degrading bacteria isolated from the tropical soil environment.

Two oil-degrading bacteria identified as Pseudomonas aeruginosa and Micrococcus luteus were isolated from crude-oil-polluted soils in Nigeria. The organisms were grown on n-hexadecane and sodium succinate and then examined for the presence of hydrocarbon inclusions. Inclusion bodies were found in n-hexadecane-grown cells and were absent in succinate-grown cells. Formation of hydrocarbon inclusion bodies appears to be a general phenomenon among hydrocarbon utilizers.