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Adenylate compartmentation and storage in coelomic cells of the polychaete Nereis virens

The concentrations of adenine nucleotides (AMP, ADP, ATP) were determined in coelomic cells (eleocytes) of the polychaete Nereis virens. In cells of immature and male animals, total ADP and AMP contents (each 10&shy;15 &micro;mol ml-1 packed cell volume) greatly exceeded that of ATP (0.8 &micro;mol ml-1 packed cell volume). 31P-nuclear magnetic resonance (NMR) studies of living eleocytes showed that the high concentrations of both AMP and ADP are free in solution. Comparisons of in vivo NMR spectra with those obtained from metabolite extracts of eleocytes suggest that the adenylate pools are compartmentalized, with a large pool being in an environment with a pH<6.0 and a small pool being in a domain where pH>6.7. This indicates that eleocytes are capable of storing high concentrations of ADP and AMP without inhibiting energy metabolism by sequestering these compounds into an acidic compartment. The large acidic vacuole characteristic of eleocytes may function as this compartment.

Journal Article↗

Direct selection of trans-acting ligase ribozymes by in vitro compartmentalization.

We have used a compartmentalized in vitro selection method to directly select for ligase ribozymes that are capable of acting on and turning over separable oligonucleotide substrates. Starting from a degenerate pool, we selected a trans-acting variant of the Bartel class I ligase which statistically may have been the only active variant in the starting pool. The isolation of this sequence from the population suggests that this selection method is extremely robust at selecting optimal ribozymes and should, therefore, prove useful for the selection and optimization of other trans-acting nucleic acid catalysts capable of multiple turnover catalysis.

Base Pairing↗

Intracellular compartmentation of metals in aquatic organisms: roles in mechanisms of cell injury.

The intracellular compartmentation of essential and toxic metals is of intense scientific interest because of its potential for adding to our understanding of both normal homeostatic mechanisms for metals and of the mechanisms which underlie metal-induced cell injury. High-affinity metal-binding proteins, lysosomes, and precipitates such as inclusion bodies or concretions, play major roles in the regulation of divalent-metal cation bioavailability. The contribution of a given compartment toward metal homeostasis is dependent upon the level exposure, cell type, organ, species, and life cycle of the organism. Toxic metals may move between these compartments, but the rates and determinants of such exchanges have not been characterized. Available data clearly indicate that sequestration of toxic metals in these specialized compartments can produce profound disturbances in the subcellular handling of essential metals. Further studies of the mechanisms by which metals partition and/or transfer among these compartments are essential to understand and predict toxicity of this important class of toxic agents.

Animals↗

Compartmental divisions of the hand revisited. Rethinking the validity of cadaver infusion experiments.

The results of a cadaver dye-infusion experiment suggested that the hand has ten muscle compartments and that the volar interossei occupy a separate anatomical compartment from the adjacent dorsal interossei. This is not supported by clinical findings. With various minor modifications, we repeated the experiment, infusing Omnipaque into the second dorsal interosseus muscle of four cadaver hands. We used real-time CT imaging to monitor the spread of contrast medium and side-ported needles to measure compartmental pressures. In all four hands, the tissue barrier between dorsal and volar interossei became incompetent at pressures of less than 15 mmHg. Our data indicate that, although cadaver infusion studies can delineate potentially significant musculoskeletal barriers, their physiological relevance must be confirmed clinically.

Cadaver↗

Lung compartmentalization of increased TNF releasing ability by mononuclear phagocytes in pulmonary sarcoidosis.

The TNF is a monokine with cytotoxic and tumor-necrosing activities; in addition, TNF may play a role in inflammatory processes. The present study evaluates spontaneous and LPS-mediated release of TNF by AMs and autologous peripheral BMs of normal subjects and patients with pulmonary sarcoidosis. A recently developed cytotoxicity assay, specific for detection of TNF activity, was applied. This study demonstrates that (1) unstimulated mononuclear phagocytes released low levels of TNF with no differences between groups; (2) when effector cells were stimulated with LPS, AMs from patients with active pulmonary sarcoidosis released more TNF than AMs recovered from normal subjects and from patients with inactive disease; (3) this increase was compartmentalized to the lungs, since comparisons of TNF production by LPS-stimulated BMs failed to show any difference between study groups. These results suggest that TNF might play a role in the pathogenesis of the alveolitis of pulmonary sarcoidosis.

Adult↗

Cellular compartmentation of lysozyme and alpha-amylase in the mouse salivary glands. Immunogold approaches at light and electron microscopy level.

The research was planned to study the subcellular distribution of enzymatic secretory products within the secretory structures of the mouse major salivary glands at light and electron microscopy level by immunogold silver stain (IGSS) technique and double-sided post-embedding immunogold binding and silver amplification in order to speculate about their compartmentation. In particular, we experimented the above immunogold labeling approaches to localize the lysozyme and to verify its distribution patterns in relation to another secretion enzyme, alpha-amylase. Co-presence of lysozyme and alpha-amylase was observed in the convoluted granular tubule cells of the submandibular gland and in the demilunar cells of the sublingual gland as well as in the electron-dense regions of the mottled secretory granules in the parotid gland. Exclusive binding patterns of lysozyme were observed in the acinar cells of the submandibular and sublingual glands where alpha-amylase did not occur.

Animals↗

Continuous photobleaching in vesicles and living cells: a measure of diffusion and compartmentation.

We present a comprehensive and analytical treatment of continuous photobleaching in a compartment, under single photon excitation. In the very short time regime (t<0.1 ms), the diffusion does not play any role. After a transition (or short time regime), one enters in the long time regime (t>0.1-5 s), for which the diffusion and the photobleaching balance each other. In this long time regime, the diffusion is either fast (i.e., the photobleaching probability of a molecule diffusing through the laser beam is low) so that the photobleaching rate is independent of the diffusion constant and dependent only of the laser power, or the diffusion is slow (i.e., the photobleaching probability is high) and the photobleaching rate is mainly dependent on the diffusion constant. We illustrate our theory by using giant unilamellar vesicles ranging from approximately 10 to 100 microm in diameter, loaded with molecules of various diffusion constants (from 20 to 300 microm2/s) and various photobleaching cross sections, illuminated under laser powers between 3 and 100 microW. We also demonstrated that information about compartmentation can be obtained by this method in living cells expressing enhanced green fluorescent proteins or that were loaded with small FITC-dextrans. Our quantitative approach shows that molecules freely diffusing in a cellular compartment do experience a continuous photobleaching. We provide a generic theoretical framework that should be taken into account when studying, under confocal microscopy, molecular interactions, permeability, etc.

Biophysics↗

The role of CXCR5 and its ligand CXCL13 in the compartmentalization of lymphocytes in thyroids affected by autoimmune thyroid diseases.

OBJECTIVE: Graves' disease (GD) and Hashimoto's thyroiditis (HT) are characterized by lymphocytic infiltrates partly resembling secondary lymphoid follicles in the thyroid. CXCR5 and its ligand CXCL13 regulate compartmentalization of B- and T-cells in secondary lymphoid organs. The aim of the study was to elucidate the role of this chemokine receptor-ligand pair in thyroid autoimmunity. METHODS: Peripheral blood and thyroid-derived lymphocyte subpopulations were examined by flow cytometry for CXCR5. CXCR5 and CXCL13 cDNA were quantified in thyroid tissues by real-time RT-PCR. RESULTS: We found no differences between the percentages of peripheral blood CXCR5+ T- and B-cells in GD patients (n=10) and healthy controls (n=10). In GD patients, the number of memory CD4+ cells expressing CXCR5 which are functionally characterized as follicular B helper T-cells is higher in thyroid-derived (18+/-3%) compared with peripheral blood T-lymphocytes (8+/-2%). The highest CXCL13 mRNA levels were found in HT (n=2, 86.1+/-1.2 zmol (10(-21) mol) cDNA/PCR) followed by GD tissues (n=16, 9.6+/-3.5). Only low amounts were determined in thyroid autonomy (TA) (n=11) thyroid tissues, irrespective of whether the autonomous nodule (0.5+/-0.1) or the surrounding normal tissue (1.8+/-0.7) had been analyzed. The same differences were found for CXCR5 (HT: 179.1+/-6.8; GD: 17.4+/-10.6; TA(nodule): 0.8+/-0.5; TA(normal): 4.4+/-3.6). In GD, there is a correlation between CXCL13 and CXCR5 mRNA levels and the number of focal lymphocytic infiltrates and germinal centers as well as anti-thyroperoxidase but not anti-TSH receptor autoantibodies. CONCLUSIONS: CXCR5 and CXCL13 play an essential role in maintaining B- and T-cells in lymphocytic infiltrates and ectopic follicles in thyroid tissue from patients affected by autoimmunity.

Adult↗

Compartmentalization gamma/delta T cells and their putative role in mucosal immunity.

gamma/delta T cells are an enigmatic group of cells and their functions still remain unknown. The epithelial-associated gamma/delta T cells, which are abundant at mucosal surfaces, are ideally situated to contribute to the initial stages of the immune response. Recent evidence suggests that they recognize stress-induced self-antigens which would enable a homogeneous population of gamma/delta T cells to monitor multiple insults to the epithelium. This could explain the observed oligoclonality and homogeneous distribution of cells carrying identical TCR within mucosal surfaces. However, the analysis of the TCR delta repertoire from different mucosal surfaces indicated that gamma/delta T cells are highly compartmentalized. Thus, gamma/delta T cells are not one homogeneous group of cells which recognize the same (stress-induced) self-antigens, but consist of different subsets that are likely to have distinct functions. It is possible that gamma/delta T cells interact with antigens that are specific for that organ or recognize foreign antigens which are limited to that site. In addition it was shown that gamma/delta T cells can have opposite functions and be proinflammatory or promote epithelial healing. This review focuses on the distribution and repertoire of mucosal gamma/delta T cells and discusses what is currently known about the functions of these cells. Furthermore, their potential role in inflammatory bowel disease is examined.

Animals↗

Characterization of uptake and compartmentalization of 3,5,3'-tri-iodothyronine in cultured neonatal rat cardiomyocytes.

The uptake of tri-iodothyronine (T(3)) in cultured neonatal rat cardiomyocytes was investigated and compared with the uptake of reverse T(3 )(rT(3)) and thyroxine (T(4)). Cellular compartmentalization of T(3) was studied by distinguishing T(3) activity associated with the plasma membrane from that in the cytosol or incorporated in the cell nucleus. T(3) and T(4) uptake displayed similar temperature dependencies which, in magnitude, differed from that of rT(3) uptake. T(3) uptake was Na(+ )independent, and sensitive to oligomycin and monodansylcadaverine (42-49% and 25% inhibition of 15-min cellular uptake respectively). Furthermore, T(3) uptake could be inhibited by tryptophan (20%) and tyrosine (12%), while 2-aminobicyclo[2,2,1]heptane-carboxylic acid had no effect. Co-incubation with tryptophan and oligomycin resulted in an additive inhibition of T(3) uptake (77%). We therefore conclude that (i) T(3) uptake is energy dependent, (ii) receptor-mediated endocytosis may be involved and (iii) the aromatic amino acid transport system T may play a role, while system L is not involved in T(3) transport in cardiomyocytes. Co-incubation with unlabeled iodothyronines showed that 3,3'-di-iodothyronine and T(3) itself were the most effective inhibitors of T(3) uptake (30% and 36% inhibition of 15-min cellular uptake respectively). At 15-min incubation time, 38% of the total cell-associated T(3) was present in the cytosol and nucleus, and 62% remained associated to the plasma membrane. Unidirectional uptake rates did not saturate over a free T(3) concentration range up to 3.9 microM. We have concluded that T(3) uptake in neonatal rat cardiomyocytes occurs by an energy- and temperature-dependent mechanism that may include endocytosis and amino acid transport system T, and is not sensitive to the Na(+) gradient. Elucidation of the molecular basis for the T(3) transporter is the subject of current investigation.

Amino Acids↗

Compartmentalization of metals within the diverse colloidal matrices comprising activated sludge microbial flocs.

Activated sludge floc from a wastewater treatment system was characterized, with regard to principal structural, chemical, and microbiological components and properties, in relation to contaminant-colloid associations and settling. Multiscale analytical microscopies, in conjunction with multimethod sample preparations, were used correlatively to characterize diverse colloidal matrices within microbial floc. Transmission electron microscopy, in conjunction with energy dispersive spectroscopy (EDS), revealed specific associations of contaminant heavy metals with individual bacterial cells and with extracellular polymeric substances (EPS). Floc structure was mapped from the level of gross morphology down to the nano-scale, and flocs were described with respect to settling properties, size, shape, density, porosity, bound water content, and EPS chemical composition; gross surface properties were also measured for correlation with principal floc features. Compartmentalization results based on 171 EDS analyses and representative high-resolution images showed that nano-scale agglomerations of (i) silver (100%) and (ii) zinc (91%) were confined almost entirely to EPS matrices while (iii) Pb (100%) was confined to intracellular granules and (iv) aluminum was partitioned between EPS matrices (41%) and intracellular matrices (59%). The results suggest that engineered changes in microbial physiology and/or in macromolecular EPS composition may influence metal removal efficiencies.

Flocculation↗

Microbial community structure and activity in a compartmentalized, anaerobic bioreactor.

The objective of this study was to evaluate staging and its effects on reactor performance in a compartmentalized bioreactor, designated the anaerobic migrating blanket reactor (AMBR). The AMBR was operated by reversing the flow several times per day, which allowed for substantial biomass migration without biomass accumulation in the final compartment. During reactor startup, the structures of the microbial communities in the five compartments were quite similar despite substantial differences in substrate types and concentrations in the different compartments. During the rest of the operational period, biomass migration was reduced by changing operating conditions and, as a result, a larger difference in the structures of the microbial communities developed for the different compartments (biomass staging). For example, after changing operating conditions, rRNA levels for the acetate-utilizing methanogen Methanosaeta concilii were approximately 35 and 10% of the total rRNA in the middle and outside compartments, respectively (before changing operating conditions these levels were approximately 20 and 12% of the total rRNA, respectively). Promoting larger differences in the structures of the microbial communities for the different compartments did not improve reactor performance as lower levels of M. concilii in the outside compartments hindered acetate removal and compromised effluent quality.

Biomass↗

Compartmental anatomy: relevance to staging and biopsy of musculoskeletal tumors.

A thorough understanding of compartmental anatomy is essential for accurate staging of a suspected musculoskeletal tumor with MR imaging and for avoiding potentially devastating biopsy-related complications. Imaging-guided, percutaneous needle biopsy is a safe and cost-effective technique but requires careful planning in conjunction with the surgeon who will perform the definitive surgery because it constitutes the final step in the staging process and the first step in surgical therapy.

Biopsy, Needle↗

Is soft-tissue inflammation in pedal infection contained by fascial planes? MR analysis of compartmental involvement in 115 feet.

OBJECTIVE: The purpose of this study was to analyze compartmental involvement and patterns of spread of soft-tissue inflammation in pedal infection. MATERIALS AND METHODS: We reviewed 115 contrast-enhanced 1.5-T MR examinations of the foot in 41 women and 74 men with a mean age of 58.4 years who had undergone bone biopsy or surgery for suspected osteomyelitis. Presence of inflammation (contrast enhancement, fat signal intensity loss on T1-weighted images, and high signal intensity on T2-weighted images) was noted by two musculoskeletal radiologists in the following foot compartments: toes, medial, central, lateral, interosseous, dorsal, hindfoot, malleoli, and lower leg. Proximal and distal extension of soft-tissue inflammation was analyzed. The compartment closest to the ulcer that showed MR signs of direct contiguous infection was designated the primarily infected compartment. RESULTS: Spread of inflammation across fascial planes into neighboring compartments originated from the following primary compartments: medial (3/10, 30%), central (7/16, 44%), and lateral (16/20, 80%). Spread from the hindfoot and malleoli into adjacent compartments was seen in only 7% of such cases (2/24). Inflammation from toe infections spread in 34% of cases to forefoot compartments (15/44). Inflammation from forefoot or toe infections spread in 4.5% of cases to the midfoot and in 2% of cases to the hindfoot; ascension into the calf was rare (1% of cases). Spread of inflammation into neighboring compartments was not correlated with the presence of diabetes (p = 0.81) or with osteomyelitis (p = 0.34). CONCLUSION: Soft-tissue inflammation of the forefoot tends to spread into neighboring compartments, with little respect for fascial planes. Hindfoot inflammation tends to stay confined. Spread from the foot to the lower leg is rare.

Adult↗

Contrasting patterns of insulin biosynthesis, compartmental storage, and secretion. Rat tumor versus islet cells.

A series of 3H-leucine pulse-labeling experiments was carried out with dispersed cells freshly isolated from transplanted rat insulinomas. After secreted fractions were separated, insulin was purified and specific activities were determined for both secreted and average cellular insulins. Labeling patterns in this line of tumor cells were compared with those previously established for isolated rat islets. With both tumors and islets, conversion of labeled proinsulin to insulin occurred to the same extent by 2.5 h, suggesting similar onset and half-time of proteolysis in these cells. However, total cellular insulin in tumors attained a threefold higher specific activity than in islets. Because total B-cell mass in these tumors was unknown, either a more rapid proinsulin biosynthesis or diminished cellular storage (or both) could lead to this faster fractional replacement of total stored insulin. Insulin secretion in these tumor cells was insensitive to high glucose but responded, albeit poorly, to leucine plus 3-isobutyl-1-methylxanthine (IBMX). Under all secretory conditions tested, tumor cells continuously secreted insulin at elevated fractional rates, which were slightly higher than fractional insulin secretory rates in maximally glucose-stimulated islets. In contrast with normal islets, newly synthesized insulin was stored homogeneously in tumor cells, and compartmental storage characteristics were not generated by incubation with either 20 mM glucose or leucine plus IBMX in the marking period. Thus, preferential secretion of insulin was never observed in tumor cells.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-3-isobutylxanthine↗

Technical note: the use of a compartmental model to estimate the de novo production rate of N tau-methylhistidine in cattle.

Urinary N tau-methylhistidine (NMH) excretion has been used as an index of muscle protein breakdown in cattle. An alternative means to estimate muscle proteolysis in cattle is to estimate the de novo production of NMH from plasma kinetics isotopically. Three crossbred steers (average 229 kg) were given a 5.0-mg bolus intravenous injection of [methyl-2H3-N tau-methylhistidine (d3-NMH), after which 16 serial blood samples and three consecutive 24-h urine samples were taken. The enrichment of NMH in plasma was determined by gas chromatography-mass spectrometry, and compartmental analysis of the kinetic data was performed using the SAAM modeling program. The NMH production rates per day (NMHPR, micromoles per day) were 732, 782, and 725, and the fractional breakdown rates (FBR, percentage per day) were 1.61, 1.72, and 1.58 as determined by urinary excretion of NMH, by a three-pool catenary model (plasma kinetics, Model A), and by a more descriptive, three-pool model with two response curves (both plasma and urine kinetics, Model B), respectively. Model A and B estimates of NMHPR and FBR were similar (P greater than .25) to those of estimates obtained from urinary NMH excretion. Kinetic modeling also allows calculation of compartment mass and flux of NMH between compartments and indicates that when NMH exists the muscle pool it is rapidly excreted via the urine. In conclusion, kinetic modeling offers an alternative approach to estimating the NMH production rate.

Animals↗

Validity of specifically applied rare earth elements and compartmental models for estimating flux of undigested plant tissue residues through the gastrointestinal tract of ruminants.

The validity of using rare earth elements as flow markers of undigested residues was evaluated by comparing mean gastrointestinal residence time (GMRT) of rare earths specifically applied to cottonseed hulls (CSH) to that of the indigestible fiber of CSH. Feces were collected from five lambs fed a mineral supplemented diet of CSH containing 52 g CP/kg DM and five lambs fed a CSH plus cottonseed meal diet (CSH+CSM) containing 123 g CP/kg DM. Rare earth elements (La, Yb, and Tb) specifically bound to CSH were included in the diet for a 5-d period and then deleted from the diet for a 3-d period. Following the last fecal collection, lambs were slaughtered for collection of digesta from segments of the gastrointestinal tract. Potentially indigestible NDF (PIF) was determined in diets and digesta from each segment of the gastrointestinal tract. Mean turnover rate, time delay, and GMRT for each rare earth element was estimated by fitting an age-dependent compartment model to profiles of markers appearing in the feces (compartmental model-marker method, CMM). The GMRT also was computed by the indigestible entity pool dilution method (IEPD) as grams of PIF in sampled segment/mean intake rate of PIF proceeding slaughter, g/h. The GMRT computed by the CMM and the IEPD methods did not significantly (P < 0.05) differ (99.6 vs 94.8 h and 58.9 vs 59.5 h for CMM vs IEPD and CSH and CSH+CSM diets, respectively). Regression of GMRT estimated for rare earths vs PIF yielded a highly significant regression (P = 0.001) with a regression coefficient of 0.94 +/- 0.016. It was concluded that rare earth elements applied to specific feeds are valid flow markers for the undigested residues derived from such marked feeds.

Animal Feed↗

Compartmental analysis of resting and activated pulmonary natural killer cells.

Conflicting reports exist regarding the activity of natural killer (NK) cells in resting lung. We speculated that use of cells from differing lung compartments in past studies might have contributed to these discrepancies. To address this issue, a compartmental analysis of relative NK activity of airspace (bronchoalveolar lavage) cells versus interstitial lung cells was conducted in guinea pigs. Non-pulmonary (spleen) cells were also assayed for NK activity. For assays, cells were obtained from each compartment in the same host, then assayed concomitantly using a chromium release assay with the tumor target cell line, K562. Virtually identical NK cell activity was demonstrated by cells derived from airspaces and lung interstitium. This activity was significantly less (p less than 0.05) than that of spleen cells, and its expression required periods of effector: target cell contact which were considerably longer (16 hr) than those needed for spleen cells (4 hr). Further experiments documented the capacity for augmentation of airspace NK cell activity by intratracheal instillation of interferon inducers poly I:C and carboxymethyl acridanone. We conclude that resting guinea pig pulmonary NK cells have low activity relative to systemic cells, that no significant difference in activity exists among NK cells in airspace and interstitial compartments, and that local activation of airspace NK cells is possible using intratracheal interferon inducers.

Acridines↗