Role of ascorbic acid in procollagen expression and secretion by human intestinal smooth muscle cells

The role of ascorbate in the production and secretion of procollagen by human intestinal smooth muscle cells and the conditions in culture for optimal ascorbate bioefficacy were studied. Procollagen synthesis and secretion were determined by the incubation of cells with L-[5-³H]proline, and the quantitation of radiolabelled procollagen bands in the cell layer and the culture medium by polycrylamide slab gel electrophoresis and densitometry. When cells were cultured without ascorbate in the culture medium, procollagen secretion into the medium was 75% less than in cells receiving fresh ascorbate daily. In the cell layer, in contrast, procollagen accumulation was fourfold greater in the scorbutic cells than in the ascorbate-replete cells. These findings contrasted with those in a control line of scorbutic human dermal fibroblasts in which a 95% decrease in procollagen secretion was not associated with any procollagen accumulation in the cells. In the intestinal smooth muscle cells, the absence of ascorbate resulted in a 25 and 50% decrease in steady-state levels of procollagen I and III mRNA, respectively, compared to a 40 and 75% decrease in fibroblasts. Heat inactivation of the serum in the culture medium augmented the promotion of procollagen secretion by ascorbate two- to fourfold. L-ascorbate phosphate did not increase the activity of L-ascorbate when replaced in medium either daily or every 4 days, and its efficacy was not augmented by serum heat inactivation. The changing of culture medium induced collagen secretion in the absence of ascorbate, but this process was markedly enhanced by ascorbate and induced a transient decrease in the steady-state levels of both procollagen and nonprocollagen mRNAs. The predominant action of L-ascorbate on HISM cells in vitro is to promote procollagen secretion and not procollagen synthesis. L-ascorbate-phosphate is not an adequate substitute for L-ascorbate in this cell line. © 1995 Wiley-Liss, Inc.     

Arsenic Bioaccumulation in Freshwater Fishes

The arsenic ambient water quality criterion (AWQC) for protection of human health via ingestion of aquatic organisms is currently 0.14 μ g/L. This AWQC is derived using a bioconcentration factor (BCF) of 44, which is a consumption-weighted average based on two data points for oysters and fish that was proposed by the U.S. Environmental Protection Agency in 1980 for broad application to freshwater and marine environments. This BCF is based on the assumption that bioaccumulation is a simple linear function of the exposure concentration. In the nearly quarter of a century since this BCF was promulgated, there have been additions to the arsenic bioaccumulation database and a broader scientific understanding of bioaccumulation mechanisms and how they can be applied to estimating tissue concentrations in aquatic organisms. From this database, we identified 12 studies of arsenic bioaccumulation in freshwater fishes in order to explore differences in laboratory-generated BCFs and field-generated bioaccumulation factors (BAFs) and to assess their relationship to arsenic concentrations in water. Our analysis indicates that arsenic concentrations in tissue and arsenic BAFs may be power functions of arsenic concentration in water. A power function indicates that the highest BCF values may occur at low background levels and may decrease as environmental concentrations increase above the ambient range.     

Temperature-shift analysis of conidial development in Aspergillus nidulans

Temperature-shift experiments have been performed on spore-originated colonies of 11 thermosensitive aconidial mutants of Aspergillus nidulans in order to determine the latest time of shift to the restrictive temperature that prevents the initiation of conidiation. This time defines the beginning of the thermosensitive period (TSP) of the mutant. Eight of the mutants have TSPs that begin in the 7-hour period (32–39 hr) just prior to the first appearance of conidia-bearing structures, while 3 of the mutants have TSPs that begin later and very close to the time of onset of conidiation (45 hr). Thus no mutant of the set has a TSP that begins during the first 32 hr of vegetative growth of spore-originated colonies. For all mutants, an upshift performed after the beginning of the TSP allows initiation of conidiation at close to the normal time and at the normal rate, but results in an abrupt cessation of conidiation at some fixed time after upshift, characteristic of the mutant. The mutant whose TSP begins the earliest (aco-49) is exceptional in that, if conidiation is suppressed by growth of colonies in submerged culture, this mutant becomes thermoinsensitive during vegetative submerged growth; in contrast, the remaining 10 mutants become thermoinsensitive only after the suppressive condition has been relieved. We discuss the possibility that this exceptional mutant is defective in a function required for initiation of the process that ultimately results in the formation of conidia.     

Imaging of Surfaces by Concurrent Surface Plasmon Resonance and Surface Plasmon Resonance-Enhanced Fluorescence

Surface plasmon resonance imaging and surface plasmon induced fluorescent are sensitive tools for surface analysis. However, existing instruments in this area have provided limited capability for concurrent detection, and may be large and expensive. We demonstrate a highly cost-effective system capable of concurrent surface plasmon resonance microscopy (SPRM) and surface plasmon resonance-enhanced fluorescence (SPRF) imaging, allowing for simultaneous monitoring of reflectivity and fluorescence from discrete spatial regions. The instrument allows for high performance imaging and quantitative measurements with surface plasmon resonance, and surface plasmon induced fluorescence, with inexpensive off-the-shelf components.     

Encapsulation of hemoglobin in phospholipid vesicles

Hemoglobin has been encapsulated in phospholipid vesicles by extrusion of hemoglobin/lipid mixtures through polycarbonate membranes. This technique avoids the use of organic solvents, sonication, and detergents which have proven deleterious to hemoglobin. The vesicles are homogeneous, with a mean size of 2400 A as determined by photon correlation spectroscopy. The encapsulated hemoglobin binds oxygen reversibly and the vesicles are impermeable to ionic compounds. Hemoglobin encapsulated in egg phosphatidylcholine vesicles converts to methemoglobin within 2 days at 4 degrees C. By contrast, when a mixture of dimyristoyl phosphatidylcholine, cholesterol and dicetyl phosphate is used there is no acceleration in methemoglobin formation, and the preparation is stable for at least 14 days at 4 degrees C.     

Audition in the praying mantis,Mantis religiosa L.: identification of an interneuron mediating ultrasonic hearing

1. The praying mantis possesses a single ear located in the ventral midline of the metathorax. We have studied the mantis' auditory nervous system using both extracellular and intracellular techniques and have identified anatomically and physiologically a mirror-image pair of interneurons (MR-501-T3) in the metathoracic ganglion that mediates ultrasonic hearing. 2. MR-501-T3 is tuned broadly to ultrasound with best sensitivity (55-60 dB SPL) between 25 and 45 kHz. Its tuning matches closely that of the whole tympanal nerve. 3. The physiological responses of MR-501-T3 are characterized by: (1) a phasic-tonic firing pattern with a distinctive initial burst at 500-800 spikes/s; (2) minimum latencies of 8-12 ms; (3) no spontaneous activity; (4) sigmoid intensity response curves with a small (10 dB) dynamic range; (5) accurate coding of stimulus duration and of repetition rates up to 60 pps. 4. The ascending axon of MR-501-T3 conducts action potentials at 4 m/s, a rate comparable with some giant fiber systems. 5. MR-501-T3 shows no directional capability. Sound from right and left produce identical responses in both cells of the pair. Neither cutting one tympanal nerve nor removing one hemi-ear leads to different responses in the two cells indicating that they must receive a common input, either from the auditory afferents or from interneurons. We present evidence that the two cells are not directly connected. 6. MR-501-T3 is a large, symmetrical cell with its processes primarily in the intermediate neuropil (lateral ring tract). Its integration segment crosses the midline in the supramedian commissure, and the cell body lies dorsally near the entrance of the leg nerve. The axon travels in the dorsal lateral tract and is one of the largest (17 microns) in the connective. 7. Given the strong anatomical similarities between MR-501-T3 and the G and B cells of the locust, these cells may be homologous. 8. We present arguments based on our physiological results and existing behavioral data that MR-501-T3 is part of an ultrasonic warning/escape system in the mantis. As in moths, lacewings, and crickets, this system may provide a defense against nocturnally foraging bats.     

Variation in the ratio of curli and phosphoethanolamine cellulose associated with biofilm architecture and properties

Bacterial biofilms are communities of bacteria entangled in a self‐produced extracellular matrix (ECM). Escherichia coli direct the assembly of two insoluble biopolymers, curli amyloid fibers, and phosphoethanolamine (pEtN) cellulose, to build remarkable biofilm architectures. Intense curiosity surrounds how bacteria harness these amyloid‐polysaccharide composites to build biofilms, and how these biopolymers function to benefit bacterial communities. Defining ECM composition involving insoluble polymeric assemblies poses unique challenges to analysis and, thus, to comparing strains with quantitative ECM molecular correlates. In this work, we present results from a sum‐of‐the‐parts ¹³C solid‐state nuclear magnetic resonance (NMR) analysis to define the curli‐to‐pEtN cellulose ratio in the isolated ECM of the E. coli laboratory K12 strain, AR3110. We compare and contrast the compositional analysis and comprehensive biofilm phenotypes for AR3110 and a well‐studied clinical isolate, UTI89. The ECM isolated from AR3110 contains approximately twice the amount of pEtN cellulose relative to curli content as UTI89, revealing plasticity in matrix assembly principles among strains. The two parent strains and a panel of relevant gene mutants were investigated in three biofilm models, examining: (a) macrocolonies on agar, (b) pellicles at the liquid‐air interface, and (c) biomass accumulation on plastic. We describe the influence of curli, cellulose, and the pEtN modification on biofilm phenotypes with power in the direct comparison of these strains. The results suggest that curli more strongly influence adhesion, while pEtN cellulose drives cohesion. Their individual and combined influence depends on both the biofilm modality (agar, pellicle, or plastic‐associated) and the strain itself.