A note on the effect of storage on the chemical resistance of spores of Bacillus subtilis SA22 and Bacillus subtilis globigii B17

Spores of Bacillus subtilis SA22, harvested from nutrient agar after 9 d at 30°C and stored in distilled water at 4°C, were unaltered in their resistance to 17.7% hydrogen peroxide or 0.04% peracetic acid after storage for up to 134 weeks. Three spore crops of B. subtilis globigii were unaffected by storage for up to 134 weeks with respect to 17.7% hydrogen peroxide resistance but were significantly more resistant to 0.04% peracetic acid following storage.     

The role of polypyrrole as charge transfer mediator and immobilization matrix for d-fructose dehydrogenase in a fructose sensor

A fructose dehydrogenase (FDH) modified electrode is produced by the electroadsorption of a layer of FDH on a platinum electrode followed by the electropolymerization of a polypyrrole (PPy) film around and over the enzyme. This immobilizes and stabilizes the enzyme as well as providing an electron transfer pathway to the electrode. The amperometric response to fructose and the enzymatic activity are measured as a function of PPy film thickness. The electrode is shown to have a maximum response at a PPy thickness of approximately the thickness of the enzyme layer. A measure of the electrode efficiency is also obtained, this is the amperometric response to fructose as a percentage of that expected on the basis of the enzyme activity. The functioning of the electrode is also dependent on the counter-ion used for PPy polymerization. This is shown to be mainly related to the nucleation and growth of the PPy film in the interfacial region.     

Effects of exercise on collateral development in myocardial ischemia in pigs

To study the effects of exercise on collateral development in myocardial ischemia, we induced coronary arterial stenosis of the left circumflex coronary artery (LCCA) in 18 of 30 pigs. During that surgery, we identified the coronary bed at risk. Nine of these pigs were then subjected to 5 mo of exercise training on a treadmill. After exercise training, we determined regional collateral and myocardial blood flow using radiolabeled microspheres. At autopsy, all animals had complete occlusion of the LCCA. Infarct size in the exercise-trained pigs was significantly less than in the sedentary pigs (5.9 +/- 1.0 vs. 11.7 +/- 1.0% of the left ventricle). The exercise-trained animals had a greater increase in collateral flow, 35.1 +/- 3.0 vs. 28.7 +/- 4.1 ml X min-1 X 100 g-1, in the noninfarcted jeopardized zone of the LCCA bed. The major findings of the study were the following: 1) chronic coronary artery stenosis progressing to occlusion stimulated development of the collateral circulation and salvaged tissue in the jeopardized myocardium of an animal model with sparse collaterals; 2) development of the collateral circulation and tissue salvage is increased by exercise training; 3) collaterals develop primarily in or near the ischemic zone; and 4) all collateral beds develop a circumferential flow gradient following occlusion.     

Basolateral plasma membrane localization of ouabain-sensitive sodium transport sites in the secretory epithelium of the avian salt gland

The distribution of Na+ pump sites (Na+-K+-ATPase) in the secretory epithelium of the avian salt gland was demonstrated by freeze-dry autoradiographic analysis of [(3)H] ouabain binding sites. Kinetic studies indicated that near saturation of tissue binding sites occurred when slices of salt glands from salt-stressed ducks were exposed to 2.2 μM ouabain (containing 5 μCi/ml [(3)H]ouabain) for 90 min. Washing with label-free Ringer's solution for 90 min extracted only 10% of the inhibitor, an amount which corresponded to ouabain present in the tissue spaces labeled by [(14)C]insulin. Increasing the KCl concentration of the incubation medium reduced the rate of ouabain binding but not the maximal amount bound. In contrast to the low level of ouabain binding to salt glands of ducks maintained on a freshwater regimen, exposure to a salt water diet led to a more than threefold increase in binding within 9-11 days. This increase paralleled the similar increment in Na+-K+-ATPase activity described previously. [(3)H]ouabain binding sites were localized autoradiographically to the folded basolateral plasma membrane of the principal secretory cells. The luminal surfaces of these cells were unlabeled. Mitotically active peripheral cells were also unlabeled. The cell-specific pattern of [(3)H]ouabain binding to principal secretory cells and the membrane-specific localization of binding sites to the nonluminal surfaces of these cells were identical to the distribution of Na+-K+-ATPase as reflected by the cytochemical localization of ouabain-sensitive and K+-dependent nitrophenyl phosphatase activity. The relationship between the nonluminal localization of Na+-K+-ATPase and the possible role of the enzyme n NaCl secretion is considered in the light of physiological data on electrolyte transport in salt glands and other secretory epithelia.     

Effect of acute exercise stress in cardiac hypertrophy: I. correlation of regional blood flow and qualitative ultrastructural changes.

Ultrastructural myocardial cell changes were determined in eight miniswine after the development of pressure-overload hypertrophy induced by supra-valvular aortic constriction. Four miniswine served as control animals. Regional myocardial blood flows were measured at rest and during exercise stress with radioactive microspheres after two days and one month of aortic constriction. Exercise stress, causing the heart rate to increase to 85 percent of its maximum, was imposed twice weekly for 7 minutes on four pressure-overloaded animals and the four control animals to elicit differences between the control and experimental groups that might not occur at rest. After one month of pressure overload the swine were killed and myocardial samples were processed for electron microscopy. Ultrastructural changes similar to those in hypertrophied hearts were present throughout the left ventricular walls of the pressure-overloaded animals. Other changes consistent with ischemic injury were present in the subendocardial regions of pressure-overloaded animals subjected to exercise stress. These changes included disorganization of myofibrils, disintegration and broadening of Z-bands, swelling and aggregation of mitochondria, electron-dense deposits in mitochondria, decreased cristal density and vacuolization of mitochondria, intracellular edema, margination and clumping of nuclear chromatin, and a decrease of glycogen granules. Regional ischemia in the subendocardium of these animals was confirmed by functional studies which showed decreased regional myocardial blood flow to the subendocardium during exercise and S-T segment elevation for the first 2-10 days after inducing pressure overload. The ischemia, as shown by flow studies, during exercise stress persisted in the compensatory stage of hypertrophy although S-T segments returned to normal. Thus, the combined effect of pressure overload and exercise stress can produce focal subendocardial ischemia in the compensated, hypertrophied heart.     

Kinetics of subcellular distribution in rat intestine of 1,25-dihydroxycholecalciferol administered in vivo. Evidence for concentration within 5 min into purified nuclei.

To better understand the initial steps in the induction of intestinal Ca2+ transport by 1,25-dihydroxycholecalciferol [1,25(OH)2D3], we studied the early subcellular localization of 1,25(OH)2D3 in rat intestine. Vitamin D-deficient rats received 300 pmol of 1,25(OH)2[3H]D3 intravenously at 5 min to 4h before being killed. Cells homogenized in buffer of I = 90 mmol/litre were fractionated by centrifugation into a crude nuclear pellet, purified nuclei, Golgi and basal-lateral membranes, cytosol and a post-nuclear pellet. Nuclear purification was established by biochemical and morphological criteria and gave a yield of 32 +/- 2% (mean +/- S.E.M.; n = 21). Although re-establishment of Ca2+ uptake by Golgi is one of the earliest reported intestinal responses to 1,25(OH)2D3, no direct localization of 1,25(OH)2D3 to Golgi was detected. Purified nuclei had the highest specific radioactivity at all times studied, with nuclear localization detectable at 5 min and peak nuclear uptake at 1 h. Relative specific radioactivity of nuclei to cytosol increased from 5 min to 30 min, at which time equilibrium between cytosol and nucleus appeared to be attained. Nuclear uptake occurred in all cells from villus to crypt. Of total nuclear binding 10% was resistant to high ionic strength buffer (I = 365 mmol/litre); peak nuclear uptake was observed at 30 min in this buffer. This tight binding may represent the active fraction of 1,25(OH)2D3. These results indicate that localization of 1,25(OH)2D3 to rat intestinal nuclei precedes the observed Golgi-membrane effects and suggest the existence of high-affinity nuclear 1,25(OH)2D3-binding sites.     

Hapten-specific B cell repertoire probed by hybridoma technology: selection and characterization of representative clonotypes from the antibody-forming cell pool

In the course of this study, more than 3000 phthalate-specific antibody-forming cell hybrids were identified using the hybridoma technology. With the aid of a rapid screening assay and an extensive library of phthalate analogs, it was possible to assign selected hapten-specific clones to one of 11 distinct fine-specificity sets. This compartmentalization of the phthalate-specific hybridomas has made it possible to focus attention upon a single manageable portion of the phthalate-specific repertoire. Fourteen clones from a single fine-specificity set were selected for further immunochemical characterization. Five of these clones were found to secrete an antibody that was indistinguishable in isoelectric focusing. Affinity-purified, high-resolution anti-idiotype antibodies were prepared with specificity for the antibodies produced by one of these clones (i.e., 4C7). A major portion of the serologically defined private idiotype (4C7 IdI) was shown to be associated with the ligand-combining site. Our results indicate that the five clones that share a common spectrotype also express the 4C7 IdI. Two other independently derived clones from two distinct fusions also share this idiotype. The 4C7 IdI was also identified in affinity-purified anti-phthalate antibodies derived from a pool of phthalate-immune serum (conventional antibody) and from affinity-purified antibodies derived from a pool of serum from unimmunized BALB/c mice (natural antibody). The 4C7 IdI is thus considered to represent a repeating clonotype in the phthalate-specific repertoire of BALB/c mice, and will serve as one of several useful clonal markers that are being developed for studies of the mechanism regulating idiotype expression.