Immunoelectron microscopic localization of fibronectin in the smooth muscle layer of mouse small intestine

We studied the ultrastructural distribution of fibronectin in the smooth muscle layer of mouse small intestine with affinity-purified antibodies using the immunogold technique. Fibronectin was present over the pericellular area extending from the cell membrane to the extracellular matrix beyond the basal lamina. Distribution of the glycoprotein over the pericellular area was heterogeneous, i.e., it was localized more abundantly in the narrow space between smooth muscle cells, the gaps having a width of 60-80 nm where the two dense bands in adjacent cells matched each other. Such localization suggests that fibronectin contributes to cell adhesion. Within the basement membrane, gold label was localized both in lamina lucida and lamina densa, more densely in the latter than in the former. Fibronectin was also co-distributed with collagen fibers in the extracellular matrix. Within smooth muscle cells, gold particles were observed on rough endoplasmic reticulum and secretory vesicle-like structures. These results suggest that smooth muscle cells synthesize fibronectin and secrete it as a component of the basal lamina and extracellular matrix.     

Blockade of intestinal lipoprotein clearance in rabbits injected with Triton WR 1339-ethyl oleate

Although Triton WR 1339 has been used to block triglyceride or cholesterol removal from plasma, no data are available on the extent to which Triton WR 1339 administered to rabbits blocks clearance of newly absorbed dietary lipids. In the present study, we have measured the efficiency of this blockade during a 24-hr interval. After the Triton WR 1339 administration, plasma Sf greater than 400 and d less than 1.019 g/ml lipoprotein lipid concentrations increased greatly, but the concentration of d greater than 1.019 g/ml lipids decreased. In the rabbits fed 0.5% cholesterol for 1 week, the increase in d less than 1.019 g/ml and the decrease in 1.019 less than d less than 1.063 g/ml lipoprotein fractions 24 hr after the Triton WR 1339 injection were much greater than in the chow-fed Tritonized rabbits. After the Triton treatment, 50% of intravenously injected LDL-125I-labeled apoB disappeared in 24 hr, but little or no apoB appeared in other lipoprotein fractions and no VLDL apoB was converted to LDL. Labeled cholesterol and retinol were fed to rabbits and 24-hr increments in plasma cholesteryl- and retinyl-ester label and mass were measured. In chow-fed Tritonized rabbits about one-half of the absorbed oral doses of both labeled lipids was recovered in plasma, indicating that Triton WR 1339 does not completely inhibit the clearance of intestinal lipoproteins. When rabbits were injected with Triton and an ethyl oleate emulsion, the blockade of dietary lipid removal from plasma was substantially improved and chylomicron cholesterol uptake by extra-hepatic tissues was completely abolished.     

Effect of Exogenous Gangliosides on Amino Acid Uptake and Na+,K+-ATPase Activity in Superior Cervical and Nodose Ganglia of Rats

The effects of some gangliosides on active uptake of nonmetabolizable alpha-aminoisobutyric acid (AIB) and Na+, K+-ATPase and Ca2+, Mg2+-ATPase activities in superior cervical ganglia (SCG) and nodose ganglia (NG) excised from adult rats were examined during aerobic incubation at 37 degrees C for 2 h. In NG, amino acid uptake was greatly accelerated with the addition of galactosyl-N-acetylgalactosaminyl-[N-acetylneuraminyl]-galactosylgluc osyl ceramide (GM1) (85%) and also with N-acetylgalactosaminyl-[N-acetylneuraminyl]-galactosylglucosyl ceramide (GM2) or [N-acetylneuraminyl]-galactosyl-N-acetylgalactosaminyl-[N-acetyl- neuraminyl]-galactosylglucosyl ceramide (GD1a) (43% each) compared with a nonaddition control at a 5 nM concentration. Under identical conditions, Na+, K+-ATPase activity was strongly stimulated with GM1 (180%) and GD1a (93%), whereas Ca2+, Mg2+-ATPase activity showed no change. In SCG, on the other hand, AIB uptake was apparently inhibited (-27%) by addition of GM1, with a slight decrease in Na+, K+-ATPase but no change in Ca2+, Mg2+-ATPase activity in the tissue. Both asialo-GM1, in which N-acetylneuraminic acid is deficient, and Forssman glycolipid, which is not present in nervous tissue, failed to produce any significant increase in both SCG and NG not only in amino acid uptake, but also in Na+, K+-ATPase activity. A kinetic study of active AIB uptake showed that GM1 ganglioside produced an increase in Km with no change in Vmax in SCG, whereas it caused a decrease in Km with a slight increase in Vmax in NG. Treatment of NG and SCG with neuraminidase from Vibrio cholerae, an enzyme that split off sialic acid from polysialoganglioside, leaving GM1 intact, caused little inhibition of the amino acid uptake.(ABSTRACT TRUNCATED AT 250 WORDS)     

Experimental candidiasis in liver injury

In an attempt to evaluate effects of liver injury and roles of iron metabolism on systemic fungal infection, experimental systemicCandida infection was produced in mice with galactosamine-induced liver injury. Survival rate and extent of fungal lesion are compared between mice with liver injury (Group 1) and ones without liver injury (Group 2). Median survival was 7 and 18 days in Group 1 and 2 respectively after 21 days observation. Mortality rate of Group 1 was significantly higher (P=0.05) than that of Group 2. This difference was reflected to the extent of fungal lesions in that they were extensive and disseminated, involving the multiple organs in Group 1 but predominantly localized to the kidneys in Group 2. UIBC (unbound iron binding capacity) and TIBC (total iron binding capacity), i.e., serum transferrin as well as serum iron levels were significantly lower in Group 1 as compared with those in Group 2. These results indicate that hepatic injury promotesCandida infectionin vivo and suggest that increased susceptibility toCandida in the presence of liver injury is, at least partially, attributable to low UIBC and/or TIBC.     

Spontaneous excretion of D-alanine in urine in mutant mice lacking D-amino-acid oxidase.

Urine from mutant mice lacking D-amino-acid oxidase contained a large amount of alanine compared with that from normal mice. Urinary alanine of the mutant mice was sensitive to D-amino-acid oxidase. H.p.l.c. showed that about 94% of the urinary alanine had the D-configuration. These results suggest that D-amino-acid oxidase functions to decompose D-amino acid(s) in normal mice.     

Identification of a novel nifH-like (frxC) protein in chloroplasts of the liverwort Marchantia polymorpha

The frxC gene, one of the unidentified open reading frames present in liverwort chloroplast DNA, shows significant homology with the nifH genes coding for the Fe protein, a component of the nitrogenase complex (Ohyama et al., 1986, Nature 322: 572–574). A truncated form of the frxC gene was designed to be over-expressed in Escherichia coli and an antibody against this protein was prepared using the purified product as an antigen. This antibody reacted with a protein in the soluble fraction of liverwort chloroplasts, which had an apparent molecular weight of 31 000, as revealed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in good agreement with a putative molecular weight of 31945 deduced from the DNA sequence of the frxC gene. In a competitive inhibition experiment, the antigenicity of this protein was indicated to be similar to that of the over-expressed protein in E. coli. Therefore, we concluded that the frxC gene was expressed in liverwort chloroplasts and that its product existed in a soluble form. The molecular weight of the frxC protein was approximately 67 000, as estimated by gel filtration chromatography, indicating that the frxC protein may exist as a dimer of two identical polypeptides analogous to the Fe protein of nitrogenase. The results obtained from affinity chromatography supported the possibility that the frxC protein, which possesses a ATP-binding sequence in its N-terminal region that is conserved among various other ATP-binding proteins, has the ability to bind ATP.     

Involvement of D-amino acid oxidase in elimination of free D-amino acids in mice.

The physiological role of D-amino acid oxidase was investigated by using mutant ddY/DAO- mice lacking the enzyme. Free D-amino acid concentrations in the mutant mice were significantly higher than those of control ddY/DAO+ mice in kidney, liver, lung, heart, brain, erythrocytes, serum and urine. The results suggest that the enzyme is involved in the catabolism of free D-amino acids in the body, and that free D-amino acids are also excreted into urine.     

Effects of inhaled cadmium on breathing in the mouse

Effects of intranasally administered cadmium (3.67 micrograms Cd approximately 36.7 mg per mouse) on breathing were investigated in mice under pentobarbital anesthesia. Cd levels found in the respiratory tract were dependent on the amount administered. Cd mainly caused degeneration and desquamation of the bronchial epithelium and pulmonary congestion, while the carrier solvent had no effects. On the other hand, the carrier solvent decreased respiratory frequency and enhanced its amplitude. These effects were absent 24 h later. However, Cd strongly affected respiration; frequency and amplitude were decreased and recovery at 24 h was not complete at the higher concentrations. These effects by Cd on respiration were dependent on the concentration of administered Cd and the Cd level in lung. Therefore, these results suggest that intranasally administered Cd has inhibitory effects on mouse respiration, perhaps owing to its acute toxicity to pulmonary tissues.     

Electroantennogram responses of the Mediterranean fruit fly, Ceratitis capitata to identified volatile constituents from calling males

Fifty-six compounds from the odor of calling, sexually mature, laboratory reared males of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) were isolated by headspace trapping on Tenax columns and identified using GC/MS techniques (69 total compounds were detected). Electroantennogram responses (EAGs) to 54 of the 56 identified compounds as well as 5 analogs were tested on both sexes. Significant differences between the sexes in their responsiveness were found in 9 of the 54 identified compounds tested. There was no correlation between the amplitude of the EAG response and the relative abundance of compound identified from headspace analysis. Of the five major identified components, three elicited relatively small EAG responses, while two elicited large EAGs compared to the hexan-1-ol standard. The relative ranking of EAG responses were: methyl and ethyl hexenoates and hexanoates > C₄–C₆ esters and/or acetates > ethyl and methyl octenoates > monoterpenes > sesquiterpenes > C₂–C₅ acetates, alcohols and ketones. Behavioral bioassays on each of the five major identified components as well as a blend of six of the compounds showed some degree of attractancy to virgin females which in some cases approached the response to a pheromonal standard (male odors absorbed onto filter paper). These results are discussed in relationship to the insect's antennal sensitivity to putative pheromone components and/or allomonal components and to other reported C. capitata pheromone studies.

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Résumé Cinquante-six composés de l'odeur de mâles de C. capitata Weidemann, élevés en laboratoire, sexuellement mûrs et en appel, ont été isolés par piégeage sur colonnes tenax et identifiés par la technique GC/MS (69 composés avaient été détectés en tout). Les électroantennogrammes (EAGs) ont été examinés chez les deux sexes pour 54 des 56 composés identifiés et 5 de leurs analogues. Des différences significatives entre les sexes ont été observées pour 9 des 54 composés identifiés. Il n'y avait pas de corrélation entre l'ampleur de l'EAG et l'abondance relative du composé lors de son isolement. Pour les 5 principaux composés identifiés, 3 ont induit des EAGs relativement faibles, tandis que 2 étaient importants, par comparaison avec l'Hexane-1-ol utilisé comme témoin. Le classement relatif des EAG a été: hexénoates et hexanoates d'éthyl et de méthyl C₄–C₆ esters et/ou acétates octénoates d'éthyl ou de méthyl monoterpènes sesquiterpènes C₂–C₅ acétates, alcools et kétones. Les expériences de comportement avec chacun des 5 composés principaux identifiés, comme avec des mélanges de 6 composés ont mis en évidence une attraction des femelles vierges qui dans quelques cas avoisine la réponse à la phéromone témoin (odeur du mâle absorbée sur papier filtre). Ces résultats sont discutés en fonction de la sensibilité de l'antenne d'insexte aux composés supposés de la phéromone et aux composés allomonaux, et en fonction des autres études connues sur les phéromones de C. capitata.