Collagen crosslinking and cartilage glycosaminoglycan composition in normal and scoliotic chickens

The amounts of lysine-derived crosslinks in collagens from tendon, cartilage, intervertebral disc, and bone and changes in the composition of sternal cartilage glycosaminoglycans were estimated in two lines of chickens, a control-isogenic line and a line that develops scoliosis. In the scoliotic line, scoliosis first appears at 3-4 weeks and progressively increases in severity and incidence so that 90% of the birds express the lesion by week 10. We have reported previously that cartilage, tendon, and bone collagens from scoliotic birds are more soluble than corresponding collagens from normal birds. Herein, collagen crosslinking and altered proteoglycan metabolism are examined as possible mechanisms for the differences in collagen solubility. At 1 week of age there were fewer reducible crosslinking amino acids (hydroxylysinonorleucine, dihydroxylysinonorleucine, and lysinonorleucine) in collagens from sternal cartilage and tendon in the scoliotic line than in the isogenic line. However, by week 3 and at weeks 5 or 7 values were similar in both groups. The amounts of hydroxypyridinium in vertebral bone and intervertebral disc collagen were also similar in both groups of birds. Consequently, differences in collagen crosslinking do not appear to be a persistent developmental defect underlying the expression of scoliosis in the model. However, differences were observed in cartilage proteoglycans and glycosaminoglycans from the scoliotic line that were not present in cartilage from the isogenic line. The average molecular weight of the uronide-containing glycosaminoglycans was 30% less in the scoliotic line than in the isogenic line, i.e., 12,000 compared to 18,000. The size distribution of cartilage proteoglycans from the scoliotic line also differed from that of proteoglycans from the isogenic line.(ABSTRACT TRUNCATED AT 250 WORDS)     

Excretion of cholera toxin from Escherichia coli: a potential oral vaccine for cholera

Escherichia coli strain N100 has been mutagenized by transposon mutagenesis and mutants with a cell surface leaky phenotype have been isolated. The mutant designated as E. coli N100::Tn5 excreted periplasmic proteins like ribonuclease and alkaline phosphatase. When this mutant strain was transformed with plasmids containing cloned cholera toxin genes, the toxin protein synthesized in the cells were excreted. The potentiality of this strain as a live oral vaccine for cholera has been discussed.     

Estimation of P-to-O ratio in Bacillus subtilis and its influence on maximum riboflavin yield.

Simultaneous growth and riboflavin overproduction were investigated using a previously developed stoichiometric model of Bacillus subtilis metabolism. A fit of model predictions to experimental data was used to obtain estimates of fundamental energetic parameters of B. subtilis. Although multiple solutions describe the experimental data, evidence for a P-to-O ratio of about 1(1/3) mole of ATP produced per atom of oxygen consumed in oxidative phosphorylation was provided by genomic analysis of electron transport components, because no homologue of the proton-translocating NADH dehydrogenase I was found in the B. subtilis genome database. These results allow us to devise a rational metabolic engineering strategy to improve riboflavin production. The potential influence of increased energy coupling in oxidative phosphorylation on riboflavin yield is discussed. Higher coupling is most significant under carbon-limiting conditions in slow-growing cells, that is, in fed-batch processes of industrial interest.     

Photoreactions of human lens monomeric crystallins

Human lens beta s- and gamma A-crystallins exhibit very similar tryptophan fluorescence emission maxima (329 nm). gamma A isolated from infant human lenses is photo-oxidized by 300 nm irradiation and forms water-insoluble aggregates; beta s or gamma A from young human lenses form a small amount of water-soluble crosslinked species. At least part of the mechanism of photodamage by 300 nm irradiation is photogeneration of the oxidant H2O2 via the generation of O2- radical, this reaction occurs via photosensitization by the tryptophan photo-oxidation product N-formylkynurenine (N-FK) or related species. These results indicate that even though the tryptophan residues of beta s- and gamma A-crystallins are in hydrophobic (buried) microenvironments as compared to those of the alpha- and beta-crystallins, the photogeneration of N-FK is sufficient to produce O2- and H2O2.     

Unexpected metabolites produced from clomethiazole

The heterocyclic drug clomethiazole is metabolized by body passage partly to small aliphatic molecules. The occurrence of such unexpected metabolites is usually overlooked. An appropriate method for their detection is the comparison of urine profiles during drug intake and after withdrawal of the drug.     

Bcl-2 phosphorylation is required for inhibition of oxidative stress-induced lysosomal leak and ensuing apoptosis.

B-cell leukemia/lymphoma 2 (Bcl-2) blocks oxidant-induced apoptosis at least partly by stabilizing lysosomes. Here we report that phosphorylation of Bcl-2 may be required for these protective effects. J774 cells overexpressing wild-type Bcl-2 resist oxidant-induced lysosomal leak as well as apoptosis, and this protection is amplified by pretreatment with phorbol 12-myristate 13-acetate (which promotes protein kinase C (PKC)-dependent phosphorylation of Bcl-2). In contrast, cells overexpressing the Bcl-2 mutant S70A (which cannot be phosphorylated) are not protected in either circumstance. Transfection with Bcl-2(S70E), a constitutively active Bcl-2 mutant which does not require phosphorylation, is protective independent of PKC activation. In contrast, C(2)-ceramide, a putative protein phosphatase 2A activator, abolishes the protective effects of wild-type Bcl-2 overexpression but does not diminish protection afforded by Bcl-2(S70E). Additional results suggest that, perhaps as a consequence of lysosomal stabilization, Bcl-2 may prevent activation of phospholipase A2, an event potentially important in the ultimate initiation of apoptosis.     

Microvascular pressures measured by micropuncture in isolated perfused lamb lungs

To investigate the influence of vasomotor tone and vessel compliance on pulmonary segmental vascular resistance, we determined the longitudinal distribution of vascular pressures in 15 isolated blood perfused lungs of newborn lambs. We measured pulmonary arterial and left atrial pressures and by micropuncture the pressures in 20- to 80-micron-diam subpleural arterioles and venules, both before and after paralyzing the vasculature with papaverine hydrochloride. In five lungs we also determined the microvascular pressure profile during reverse perfusion. In lungs with baseline vasomotor tone, approximately 32% of the total pressure drop was in arteries, approximately 32% in microvessels, and approximately 36% in veins. With elimination of vasomotor tone, arterial and venous resistances decreased to one-fifth and one-half of base-line values, respectively, indicating that vasomotor tone contributed mainly toward arterial resistance. During reverse perfusion, the pressure drop in veins was similar to that in arteries during forward perfusion, suggesting that the compliance of arteries and veins is comparable. We conclude that vascular tone and compliance are important factors that determine the distribution of segmental vascular resistance in lungs of the newborn.