Lung tissue mechanics and extracellular matrix composition in a murine model of silicosis.

The dynamic mechanical properties of lung tissue and its contents of collagen and elastic fibers were studied in strips prepared from mice instilled intratracheally with saline (C) or silica [15 (S15) and 30 days (S30) after instillation]. Resistance, elastance, and hysteresivity were studied during oscillations at different frequencies on S15 and S30. Elastance increased from C to silica groups but was similar between S15 and S30. Resistance was augmented from C to S15 and S30 and was greater in S30 than in S15 at higher frequencies. Hysteresivity was higher in S30 than in C and S15. Silica groups presented a greater amount of collagen than did C. Elastic fiber content increased progressively along time. This increment was related to the higher amount of oxytalan fibers at 15 and 30 days, whereas elaunin and fully developed elastic fibers were augmented only at 30 days. Silicosis led not only to pulmonary fibrosis but also to fibroelastosis, thus assigning a major role to the elastic system in the silicotic lung.     

Models of fibronectin.

The radius of gyration of human plasma fibronectin was determined by light scattering both under conditions in which the molecule is in an extended conformation (ionic strength 1.01 M, pH 8) and close to its native, more compact conformation (ionic strength 0.16 M, pH 8). These values were found to be 17.5 +/- 0.8 nm and 10.7 +/- 0.9 nm respectively, for a constant mol. wt of 533,000 +/- 8000, in excellent agreement with the value of 520,000 deduced from its known composition. A set of models, each made of two identical, end-to-end joined chains of 28 beads, was then constructed, and their calculated physico-chemical parameters were compared with those available for the whole fibronectin molecule and for some of its proteolytic fragments in both conformations. Two possible models for the circulating form are presented here: in both, the fibronectin molecule is in a compact, tangled conformation, with the amino-terminal end of one chain folded over to the carboxy end of itself or of the other chain either in a hairpin or in a circular fashion. With the exception of the carboxy-terminal fibrin(ogen)-binding domains, all the domains appear to be well exposed to the solvent, and thus free to interact with potential ligands.     

Effect of verapamil on the aldosterone-stimulating properties of metoclopramide: in vitro and in vivo studies

The role of calcium in the regulation of aldosterone secretion has been recently clarified. Angiotensin II and potassium stimulate aldosterone secretion through a calcium-entry dependent mechanism, while ACTH action is both calcium and cyclic AMP dependent. To establish whether also the so-called aldosterone dopaminergic regulatory system is calcium-dependent we have studied, in vitro and in vivo, the effect of verapamil, a calcium entry blocker agent, on the aldosterone-stimulating properties of the antidopaminergic drug, metoclopramide. In the rat adrenal cells perfusion system, verapamil blocked both angiotensin II and metoclopramide-stimulated aldosterone. This effect on metoclopramide action seems to be present also in vivo in normal subjects: in fact aldosterone response was slightly but significantly reduced after pretreatment with verapamil. In conclusion the results suggest that also the dopaminergic system could regulate aldosterone secretion through calcium-mediated mechanisms.     

TxA2 production by human arteries and veins

Human arterial and venous segments from patients under-going operations when incubated in Tris buffer both alone and with arachidonic acid were able to produce thromboxane B2 (assessed by radioimmunoassay). Thromboxane B2 (TxB2) production was progressive in time (till 40 min.) and was enhanced by the addition of 1mM norepinephrine. Contamination of tissues by platelet was checked and platelets did not contribute to thromboxane formation. The investigation of the conversion of 1-14C arachidonic acid by vascular tissue indicated that human vascular tissues produce the metabolites of the cyclooxygenase dependent pathway and that prostacyclin is the main metabolite with a PGI2/TxA2 ratio of 4:1. The arterial wall was found to possess an active lipoxygenase dependent pathway. Thromboxane production by intimal cells was negligible and the main source of thromboxane was the media. The production of thromboxane did not change in relation to age, but arterial segments from men produced significantly larger amounts of thromboxane than those from women.     

Tibenelast, 5,6-diethoxybenzo(B)thiophene-2-carboxylic acid, sodium salt (LY186655), is an orally active anti-asthma compound in the guinea pig

Anaphylactic shock was induced in actively sensitized guinea pigs by free inhalation of a high dose of ovalbumin (10 mg/ml) aerosol. Tibenelast (LY186655), 5,6-diethoxybenzo(b)-thiophene-2-carboxylic acid, sodium salt, proved to be a potent orally active compound against anaphylactic shock induced by high dose antigen aerosol. When a lower aerosol challenge (0.05 mg/ml) was employed, bronchoconstriction was observed with a concomitant increase in lung resistance (RL) and a fall in dynamic compliance (Cdyn). Tibenelast at 25 mg/kg p.o. prevented these changes. Tibenelast was 10 times more potent than aminophylline by i.v. administration; normalization of pulmonary function was achieved at 1 mg/kg i.v. Tibenelast was synergistic with epinephrine. Combination of no-effect doses of epinephrine (0.025 mg/kg s.c.) and tibenelast (0.1 mg/kg i.v.) normalized pulmonary function. The oral dose response curve of tibenelast was enhanced with the co-administration of epinephrine. These data suggest that tibenelast may act at a site different from that of epinephrine, although the mechanism of action of tibenelast is unclear at present. Tibenelast may be of significant value in the treatment of asthma and other respiratory diseases.     

Low temperature optical spectroscopy of cobalt-substituted hemocyanin from Carcinus maenas

In this work we report the optical absorption spectra of three cobalt-substituted derivatives of hemocyanin (He) from Carcinus maenas, in the temperature range 300–20 K. The derivatives studied are the mononuclear (Co²⁺)-He with a single cobalt ion in the Cu_A site, the binuclear (Co²⁺)₂-He and the binuclear mixed metal (Co²⁺-Cu¹⁺)-He. At low temperature three main bands are clearly resolved; the temperature dependence of their zeroth, first and second moments sheds light on the stereodynamic properties in the surroundings of the chromophore. Within the limits of the reported analysis, in the binuclear derivatives the motions coupled to the chromophore appear to be essentially harmonic in the whole temperature range investigated; moreover the data are consistent with the presence of an exogenous ligand strongly bound to the two metal ions. For the mononuclear derivative an essentially harmonic behavior is evident only up to 200 K where the data are consistent with the presence of an exogenous ligand much less strongly bound, while at higher temperatures the behavior of the spectra indicates the onset of very large anharmonic contributions to motions, that plausibly involve the above exogenous ligand and, quite likely, the entire active site.Abbreviations He Hemocyanin - M₀ zeroth moment - M₁ first moment - M₂ second moment - (Co^2–)₂-He binuclear bicobalt hemocyanin derivative - (Co²⁺)-He mononuclear monocobalt hemocyanin derivative - (Co²⁺-Cu¹⁺)-He binuclear mixed metals hemocyanin derivative - LFT ligand field theory - CT charge transfer - EPR electronic paramagnetic resonance - XANES X-ray absorption near edge structure Correspondence to: L. Cordone     

Amino acid sequence of the minor isomorph of the crustacean hyperglycemic hormone (CHH-II) of the Mexican crayfish Procambarus bouvieri (Ortmann): Presence of a d-amino acid

The primary structure of the neurohormone crustacean hyperglycemic hormone (CHH-II) was determined by means of enzymatic digestions, manual Edman degradation, and mass spectrometry. CHH-II is a 72 residue peptide (molecular mass 8388 Da), with six cysteines forming three disulfide bridges that connect residues 7–43, 23–39, and 26–52. The peptide has blocked N- and C-termini, and lacks tryptophan, histidine, and methionine. The CHH-I and CHH-II of Procambarus bouvieri have identical sequences and elicit levels of hyperglycemia that are not distinguishable. The difference between the two isomorphs consists in a posttranslational modification of a l-Phe in CHH-I to a d-Phe in CHH-II at the third position from the N-terminus.     

Glycosylation and structure of the yeast MF alpha 1 alpha-factor precursor is important for efficient transport through the secretory pathway

The MF alpha 1 gene encodes a precursor, prepro-alpha-factor, that undergoes several proteolytic processing steps within the classical secretory pathway to produce the mature peptide pheromone, alpha-factor. To investigate the role of structural features of the MF alpha 1 precursor in alpha-factor production, we analyzed the effect of mf alpha 1 mutations that alter precursor structure in a number of ways. These mutations resulted in decreased alpha-factor secretion and intracellular accumulation of pro-alpha-factor. With the exception of the mutant lacking all three N glycosylation sites, the pro-alpha-factor forms that accumulated were core glycosylated but had not yet undergone the addition of outer chain carbohydrate. The delay, therefore, occurred at a step prior to the first proteolytic processing step involved in maturation of the precursor and was probably due to inefficient endoplasmic reticulum-to-Golgi transport. Elimination of all three N-glycosylation sites caused a delay in disappearance of intracellular precursor, and alpha-factor secretion was also slowed. These data indicate that N glycosylation is important but not essential for transport of the precursor through the secretory pathway. The decreased alpha-factor secretion and increased precursor accumulation seen with many different structural changes of pro-alpha-factor indicate that the secretory pathway is extremely sensitive to changes in precursor structure. This sensitivity could cause inefficient secretion of heterologous proteins and hybrids between MF alpha 1 and heterologous proteins in yeast cells.     

Age-related changes in the morphology and function of the zona glomerulosa of the rat adrenal cortex

The age-related changes in the morphology and function of rat adrenal zona glomerulosa (ZG) were investigated by coupled stereological and radioimmunological techniques. For this purpose 4-, 8-, 16- and 24-month-old rats were studied. Aging caused a notable lowering in the plasma aldosterone concentration and a marked decrease in both basal and ACTH- or angiotensin II (ANG-II)-stimulated secretion of collagenase-dispersed ZG cells. Plasma renin activity (PRA) underwent an age-dependent decrease, while the plasma level of ACTH displayed a significant rise. ZG and its parenchymal cells did not evidence any age-related morphologically demonstrable alteration in their growth, nor ZG cells showed any marked ultrastructural change, with the exception of a severe lipid-droplet repletion. This last finding is in keeping with the aging-induced decrease in the secretory activity of ZG cells, inasmuch as lipid droplets are the intra-cellular stores of cholesterol esters, the obligate precursors of steroid hormones in rat adrenals. ACTH and ANG-II are well known to be involved in the maintenance of the growth of rat ZG; thus, the combined impairment of ANG-II production (as evidenced by PRA lowering) and increase in ACTH secretion may maintain unchanged ZG growth during aging.