Platelet aggregability in apparently healthy humans of different ages

Platelet aggregability was studied in 154 apparently healthy subjects aged 18–60 years. Both spontaneous platelet aggregation and aggregation induced by inducers with different mechanisms of action were studied. In 25-to 27-year-old subjects, the parameters of both the spontaneous aggregation, which characterized the platelet activation, and ADP-induced platelet aggregation, which characterized the total platelet aggregability, were lower than in older subjects (45–50 and 60–70 years of age). The parameters of the platelet activity and aggregation indicated the absence of age-related changes in the structure and functions of platelets in response to inducers, including epinephrine, collagen, and ristocetin.     

Effects of establishing cell cultures and cell culture conditions on the proliferative life span of human fibroblasts isolated from different tissues and donors of different ages

It is well known that normal human cells placed in a culture environment exhibit a limited proliferative capacity. The extent to which the culture environment influences proliferative life span is not understood. This study evaluated the effects of the standard procedures used to establish and maintain cultures on the proliferative life spans of different types of human fibroblast cells established from fetal and adult skin and lung. The results of this study demonstrate that procedures to establish cell cultures use only one of several subpopulations of cells present in biopsy pieces and that the culture conditions routinely employed by most laboratories can exert significant effects on proliferative life-span determinations. The maximum proliferative life span differed significantly when obtained by growing the cells in two commonly used commercial media. Proliferative life span was inversely related to ambient oxygen tension and directly related to seeding density in all of the lines examined although lines established from adult skin were much more resistant to toxicity. Enzymatic antioxidant defense levels of fetal skin fibroblasts were much lower than those observed in adult skin fibroblasts, but the effects of oxygen on their life spans were similar. Hyperoxia induced larger increases in glutathione concentration in cell lines with low antioxidant enzyme levels.     

Variations in insecticidal activity and chemical compositions of leaf essential oils from Cryptomeria japonica at different ages

The larvicidal effects of the essential oils extracted from the leaves of Cryptomeria japonica at different ages (58, 42, and 26 years old) against 2 mosquito species, Aedes aegypti and Aedes albopictus, were studied. The analysis of major constituents of these essential oils was also investigated. Results obtained from the larvicidal tests, using essential oil from the leaves of 58-year-old C. japonica was found to be most effective against both A. aegypti and A. albopictus larvae, indicating tree age has significant influence on mosquito larvicidal activity. In addition, the eleven pure constituents from C. japonica leaf essential oil were also tested individually against the two mosquito larvae. Among them, alpha-terpinene, gamma-terpinene, p-cymene, 3-carene, terpinolene, and beta-myrcene shows strong larvicidal effect against the two mosquito larvae. Among these pure constituents, 3-carene exhibits the best larvicidal effect against A. aegypti and terpinolene shows an excellent inhibitory action against A. albopictus larvae. The results of this study show that the leaf essential oil and its effective constituents might be considered as a potent source for the production of fine natural larvicides.     

Lipid fluidity and composition of intestinal microvillus membranes isolated from rats of different ages

The lipid composition and fluidity of microvillus (luminal) membranes isolated from the small intestines of Fisher 344 rats aged 6, 17, and 117 weeks were compared. Lipid fluidity, as assessed by the fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene, was significantly greater in rats aged 6 weeks as compared to 17 or 117 weeks. A lipid thermotropic transition was observed at 17.5 +/- 1.3 degrees C in the membranes of the youngest group, approx. 5-6 degrees C lower than that of the older animals. The differences in lipid composition which account for the higher fluidity of the youngest preparations include a decreased cholesterol/phospholipid molar ratio in both the proximal and distal halves of the small intestine and, in the proximal half alone, increases in the lipid/protein ratio and double bond index. The foregoing reduction in cholesterol/phospholipid ratio derives mainly from a higher content of total phospholipid, and the increment in double bond index results from an increase in arachidonic acid residues. The results demonstrate an age-dependent decrease in fluidity of intestinal microvillus membranes in the early post-weaning period in the rat. This pattern was unlike that of the microvillus membrane p-nitrophenylphosphatase, whose specific activity declined progressively in the older age groups.     

Mechanisms of calcification by vesicles isolated from atherosclerotic rabbit aortas

Although several lines of evidence support the role of calcifiable vesicles in dystrophic vascular calcification, the mechanisms whereby vesicles promote aortic calcification remain incompletely understood. Previous reports indicate that ATP promotes in vitro vesicle calcification. Whether ATP-initiated calcification is simply mediated through increased Pi concentrations or by other unknown mechanisms related to ATP hydrolysis is unclear. To determine whether high Pi levels resulting from ATP hydrolysis may cause CaxP ion products to surpass the threshold for calcium phosphate precipitation, 3 mM Pi instead of 1 mM ATP was added to calcifying media. The inclusion of 1 mM ATP in calcifying media with an initial serum level of Ca2+ (1.45 mM) and Pi (2.3 mM) was much more effective in promoting calcification than the addition of 3 mM Pi. The higher effectiveness of ATP over Pi in promoting calcification was consistent throughout various incubation periods and vesicle protein ranges. To minimize the effect of CaxPi ion products on calcification, the ion product was kept within the physiological ranges throughout the incubation period by reducing initial Pi or ATP concentrations in calcifying media. At these low levels of ion products, ATP was still more effective than Pi in promoting calcification. Both ATP- and Pi-stimulated calcifications were found to increase with increasing levels of ion products whereas greater effectiveness of ATP over Pi remained unaltered. These observations indicate that ATP hydrolysis may initiate calcification through some mechanisms other than a simple provision of Pi in order to surpass the solubility products. Concanavalin A (Con A) was found to bind to vesicles and to enhance both ATP- and Pi-promoted calcification. Taken together, these observations suggest that ATP hydrolysis, CaxP ion products, and vesicle-associated carbohydrates are implicated in vesicle-mediated calcification.     

Comparative antibiotic sensitivity of different species of staphylococci isolated from humans]

A total of 206 strains of various staphylococcal species isolated from various sources were studied with respect to their sensitivity to 18 antibiotics. The number of strains poly-resistant to the antibiotics was almost the same among Staph. aureus and Staph. epidermidis, i. e. 54.8 and 51.3 per cent respectively. The coagulase-negative and mannitol-negative variants of Staph. aureus and Staph. epidermidis possessing high biological activity (10-14 properties) were resistant to more antibiotics as compared to the low active strains.     

Chemical compositions and antimicrobial activities of four different Anatolian propolis samples

Propolis means a gum that is gathered by bees from various plants. It is known for its biological properties, having antibacterial, antifungal and healing properties. The aims of this study were to evaluate the antimicrobial activity of four different Anatolian propolis samples on different groups of microorganisms including some oral pathogens and comparison between their chemical compositions. Ethanol extracts of propolis (EEP) were prepared from four different Anatolian propolis samples and examined whether EEP inhibit the growth of the test microorganisms or not. For the antimicrobial activity assays, minimum inhibitory concentrations (MIC) were determined by using macrodilution method. The MIC values of the most effective propolis (TB) were 2 microg/ml for Streptococcus sobrinus and Enterococcus faecalis, 4 microg/ml for Micrococcus luteus, Candida albicans and C. krusei, 8 microg/ml for Streptococcus mutans, Staphylococcus aureus, Staphylococcus epidermidis and Enterobacter aerogenes, 16 microg/ml for Escherichia coli and C. tropicalis and 32 microg/ml for Salmonella typhimurium and Pseudomonas aeruginosa. The chemical compositions of EEP's were determined by high-temperature high-resolution gas chromatography coupled to mass spectrometry. The main compounds of four Anatolian propolis samples were flavonoids such as pinocembrin, pinostropin, isalpinin, pinobanksin, quercetin, naringenin, galangine and chrysin. Although propolis samples were collected from different regions of Anatolia all showed significant antimicrobial activity against the Gram positive bacteria and yeasts. Propolis can prevent dental caries since it demonstrated significant antimicrobial activity against the microorganisms such as Streptococcus mutans, Streptococcus sobrinus and C. albicans, which involves in oral diseases.     

Carbohydrate content of insoluble elastins prepared from adult bovine and calf ligamentum nuchae and tropoelastin isolated from copper-deficient procine aorta

1. Insoluble elastin has been prepared by several different methods from adult bovine and calf ligamentum nuchae. Highly purified tropoelastin has been prepared from copper-deficient porcine aorta. 2. Amino acid analyses indicated that all preparations, except that obtained from calf ligamentum nuchae by using an EDTA extraction followed by collagenase digestion (preparation E6), were typical of pure elastin having high concentrations of hydrophobic and low concentrations of hydrophilic amino acids. Preparation E6 was found to contain approx. 40% collagen. 3. The determination and composition of the carbohydrates associated with these preparations is reported. With the exception of preparation E6, the insoluble elastins contained only trace amounts of neutral sugars (0.13-0.35%, w/w) and amino sugars (0.01-0.06%, w/w). The porcine tropoelastin contained virtually no carbohydrate. 4. The results suggest that carbohydrate analyses can yield valuable information about the purity of elastin preparations.     

Chemical characterization of glomerular and tubular basement membranes of cattle of different ages

Glomerular and tubular basement membranes were isolated from fetal, neonatal, young and adult bovine kidneys.An isolation method with sieves for both glomeruli and tubules from the same kidney was developed. A detergent procedure appeared to give purer glomerular and tubular basement membrane preparations than the generally used sonication method. No large differences were found in the composition of glomerular and tubular basement membrane of adult animals.Glomerular and tubular basement membrane preparations of the four age groups showed an increase with age of hydroxylysine and both 3- and 4-hydroxyproline. The most marked increases appeared at different stages of development, that of tubular basement membrane being between fetal and neonatal stages and glomerular basement membrane between 18 weeks old and adult animals. The ratio of 3- to 4-hydroxyproline increased considerably during development. Total imino acid content was higher for both types of basement membrane from adult than from young animals, while total content of hydroxylysine plus lysine remained fairly constant.The increase in hydroxylation of lysine was accompanied by a corresponding change in glucose and galactose content so that the ratio of galactose to hydroxylysine or glucose to galactose remained constant. Fucose content of both types of basement membranes was the same for all age groups but content of aminosugars and mannose gradually increased with age.     

Chemical and structural characterisation of iron cores of haemosiderins isolated from different sources.

The elemental content of the iron cores of haemosiderins isolated from animal and human tissues has been determined to ascertain whether changes in composition are correlated with structural differences previously identified in these mineralisation products. Significant differences were observed in the elemental composition of haemosiderins isolated from patients subjected to desferrioxamine-chelation therapy compared to patients who had been venesected. The P/Fe molar ratio was considerably higher in haemosiderin isolated from treated primary haemochromatosis (0.83), compared to untreated primary haemochromatosis (0.10) and treated secondary haemochromatosis (0.25), and this could account for the amorphous nature of these iron cores. The levels of M/Fe (M = Ca, Cu, Zn) were reduced in the haemosiderins derived from treated secondary haemochromatosis patients, possibly due to the chelation of these ions by desferrioxamine therapy. In an experimentally iron-loaded rat, receiving either desferrioxamine or 1,2-diethyl-3-hydroxypyrid-4-one, selective decreases in these three elements were also observed after two weeks of desferrioxamine therapy. Such changes may be important determinants in the modification of biomineralisation of the iron cores.     

Effects of lectins on calcification by vesicles isolated from aortas of cholesterol-fed rabbits

Advanced vascular calcification in atherosclerosis weakens arterial walls, thereby imposing a serious rupturing effect. However, the mechanism of dystrophic calcification remains unknown. Although accumulating morphological and biochemical evidence reveals a role for calcifiable vesicles in plaque calcification, the mechanism of vesicle-mediated calcification has not been fully explored. To study whether vesicles' membrane components, such as carbohydrates, may have a role in vesicle-mediated calcification, the effect of sugar-binding lectins on calcification was investigated. Atherosclerosis was developed by feeding rabbits with a diet supplemented with 0.5% cholesterol and 2% peanut oil for 4 months. Calcifiable vesicles were then isolated from thoracic aortas by collagenase digestion. The histological examination of aortas with hematoxylin counter-staining indicated abnormal formation of large plaques enriched with macrophage-derived foam cells. Fourier transform spectroscopy revealed mild calcification in aortas indicating that advanced stages of heavy calcification have yet to be reached. However, vesicles isolated from the aortas were capable of calcification in the presence of physiological levels of Ca(2+), Pi, and ATP. Thus, at this stage of atherosclerosis, aortas may start to produce calcifiable vesicles, but at a level insufficient for substantial formation of mineral in aortas. The assessments by FT-IR analysis and Alizarin red staining indicated that concanavalin A (Con A) substantially increased mineral formation by isolated vesicles. Con A also exerted a marked stimulatory effect on (45)Ca and (32)Pi deposition in a dose-dependent fashion with a half-maximal effect at 6-10 microg/ml. Either alpha-methylmannoside or alpha-methylglucoside, but not mannitol, at 10 mM abolished the stimulation. Con A stimulation was abolished after Con A was removed from calcifying media, suggesting that covalent binding may not be involved in the effect. Galactosides appear to also be implicated in (45)Ca and (32)Pi deposition since Abrus precartorius agglutinin, which specifically binds galactosides, enhanced the deposition. Neither wheat-germ agglutinin that binds N-acetylglucoside nor N-acetylgalactoside-specific Helix pomatia agglutinin was effective, suggesting that the acetylated forms of carbohydrate moieties are either absent in vesicles or may not be involved in calcification. None of these lectins exerted an effect on ATPase. Thus, the effects of lectins appeared to be mediated through interactions with carbohydrate moieties of calcifiable vesicles. Whether stimulation of vesicle-calcification by lectins is of pathological significance in atherosclerotic calcification requires further investigation.     

Chemiluminescence response of beta-glucan stimulated leukocytes isolated from different tissues and peritoneal cavity of Dicentrarchus labrax

The respiratory burst of leukocytes isolated from sea bass (Dicentrarchus labrax) pronephros, peritoneal cavity (P.C.), spleen and blood, was measured by a chemiluminescence (CL) assay after stimulation with beta-glucan. The CL response by P.C. and pronephros leukocytes was significantly higher than that expressed by a similar number of cells separated from spleen and blood. This probably reflects the observation that the proportion of macrophages and neutrophils was highest in the populations of leukocytes from peritoneal cavity and pronephros. Comparative observations showed a higher degree of yeast phagocytosis by leukocytes taken from peritoneal cavity than the pronephros. Moreover phagocytic index evaluated by microscopical observations, indicated that peritoneal macrophages internalised more yeast cells than neutrophils (identified by the peroxidase reaction). Scanning electron microscopy observations were also carried out.Inhibition experiments by a myeloperoxidase inhibitor sodium azide, iodonium-diphenyl-chloride which inhibits NADPH-oxidase, and exogenous superoxide dismutase, which catalyses O-2 dismutation to H(2)O(2), supported the correlation between CL and respiratory burst. Treatment with ouabain and DNP suggested that in this response, Ca(++) pump channels and calmodulin are involved in a metabolic energy-dependent pathway.