Time-course of changes in plasma levels of trace elements after thrombolysis during the acute phase of myocardial infarction in humans

It has been suggested that the injury induced by reperfusion of the ischemic myocardium could result, in part, from the cytotoxic effects of oxygen free radicals. Since various trace elements are involved in several of the reactions leading to free radical production, we have measured plasma levels of copper, zinc, selenium, and iron:

1. In 18 patients (mean age 60 yr old) subjected to thrombolytic therapy within 6 h after the onset of a myocardial infarction (G1);
2. In 16 patients with coronary artery disease, but without a history of a previous myocardial infarction (MI) (mean age 50 yr old, G2); and
3. In 50 healthy volunteers divided into two subgroups according to age (mean age 33 yr old, G3 and 55 yr old, G4).

Plasma myosin levels were used to estimate quantitatively the extent of the infarcted mass. Plasma trace element levels were measured in blood samples following centrifugation and storage at ?80°C. The main results were as followed: In G1 patients who have been subjected to thrombolysis, an important release of myosin was measured in plasma, with a peak at D6 (1678 vs 95 μU/L at H0). In those G1 patients after MI:

1. A significant increase in plasma copper levels was observed from day 4 to day 10 postinfarction (×1.15 in reference to the baseline data at H0);
2. A decrease in plasma zinc levels was observed and was maximum 12 h after the onset of the thrombolytic treatment;
3. A decrease in selenium concentration was observed in G1, as well as in G2 patients, compared to the control groups (80% of G3 and G4 values); and
4. A significant decrease in plasma iron levels was observed in G1 (67.8% of G3 and G4 control values) and was significant from H0 to day 7 (p<0.01).

In conclusion, this study underlined the time-course evolution of plasma trace element levels in the followup of patients who have been subjected to thrombolysis following a MI and the potential prognostic implication of such variations.     

Test-tube fertilization inNicotiana tabacum by means of an artificial pollen tube culture

Three techniques of the test-tube fertilization by means of an artificial pollen tube culture are described. The essentials for each individual method are:

1. Transfer of pollen tubes from the sugar solution onto the placentae in the test tube.
2. Placing the placentae with ovules onto pollen tube culture.
3. Precultivation of pollen tubes on small cellophane squares placed on the surface of a semi-liquid medium and transfer of cellophane squares along with the pollen culture to the solid medium, over which finally the placentae with ovules are placed.

By means of all the three techniques viable seeds were obtained in vitro. In an artificial medium pollen tubes are able to maintain their fertilizing ability even after the 24 hours cultivation, i.e. at the time after the formation of the gametes.     

Accumulation,mobilization and turn-over of glycogen in the blue-green bacterium Anacystis nidulans

1. Accumulation of glycogen up to a constant amount per cell was observed during the post-exponential phase of growth, in the presence of an excess of a utilizable carbon source. Cell multiplication was reproducibly controlled by growth of the organism in a nitrogen-limiting medium under photoautotrophic conditions (presence of light, air plus CO₂).
2. Temporary starvation, i.e. by removal of light or by the addition to an illuminated culture of DCMU, 3-(3′,4′-dichlorophenyl)-1,1′-dimethylurea, a specific inhibitor of photosystem II, lead to a mobilization of glycogen in the cell. Furthermore, Anacystis nidulans, having accumulated glycogen by virtue of preculture under nitrogen-limiting conditions, will resume cell division when the culture medium is complemented with a nitrogen source. The ability of the organism to use glycogen as an endogenous carbon source for growth was observed by addition of a nitrogen source to nitrogen-starving cells and simultaneous removal of CO₂.
3. During the period of constant amount of glycogen per cell the reserve polysaccharide was subject to turnover as demonstrated with a pulse chase-labelling technique. The demonstration of a turnover—for the first time with a bacterial species—indicated a strict balance in the relative rate of synthesis and degradation.

     

Molybdenum and iron as functional constituents of the enzymes of the nitrate-reducing system of Azotobacter chroococcum

The roles of molybdenum and iron in the enzymes of the assimilatory nitrate-reducing system from Azotobacter chroococcum have been investigated.

1. By adding ⁹⁹Mo-molybdate to a cell culture of A. chroococcum with nitrate as the nitrogen source, it has been possible to inccrporate the radioactive metal into a purified preparation of the enzyme nitrare reductase.
2. When ¹⁸⁵W-tungstate was supplied to a culture medium lacking added molybdate, a ¹⁸⁵W-labelled nitrate reductase preparation with negligible activity could be obtained. This in vivo incorporation of tungsten was competitively hindered by molybdenum.
3. The cellular level of nitrite reductase activity gradually increased in response to the addition of increasing amounts of iron to the culture medium. Under the same conditions, the level of nitrate reductase activity was not affected.

     

Galanthamine production in “shoot-clump” cultures of Narcissus confusus in liquid-shake medium

Galanthamine (GAL) is increasingly used in the treatment of Alzheimer's disease. We have attempted to develop a method of producing this alkaloid using in vitro cultures of Narcissus confusus plants. The “shoot-clump” culture in liquid medium was shown to be an appropriate method for the micropropagation of this bulbous plant. The complete process included three steps:

1. culture of “twin-scales” starting from the bulbs;
2. culture of the newly formed shoots in a medium for bud proliferation (Murashige Skoog+1 mg l^-1 of 2,4-dichlorophenoxyacetic acid+5 mg l^-1 of benzyladenine), and
3. culture of “shoot-clumps” in a liquid-shake medium. Here we describe the effect of the addition of trans-cinnamic acid, a precursor in the biosynthesis of the Amaryllidaceae alkaloids, on the production of galanthamine and related alkaloids, and also on the growth of the “shoot-clump” culture. The production of galanthamine was found to be inhibited by the addition of the precursor, which promoted the production of the other alkaloid in the same biosynthetic pathway, N-formyl-norgalanthamine. The total production of galanthamine in the control cultures in day-long photoperiod was 2.50 mg per culture, of which 1.97 mg per culture were released into the medium.

     

Somatic embryogenesis and plant regeneration in garden leek

High frequency plant regeneration via somatic embryogenesis has been induced from in vitro shoot-base cultures of seedlings of garden leek (Allium porrum L.). Four main steps are involved in the procedure using BDS medium:

- shoot multiplication with 17.6 mM benzyladenine;

- induction of nodular callus from the in vitro shoot base with 9 mM 2,4-dichlorophenoxyacetic acid;

- initiation of embryogenic callus from nodular callus with 9 mM 2,4-dichlorophenoxyacetic acid +7.6 mM abscisic acid;

- plant regeneration from embryogenic callus with 9.8 mM N⁶-(2-isopentenyl)adenine.

The presence of 2,4-dichlorophenoxyacetic acid in the medium and light conditions were shown to be essential for nodular callus induction and somatic embryogenesis. Abscisic acid was not a prerequiste for somatic embryogenesis, but it significantly increased the frequency.     

Studies on the enzymatic activities related to varied pattern of diets in the air-breathing cat fish,clarias batrachus (Linn.)

1. Specific activity of amylase, cellulase, protease and lipase in the intestines of the air-breathing catfish, Clarias batrachus (Linn.) has been studied.
2. Excepting amylase and protease, the activity of lipase and cellulase showed practically no changes with change in the nutritional status of the diets.
3. pH optima of all enzymes were between 6.9 and 7.6
4. There is reason to believe from cellulase and high amylase activity in the intestine of the species that its culture operation could be done more economically by giving them a supplementary diet from indigeneously available raw material particularly from plant origin.

     

Intracellular proteolytic activity in developing myxospores of Myxococcus xanthus

1. Cell-free extracts from vegetative cells and developing myxospores of Myxococcus xanthus were found to contain similar amounts of proteolytic activity, approximately 80% of which was due to one or more neutral metal proteases.
2. Sixty per cent of the proteolytic activity was particulate.
3. The specific activity of the proteases was high throughout all stages of myxospore formation and displayed small increases in activity at two stages of development: (1) during cell shortening and (2) immediately following the conversion to spheres. The first peak in activity was apparent in assays conducted at pH 8 or 10 whereas the second peak was obvious only at pH 6.
4. A mutant which develops into myxospores only after a lag of approximately 7–8 h possessed levels of proteases similar to the wild type and displayed a peak in proteolytic activity after a delay of 7–8 h.
5. Low levels of serine protease activity were occasionally detected in both vegetative cells and myxospores; no sulfhydryl proteases were detectable in either cell type.
6. Extracellular proteases accumulated in the medium throughout myxospore development but differed from the intracellular proteases in pH optima and sensitivity to inhibitors.

     

Pineapple (Ananas comosus L. Merr) micropropagation in temporary immersion systems

A procedure for the mass propagation of pineapple plants (Ananas comosus L. Merr) using a temporary immersion technique is described. This procedure involved three distinct phases in the automated temporary immersion system: shooting, bud differentiation and elongation. To establish this protocol, we used in vitro shoots obtained from established liquid culture as starting materials. Three culture methods (solid, liquid and temporary immersion) were compared. Temporary immersion increased the multiplication rate and fresh and dry weight after 42 days. Conventional micropropagation (liquid medium) and temporary immersion were compared in combination with paclobutrazol. Paclobutrazol promoted the formation of compact bud clusters with limited leaf development. The highest multiplication rate (106) was found when ex-plants were cultured in shooting medium (MS+2.1 mg/l BA+0.3 mg/l NAA) supplemented with 1 mg/l PB for 7 weeks. A 10-l temporary immersion bioreactor was used to test two approaches during elongation stage: reduction of the shoot-formation period or decrease of the initial number of explants. The highest number of competent and uniform plants (191.8 plant/l) was achieved when shoots were cultured for 4 weeks in shooting medium supplemented with PB. Received: 4 February 1998 / Revision received: 22 June 1998 / Accepted: 14 August 1998     

A novel method for regenerating plants from mesophyll protoplasts of Arabidopsis thaliana line WS

A procedure has been developed for the successful regeneration of plants from mesophyll protoplasts of Arabidopsis thaliana line WS. The protocol is an improved version of that of Damm and Willmitzer (1988). The main changes in original procedure are as follows:

1. A mixture of Cellulase Y-C (0.5%) and Pectolyase Y-23 (0.05%) is used for the isolation of protoplasts. Use of these enzymes reduces the incubation time to 50 min.
2. α-Naphthaleneacetic acid is used as the auxin throughout cultures of protoplasts and calli.
3. Protoplasts and calli are incubated under dim white light (0.8–8 μW/cm²) during culture.

With these modifications, we were able consistently to obtain regenerated shoots from about 70% of calli that had been transferred to shoot-forming medium even though the plating efficiency was rather low (about 0.5–1.5%).     

Stearoyl-CoA desaturase activity in primary culture of chicken hepatocytes. influence of insulin,glucocorticoid, fatty acids and cordycepin

• 1.1. Stearoyl-CoA desaturase (Δ9-desaturase) activity was measured in chicken primary hepatocytes, as a function of time in culture.
• 2.2. When using fasted donor animals, the desaturase activity was low at the beginning of culture and then increased steadily to a maximum value between 30 and 70 hr of culture. When hepatocyte cultures were prepared from fed animals, enzyme activity was high at the beginning of culture and maintained thereafter at similar values to those obtained in cultured hepatocytes from fasted animals after 30 hr of culture.
• 3.3. Insulin significantly enhanced enzyme activity when added to the culture medium at a 10⁻⁹M concentration, and a small stimulating effect was also observed with 10⁻⁶M dexamethasone.
• 4.4. Linoleic acid (0.5 mM) added to the culture medium as albuminic complex partly inhibited Δ9-desaturase activity.
• 5.5. Cordycepin (3' deoxyadenosine) decreased enzyme activity when present at a 3 μg/ml concentration in the culture medium.
• 6.6. Taken together, the induction of enzyme activity in culture, its impairment by cordycepin and response to insulin and linoleic acid strongly suggest that synthesis and translation of the Δ9-desaturase mRNA occur in chicken hepatocytes in primary culture, and that this cellular model may be a useful tool for further studies on Δ9-desaturase regulatory mechanisms.

     

Antioxidant enzymes and trace elements in hemodialyzed patients

Antioxidant enzymes together with trace elements in 26 patients with chronic renal failure treated with hemodialysis and 25 healthy subjects were investigated. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSHPx) in plasma and erythrocytes were examined immediately before and after hemodialysis. The results are summarized as follows:

1. A significant decrease in plasma SOD, CAT, and GSHPx and erythrocyte GSHPx were found in patients before hemodialysis.
2. Erythrocyte CAT and GSHPx were significantly lower in the patients after hemodialysis than in the controls.
3. Plasma GSHPx was significantly higher after a single hemodialysis than before hemodialysis.
4. A good correlation between erythrocyte SOD and copper (Cu) in patients before hemodialysis was found.
5. A good correlation of GSHPx in erythrocytes and plasma was found before hemodialysis, whereas an even better correlation was found after hemodialysis.
6. Abnormalities of trace elements were found in hamodialyzed patients.
7. There is indirect evidence for increased oxidizing stress in uremic patients with hemodialysis. Dialysis treatment may improve some abnormalities (e.g., Hb, P), but may also induce some deleterious effects of free radicals or lipid peroxidation.

     

The use of toxicokinetics for the safety assessment of drugs acting in the brain

Pharmacological and toxicological studies undertaken on drugs that affect the brain are frequently performed in disparate species under various experimental conditions, at doses often greatly in excess of those expected to be administered to humans, and the findings are extrapolated implicitly or explicitly with scant regard to differences in the biodisposition of the drugs. Such considerations are necessary since:

1. Species;
2. Strain;
3. Gender;
4. Route;
5. Dose;
6. Frequency and time of administration;
7. Temperature;
8. Coadministration of drugs; and
9. Surgical manipulation

are but some of the factors that have been shown to influence the kinetics and metabolism of drugs. This article, using MDMA and other phenylethylamines as examples, provides evidence for the need to measure the exoosure of the drugs and their active metabolites in blood and brain (toxicokinetics) in order that conclusions based only on dynamic, biochemical, or histological evidence are more pertinent. Further, the combined use of toxicokinetic-dynamic modeling can lead to a better appreciation of the mechanisms involved and a more useful approach to the calculation of safety margins.     

Untersuchungen zur individuellen Entwicklungskinetik vonGallionella ferruginea in statischer Mikrokultur

The velocity of individual development of the iron bacteriumGallionella jerruginea starting from the point of stalk elongation was determined quantitatively in microculture. In vivo measurements were done by determining stalk elongation, generation time and velocity of cell rotation on individualGallionella-stalks with apical cells developing up to 3-4 generations. The experiments led to the following results:

1. In the presence of ferrous iron and under microaerophilic growth conditions, the development ofGallionella-stalks in microcultures starts immediately after closing the small culture chambers (slide culture). TheGallionella individuals grow in one-layered microzones nearly parallel to the surface of the liquid.
2. The velocity of stalk elongation, up to the first cell division, is 40 to 60 μm/h; in well growing cultures it increases to 80–90 μm/h. The velocity of development in microculture is 2–4 times greater than in natural environments (Hanert, 1973).
3. The stalk development starting from an individualGallionella-thread with apical cell is highest limited to the 3rd–4th generation. It stops after 22–26 h. The maximal total length of all stalk branches formed during this time is 2200–2900 μm.
4. The limitation of individual stalk development after 3–4 division cycles is not caused by limited nutrition or exocellular products in the batch cultures. It is not a function of the special environment because it was found that the stalk development in a natural flowing system is also limited to 2 generations (unpublished). Therefore the limitation must be effected by a special intracellular limiting factor.
5. The average generation time of individualGallionella development requires 5–7 h. In the first generation the cells divide nearly synchronously, in the second there are differences of 2–5 h even among daughter cells. The period up to the first cell division is 1.5–2 h on an average. Compared with development under natural conditions, cell division in microculture occurs more rapidly.
6. It was shown that the developingGallionella-stalks with apical cells elongate only unipolarly at the free non-sessile end of the stalks which extends into the culture medium. The apical cell rotates round its y-axis. At each rotation of 360°, the stalk elongates in a length of λ, i.e. twice the distance between two twists of a stalk.

     

The influence of EDTA on the composition of alginate synthesized by Azotobacter vinelandii

1. During an investigation of the physiology of Azotobacter vinelandii with particular reference to polysaccharide formation, a suitable medium which was precipitate-free was developed by adding EDTA at a concentration of 50 mg/l to a basal medium containing one of eight different carbohydrates as sole carbon source.
2. Acetylated alginate was always produced by the organism when cultured under defined conditions, regardless of the carbohydrate source incorporated in the basal medium.
3. When EDTA was added to the medium, the bacteria produced acetylated polyuronides with a preponderance of mannuronic acid residues.
4. A comparison of the infrared spectra of the alginate produced by Azotobacter vinelandii and the affect of EDTA upon the mannuronic acid/guluronic acid ratios of the alginate are reported.

     

Growth of Nitrobacter in the presence of organic matter

1. Culture filtrates of heterotrophic bacteria were tested for their stimulatory effect on nitrification of three strains of Nitrobacter.
2. Yeast extract-peptone solution, in which Pseudomonas fluorescens had grown, after removal of the cells was added to autotrophically growing cultures of Nitrobacter agilis; it caused a stimulated nitrite oxidation and growth of Nitrobacter agilis.
3. The degree of stimulation depended on: a) the proportion of the culture filtrate to the autotrophic medium; b) the composition of the complex medium in which Pseudomonas fluorescens had been grown; c) the time the heterotrophic bacterium had been grown in the complex medium.
4. The stimulatory effect was highest with Nitrobacter agilis, less with Nitrobacter winogradskyi and negligible with Nitrobacter K ₄.
5. It was possible to adapt nitrifying cells of Nitrobacter agilis to higher concentrations of yeast extract and peptone. After the nitrite had been completely oxidized the cell-N still increased up to 30% before growth stopped.

     

The use of a continuous assay system to show the stimulation of phosphoenolpyruvate production in intact mitochondria by valinomycin and by Mn2+

1. A method for the direct recording of the PEP efflux from isolated mitochondria is described.
2. This method has been used to show the stimulation of PEP efflux by externally added Mn⁺⁺ ions.
3. Valinomycin, uncoupler and oleate were also shown to stimulate PEP efflux.
4. Valinomycin caused an increase in the internal concentration of both PEP and citrate.
5. The results indicate that the major pathway of PEP synthesis in isolated mitochondria is via PEP carboxykinase and the results do not call for an unknown pathway of metabolism.
6. Two interactions between PEP and citrate are described; competition for the mitochondrial interior and the stimulation of PEP production by citrate.

     

The chemical identification of root promoters extracted from avocado tissues

From the Avocado Rooting Promoter (ARP) 4 compounds were isolated and identified as:

1. 1 acetoxy - 2,4 dihydroxy-n-heptadeca-16-en;
2. 1 acetoxy - 2,4 dihydroxy-n-heptadeca-16-yn;
3. 1,2,4 trihydroxy-n-heptadeca-16-en;
4. 1,2,4 trihydroxy-n-heptadeca-16-yn.

The rooting activity of the pure compounds was verified using the mung bean rooting bioassay. Compound 2II is the most active.     

Seasonal variation in the biochemical composition of Mytilus viridis at Ratnagiri on the West Coast of India

1. The seasonal variation in the water, protein, fat and glycogen contents of the mussel, Mytilus viridis has been studied for the year March, 1974 to March, 1975.
2. The water level increased during the monsoon season and decreased in summer.
3. The level of protein, fat and glycogen showed correlation with the reproductive cycle of the mussel.
4. The protein level was high when the mussels were mature and dropped during the breeding period.
5. During sex change from male to female in May the protein level remained high whereas during sex change from female to male in October and November it was low.
6. The fat level was high in mature mussels and declined on spawning.
7. The glycogen level was at its peak in immature mussels and low in mature.

     

A model for a non-chemical form of life: Crystalline physiology

A definition of fundamental living units is given according to which they are constituted by the material support of some ‘memory’ the latter is required

- to be stable,

- to contain rich information,

- to diffuse it into the surrounding medium.

It is then shown that the complex dislocation networks encountered in crystals can in some cases follow these criteria and lead to a crystalline physiology. The places of possible occurrence in nature of this kind of physiology, terrestrial and extraterrestrial rocks, interplanetary dust, white dwarfs and neutron stars are then discussed.