Warm blood cardioplegia reduces the fall in the intracellular concentration of taurine in the ischaemic/reperfused heart of patients undergoing aortic valve surgery

Summary The effect of cold and warm intermittent antegrade blood cardioplegia, on the intracellular concentration of taurine in the ischaemic/ reperfused heart of patients undergoing aortic valve surgery, was investigated. Intracellular taurine was measured in ventricular biopsies taken before institution of cardiopulmonary bypass, at the end of 30 min of ischaemic arrest and 20 min after reperfusion. There was no significant change in the intracellular concentration of taurine in ventricular biopsies taken after the period of myocardial ischaemia in the two groups of patients (from 10.1 ± 1.0 to 9.6 ±0.9mol/g wet weight for cold and from 9.3 ± 1.3 to 10.0 ± 1.3mol/g wet weight for warm cardioplegia, respectively). Upon reperfusion however, there was a fall in taurine in both groups but was only significant (P 0.05) in the group receiving cold blood cardioplegia (6.9 ± 0.8mol/g wet weight after cold blood cardioplegia versus 8.0± 0.8mol/g wet weight following warm blood cardioplegia). Like taurine, there were no significant changes in the intracellular concentration of ATP after ischaemia in the two groups of patients (from 3.2 ± 0.32 to 2.95 ± 0.43mol/g wet weight for cold and from 2.75 ± 0.17 to 2.62 ± 0.21mol/g wet weight for warm cardioplegia, respectively). However upon reperfusion there was a significant fall in ATP in both groups with the extent of the fall being less in the group receiving warm cardioplegia (1.79 ± 0.19mol/g wet weight for cold and 1.98 ± 0.27mol/g wet weight for warm cardioplegia, respectively). This work shows that reperfusion following ischaemic arrest with warm cardioplegia reduces the fall in tissue taurine seen after arrest with cold cardioplegia. Accumulation of intracellular sodium provoked by hypothermia and a fall in ATP, may be responsible for the fall in taurine by way of activating the sodium/taurine symport to efflux taurine.     

Method of isolation of cysteine constitutive mutants of the cysteine regulon in Salmonella typhimurium.

Summary Size and shape of mitochondrial DNA molecules of Schizosaccharomyces pombe were analyzed by electron microscopy. Besides numerous linear molecules, circular molecules ranging from 0.83 m to 12.81 m were found. Depending on the method of preparation, both closed and open circular molecules were found. Most of the circular molecules could be assigned to five major size classes of 0.83±0.05 m, 1.7±0.05 m, 4.74±0.04 m, 5.74±0.04 m, and 8.32±0.07 m. Possible explanations for the different size classes of mitochondrial DNA molecules are discussed.     

Localization of sodium absorption and chloride secretion in an intestinal epithelium

Summary Hen coprodeum absorbs sodium electrogenically and, when stimulated by theophylline, secretes chloride. In this study the vibrating microprobe technique was used to localize the transport of these ions to intestinal villi/folds and crypts. With the isolated, stretched epithelium, controlled by light microscopy and scanning electron microscopy, in open circuit, currents were inward, 40±7 A/cm², 50 m vertically above villi, and outward, 36±7 A/cm² above crypts. The currents decayed exponentially to near zero at 300 m with the same length constant. A physical model simulating the observed loci of current sources and sinks predicts potential profiles consistent with our data. Extrapolation of the currents gives a surface potential of 45 V, negative on villi and positive above crypts. Short circuiting increased villus current to 86±27 A/cm² at 50 m, and amiloride treatment reduced it to –8 A/cm²; in both cases crypt currents were abolished. The inward currents are compatible with sodium absorption. Induction of chloride secretion after amiloride treatment, resulted in current circuits similar to those induced by sodium absorption, with villus currents of 23±7 A/cm². This is in accord with chloride secretion at the villi. Quantitative estimates of crypt number (860/cm²) and opening diameter (15 m), in conjunction with isotopic measurements of active and electrical potential-driven ion fluxes demonstrate, however, that only 4% of the potential-driven co-ion transport occurs through the crypts. This indicates that nearly all chloride secretion comes from the sodium-absorbing villar area. Were the chloride secretion to occur solely from the crypts, the current should have been in the opposite direction and 10,000-fold larger.     

Determination of copper, manganese and zinc in human liver

Autopsied liver tissue samples collected from 42 males and 31 females were analyzed for copper, manganese and zinc using atomic absorption spectrometry (AAS). With the exception of two liver samples for which the copper levels were determined to be 74.8 and 104.0 g/g (dry weight), hepatic copper concentrations were found to range from 1.7 to 32.4 g/g with a mean concentration of 14.2 g/g and standard deviation of 7.0 g/g. Manganese concentrations (with the exception of one sample having 12.9 g/g) ranged from 0.22 to 4.6 g/g with a mean of 2.26 ± 1.00 g/g. Hepatic zinc levels averaged 118.3 ± 44.4 g/g and ranged from 38.5 to 231.3 g/g. There were no apparent trends for the levels of any metals versus age nor were there any differences in average hepatic metal concentrations for males and females. © Rapid Science 1998.     

Regional and subcellular distribution of taurine-synthesizing enzymes in the rat central nervous system

The distribution of cysteine oxidase (CO) and cysteine sulfinate decarboxylase (CSD) was examined in 12 regions of the rat central nervous system (CNS). The distribution of CO activity, expressed as mol of cysteine sulfinate formed per h per g, was the following: hypothalamus, superior and inferior colliculi, 94–99 mol/h/g; olfactory bulbs, cerebral cortex, striatum, and hippocampus, 44–51 mol/h/g; cerebellum, 71 mol/h/g; pons-medula and spinal cord, 94 and 60 mol/h/g, respectively. The distribution of CSD activity expressed as mol of cysteine sulfinate decarboxylated per h per g was the following: hypothalamus and colliculi, 14–21 mol/h/g; olfactory bulbs, cerebral cortex, striatum, hippocampus, and cerebellum, 8–13 mol/h/g; pons-medulla, 7.3; and spinal cord, 3.6 mol/h/g. No CSD activity was detected in sciatic nerve. The subcellular distribution of CO and CSD activities was studied in hypothalamus, colliculi, and cerebral cortex. CO activity was localized in synaptosomes, mitochondria, and microsomes. CSD was primarily confined to the crude mitochondrial fraction and after subfraction, recovered mainly in the synaptosomal fraction.     

Methylmercury-injury effect on tube formation by cultured human vascular endothelial cells

The effect of methylmercury chloride (MeHg) on growth and tube formation by cultured human umbilical vein endothelial cells (HUVECs) was investigated. HUVECs were collected by enzymatic digestion with collagenase. Precultivation of HUVECs with MeHg at concentrations of 1.0–50.0 mol/L exerted negligible effects on the viable cell number, while the viable cell number was slightly reduced at 100 mol/L and fell to zero at concentrations exceeding 500.0 mol/L MeHg. The viable cell number was depressed in a concentration-dependent manner. Tube formation was studied by culturing the cells on gelled basement membrane matrix (Matrigel). Treatment of HUVECs with 0.1–5.0 mol/L MeHg for 24 h inhibited tube formation dose-dependently. Fetal bovine serum (FBS) increased tube formation in a dose-dependent manner, with half-maximum stimulation of tube formation at approximately 3.4% FBS. The length of tube formation decreased time-dependently at concentrations of 0.1 and 1.0 mol/L MeHg. Pretreatment of Matrigel with 1 mol/L MeHg before the cell seeding reduced the tube formation by HUVECs. These results suggest that the growth and tube formation by HUVECs is susceptible to MeHg cytotoxicity, and that MeHg could be injurious to endothelial cell function.Abbreviations MeHg methylmercury chloride - HUVECs human umbilical vein endothelial cells     

Characterization and localization of an ATP diphosphohydrolase activity (EC 3.6.1.5) in sarcolemmal membrane from rat heart

In the present report we describe an ATP diphosphohydrolase (apyrase EC 3.6.1.5) in rat cardiac sarcolemma. It is Ca2+ dependent and is insensitive to ouabain, orthovanadate, N-ethylmaleimide (NEM), lanthanum, and oligomycin that are classical ATPase inhibitors. Sodium azide that is a mitochondrial inhibitor at low concentrations, did not affect the enzyme activity at 5.0 mM or below. In contrast, at high concentrations (> 10 mM) sodium azide inhibited the enzyme. Levamisole, a specific inhibitor of alkaline phosphatase and P1, P5-di(adenosine 5-)pentaphosphate (Ap5A), a specific inhibitor of adenylate kinase did not inhibit the enzyme. Mercury chloride showed a parallel inhibition of the hydrolysis of both substrates of apyrase. Similar inhibition profiles are powerful evidence for a common catalytic site for the hydrolysis of both substrates. The enzyme has an optimum pH range of 7.5–8.0 and catalyzes the hydrolysis of triphospho- and diphosphonucleosides other than ATP or ADP. The apparent Km (Michaelis constant) and Vmax (maximal velocity) are 62.1 ± 5.2 M and 1255.7 ± 178 mol inorganic phosphate liberated/min/mg with ATP and 59.4 ± 4.3 M and 269.2 ± 39 mol inorganic phosphate liberated/min/mg with ADP. Enzyme markers indicated that this apyrase is associated with the plasma membrane. A deposition of lead phosphate granules on the outer surface of the sarcolemmal vesicles was observed by electron microscopy in the presence of either ATP or ADP as substrate. It is suggested that the ATP diphosphohydrolase could regulate the concentration of extracellular adenosine, and thus is important in the control of vascular tone and coronary flow.     

In vitro meristem cloning of Eucalyptus tereticornis Sm.

Rapid multiplication of axillary meristems and direct shoot development occurred from nodal explants of mature Eucalyptus tereticornis Sm. with 5.3 M NAA, 1.1 M IAA and 4.4 M BA in Murashige-Skoog medium. Repeated subcultures of the second generation shoot cultures into low cytokinin-auxin containing media (0.44–0.88 M BA+0.1 M NAA) yielded axillary microshoots in large numbers. Half-strength MS liquid medium with 4.9 M IBA, 5.5 M IAA and 5.3 M NAA for four days, half-strength semi-solid hormonefree MS medium with charcoal, and MS liquid medium without charcoal and hormones, in sequence, induced rooting of shoots in the dark. This system is suitable for the mass propagation of this difficult-to-root eucalypt.Abbreviations BA 6-benzyladenine - IAA indole-3-acetic acid - IBA -indole-3-butyric acid - 2-iP isopentyl adenine - Kn kinetin - MS Murashige-Skoog - NAA -naphthalene acetic acid - PVP polyvinylpyrrolidone     

Antifibrotic effects of <Emphasis Type="Italic">Salvia miltiorrhiza </Emphasis>on dimethylnitrosamine-intoxicated rats

Excessive oxidative stress is implicated in hepatic fibrogenesis. Extracts of Salvia miltiorrhiza (Sm) have been shown to protect cells against oxidative stress. In this study we investigated the in vitro and in vivo effects of Sm on hepatic fibrosis. A cell line of rat hepatic stellate cells (HSC-T6) was stimulated with transforming growth factor-1 (TGF-1). The inhibitory effects of Sm (50~400 g/ml) on TGF-1-induced -smooth muscle actin (-SMA) secretion and the mRNA expressions of fibrosis-related genes, including -SMA, connective tissue growth factor (CTGF), and tissue inhibitor of metalloproteinase-1 (TIMP-1), were assessed. Fibrosis was induced by dimethylnitrosamine (DMN) administration in rats. DMN-treated rats were randomly assigned to 1 of 4 groups: saline, Sm (20 mg/kg), Sm (100 mg/kg), or silymarin (100 mg/kg), each given by gavage twice daily for 5 weeks starting from the onset of DMN administration. Sm (200 and 400 g/ml) significantly inhibited TGF-1-stimulated -SMA secretion and the mRNA expressions of -SMA, CTGF, and TIMP-1 in HSC-T6 cells. Fibrosis scores of livers from DMN-treated rats with either a low (1.8 ± 0.2) or high (1.8 ± 0.1) dose of Sm, or silymarin (1.4 ± 0.2) were significantly reduced in comparison with DMN-treated rats receiving saline (3.1 ± 0.1). Hepatic collagen contents were also significantly reduced by either Sm or silymarin treatment. The mRNA expression levels of -SMA, TGF-1, and procollagen I were all attenuated in Sm- and silymarin-treated rats. Moreover, levels of plasma aspartate transaminase activities were reduced by Sm and silymarin treatment. In conclusion, our results show that Sm exerted antifibrotic effects in both HSC-T6 cells and in rats with DMN-induced fibrosis.     

Factors affecting chloride conductance in apical membrane vesicles from human placenta

Summary Apical membrane vesicles from human term placenta were isolated using a magnesium precipitation technique, and the purity of the vesicles was assessed morphologically using scanning and transmission electron microscopy, and biochemically, using marker enzymes. The vesicles were found to be morphologically intact and significantly enriched in enzymes associated with apical membranes. ³⁶Cl^– uptake into these vesicles was studied in the presence of an outwardly directed Cl^– gradient. This uptake was found to be time dependent, with an initial rapid uptake tending to peak between 10 and 20 min and thereafter decline. Uptake was found to be voltage dependent since 5 m valinomycin caused a decrease in uptake. The effects of N-phenylanthranilic acid (NPA) and 4,4-diisothiocyanostilbene-2,2-disulphonic acid (DIDS) and bumetanide on the initial rate of Cl^– were examined in the presence and absence of 5 m valinomycin. NPA and DIDS inhibited isotope uptake strongly with IC₅₀ values of 0.83±0.35 m and 3.43±0.37 m, respectively, in the absence of valinomycin. Although valinomycin reduced ³⁶Cl^– uptake by about 80% when added before the isotope, DIDS reduced the uptake which remained in a concentration-dependent fashion with an IC₅₀ of 5.6±2.1 m. Under these conditions, NPA was without effect at concentrations below 100 m. Bumetanide was without effect at the concentrations used in the absence of valinomycin. However, following valinomycin pretreatment, bumetanide reduced ³⁶Cl^– uptake significantly at 100 m concentration. Vesicle diameter, as assessed by flow cytometry, did not change under the conditions employed.The effects of some fatty acids were also investigated. Arachidonic acid and linoleic acid inhibited Cl^– uptake with IC₅₀ values of 37.6±14.9 m and 4.59±0.51 m, respectively. Arachidonyl alcohol and elaidic acid were found to be without effect. These studies show that human placental brush border membrane vesicles possess a chloride conductance channel, the activity of which can be measured in the presence of an outwardly directed Cl^– gradient and this channel is sensitive to Cl^– channel inhibitors, especially N-phenylanthranilic acid, and can be inhibited by unsaturated fatty acids such as arachidonic acid and linoleic acid.This work was supported in part by the Cystic Fibrosis Association of Ireland and Eolas, The Irish Science and Technology Agency. The technical assistance of Mr. Cormac O' Connell in the preparation of the electron micrographs and of Mr. Roddy Monks in the flow cytometric analysis is gratefully acknowledged.     

Accumulation of silver by growing and non-growing populations of Citrobacter intermedius B6

Summary Accumulation of silver is reported for growing and non-growing populations of Citrobacter intermedius B6. In non-growing cultures a maximum uptake of 4.35% (w/w) was observed at an initial silver concentration of 2111.2 mol Ag⁺ l^-1. In contrast the maximum uptake of silver by growing bacteria was 2.81% (w/w) at an effective concentration of silver of 217.8 mol·l^-1. Silver accumulation rates in both resting (460 mol Ag⁺ g^-1 per hour) and continuously grown (41 mol Ag⁺ g^-1 per hour) bacteria are higher than previously reported. Cell fractionation and electron microscopy of continuously grown bacteria indicated that accumulation of silver was associated with the cell envelope, in the form of dense deposits of macromolecular proportions. This observation discounts simple surface adsorption as the process of accumulation in growing cultures.     

Plasma α- and γ-tocopherol have different pattern during normal human pregnancy

Plasma - and -tocopherol were monitored in pregnant women throughout healthy gestational periods and after delivery and were compared with that of non pregnant women. The mean plasma -tocopherol and -tocopherol concentrations in non pregnant Saudi women (15.2 ± 1.3 and 1.8 ± 0.2 ol/l respectively) were found within normal range. The maternal plasma -tocopherol level steadily increased reaching maximum level (19.1 ± 1.6 mol/l) at late gestation and then gradually decreased after delivery. On the contrary, the optimum level of -tocopherol (2.1 ± 0.2 mol/l) was at mid gestation, followed by a progressive decrease until one month after delivery (1.5 ± 0.1 ol/l). This study shows that the maternal plasma - and -tocopherol have different profiles that may be attributed to their different responses to the changes in maternal lipids during pregnancy.     

A lyophilized aqueous extract of<Emphasis Type="Italic"> Maytenus ilicifolia</Emphasis> leaves inhibits histamine-mediated acid secretion in isolated frog gastric mucosa

Lyophilized aqueous extract of Maytenus ilicifolia leaves (LAEMIL) is commonly used in Brazilian folk medicine in the treatment of dyspepsia as well as gastric ulcers. We have investigated the effect and the possible mechanism of action of the LAEMIL on acid secretion in isolated frog gastric mucosa incubated in an Ussing chamber. It was observed that LAEMIL (7–28 mg%) as well as cimetidine (125–4,000 M), a well-known histamine H₂ receptor antagonist, decreased basal acid secretion in a concentration-dependent manner. Similarly to cimetidine (190 M), LAEMIL (21 mg%) also inhibited gastric acid secretion induced by increasing concentrations of histamine (50–800 M). The EC₅₀ values for histamine alone and histamine in the presence of LAEMIL or cimetidine were 94.6 M (71.1–125.9 M), 244.9 M (209.4–286.4 M) and 142.2 M (23.6–855.0 M), respectively. LAEMIL, histamine and cimetidine were effective on acid secretion only when added to the serosal surface of the mucosa. Furthermore, simultaneous addition of LAEMIL and cimetidine at concentrations, per se, ineffective, caused a 16% reduction in the basal acid secretion [from 8.3±0.3 to 6.9±0.2 Eq g^–1 (15 min)^–1, n=4]. Although effects such as inhibition of histamine biosynthesis and/or histamine release can not be ruled out, our data suggest that LAEMIL, like cimetidine, reduces acid secretion in the isolated frog gastric mucosa by antagonising histamine H₂ receptors.Abbreviations LAEMIL Lyophilized aqueous extract of Maytenus ilicifolia leaves - Hist Histamine - Cim Cimetidine     

Heparin potentiates in vivo neutrophil migration induced by IL-8

Chemokine IL-8 attracts neutrophils by a haptotactic gradient, made possible by its interaction with proteoglycans of the extracellular matrix. Heparan sulfate, but not heparin, potentiates the attraction exerted in vitro by IL-8. In the present study we first confirmed this in vitro phenomenon, observing that IL-8 activity was potentiated 100% by heparan sulfate, but not by heparin. Then, we evaluated the interference of heparan sulfate or heparin on in vivo neutrophil migration induced by IL-8. The activity of rat IL-8 (3.5 g/animal) preincubated with heparan sulfate (50 g/animal) or heparin (77 g/animal) was assayed on the rat dorsal air pouch. Contrary to in vitro experiments, heparin, but not heparan sulfate, potentiated the in vivo IL-8 activity two-fold. We investigated the relationship between this observation and that reported by others, that IL-8-induced migration depends on the presence of mast cells, which contain heparin-rich granules. We studied the neutrophil migration induced by IL-8 (3.5 g/animal) into the rat peritoneal cavity depleted of mast cells. Neutrophil migration was reduced by 32% when compared to that observed in normal animals. The response of depleted rats was reconstituted by preincubation of IL-8 with heparin (77 g/animal). These data suggest that heparin released from cytoplasmic granules may be the contribution of mast cells to IL-8-induced neutrophil migration.     

Salutary effect of tedisamil on post-ischemic recovery rat heart: Involvement of sarcolemmal (Na,K)-ATPase

The in vitro effect of tedisamil on the specific activity and kinetic parameters of the sarcolemmal (Na,K)-ATPase as well as its ex vivo effect on the (Na,K)-ATPase in the isolated, perfused rat hearts was determined. Five mol/l of tedisamil was added 5 min before the onset of 30 min global normothermic ischemia followed by 10 min reperfusion. At the conditions of its maximal cardioprotective effect (heart rate reduction, improved postischemic recovery of left ventricular developed pressure), the hearts were immediately used for isolation of sarcolemmal vesicles. In vitro, 1–100 mol/l of tedisamil produced a concentration-dependent stimulatory effect on (Na,K)-ATPase activity, with a peak seen at 20 mol/l (p < 0.01), while Mg-dependent ATPase was almost unchanged. Kinetic analysis revealed a significant increase in the affinity of the Na-binding sites on ATPase molecule at 20 mol/l of tedisamil. These biochemical findings were confirmed by cytochemistry. Moreover, ex vivo experiments revealed that tedisamil rendered the sarcolemmal (Na,K)-ATPase activity to be a more resistant to detrimental effects of ischemia. In conclusion, the cardioprotective action of tedisamil was accompanied with a better preservation of the specific activity of (Na,K)-ATPase.     

Immunolocalization of synexin (annexin VII) in adrenal chromaffin granules and chromaffin cells: evidence for a dynamic role in the secretory process

Summary Synexin (annexin VII) is a Ca²⁺- and phospholid-binding protein which has been proposed to play a role in Ca²⁺-dependent membrane fusion processes. Using a monoclonal antibody against synexin, Mab 10E7, and immunogold, we carried out a semiquantitative localization study of synexin in bovine adrenal medullary chromaffin granules, and in resting and nicotine-stimulated adrenal chromaffin cells. Isolated chromaffin granules contained very little synexin, whereas chromaffin granules aggregated with synexin (24 g/mg) and Ca²⁺ (1 mM) clearly showed synexin-associated immunogold particles in the vicinity of the granule membrane (1.88 gold particles per granule profile). In isolated, cultured adrenal chromaffin cells, synexin was present in the nucleus (5.5 particles/m²) and in the cytosol (5.3 particles/m²), but mainly around the granule membrane in the granular cell area (11.7 particles/m²). During the active phase of cholinergically stimulated catecholamine secretion, the amount of synexin label was reduced by 33% in the nucleus, by 23% in the cytosol, and by 51% in the granule area. The plasma membrane contained a small amount of synexin, which did not significantly change upon stimulation of the cells. We conclude that synexin is involved in the secretory process in chromaffin cells.     

The Effect of Phenylalanine on DOPA Synthesis in PC12 Cells

DOPA synthesis from phenylalanine was studied in PC12 cells incubated with m-hydroxybenzylhydrazine, to inhibit aromatic L-amino acid decarboxylase. DOPA synthesis rose with increasing concentrations of either phenylalanine or tyrosine; maximal rates (~55 pmol/min/mg protein for tyrosine; ~40 pmol/min/mg protein for phenylalanine) occurred at a medium concentration of ~10 M for either amino acid. The K_m for either amino acid was about 1 M (medium concentration). At tyrosine concentrations above 30 M, DOPA synthesis declined; inhibition was observed at higher concentrations for phenylalanine (300 M). These effects were most notable in the presence of 56 mM potassium. Measurements of intracellular phenylalanine and tyrosine suggested the K_m for either amino acid is 20–30 M; maximal synthesis occurred at 120–140 M. In the presence of both phenylalanine and tyrosine, DOPA synthesis was inhibited by phenylalanine only at a high medium concentration (1000 M), regardless of medium tyrosine concentration. The inhibition of DOPA synthesis by high medium tyrosine concentrations was antagonized by high medium phenylalanine concentrations (100, 1000 M). Together, the findings indicate that for PC12 cells, phenylalanine can be a significant substrate for tyrosine hydroxylase, is a relatively weak inhibitor of the enzyme, and at high concentrations can antagonize substrate inhibition by tyrosine.     

Fuel smoke condensate induced DNA damage in human lymphocytes and protection by turmeric (Curcuma longa)

Summary Twigs-dry leaves smoke condensate (TDS) was investigated for its DNA damaging activity in human peripheral lymphocytes, by using a sensitive method, fluorescence analysis of DNA unwinding (FADU). An aqueous turmeric component (Aq.T) was studied as a protective agent. TDS at one to 100 folds dilution induced 55% DNA damage at 20 min, while 12-0-tetradecanoylphorbol-13-acetate (TPA) at 10 ng/ml induced only 25% damage. Aq.T at 300 ng/1 afforded 90% protection to DNA against TPA and 65% against TPA. The mechanism of Aq.T protection was investigated by using (i) inhibitors of arachidonate cascade, viz., indomethacin (28 M), NDGA (10 M), DBAP (36 M), (ii) antioxidant enzymes viz., CAT (0.2 U/l), SOD (0.6 U/1), (iii) antioxidants - BHA, curcumin (40 M), mixed gangliosides (20 nM) and protease inhibitor TLCK (100 M). These compounds offered the following extents of protection to DNA against TDS: indomethacin-40%, NDGA-83%, DBAP-70%, SOD-38%, CAT-40%, BHA-38%, curcumin-60%, mixed gangliosides-88%, TLCK-85%. Against TPA as clastogenic agent, the extents of protection were: indomethacin-73%, NDGA-32%, DBAP-72%, SOD-60%, CAT, BHA-negligible, curcumin-23%, mixed gangliosides - 60%, TLCK - 59%. These results indicate that (i) TDS and TPA induce DNA damage possibly by different mechanisms, (ii) Aq.T is a more effective protectant against TDS whereas it is on par with other inhibitors against TPA.Abbreviations FADU Fluoroscence Analysis of DNA Unwinding - Aq.T Aqueous component of turmeric - TDS Twigs-Dry leaves Smoke condensate - PBS Phosphate Buffered Saline, 20 mM, 150 mM NaCl, pH 7.4 - TPA 12-O-Tetradecanoyl Phorbol-13-Acetate - NDGA Nordihydroguaiaretic Acid - DBAP 2,4-Dibromo Acetophenone - CAT Catalase - SOD Superoxide Dismutase - BHA Butylated Hydroxyanisole - TLCK Tosyl Lysyl Chloromethyl Ketone - ROS Reactive Oxygen Species - PAH Polycyclic Aromatic Hydrocarbons - DMSO Dimethyl Sulfoxide - Buffer B 250 mM m-inositol, 10 mM sodium phosphate, 1 mM magnesium chloride, pH 7.3 - BSC Beedi Smoke Condensate - CSC Cigarette Smoke Condensate     

The contribution of size-fractionated plankton to biomass and primary production of phytoplankton in saline–alkaline ponds

Primary productivity, biomass and chlorophyll-a of size fractionated phytoplankton (<0.22 m, <3 m, <8 m, <10 m, <40 m, <64 m, <112 m and <200 m) were estimated in 6 ponds and 5 experimental enclosures. The results showed that the planktonic algae less than 10 m are important in the biomass and production of phytoplankton in saline–alkaline ponds. The production of size fractionated phytoplankton corresponding to <112 m, <10 m and <3 m in saline–alkaline ponds were 10.5 ± 6.6 , 8.6 ± 5.4 and 0.33 ± 0.1 mgC l^–1 d^–1, respectively. Mean community respiration rate was 1.80 ± 0.73, 1.69 ± 0.90 and 1.38 ± 1.12 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) were 1.61, 8.30 and 0.33 mgC l^–1 d^–1, respectively. The ratio of those to the total phytoplankton production was 15%, 79% and 3%, respectively. The mean respiration rate of the different size groups was 0.11, 0.31 and 1.38 mgC l^–1 d^–1; the ratio of those to total respiration of phytoplankton was 6%, 17% and 77%, respectively. The production of size-fractionated phytoplankton corresponding to <200 m, <10 m and <3 m in enclosures was 2.19 ± 1.63, 2.08 ± 1.75 and 0.22 ± 0.08 mgC l^–1 d^-1, respectively. Mean community respiration rates were 1.25 ± 1.55, 1.17 ± 1.42 and 0.47 ± 0.32 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton was 0.11, 1.86 and 0.22 mgC l^–1 d^–1, respectively. The ratio of those to the total production of phytoplankton was 5%, 85% and 10%, respectively. The mean respiration rate of different size groups were 0.08, 0.72 and 0.46 mgC l^–1 d^–1, the ratio of those to total respiration of phytoplankton was 6%, 57% and 37%, respectively. The concentrations of chlorophyll-a of the phytoplankton in the corresponding size of micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) plankton in the experimental ponds were 19.3, 98.2 and 11. 9 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 15%, 76% and 9%, respectively. The concentrations of chlorophyll-a of phytoplankton micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton in enclosures were 1.7, 34.3 and 3.0 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 4%, 88% and 8%, respectively.     

The role of intracellular zinc in modulation of life and death of Hep-2 cells

Varying intracellular concentrations of zinc in laryngeal Hep-2 cells in relation to changing cultivation conditions in vitro were determined by atomic absorption spectrophotometry. Upon standard cultivation in DMEM with 10% serum, the mean concentration of zinc was determined at 0.88 ± 0.09 g/mg protein, with substantially decreased values in the cells exposed to a low-serum medium. Next, the study of the effects of a series of physiological and supraphysiological concentrations of ZnSO₄ on laryngeal cells and their correlation with determined intracellular concentrations of zinc was performed. It was found that zinc concentrations above 100 M were toxic to Hep-2 cells, inducing cell death in the interval of 96 h as determined by videomicroscopy, selective nuclear staining, and immunofluorescence detection of caspase-3 and specific cytokeratin 18 fragment. Both types of cell death were observed, with apoptosis being induced at moderately toxic zinc concentration of 150 M and necrosis at higher zinc concentrations of 300 M and 750 M, respectively. Lower concentrations (1.5–100 M), on the other hand, did not produce any measurable changes in cell morphology and function in the same time interval. Zinc at concentration of 1.5 M was found to slightly enhance proliferation of Hep-2 cells up to the certain time point, which seemed to correlate with maximal tolerable momentary intracellular level of zinc. These results illustrate the importance of determining the intracellular levels of zinc when trying to characterize the effect of exogenous zinc on life and death of laryngeal cells.