Vitamin B12-induced alterations in activities of α-glycerophosphate dehydrogenase and malic enzyme in brain of singi fish, Heteropneustes fossilis (Bloch)

Different doses of vitamin B₁₂ (0.25, 0.5, 1, 2 and 4 μg/g, injected intraperitoneally for three consecutive days) altered the activities of mitochondrial-α-glycerophosphate dehydrogenase (α-GPD) and NADP-dependent cytosolic malic enzyme (ME) in the brain of singi fish. The α-GPD activity increased at doses of 0.5, 1, 2 and 4 μg/g vitamin B₁₂. A dose of 0.5 μg/g vitamin B₁₂ induced less activity than higher doses. ME activity increased with 1, 2 and 4 μg/g of vitamin B₁₂/g. The mitochondrial and cytosolic protein content remained unchanged after vitamin B₁₂ administration. Cycloheximide treatment inhibited the vitamin B₁₂-induced increase in α-GPD and ME activity. Thus, vitamin B₁₂ is involved in the induction of some enzymes in fish brain.     

Vitamin B12-induced alterations in activities of alpha-glycerophosphate dehydrogenase and malic enzyme in brain of singi fish, Heteropneustes fossilis (Bloch)

Different doses of vitamin B12 (0.25, 0.5, 1, 2 and 4 micrograms/g, injected intraperitoneally for three consecutive days) altered the activities of mitochondrial-alpha-glycerophosphate dehydrogenase (alpha-GPD) and NADP-dependent cytosolic malic enzyme (ME) in the brain of singi fish. The alpha-GPD activity increased at doses of 0.5, 1, 2 and 4 micrograms/g vitamin B12. A dose of 0.5 microgram/g vitamin B12 induced less activity than higher doses. ME activity increased with 1, 2 and 4 micrograms/g of vitamin B12/g. The mitochondrial and cytosolic protein content remained unchanged after vitamin B12 administration. Cycloheximide treatment inhibited the vitamin B12-induced increase in alpha-GPD and ME activity. Thus, vitamin B12 is involved in the induction of some enzymes in fish brain.     

Demonstration of hepatic cytosolic malic enzyme activity as a thyroid hormone sensitive physiologic parameter in a teleost, Heteropneustes fossilis bloch

Single injections of various doses (0.1, 0.25, 0.5, 5 and 20 micrograms/g) of T3 significantly increased the cytosolic malic enzyme activity (delta OD/min/mg cytosolic protein) in liver of Singi fish Heteropneustes fossilis Bloch, in a dose-dependent nature, maximum up to 5 micrograms/g dose on the 3rd day in comparison to the control. There was no difference in the enzyme activity between 5 and 20 micrograms/g of T3 doses. When the enzyme activity was expressed per mg DNA, the dose-dependent increase in the malic enzyme activity was observed upto 0.5 microgram/g of T3, whereas a fall in the enzyme activity was noticed with 5 and 20 micrograms/g of T3 doses. Lowering the dose of T3 to 0.05 microgram/g was without any effect on the malic enzyme activity (delta OD/min/mg cytosolic protein or DNA). Hepatic cytosolic protein content showed a biphasic nature of variation, significant increase with single injections of 0.05, 0.1, 0.25 and 0.5 microgram/g and a fall with 5 and 20 micrograms/g of T3 doses in comparison to the untreated control. Cycloheximide treatments of the Singi fishes counteracted both the T3-induced rise in the hepatic cytosolic malic enzyme activity (delta OD/min/mg cytosolic protein or DNA) and the hepatic cytosolic protein contents. Thiourea-treated hypothyroid fishes showed significantly decreased level of malic enzyme activity (delta OD/min/mg cytosolic protein or DNA) and cytosolic protein content in liver. A single injection of T3 at 0.25 microgram/g to the thiourea-treated fishes not only recovered but also increased the enzyme activity and cytosolic protein content above the untreated control values.(ABSTRACT TRUNCATED AT 250 WORDS)     

Potentiation of bradykinin-induced vascular permeability increase by prostaglandin E2 and arachidonic acid in rabbit skin

The activity of prostaglandins (PG) in producing vascular permeability was quantitated by dye extraction method in skin of anaesthetized rabbits. PGE₁ and PGE₂ (0.01–10 μg) produced increase in vascular permeability. Activity was approximately equal to that of histamine (Hist) and 1/20 of that of bradykinin (BK) on a weight basis. The activity of PGF₁ and PGF₂ was only 1/20 of that of PGE₁ or PGE₂.

In spite of the relatively low potency of PGE₁ and PGE₂ in the rabbit, near threshold doses (0.1 or 1 μg) of PGE₂ could potentiate permeability responses to bradykinin (0.1 μg) by 10 or 100-fold, respectively. Equivalent doses (0.1 or 1 μg) of histamine could not potentiate the bradykinin responses. Arachidonic acid (AA) at 1 μg, produced a 10-fold potentiation in the permeability response to bradykinin (0.1 μg). Pretreatment of the rabbits with indomethacin (20 mg/kg, i.p.) reduced the responses of BK (0.1 μg) + AA (1 μg) down to a similar magnitude of those seen with bradykinin alone. However, indomethacin did not block responses to either, BK alone, BK + PGE₂, or BK + Hist. Various doses (1, 10, 100 and 300 μg) of arachidonic acid alone also produced increase in cutaneous vascular permeability, although its potency was only 1/3–1/8 of that of PGE₂. This activity of arachidonic acid was attributed in part to its bioconversion to PGE₂, since its activity was significantly reduced by the prostaglandin antagonist, diphloretin phosphate (DPP) (60 mg/kg, i.v.) and by indomethacin (20 mg/kg, i.p.), which blocks conversion of arachidonic acid to prostaglandins. Arachidonic acid may owe some of its permeability increaseing effects to histamine release, since its effects were also reduced by the anti-histamine, pyrilamine (2.5 mg/kg, i.v.).     

Annona squamosa seed extract in the regulation of hyperthyroidism and lipid-peroxidation in mice: Possible involvement of quercetin

Annona squamosa (Custard apple) seeds are generally thrown away as waste materials. The extract of these seeds was evaluated for its possible ameliorative effect in the regulation of hyperthyroidism in mouse model. Serum triiodothyronine (T₃), thyroxine (T₄) concentrations, hepatic glucose-6-phospatase (G-6-Pase) and 5′-mono-deiodinase (5′DI) activity were considered as the end parameters of thyroid function. Simultaneously hepatic lipid peroxidation (LPO), superoxide dismutase (SOD) and catalase (CAT) activities were investigated to observe its hepatotoxic effect, if any.

L-T₄ administration (0.5 mg/kg/d for 12 days, i.p.) increased the levels of serum T₃ and T₄, activity of hepatic G-6-Pase, 5′DI and LPO with a parallel decrease in SOD and CAT activities. However, simultaneous administration of the Annona seed extract (200 mg/kg) or quercetin (10 mg/kg) to T₄-induced hyperthyroid animals for 10 days, reversed all these effects indicating their potential in the regulation of hyperthyroidism. Further, the seed extract did not increase, but decreased the hepatic LPO suggesting its safe and antiperoxidative nature. Quercetin also decreased hepatic LPO. When relative efficacy was compared with that of propyl thiouracil (PTU), a standard antithyroidic drug, experimental seed extract appeared to be more effective. Phytochemical analyses including HPLC revealed the presence of quercetin in the seed extract and the results on the effects of quercetin suggested the involvement of this phytochemical in the mediation of antithyroidal activity of Annona squamosa seed extract.     

Interaction of thyroid hormone and steroidogenic acute regulatory (StAR) protein in the regulation of murine Leydig cell steroidogenesis

The steroidogenic acute regulatory (StAR) protein, a novel phosphoprotein, is a crucial factor involved in intramitochondrial cholesterol transportation, the rate-limiting step in steroidogenesis. The present investigations were undertaken to elucidate involvement of thyroid hormone and StAR protein in the regulation of steroidogenesis in mouse Leydig cells. Treatment of cells with triiodothyronine (T₃) coordinately augmented the levels of StAR protein, StAR mRNA, and steroid production, and these responses were progressively dependent on expression of steroidogenic factor 1 (SF-1). With regard to steroidogenesis and StAR expression, the T₃ response requires both on-going mRNA and protein synthesis. In addition, the effects of T₃ were acutely modulated at the steroidogenic machinery and luteinizing hormone receptor (LHR) function, while these levels were suppressed following longer periods of exposure to T₃. Furthermore, the inhibition of SF-1 expression by DAX-1 markedly abolished T₃-mediated StAR expression in a time frame, which was consistent with decreased steroid biosynthesis. Specific involvement of SF-1 was further confirmed by assessing the 5′-flanking region of the mouse StAR gene, which identified a region between −254 and −110 bp that was essential for T₃ function. Importantly, it was found that the SF-1 binding site at position −135 bp of the 5′-flanking region was greatly involved in T₃-mediated reporter activity. Electrophoretic mobility shift assays (EMSA) also demonstrated involvement of SF-1 in T₃ function. The relevance of T₃-mediated LHR function was investigated in mice rendered hypo-and hyperthyroid, which accounted for up-regulation in the former and down-regulation in the latter group, respectively. These findings demonstrate a key role of thyroid hormone in maintaining mouse Leydig cell function, where thyroid hormone and StAR protein coordinately regulate steroid hormone biosynthesis.     

Enantiorecognition of triiodothyronine and thyroxine enantiomers using different chiral selectors by HPLC and micro-HPLC

This paper deals with the chiral separation of triiodothyronine (T₃) and thyroxine (T₄) by HPLC and micro-HPLC. The separation of T₃ and T₄ is of great pharmaceutical and clinical interest, since the enantiomers exhibit different pharmacological activities. The HPLC measurements were performed on a chiral stationary ligand-exchange phase using l-4-hydroxyproline bonded via 3-glycidoxypropyltrimethoxysilane to silica gel as a selector. Also a chiral teicoplanin (Chirobiotic ™®) phase was used.

In micro-HPLC the chiral separation behaviour of l-4-hydroxyproline, and of the macrocyclic antibiotics teicoplanin and teicoplanin aglycone was investigated for the enantioseparation of T₃ and T₄. l-4-Hydroxyproline was bonded to 3 μm and the glycopeptide antibiotics were bonded to 3.5 μm silica gel and separations were accomplished by microbore HPLC columns (10 cm × 1 mm I.D.). With both techniques and all chiral selectors investigated T₃ and T₄ were baseline resolved. micro-HPLC was found to be superior to analytical HPLC with respect to low consumption of packing material, mobile phase and analyte.     

Zymosan-induced luminol-amplified chemiluminescence of whole blood phagocytes in experimental and human hyperthyroidism

Luminol-amplified CL of whole blood phagocytes was studied in rats given 3 consecutive doses of 0.1 mg L-triiodothyronine T₃/kg or in hyperthyroid patients, after stimulation by zymosan. In both cases, CL was significantly increased, in effect which was produced independently of the opsonization of the zymosan particles and markedly inhibited by azide. The in vitro addition of T₃ or L-thyroxine (T₄) to whole blood phagocytes from normal rats did not modify the opsonized zymosan-dependent CL, when assayed at the concentrations found in eutrhyroid subjects or in hyperthyroid patients. Administrations of propylthiouracil (400 mg/day for 2–3 months) to hyperthyroid patients reduced the CL response observed prior to treatment, to values comparable to those found in the euthyroid group. These data indicate that hyperthyroidism elicits an enhanced respiratory burst activity of whole blood phagocytes, probably related to adaptive changes induced by thyroid hormone on the mieloperoxidase-H₂O₂ system, rather than to direct actions of the hormone molecule or changes in the opsonic capacity of plasma.     

Seasonal Thyroid Cycle of the Soft-shelled Turtle, Lissemys p. punctata (Bonnoterre)

The current study was undertaken to ascertain the seasonal influence of thyroid activity in female soft-shelled turtles, Lissemys p. punctata. Thyroid gland was studied month-wise throughout the year from relative gland weight, histology, epithelial height, glandular peroxidase activity, and RIA of T₃ and T₄ levels from blood serum and the thyroid gland. The values of all the parameters, except those of T₃ and T₄, were higher during March through May, decreased from June through August and began to rise thereafter (September through February). Whereas T₃ and T₄ levels were highest in May, lower during June to November and began to rise thereafter. The difference in the peaks between T₃ and T₄ levels and other parameters have been explained. The findings suggest that thyroid activity of Lissemys turtles varies seasonally and that seasonal factors like temperature play an important role in influencing thyroid activity in soft-shelled turtles.     

Circadian Dependent Effect of Epidermal Growth Factor, Insulin and Glucagon on Hepatic Pyruvate Kinase and Malic Enzyme of Mice

The influence of epidermal growth factor (EGF), 0.75 μg g^-1; insulin, 1.5 μg g^-1; glucagon, 1.25 ygg^-1 and their combinations on the activities of hepatic pyruvate kinase (PK) and malic enzymes (ME) was monitored. Male CD2F₁ mice were treated toward the end of the light or dark periods, 9 or 23 /tours after /ights on (9 or 23 HALO), and subgroups of six mice were killed at 4,8 or 12 hr post-treatment. PK and ME activities from control mice were well characterized by cosine curves. The PK activity was maximal when ME activity was minimal at the transition from light to dark (9 HALO plus 4 hr) and PK was at a minimum when ME was highest (23 HALO plus 4 hr). Both enzymes were influenced by at least one peptide hormone, and the effects were strongly circadian -stage dependent. The only effect attributed to EGF was an increase of PK activity (23%) 12 hr after injection at 23 HALO. PK activity was increased by insulin (23%) at 23 HALO (4 hr after injection), but not at 9 HALO, and decreased (17%) by glucagon 12 hr after injection at 9 HALO. Several reductions in PK activity in response to various combinations of peptides were observed, and appeared to be caused by glucagon but influenced by insulin. The activity of ME was decreased (33%) in response to insulin 4 hr after injection at 23 HALO but not at 9 HALO and increased (60-70%) by glucagon alone or in combinations with insulin or EGF, or both, at 4 hr after injection at 9 HALO but not at 23 HALO. In general, when ME activity was altered by either insulin or glucagon, PK activity was also altered in the opposite direction, and the effects of glucagon were opposed by insulin.     

Ontogeny and Cellular Localization of the Pyruvate Recycling System in Rat Brain

Abstract: The ontogeny of the cerebral pyruvate recycling pathway and the cellular localization of associated enzymes, malic enzyme (ME) and phosphoenolpyruvate carboxykinase (PEPCK), have been investigated using a combination of ¹³C NMR spectroscopy, enzymatic analysis, and molecular biology approaches. Activity of the pathway, using [1,2-¹³C₂]acetate as a substrate, was detected by ¹³C NMR in brain extracts 3 weeks after birth, increasing progressively up to the third month of age. In whole-brain homogenates, ME activity increased to adult levels with the same time course as the recycling pathway. PEPCK activity was low during the first 2 weeks of life and decreased further toward adulthood. ME and PEPCK activity were found in primary cultures of astrocytes and in synaptosomal fractions of adult brain. Primary cultures of cortical neurons showed PEPCK activity but no detectable ME activity. The cytosolic ME gene was expressed in primary cultures of neurons and in astrocytes as well as in the neonatal and adult brain. The PEPCK gene was expressed both in primary cultures of cortical neurons and in astrocytes, but the level of its expression in the neonatal and adult brain was undetectable.     

Effects of Acacia nilotica, A. polyacantha and Leucaena leucocephala leaf meal supplementation on performance of Small East African goats fed native pasture hay basal forages

Optimal utilisation of tannin-rich browse tree fodders including Acacia spp. foliages as crude protein (CP) supplements to ruminants in the tropics is limited by less available information on their feed nutritive potential. Two studies were conducted to: (1) determine rate and extent of ruminal dry matter (DM) degradability (DMD) and (2) investigate effect of sun-dried Acacia nilotica (NLM), A. polyacantha (PLM) and Leucaena leucocephala leaf meal (LLM) supplementation on growth performance of 20 growing (7–9 months old) Small East African male goats (14.6 ± 0.68 kg) fed on native pasture hay (NPH) basal diet for 84 days in a completely randomised design experiment in north-western Tanzania. The goats were randomised into four treatment groups consisting of five animals each. Three supplement diets: 115.3 g NLM (T₂), 125.9 g PLM (T₃) and 124.1 g LLM (T₄), which was used as a positive control, were supplemented at 20% of the expected DM intake (DMI; i.e., 3% body weight) to the three animal groups fed on NPH (basal diet) compared to the animals in a control group that were fed on NPH without browse supplementation (T₁).

NPH had significantly the lowest (P < 0.05) CP of 45.5 g kg⁻¹ DM compared to NLM, PLM and LLM (159, 195 and 187 g kg⁻¹ DM, respectively). NPH had higher (P < 0.05) fibre fractions; lower ruminal DM degradability characteristics and ME than NLM, PLM and LLM. Supplementation of the animals with browse resulted to (P < 0.05) higher average daily weight gains (ADG) of 157.1 g day⁻¹ in T₄ than the animals fed on T₂ (114.3 g day⁻¹) and T₃ (42.9 g day⁻¹), and even to those fed on T₁ (control), which lost weight (−71.4 g day⁻¹). Improved weight gains were mainly due to corrected feed nitrogen (N) or CP due to supplementation of the animals with browse fodder. Too low CP of the NPH would not meet the normal requirements of CP (80 g CP kg⁻¹ DM) for optimal rumen microbial function in ruminants. Higher ADG due to LLM (T₄) and NLM (T₂) supplementation suggest optimised weight gains due to browse supplementation (20% of expected DMI); while lower weight gains from supplementation with PLM (T₃) indicate the possible utilisation of A. polyacantha leaves to overcome weight losses especially during dry seasons.     

Kupffer Cell Function in Thyroid Hormone-Induced Liver Oxidative Stress in the Rat

The influence of thyroid hormone (L-3, 3', 5-triiodothyronine, T₃) on Kupffer cell function was studied in the isolated perfused rat liver by colloidal carbon infusion. Rates of carbon uptake were determined from the influent minus effluent concentration difference and the flow rate, and the respective carbon-induced respiratory activity was calculated by integration of the area under the O₂ curves during carbon infusion. In the concentration range of 0.2 to 2.0 mg of carbon/ml, livers from euthyroid rats exhibited a sigmoidal-type kinetics of carbon uptake, with a Vmax of 4.8 mg/g liver/min and a concentration of 0.82 mg/ml for half-maximal rate; carbon-induced O₂ uptake presented a hyperbolic-type kinetics, with a Vmax of 4.57 μmol of O₂/g liver and a Km of 0.74 mg of carbon/ml, which significantly correlates with the carbon uptake rates. Light-microscopy showed that carbon was taken up exclusively by non-parenchymal cells, predominantly by Kupffer cells. Thyroid calorigenesis was found in parallel with increased rates of hepatic O₂ consumption and thiobarbituric acid reactive substances (TBARS) formation, glutathione (GSH) depletion, and higher sinusoidal lactate dehydrogenase (LDH) efflux compared to control values. In the concentration range of 0.25 to 0.75 mg/ml, carbon infusion did not modify liver LDH efflux in control rats, while it was significantly enhanced in T₃-treated animals. In this latter group, higher carbon concentrations (1 and 1.3 mg/ml) led to loss of viability of the liver. At 0.25 to 0.75 mg of carbon/ml, both the rates of carbon uptake and the associated carbon-induced respiratory activities were significantly increased by T₃ treatment, effects that were abolished by pretreatment of the rats with gadolinium chloride (GdCl₃). In addition, GdCl₃ decreased by 50% the changes induced by T₃ in hepatic GSH content and TBARS formation. It is concluded that hyperthyroidism enhances Kupffer cell function, correlated with the increased number of liver macrophages observed histologically, which may represent an alternate source of reactive O₂ species to that induced in parenchymal cells, thus contributing to the enhanced oxidative stress status developed.     

Effect of clomiphene on Ca movement in human prostate cancer cells

The effect of clomiphene, an ovulation-inducing agent, on cytosolic free Ca²⁺ levels ([Ca²⁺]_i) in populations of PC3 human prostate cancer cells was explored by using fura-2 as a Ca²⁺ indicator. Clomiphene at concentrations between 10-50 μM increased [Ca²⁺]_i in a concentration-dependent manner. The [Ca²⁺]_i signal was biphasic with an initial rise and a slow decay. Ca²⁺ removal inhibited the Ca²⁺ signal by 41%. Adding 3 mM Ca²⁺ increased [Ca²⁺]_i in cells pretreated with clomiphene in Ca²⁺-free medium, confirming that clomiphene induced Ca²⁺ entry. In Ca²⁺-free medium, pretreatment with 50 μM brefeldin A (to permeabilize the Golgi complex), 1 μM thapsigargin (to inhibit the endoplasmic reticulum Ca²⁺ pump), and 2 μM carbonylcyanide m-chlorophenylhydrazone (to uncouple mitochondria) inhibited 25% of 50 μM clomiphene-induced store Ca²⁺ release. Conversely, pretreatment with 50 μM clomiphene in Ca²⁺-free medium abolished the [Ca²⁺]_i increase induced by brefeldin A, thapsigargin or carbonylcyanide m-chlorophenylhydrazone. The 50 μM clomiphene-induced Ca²⁺release was unaltered by inhibiting phospholipase C with 2 μM 1-(6-((17β-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122). Trypan blue exclusion assay suggested that incubation with clomiphene (50 μM) for 2-15 min induced time-dependent decrease in cell viability by 10-50%. Collectively, the results suggest that clomiphene induced [Ca²⁺]_i increases in PC3 cells by releasing store Ca²⁺ from multiple stores in an phospholipase C-independent manner, and by activating Ca²⁺ influx; and clomiphene was of mild cytotoxicity.     

Manganese complexes of curcumin and its derivatives: evaluation for the radical scavenging ability and neuroprotective activity

In this study, three manganese complexes of curcumin (Cp) and related compounds, diacetylcurcumin (AcylCp) and ethylenediamine derivative (CpED), were synthesized and evaluated in vitro for antilipid peroxidation and superoxide dismutase activity. The manganese complexes exhibited a great capacity to protect brain lipids against peroxidation with IC₅₀ of 6.3–26.3 μM. All manganese complexes showed much greater SOD activity than their corresponding antioxidant ligands as well as trolox with IC₅₀ values of 8.9–29.9 μM. AcylCp and curcumin manganese complexes (AcylCpCpx and CpCpx) also gave the highest inhibitory activity to H₂O₂-induced cell damage (oxidative stress) at 0.1 μg/ml (< 0.2 μM) in NG108-15 cells, which were more potent than curcumin and related compounds. The neuropharmacological tests in mice supported the idea that the SOD mimicking complexes were able to penetrate to the brain as well as their role in the modulation of brain neurotransmitters under the aberrant conditions. The complexes significantly improved the learning and memory impairment induced by transient ischemic/reperfusion. AcylCpCpx, CpCpx, and CpEDCpx showed significant protection at 6.25, 25, and 50 mg/kg (i.p.), respectively, whereas manganese acetate and curcumin had no effect at doses of 50 mg/kg. In addition, treatment of AcylCpCpx and curcumin significantly attenuated MPTP-induced striatal dopamine depletion in mice, which was in accordance with the increase in the density of dopaminergic neurons when compared with MPTP-treated mice. These results support the important role of manganese in importing SOD activity and consequently, the enhancement of radical scavenging activity. AcylCpCpx and CpCpx seem to be the most promising neuroprotective agents for vascular dementia.     

Regulation of ganglioside and sialoglycoprotein biosynthesis—Effect of drugs affecting membrane flow

The labelling of gangliosides and sialoglycoproteins (SGP) after injecting intraocularly 30 μCi of [³H]ManNAc was studied in chickens that had previously received, also intraocularly, 30 or 70 nmol of monensin in 10 μl of ethanol or 24 μg of colchicine in 10 μl saline. Controls received ethanol or saline only. Colchicine at doses that almost completely block the axonal transport to the optic tectum does not affect the labelling of those sialocompounds. The translocation of gangliosides and SGP, measured as a percentage of accessibility to the sialidase was inhibited by colchicine, but this effect was different for gangliosides and for SGP. In contrast, monensin, also at doses that block axonal transport, inhibits the synthesis and the translocation of gangliosides and SGP. It also modifies the labelling pattern of retinal gangliosides, which with respect to the normal pattern shows an increase in the relative labelling of GM₃, GD₃, GM₁ and probably GD_1b and a decrease in GD_1a and GT_1b. The activity of UDP-GalNAc: GM₃, GalNAc transferase (EC 2.4.1.79) of retina membranes of chickens that received an intraocular injection of monensin 4 h before being sacrificed showed a dose-dependent decrease with respect to the controls. In experiments in vitro monensin at a concentration of 3 mM failed to inhibit this activity.     

Hemoglobin and iron-evoked oxidative stress in the brain: protection by bile pigments, manganese and S-nitrosoglutathione

In the present in vitro and in vivo study we investigated the pro-oxidant effects of hemoglobin, as well as the antioxidant effects of its metabolites, in the brain. Incubation of rat brain homogenates with hemoglobin (0-10 μM) but not hemin induced lipid peroxidation up to 24 h (EC₅₀ = 1.2 μM). Hemoglobin's effects were similar to ferrous ion (EC₅₀ = 1.7 μM) and were blocked by the chelating agent deferoxamine (IC₅₀ = 0.5 μM) and a nitric oxide-releasing compound S-nitrosoglutathione (IC₅₀ = 40 μM). However, metabolites of hemoglobin — biliverdin and bilirubin — inhibited brain lipid peroxidation induced by cell disruption and hemoglobin (biliverdin IC₅₀ = 12-30 and bilirubin IC₅₀ = 75-170 μM). Biliverdin's antioxidative effects in spontaneous and iron-evoked lipid peroxidation were further augmented by maganese (2 μM) since manganese is an antioxidative transition metal and conjugates with bile pigments. Intrastriatal infusion of hemoglobin (0-24 nmol) produced slight, but significant 20-22% decreases in striatal dopamine levels. Whereas, intrastriatal infusion of ferrous citrate (0-24 nmol) dose-dependently induced a greater 66% depletion of striatal dopamine which was preceded by an acute increase of lipid peroxidation. In conclusion, contrary to the in vitro results hemoglobin is far less neurotoxic than ferrous ions in the brain. It is speculated that hemoglobin may be partially detoxified by heme oxygenase and biliverdin reductase to its antioxidative metabolites in the brain. However, in head trauma and stroke, massive bleeding could significantly produce iron-mediated oxidative stress and neurodegeneration which could be minimized by endogenous antioxidants such as biliverdin, bilirubin, manganese and S-nitrosoglutathione.     

Metabolic effects of thyroid hormones at a low temperature in the snake

1. 1.|The metabolic role of the thyroid gland was studied in intact snakes, Naja naja and Ptyas korros treated with tri-iodo-thyronine (T₃) and thyroxine (T₄) and in thyroidectomized (Tx) N. naja kept at 21°C by analyzing tissue composition and glycogen phosphorylase a activity.

2. 2.|Liver weight was unaffected by thyroid hormone injection in both species but decreases in liver glycogen followed T₃ or T₄ injection, and there was an increase in liver glycogen in N. naja. These changes in liver glycogen were accompanied by a decrease in glycogen phosphorylase a activity with T₃ injection. T₃ decreased muscle glycogen in Ptyas and Tx increased it in N. naja.

3. 3.|T₃ increased % liver lipid in Ptyas but not in Naja.

4. 4.|Between species, there were differences in liver weight, blood glucose level, cholesterol level and % muscle lipid.

5. 5.|The results showed that thyroid hormones affected carbohydrate and lipid metabolism at a low temperature of 21°C, although the significance is not known.

Poly(methylene co-guanidine) coated alginate as an encapsulation matrix for urease

Urease was encapsulated within alginate beads, coated with poly(methylene co-guanidine) membranes via polyelectrolyte complexation. Membrane thickness increased with reaction time to 53 μm after 80 min, and to 59 μm with an increase in co-guanidine concentration from 2.5 to 20 mg ml⁻¹. A 70% mass and 31% activity yield of urease resulted following encapsulation. Although co-guanidine strongly inhibited freely soluble urease (I_0.5=5.8 μg ml⁻¹ co-guanidine), immobilization stabilized the enzyme against inactivation. Encapsulated activity declined as the polycation concentration used for membrane formation increased; however an activity loss of only 35% was observed when the co-guanidine concentration was as high as 5 mg ml⁻¹. Glucose protected against inactivation, with 0.5 increasing to 28.5 μg ml⁻¹ for the freely soluble enzyme. When the beads were coated with co-guanidine in the presence of glucose, encapsulated urease activity was fully retained.